Aaina Kochhar, MD¹, J.Paige Souder, MD,PhD¹,Emily Muth, MS, CGC¹,Samantha DeMarsh, DO², Molly Hemenway, PNP²  , Nicholas K. Foreman, MD^2.

¹Children’s Hospital Colorado, Department of Pediatrics, Division of Genetics and Metabolism.

²Childrens Hospital Colorado, Department of Pediatrics, Center for Cancer and Blood Disorders.

Title – Congenital bilateral plexiform neurofibromas of the cavernous sinuses presenting as buphthalmos at birth.

Purpose – We describe a term newborn presenting with congenital glaucoma, buphthalmos and poor vision, who was found to have bilateral cavernous sinus plexiform neurofibromas extending through the skull base foramina into the left, greater than right, orbits and face. Treatment with a MEK inhibitor was initiated given progressive vision impairment. 

Method – A full term newborn female infant presented with left-sided buphthalmos and congenital glaucoma. Brain MRI showed bilateral enhancing cavernous sinus masses, extending along both trigeminal nerves, and nodular enhancement of right oculomotor nerve. These lesions were initially suspected to be atypical intracranial hemangiomas or plexiform neurofibromas. The patient had multiple cafe au lait macules on exam, but did not meet clinical diagnostic criterion for Neurofibromatosis type 1 (NF1) in the newborn period.  She received medical treatment for glaucoma. Genetic testing and follow up imaging were done. 

Results - Whole exome sequencing showed a heterozygous denovo pathogenic variant in the NF1 gene c.1756-1759del, p.(Thr586ValfsTer18). Follow up MRI brain, orbits and face at 3 months of age, showed increasing size of bilateral plexiform neurofibromas expanding the cavernous sinuses and extending through skull base foramina into left greater than right orbit and face. Given the growth in size of the masses, impact on vision and potential other cranial nerve functions, therapy with the MEK inhibitor, Trametinib, was recommended.

Conclusion – We describe aggressive bilateral cavernous sinus plexiform neurofibromas with intraorbital extension and vision loss. It is an unusual clinical manifestation of NF1 and we offered treatment with a MEK inhibitor in this scenario, to stabilize/prevent further growth of the plexiform neurofibroma. 

References

1.Congenital bilateral plexiform neurofibromas of the cavernous sinuses, Pediatr Radiol (2003) 33: 272–274.

2.Congenital ectropion uvea and mechanisms of glaucoma in neurofibromatosis type 1: new insights, Ophthalmology 2012 Jul;119(7):1485-94. 

Characteristics, Cycle of First Onset, and Time to Resolution of Commonly Reported (≥15%) Treatment-Related Adverse Events (TRAEs) in the ReNeu Trial of Mirdametinib in Children and Adults With Neurofibromatosis Type 1 (NF1)-Associated Plexiform Neurofibroma (PN)

Presenting Author: Laura J. Klesse, MD, PhD

University of Texas Southwestern, Dallas, TX

Laura J. Klesse, MD, PhD¹; Christopher L. Moertel, MD²; Prakash Ambady, MD³; Stefania Maraka, MD⁴; Phioanh (Leia) Nghiemphu, MD⁵; Zsila Sadighi, MD⁶; David Schiff, MD⁷; Molly Nickerson, PhD⁸; Michael D. Weber, PharmD⁸; Armend Lokku, PhD⁸; Stacie Stapleton, MD⁹; Tobias Walbert, MD, PhD¹⁰; Julia Meade, MD¹¹; Reuben Antony, MD¹²; Kathryn Nevel, MD¹³; Timothy R. Gershon, MD, PhD¹⁴; Dusica Babovic-Vuksanovic, MD¹⁵

¹University of Texas Southwestern, Dallas, TX; 

²University of Minnesota, Minneapolis, MN; 

³Oregon Health & Science University (OHSU), Portland, OR;

⁴University of Illinois at Chicago, Chicago, IL;

⁵University of California, Los Angeles (UCLA), Los Angeles, CA; 

⁶MD Anderson Cancer Center, Houston, TX; 

⁷University of Virginia - Emily Couric Cancer Center, Charlottesville, VA;

⁸SpringWorks Therapeutics Inc., Stamford, CT; 

⁹Johns Hopkins All Children's Hospital, St. Petersburg, FL;

¹⁰Henry Ford Health, Wayne State University and Michigan State University, Detroit, MI; ¹¹University of Pittsburgh School of Medicine and UPMC Children's Hospital of Pittsburgh, Pittsburgh, PA;

¹²University of California - Davis, Sacramento, CA; 

¹³Indiana University School of Medicine and IU Health, Indianapolis, IN;

¹⁴Children’s Center for Neurosciences Research, Emory University School of Medicine, Atlanta, GA; 

¹⁵Mayo Clinic - Rochester, Rochester, MN. 

Supported by: SpringWorks Therapeutics, Inc.

DISCLOSURES

LJK: Uncompensated relationships with Alexion Pharmaceuticals;

CLM: Employment with OX2 Therapeutics; leadership role with OX2 Therapeutics; equity interest in OX2 Therapeutics; consultancy/advisory role with Alexion Pharmaceuticals and SpringWorks Therapeutics; patents, royalties, or other intellectual property with OX2 Therapeutics; travel expenses from Alexion Pharmaceuticals;

PA: Honoraria with SpringWorks Therapeutics, SERVIER; Consulting or Advisory Role with  SpringWorks Therapeutics, SERVIER; Patents, Royalties, Other Intellectual Property for Morpholino oligonucleotides useful in cancer treatment; Patent number: 11679121; Abstract: Disclosed are morpholino oligonucleotides that can be used to silence expression of MGMT, pharmaceutical compositions that include said morpholino oligonucleotides, and methods of using said morpholino oligonucleotides in the treatment of cancer, particularly methods that involve the use of radiation to deliver said morpholino oligonucleotides. Type: Grant Filed: December 7, 2020; Date of Patent: June 20, 2023;

SM: Consulting or advisory role with SpringWorks Therapeutics;

PN: Honoraria with Alexion Pharmaceuticals; Consulting or Advisory Role with SpringWorks Therapeutics; Research Funding with Chimerix (Inst), Recursion Pharmaceuticals (Inst), NCCN (Inst), SpringWorks Therapeutics (Inst), Millennium (Inst), Erasca, Inc (Inst), Global Coalition for Adaptive Research (Inst), Children's Tumor Foundation (Inst);

ZS: Employment with Adjuvant Behavioral Health; Leadership with Adjuvant Behavioral Health; Speakers' Bureau with Alexion Pharmaceuticals;

DS: Consultancy/advisory role with Orbus Therapeutics, PRA, Anheart Therapeutics, Servier (I), SymBio Pharmaceuticals; royalties for submissions to UpToDate;

MN, MDW, AL: employment with SpringWorks Therapeutics, Inc; equity interest in SpringWorks Therapeutics, Inc;

SS: No relevant disclosures;

TW: Consulting or Advisory Role with Novocure, Alexion Pharmaceuticals, SpringWorks Therapeutics, SERVIER, Anheart Therapeutics; Travel, Accommodations, Expenses with SERVIER;

JM: Consultancy/advisory role with Alexion Pharmaceuticals; travel expenses from Alexion Pharmaceuticals;

RA: Consultancy/advisory role with Alexion Pharmaceuticals; honoraria from Alexion Pharmaceuticals;

KN: Consultancy/advisory role with Servier Pharmaceuticals, royalties for editorial role with Oxford University Press;

TRG: Research funding from SpringWorks Therapeutics, Inc;

DBV: Honoraria from Alexion Pharmaceuticals and SpringWorks Therapeutics, Inc; consultancy/advisory role with AstraZeneca; research funding from SpringWorks Therapeutics, Inc; employment with Mayo Clinic; consultancy/advisory role with Alexion Pharmaceuticals; research funding from Alexion Pharmaceuticals, Recursion, and SpringWorks Therapeutics, Inc.

ABSTRACT 

Background: Mirdametinib is the first FDA-approved MEK1/2 inhibitor for both adults and children (≥2y) with symptomatic NF1-PN not amenable to complete resection. This analysis evaluated cycle of first onset and time to resolution of commonly reported TRAEs in the pivotal ReNeu trial, where patient- and parent proxy-reported outcome improvements and PN volume reductions were demonstrated.

Methods: Safety results are reported as of the data cutoff (September 20, 2023). Median cycle (1-cycle=28 days) of first onset and time to resolution (days) for common TRAEs (≥15%) were evaluated for prepubescents (2 to <12y, n=32), postpubescents (12 to <18y, n=24), young adults (18 to ≤50y, n=48), and older adults (>50y, n=10). Time to resolution was calculated from the start of the first event to the end of the last event. The protocol provided strategies for AE supportive care, but no AE prophylaxis was mandated at treatment initiation.

Results: The most frequent cycle of onset for TRAEs was Cycle 1 (81%). Common TRAEs were mostly grades 1 to 2 (83%, Table 1) and did not occur with greater frequency or severity over time. Median cycle of onset (Table 2) and time to resolution (Figure) for common TRAEs are reported. In prepubescents, median cycle of onset was Cycle 1 for vomiting; Cycle 3 for dermatitis acneiform; and Cycle 5 for paronychia, diarrhea, and nausea. For postpubescents, median cycle of onset was Cycle 1 for dermatitis acneiform, diarrhea, nausea, and vomiting; and Cycle 4 for paronychia. In young adults, median cycle of onset was Cycle 1 for dermatitis acneiform, diarrhea, and nausea; and Cycle 2 for vomiting. For older adults, median cycle of onset was Cycle 1 for dermatitis acneiform, diarrhea, nausea, and vomiting. Time to resolution was variable between TRAEs and among patients in all age groups. Eighteen patients had a TRAE of asymptomatic ejection fraction (EF) decrease, median (range) time to onset was 127 (41, 1361) days, and all except one prepubescent patient experienced resolution. No patients experienced a symptomatic EF decrease.

Conclusions: In ReNeu, symptomatic improvements and PN volume reductions were observed at the earliest measured timepoint (Cycles 3 and 5, respectively) and were sustained over time. In all age groups, common TRAEs were detected earlier than treatment benefits and were generally resoluble. It is important to manage patients’ expectations and proactively manage TRAEs to improve treatment experience and adherence.

Figures/Tables: 

Table 1. TRAEs (Incidence of ≥15%) in the ReNeu Trial

TRAEs, treatment-related adverse events.

TRAEs have been reported in order of decreasing incidence.

Table 2. Median (Range) Cycle of TRAE (Incidence of ≥15%) First Onset in the ReNeu Trial

TRAEs, treatment-related adverse events.

TRAEs have been reported in order of decreasing incidence.

Figure. Median (Range) Time to TRAE (Incidence of ≥15%) Resolution in the ReNeu Trial

NE, not evaluable; TRAEs, treatment-related adverse events.

^aIndicates no event of paronychia for older adults 

Time to resolution was calculated as the start date of the first event of the preferred term to the end date of the last event of the preferred term. If the last event of the preferred term was ongoing, then the event was considered unresolved and was not

included in the summary statistics.

Yuichi Yoshida, MD

Division of Dermatology, Department of Medicine of Sensory and Motor Organs, Faculty of Medicine, Tottori University

Purpose: This retrospective study aimed to assess the prevalence and distribution of superficial and deep-seated plexiform neurofibromas (PNs) in patients with neurofibromatosis 1 (NF1) using whole-body magnetic resonance imaging (WB-MRI).

Methods: We conducted a retrospective analysis of NF1 patients who underwent WB-MRI in 2024 and 2025 at the Division of Dermatology, Tottori University Hospital, as part of the 12-member NF1 Japan Clinical Network.

Results: A total of 35 patients (22 females and 13 males) who met the 2021 diagnostic criteria underwent WB-MRI, regardless of the presence or absence of clinically recognized PNs. The median age at the time of WB-MRI was 37.5 years (range: 9-76 years). A total of 42 PNs larger than 4 cm were identified, with an average of 1.2 tumors per patient. Among the 35 patients, 22 (62.9%) had at least one superficial or deep-seated PN (Figure 1): 15 patients (42.9%) had superficial PNs, 12 patients (34.3%) had deep-seated PNs, and 2 patients (5.7%) exhibited a superficial PN contiguous with deep-seated PNs. Among the 19 superficial PNs, 8 (42.1%) were located in the limbs, 6 in the trunk, and 5 in the head and neck. In contrast, among the 21 deep-seated PNs, 8 (38.1%) were found in peripheral nerves of the limbs, 7 in deep anatomical structures (paraspinal regions, mediastinum, and retroperitoneum), 4 in the trunk, and 2 in the head and neck.

Conclusion: In this study, 62.9% of NF1 patients had either superficial or deep-seated PNs, and 11 of 35 patients (31.4%) exhibited multiple PNs in various anatomical locations. These findings underscore the utility of WB-MRI in the comprehensive detection and characterization of both superficial and deep-seated PNs in patients with NF1.

Figure 1. Whole-body magnetic resonance imaging revealed multiple superficial plexiform neurofibromas located in the right shoulder and left lower leg, as well as a deep-seated plexiform neurofibroma in the popliteal fossa.

Full List of Authors: Yuichi Yoshida, MD, Yuko Ehara, MD

Funding: This work was supported by Health and Labour Sciences Research Grants from the Ministry of Health, Labour and Welfare of Japan (23FC1037).

Disclosure: YY received consultancy fees and honoraria for lectures from Alexion Pharmaceuticals, Inc.

{{Plexiform neurofibroma}}

Authors: Belinda B. Garana, Ph.D.¹; Simge Acar, M.D.²; Gorkem Oztosun, M.D.²; Dana C. Borcherding, Ph.D.²; Jiawan Wang, Ph.D.³; Chelsea Hutchinson-Bunch, M.S.¹; Marina A. Gritsenko, M.S.¹; Paul D. Piehowski, Ph.D.¹; Christine A. Pratilas, M.D.³; Angela C. Hirbe, M.D., Ph.D.^2,5; Sara J.C. Gosline, Ph.D.^1,4,5

¹Pacific Northwest National Laboratory, Richland, Washington, USA; ²Washington University of St. Louis, St. Louis, Missouri, USA; ³Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; ⁴Oregon Health and Sciences University, Portland, Oregon, USA; ⁵Co-corresponding authors

Funding: United States Department of Defense Office of the Congressionally Directed Medical Research Programs (CDMRP) Neurofibromatosis Research Program (NFRP)

Disclosures: None.

Title: Multi-omics integration of malignant peripheral nerve sheath tumors identifies potential targets for Chr8q-amplified clones

Purpose: Chromosome 8q (Chr8q) amplification in malignant peripheral nerve sheath tumors (MPNST) is associated with high-grade transformation in MPNST, which has a five-year survival rate of only 20-50%. Despite Chr8q gain being a common event in MPNST, there is not currently a treatment approach that allows for personalization based on copy number status. In this work, we employed a proteogenomic approach to interrogate the molecular differences between Chr8q-wildtype and -amplified tumors to both better understand tumor progression and identify drugs which may target vulnerabilities unique to each set of tumors.

Methods: To do so, we leveraged our growing library of fully characterized MPNST patient-derived xenografts (PDXs) and collected LC-MS/MS global and phospho-proteomics measurements for six of these samples. We then paired these data with transcriptomics and copy number data to identify specific molecular changes that corresponded with Chr8q gain across the samples. By integrating these omics measurements, we identified specific pathways that were distinctly active in either Chr8q-amplifed or wild type samples as well as potential drugs that could be uniquely sensitive in each set of tumors. 

Results: In addition to changes caused by the genes on Chr8q such as over-expression of MYC, our results identified dramatic changes in transcriptional regulation and protein signaling upon Chr8q amplification that predict differences in drug sensitivity across Chr8q status. CDK4 and PLK1 were among potential targets identified for Chr8q-amplified MPNSTs whereas MEK and EGFR were predicted to be vulnerabilities in Chr8q-wildtype MPNSTs. 

Conclusion: This study paves the way for future studies of MPNST treatments stratified by Chr8q status. Furthermore, these results represent a new way to target tumor heterogeneity in MPNST through the proteogenomic integration and drug sensitivity prediction in distinct tumor subpopulations.

{{BIOINFORMATICS AND COMPUTATIONAL OMICS}}

Senmin Chen MD.¹, Xufeng Luo MD.², Zhihao Xing, Ph D.³, Huanli Xu MD.¹, Chao Liu MD.¹, Xiuli Yuan MD.¹

1. Hematology and Oncology Department, Shenzhen Children’s Hospital

2. Neurology Department, Shenzhen Children’s Hospital

3. Biobank, Shenzhen Children's Hospital

Background: Neurofibromatosis Type 1 (NF1) is an autosomal dominant hereditary disease caused by NF1 gene mutations. The coexistence of NF1 and other genetic disorders in the same person are rare.

Case Presentation: We present a case of NF1 with Becker muscular dystrophy(BMD) in an 8-year-old boy. He had history of multiple café au lait spots since birth and a painless lump at the left chest for over 1 year. The child's past growth and development were similar to those of children of the same age. The patient's brother and parents have normal phenotypes. On physical examination, we found six café au lait spots with a diameter greater than 1.5cm and auxillary freckling. The maximum diameter of the café au lait spot is about 12cm, located on the left chest wall. Hair growth can be seen on the local skin, and a granular mass can be palpated below the skin. 

Peripheral blood samples were collected from the child and his family members. Genomic DNA was extracted and whole exome sequencing was performed using high-throughput sequencing, followed by Sanger sequencing validation. A mutation was found in the NF1 gene and DMD gene of the patient, with NF1 c.6855C>A, which was a de novo mutation (Figure 1); DMD EX51-EX52 Del, which originated from the mother (Figure 2). Both of the above variants had been identified as pathogenic variants.

Since the child has no abnormal neurological manifestations such as delayed major motor development or muscle weakness, and no family history of Duchenne muscular dystrophy (DMD), peripheral blood myocardial enzyme spectrum testing was performed, and the serum creatine kinase (CK) concentration was found to be 572IU/L (reference value 22-200IU/L), and creatine kinase MB (CK-MB) was 29IU/L (reference value 0-20IU/L). Combining clinical manifestations and genetic testing results, Becker muscular dystrophy (BMD) can be diagnosed. 

Conclusion: The association of NF1 and Becker phenotype MD is quite rare. Although both conditions may lead to neurological damage, no significant neurological impairment was observed in this patient, suggesting that the two genes may function through different pathways.

Figure1 NF1 gene of the proband and parents

Figure2 DMD gene mutations of the proband (left) and his mother (right) 

{{Other}}

Authors: Liyan Yu, PhD., Florida State University, USA; Dan Liu, PhD., Florida State University, USA; Jonathan M. Payne, Psy.D., Murdoch Children's Research Institute, Australia; Heather L. Thompson, PhD., California State University, USA; Xian Wu, PhD., University of Kentucky, USA; Julia Moreira, BS., Florida State University, USA; Lauren Morey, BS., Florida State University, USA; Yang Hou, PhD., Florida State University, USA.

Funding: This project was funded by (a) the Department of Defense, Congressionally Directed Medical Research Programs, Neurofibromatosis Research Program (W81XWH2110504); (b) the Florida State University Faculty Startup Funding; and (c) the University of Kentucky Faculty Startup Funding.

Disclosures: The authors declare no financial conflict of interest.

Purpose: Previous studies have yielded inconsistent findings regarding the extent of oral language delays among individuals with Neurofibromatosis type 1 (NF1) compared with the general population. This study aims to provide a robust estimate of mean differences in oral language skills between NF1 and control groups and examine potential moderators contributing to variability in group differences.

Methods: Comprehensive literature searches were conducted across Scopus, PsycINFO, Web of Science, PubMed, and ProQuest using NF1-related and neurobehavioral function-related terms (e.g., language) on March 26, 2024. Studies assessing oral language skills in children/adolescents with NF1 and including control groups or standardized scores were analyzed. Hedges’ g was calculated to quantify group difference. The robust standard error estimation and random-effects model were used to estimate the mean effect size. Meta-regression and subgroup analyses were conducted to explore potential moderators of group differences.

Results: Forty-three studies involving 2,360 children/adolescents with NF1, ranging from 0.53 to 15.5, met the inclusion criteria. We found that children/adolescents with (versus without) NF1 performed significantly worse in overall oral language skills (n = 43, k = 122, g = -0.65, 95% CI [-0.77, -0.53], p < .001, I² = 82.01%) and in subskills, including core language skills (n = 16, k = 20, g = -0.78, 95% CI [-0.98, -0.59], p < .001), expressive language (n = 23, k = 35, g = -0.67, 95% CI [-0.83, -0.51], p < .001), receptive language (n = 21, k = 28, g = -0.56, 95% CI [-0.74, -0.37], p < .001), and phonological awareness (n = 11, k = 20, g = -0.72, 95% CI [-0.93, -0.52], p < .001), but not in pragmatic language (n = 4, k = 17, g = -0.45, 95% CI [-1.09, 0.19], p = .109). Significant publication bias was found in receptive language, and group differences remained significant after adjusting for publication bias. Overall group differences were not significantly moderated by age, sex composition, NF1 inheritance mode, ADHD diagnosis, learning disability diagnosis, intelligence, working memory, measure type, comparison group type, sampling resources, or measurement instruments. 

Conclusions: Children/adolescents with NF1 exhibit significant deficits in oral language skills, particularly in core language, expressive language, receptive language, and phonological awareness, highlighting the need for targeted interventions. Substantial between-study heterogeneity in effect sizes highlights the importance to further investigate factors contributing to group differences using individual-level data and large sample size.

Cognition / Behavioral / Learning

Authors: Xiaojie Hu, MD¹, Xiaoxi Lin, MD¹, Kang Zeng, MD², Wenbin Li, MD³, Yanling Li, MD⁴, Juan Tao, MD⁵, Zhongyuan Xu, MD², Zhuang Kang, MD³, Yaohui Ma, MD⁴, Liu Yang, MD⁵, Zhuli Wu, MD⁶, Xingli Wang, MD⁶, Hongmei Lin, MD⁷, Yu Liu, M.S.⁷, Wen Zhong, M.S.⁷

Affiliations:

¹Shanghai Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China

² Nanfang Hospital, Southern Medical University, Department of Dermatology, Guangzhou, China

³ Beijing Tiantan Hospital, Capital Medical University, Beijing, China

⁴ The Second Hospital of Hebei Medical University, Shijiazhuang, China

⁵ Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China

⁶ Shanghai Fosun Pharmaceutical Industrial Development Co., Ltd., Shanghai, China

⁷ Beijing Fosun Pharmaceut ical Research and Development Co., Ltd., Beijing, China

Funding: This research was supported by Shanghai Fosun Pharmaceutical Industrial Development Co., Ltd.

Disclosures: All authors declare no relevant financial relationships.

Title: Luvometinib (FCN-159) in Adult NF1 Patients: Efficacy and Safety Outcomes from a Multi-center, Open-label, Single-arm Phase II Trial

Purpose: Neurofibromatosis type 1 (NF1) is a genetic disorder caused by autosomal dominant mutations with constitutive MAPK pathway activation. Plexiform neurofibromas (PN) occur commonly in NF1 patients and are associated with severe morbidity and risk of malignant transformation. Luvometinib, a novel MEK1/2 inhibitor with high selectivity was evaluated in the Phase II trial. 

Methods: This multi-center, open-label, single-arm Phase II trial enrolled patients with NF1-related PN deemed inoperable, incompletely resected, or recurrent post-surgery. In the adult cohort, patients received continuous 8 mg luvometinib once daily (RP2D from Phase I) in 28-day cycles. Primary endpoint was investigator-assessed and confirmed objective response rate (ORR) per REiNS criteria. Key secondary endpoints included clinical benefit rate (CBR), disease control rate (DCR), progression-free survival (PFS), duration of response (DOR), and safety. Here we present the efficacy and safety results in the adult cohort.

Results: As of September 23 2024, 63 adult patients were enrolled and treated with luvometinib. With a median follow-up of 29.3 months, the investigator-assessed and confirmed overall response rate (ORR) reached 42.9% (27/63, 95% CI: 30.5-56.0), all partial responses. The CBR was 84.1% (53/63, 95% CI: 72.7-92.1) and the DCR was 95.2% (60/63, 95% CI: 86.7-99.0). The median PFS was 31.4 months (95% CI: 31.38-NE), with 1-year and 2-year PFS rates of 91.0% and 75.2%, respectively. The median DOR was 25.8 months (95% CI: 20.27-NE), with 1-year and 2-year DOR of 95.8 % and 69.6% respectively. Pain reduction was notable: 78.3% (18/23) patients with baseline tumor pain (NRS≥2) achieved ≥2 points decrease. Regarding safety, treatment-related adverse events (TRAEs) were observed in 63 patients, with majority were grade 1-2. Grade ≥3 TRAEs occurred in 24 (38.1%) patients, with folliculitis (25.4%), paronychia (6.3%), elevated creatine phosphokinase (3.2%), and hypertension (3.2%) occurred in more than 1 patient. Serious TRAEs occurred in 2 (3.2%) patients. TRAEs-induced dose interruption and discontinuation occurred in 41 (65.1%) and 9 (14.3%) patients, respectively. No TRAEs led to dose reduction or death.

Conclusion: Luvometinib demonstrates notable clinical activity and a favorable tolerability profile in adults with NF1-related PN. ClinicalTrials.gov: NCT04954001.

{{Plexiform neurofibroma}}

Yoshihiro Nishida, MD, PhD¹, Akiyo Kitajima², Tomonori Ishii²

¹Department of Rehabilitation, Nagoya University Hospital, Nagoya, Japan

²Alexion Pharma GK, Tokyo, Japan

Funding

This study was sponsored by Alexion Pharma GK.

Disclosures

Yoshihiro Nishida has received honoraria from Alexion Pharma GK and has advisory roles in AstraZeneca and Alexion Pharma GK. Akiyo Kitajima and Tomonori Ishii are full-time employees of Alexion Pharma GK.

Title

Characteristics of patients with neurofibromatosis type 1 and plexiform neurofibromas in Japan and real-world safety and effectiveness of selumetinib: A 1-year interim analysis of an all-case, postmarketing surveillance study

Purpose: This postmarketing surveillance study aimed to assess the characteristics of patients with neurofibromatosis type 1 (NF1) and plexiform neurofibromas (PN) treated with selumetinib and the real-world safety and effectiveness of selumetinib in Japan.

Methods: All patients who were prescribed selumetinib on or after the drug launch date in Japan (November 16, 2022) were enrolled in this study. The observation period was set to 3 years from drug initiation, and patient data were captured using case report forms (data cutoff for 1-year analysis: October 9, 2024). Safety was assessed through the incidence of adverse drug reactions (ADRs), and the effectiveness (overall patient evaluation of PN and target PN size on imaging) was assessed based on physician’s global assessment.

Results: In total, 52 patients were included in the analysis (median [range] age, 13.0 [5–20] years; age <19 years, 49 [94.2%] patients; female, 28 [53.8%] patients; dermatological, neurological, and bone manifestations [DNB: severity classification created by the Neurocutaneous Syndrome Research Group in Japan, wherein stage 5 is the most severe] classification stage 5, 30 [57.7%]; median disease duration [n=49], 118.0 months). Target PN lesions were most commonly observed in the head and neck (30 [57.7%] patients), followed by the trunk (20 [38.5%] patients). The median (range) treatment duration was 52.1 (21.0–65.1) weeks, and 46 (88.5%) patients continued selumetinib for 1 year. ADRs and serious ADRs were observed in 46 (88.5%) and 9 (17.3%) patients, respectively. The most common ADR was diarrhoea (15 [28.8%] patients), followed by dermatitis acneiform (14 [26.9%] patients). No fatal ADRs were reported. At 1 year, 30 (61.2%) of 49 evaluable patients showed improvement in PN, and 16 (41.0%) of 39 evaluable patients showed a decrease in the target PN size on imaging.

Conclusion: The safety and effectiveness profile of selumetinib was generally consistent with the findings of the phase 2 SPRINT and Japanese phase 1 trials. To date, no new safety concerns have been identified. We continue to assess the safety and effectiveness of selumetinib in patients with NF1 and PN in Japan.

{{Plexiform neurofibroma}}

Background: Plexiform neurofibromas (PN) present a significant clinical challenge, with a notable unmet need for effective and well-tolerated therapies, particularly for the adult population.

Methods: A single-arm, open-label, Phase IIa trial of tunlametinib was conducted to assess its efficacy and safety in adults with inoperable, radiologically measurable PN. Patients received tunlametinib at a dose of 9 mg twice daily, following a continuous 21-day cycle regimen. The primary endpoint was the confirmed objective response rate (ORR), and secondary endpoints included disease control rate (DCR), duration of response (DoR), progression-free survival (PFS), and improvements in patient-reported outcomes (PROs) including quality of life, pain, physical function, and other PN-related symptoms compared to baseline.

Results: Of 15 adults (10 males, 5 females; median age, 27 years, range 18–45), 8 patients (53.3%, 95% CI, 26.69–78.73) achieved a confirmed partial response (PR), with a median neurofibroma volume reduction of -23.5% (range 1.6% to -49.1%). The median time to initial response was 19.6 months (range, 2.7 to 31.1 months), and the median time to best response was 22.4 months (range, 11.2 to 31.7 months). Significant improvements in multiple PRO domains were observed. All patients experienced at least one treatment-related adverse event (AE), the majority of which were grade 1 or 2. Notably, a marked age-related difference in treatment response and tumor heterogeneity were observed.

Conclusion: Tunlametinib demonstrated promising efficacy and an acceptable safety profile in adults with PN. The majority of patients experienced sustained tumor shrinkage and clinically meaningful improvements. The manageable toxicity profile, along with the absence of cumulative toxic effects, supports its potential for long-term treatment. Further exploratory analysis revealed the heterogeneity of responses, emphasizing the need for personalized therapeutic approaches.

{{Plexiform neurofibroma}}

Xuan Yu^1,2#, Yihui Gu^1,2#, Jun Liu^1,2#, Qingfeng Li^1,2*, Zhichao Wang^1,2*

1. Department of Plastic and Reconstructive Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China

2. Neurofibromatosis Type 1 Center and Laboratory for Neurofibromatosis Type 1 Research, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, China

#Contributed equally

*Corresponding author

Abstract

Neurofibromatosis Type 1 (NF1) is a genetic disorder resulting from mutations in the NF1 gene, which increases susceptibility to various nervous system tumors, including plexiform neurofibromas, malignant peripheral nerve sheath tumors, and optic pathway gliomas. Recent research has shown that these tumors are intricately connected to the complex, dynamic interactions within neurons, culminating in neuronal signaling that fosters tumor growth. These interactions offer crucial insights into the molecular mechanisms underpinning tumor development, as well as broader implications for therapeutic strategies. This review summarizes the mechanisms through which mutations in the NF1 gene within neural tissues trigger tumorigenesis, while examining the role of the neuron—via factors such as visual experience, neurotransmitter, tumor microenvironment, and psychological influences—in both promoting tumor progression and being affected by the tumors themselves. By investigating the dynamic relationship between NF1-associated nervous system tumor cells and neurons, we aim to shed light on novel biological pathways and disease processes, emphasizing the potential of interdisciplinary approaches that combine neurobiology, oncology, and pharmacology to enhance treatment strategies and even inhibit the tumorigenesis.

{{THERAPEUTICS}}

Yuehua Li ^a,b , Jun Liu ^a,b , Jingxuan Huang ^a,b , Qingfeng Li ^a,b, Zhichao Wang ^a,b,

Plexiform neuroffbromas (PNFs) are a prevalent and severe phenotype associated with NF1, characterized by a high teratogenic rate and potential for malignant transformation. The growth and recurrence of PNFs are attributed to aberrant proliferation and migration of Nf1-deffcient Schwann cells. Protein tyrosine phosphatase receptor S (PTPRS) is believed to modulate cell migration and invasion by inhibiting the EMT process in NF1-derived malignant peripheral nerve sheath tumors. Nevertheless, the speciffc role of PTPRS in NF1-derived PNFs remains to be elucidated. The study utilized the GEO database and tissue microarray to illustrate a decrease in PTPRS expression in PNF tissues, linked to tumor recurrence. Furthermore, the down- and overexpression of PTPRS in Nf1-deffcient Schwann cell lines resulted in the changes of cell migration and EMT processes. Additionally, RTK assay and WB showed that PTPRS knockdown can promote EGFR expression and phosphorylation. The restoration of EMT processes disrupted by alterations in PTPRS levels in Schwann cells can be achieved through EGFR knockdown and EGFR inhibitor. Moreover, high EGFR expression has been signiffcantly correlated with poor prognosis. These ffndings underscore the potential role of PTPRS as a tumor suppressor in the recurrence of PNF via the regulation of EGFR-mediated EMT processes, suggesting potential targets for future clinical interventions.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Libby Hasell-Williams BSc, Liyam Laraba PhD, Emanuela Ercolano PhD, C. Oliver Hanemann MD PhD, David Parkinson PhD.

Peninsula Medical School, Faculty of Health, University of Plymouth, UK.

Funding: Brain Tumour Research Charity, UK.

Disclosures: None.

Title: Analysing the role and therapeutic potential of aldehyde dehydrogenase isoforms in NF2-null schwannoma and meningioma.

Purpose: To analyse isoforms of the cancer stem cell marker aldehyde dehydrogenase in NF2-null tumours and target their activity to reduce tumour growth.

Methods: We used both in vitro analysis of meningioma and schwannoma primary human tumour cells and cell lines to study ALDH expression and activity. Following this analysis, we proceeded to test the roles for ALDH activity in schwannoma tumours in vivo using the PostnCRE-Nf2^fl/fl model and in meningioma using the orthotopic convexity meningioma model with bioluminescent imaging. 

Results: Following up on our previous work (Laraba et al 2023, Brain 146: 1697-1713. Doi: 10.1093/brain/awac342), showing strong ALDH1A1 expression in human and mouse schwannoma tissue, we have used the ALDH1A1/A3 inhibitor nifuroxazide and shown it significantly reduces proliferation of schwannoma cells in vitro and in vivo. For meningioma, we have tested for the activity of ALDH isoforms and have identified ALDH1A3 as the major isoform. Knockdown experiments in vitro show that ALDH1A3 is required for meningioma cell growth and survival and that the downstream Hippo pathway TEAD transcription factors drive ALDH1A3 expression in tumour cells. Analysis of ALDH^High and ALDH^Low meningioma cell populations show that ALDH^High tumour cells possess higher proliferative capacity and colony formation. Studies are currently underway to target meningioma growth in vitro and in vivo using additional ALDH1A1 and 1A3 inhibitors (eg DIMATE) to show proof of principle for their future clinical use.

Conclusion: We find that differing isoforms of ALDH1 are driving the proliferation of NF2-null schwannomas and meningiomas in vitro and in vivo. We believe that these proteins provide novel targets for future treatments for these two important tumour types. 

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Judit Farrés-Casas, MS

Germans Trias i Pujol Research Institute (IGTP)

Malignant peripheral nerve sheath tumors (MPNSTs) are a heterogeneous group of aggressive soft tissue sarcomas arising in both, the general population and, with a special high prevalence, neurofibromatosis type 1 (NF1) individuals. MPNST initiation is highly marked by the combined inactivation of tumor suppressor genes (TSGs) like NF1, CDKN2A, PRC2 (SUZ12 or EED) and, at a lesser frequency, TP53. In our group, we performed a comprehensive genomic characterization of NF1-associated MPNSTs and identified different groups bearing a distinct TSG inactivation signature. To better understand the molecular mechanisms driving the initiation of the different MPNSTs, we used an iPSC-derived neural crest (NC)-based NF1(-/-) model to reproduce the different TSG inactivation combinations idenitified.

We used CRISPR/Cas9 technolgy to knock out key TSG in NF1(-/-) iPSC and derived NC cells. TSG inactivation was assessed by genetic analysis and confirmed by western blot, and selected clones underwent physiological and functional characterization. Cell identity markers were analyzed and proliferation capacity and differentiation potential was tested. These cell lines were amenable to 3D culturing as spheroids. To further investigate their tumorigenic capacity, we will engraft these spheroids into nude mice and potential tumors formed will be histologically characterized by an expert pathologist.  

We will use these new NC-based cell lines to characterize the impact of different TSG losses on tumorigeneicity and cell identity in light of the existing MPNST heterogeneity. At the same time, these new cell lines may serve as a valuable preclinical resource for testing novel therapeutic strategies and comparing treatment responses in the genotypically different backgrounds.

Full author list: Judit Farrés-Casas¹, Itziar Uriarte-Arrazola¹, Miriam Magallón-Lorenz¹, Helena Mazuelas¹, Sara Ortega-Bertran², Edgar Creus-Bachiller², Juana Fernández-Rodríguez^2,3, Conxi Lázaro^2,4, Bernat Gel¹, Meritxell Carrió^1*, Eduard Serra^1*

1) Hereditary Cancer Group, Germans Trias i Pujol Research Institute (IGTP), CARE Program; Can Ruti Campus, Badalona, Barcelona, 08916; Spain

2) Hereditary Cancer Program, Catalan Institute of Oncology (ICO-IDIBELL), L’Hospitalet de Llobregat, 08098 Barcelona, Spain

3) Mouse Lab, IDIBELL, Hospitalet de Llobregat, Barcelona, Spain

4) Centro de Investigacion Biomedica en Red de Cancer (CIBERONC), Madrid, Spain

* Co-last authors

Funding: This work has been supported by the Instituto de Salud Carlos III National Health Institute funded by FEDER funds - a way to build Europe -[PI23/00422]. The work has also been supported by the Generalitat of Catalonia (2021 SGR 00967). We would like to thank the constant support of Fundación Proyecto Neurofibromatosis and the AANF and AcNefi patient associations.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Helena Mazuelas, PhD 

Hereditary Cancer Group, Germans Trias & Pujol Research Institute

The development of cutaneous neurofibromas (cNFs) constitutes one of the major concerns of persons affected by Neurofibromatosis type 1 (NF1). Complete loss of NF1 in a boundary cap-derived Schwann cells (SC) is required for cNF development. MEK inhibitors (MEKis) are being analyzed as a potential therapy for cNFs, but as for plexiform neurofibromas, despite their beneficial effect, it seems that a complete remission of these tumors will require additional therapeutic players.

We showed that the activation of the proton-sensing G protein-coupled receptor GPR68 combined with the inhibition of the Ras/MAPK pathway by the MEKi Selumetinib, induces arrest and activate myelination program of cNF-derived SCs, as well as increased cell death in cNF-derived SC cultures and human iPSC-derived NF1(-/-) neurofibromaspheres (Mazuelas et al. 2022, 2024). At least, part of this response is mimicked by replacing GPR68 activators (Ogerin, PAM71) by cAMP elevators (forskolin, 8CPT-cAMP) in combination with MEKi (Mazuelas et al. 2024). 

The aim of this study is to exploit this therapeutic strategy for targeting cNFs by identifying the best combination of cAMP elevators and MEKis in vitro and in vivo. For that, we analyzed the expression of cAMP pathway components present in cNF-derived SC cultures and identified compounds already in the clinic that elevate intracellular cAMP levels by modulating activity of these cAMP pathway proteins. We tested eight candidates in combination with Selumetinib in cNF-derived SC cultures and in the human iPSC-based neurofibromasphere model, assessing SC differentiation, viability, and apoptosis. Top compounds are currently under further analyses. 

In addition, to advance towards in vivo mouse experiments and to evaluate the specificity of these compounds in heterozygous vs null neurofibromin contexts, we are currently testing top candidate combinations in vitro in 3D neurospheres model derived from a Prss56 Nf1-KO cNF mouse model (Gresset, et al 2015). These neurospheres consists in sphere cultures of Nf1(-/-) or Nf1(+/-) Tomato-expressing stem like glial cells present in the skin and derived from Prss56-expressing boundary caps. After in vitro selection of best combinations, we aim to assess the best combo candidate in vivo in different mouse models, obtaining robust pre-clinical information for potential clinical assays.

Helena Mazuelas¹, Fanny Coulpier², Míriam Magallón-Lorenz⁸, Itziar Uriarte-Arrazola¹, Judit Farrés, Juana Fernandez-Rodriguez^3,4, Conxi Lázaro^4,5, Ignacio Blanco⁶, Isabel Bielsa⁷, Adrià Plana-Pla⁷, Bernat Gel⁸, Meritxell Carrió¹, Piotr Topilko², Eduard Serra^1,4

1) Hereditary Cancer Group, Germans Trias & Pujol Research Institute (IGTP)-CARE Program; Can Ruti Campus, Badalona, Barcelona, 08916; Spain

2) Mondor Institute for Biomedical Research, Inserm U955-Team 9, Créteil, France

3) Mouse Lab, IDIBELL, Hospitalet de Llobregat, Barcelona, Spain

4) Centro de Investigacion Biomedica en Red de Cancer (CIBERONC), Madrid, Spain

5) Hereditary Cancer Program, Catalan Institute of Oncology (ICO-IDIBELL), l'Hospitalet de Llobregat, Barcelona, 08098; Spain

6) Clinical Genomics Unit, Germans Trias i Pujol University Hospital (HUGTiP), Can Ruti Campus, Badalona, Barcelona, 08916; Spain 

7) Dermatology Department, Germans Trias i Pujol University Hospital (HUGTiP), Can Ruti Campus, Badalona, Barcelona, 08916; Spain

Funding: This work was mainly supported by a Subagreement from the Johns Hopkins University via the Neurofibromatosis Therapeutic Acceleration Program (NTAP) with funds provided by Grant Agreement from the Bloomberg Family Foundation. Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the Bloomberg Family Foundation or the Johns Hopkins University. The work has also been partially supported by the Spanish Ministry of Science and Innovation, Carlos III Health Institute (ISCIII) (PI20/00228) Plan Estatal de I + D + I 2013–2016, co-financed by the FEDER program – a way to build Europe, and the Generalitat of Catalonia (2021 SGR 00967).

References

Mazuelas, H., Magallón-Lorenz, M., Fernández-Rodríguez, J., Uriarte-Arrazola, I., Richaud-Patin, Y., Terribas, E., Villanueva, A., Castellanos, E., Blanco, I., Raya, Á,. et al. (2022) Modeling iPSC-derived human neurofibroma-like tumors in mica uncovers the heterogeneity of Schwann cells within plexiform neurofibromas. Cell Rep. 38, 110385 https://doi.org/10.1016/j.celrep.2022.110385

Mazuelas, H., Magallón-Lorenz, M., Uriarte-Arrazola, I., Negro, A., Rosas, I., Blanco, I., Castellanos, E., Lázaro, C., Gel, B., Carrió, M., & Serra, E. (2024)

Unbalancing cAMP and Ras/MAPK pathways as a therapeutic strategy for cutaneous Neurofibromas. JCI insight. 9(3):e168826

https://doi.org/10.1172/jci.insight.168826.

Gresset, A., Coulpier, F., Gerschenfeld, G., Jourdon, A., Matesic, G., Richard, L., Vallat, J.-M., Charnay, P., & Topilko, P. (2015). Boundary Caps Give Rise to Neurogenic Stem Cells and Terminal Glia in the Skin. Stem Cell Reports, 5(2), 278–290. https://doi.org/10.1016/j.stemcr.2015.06.005

{{THERAPEUTICS}}

Rosalie E Ferner¹ MD, John F Golding² DPhil, Una-Marie Sheerin¹ MD PhD, Joshua Hersheson¹ MD PhD, Karine Lascelles¹ MD, Jill Cadwgan¹ MD, Mary Thomas¹ BN, Mandy Myers¹ RGN, RSCN, Katrina Kettle¹ RN

¹ Neurofibromatosis Service, Department of Neurology, Guy’s and St. Thomas’ NHS Foundation Trust, London UK

² University of Westminster, London UK

Funding No funding for this study

Disclosures: No disclosures relevant to this study

Title Clinical presentation, risk factors and survival in 132 patients with neurofibromatosis 1 associated malignant peripheral nerve sheath tumor, 2009-2025. 

Purpose There is a 15.8% lifetime risk of developing neurofibromatosis 1 (NF1) associated malignant peripheral nerve sheath tumor (MPNST). Diagnosis and treatment are challenging, high-grade tumors metastasize widely and many have a poor outcome.

We evaluated clinical presentation, risk factors and survival in individuals with NF1 associated MPNST in a single national neurofibromatosis center.

Method We carried out a retrospective case record study of 132 NF1 patients with histologically proven MPNST, who attended our national NF1 Center between 2009-2025. We evaluated: gender, NF1 family history, age at diagnosis, presenting symptoms, MPNST location, and size. Metastatic, new or recurrent MPNST was noted. We also documented prior radiotherapy, atypical neurofibromatous neoplasm of uncertain biologic potential (ANNUBP), NF1 microdeletion, disease burden, NF1 neuropathy, other malignancy and family history of MPNST. Survival at five years and ten years post diagnosis was calculated.

Results 66 males and 66 females were assessed, age range 9-74 years, mean age 31 years, (SD 13.1) at diagnosis.  Seven patients had clinically segmental NF1; 57/127 (44.8%) had an NF1 family history; Predominant symptoms were pain 111/127 (87.4%), tumor growth 98/113 (86.7%), hard texture 52/74 (70.3%), neurological deficit 57/114 (50%). Weight loss was detected in 24/97 (24.7%), n varied due to missing / not evaluable data. Tumor size was 5-230 mm, mean 94.0 (SD 46.7). The commonest MPNST locations were lower limb 36/132, (27.3%) and abdomen/pelvis 29/132 (22%). Metastases were diagnosed in 43/132 (32.6%) and new or recurrent MPNST in 13/132 (9.8%). Radiotherapy was given prior to MPNST diagnosis in 9/121 (7.4%); ANNUBP was present in 13/132, (9.8%); NF1 microdeletion was detected in 7/74 (9.5%); a heavy internal disease burden was detected in 61/99 (61.6%); NF1 neuropathy was diagnosed on neurophysiology in 8/17 (47%) and other malignancy was found in 31/131 (23.7%) patients. A family history of MPNST was present in 11/126 (8.7%). Weight loss (p = .002), pain (p =.041) and MPNST size (p =.046) were associated with a worse outcome. Five year survival was 62% and ten year survival was 58.5%; median survival time was 20.1 years. 

Conclusion Our large, single center cohort of 132 patients with NF1-MPNST had a high frequency of known presenting manifestations and risk factors.  Weight loss, pain and tumor size were significantly associated with a worse outcome. Our current survival data show improved survival, compared with historical studies, underscoring the importance of coordinated neurofibromatosis centers and patient and clinician education.

{{Malignant peripheral nerve sheath tumors (MPNST)}}

1.Mandy Myers - BSc(Hons), RGN, RSCN, MSc., SCPHN

Paediatric Clinical Nurse Specialist for Neurofibromatosis for Complex NF1 Service

Guy’s and St Thomas’ NHS Foundation Trust.

2. Carol Irving - BSc(Hons), RN Child, Specialist Practitioner Child Health

NF1 Paediatric Clinical Nurse Specialist

Complex NF1 Service

Genomic Medicine

Manchester University NHS Foundation Trust

3. Katrina Kettle 

BSc(Hons), RN, 

Paediatric Clinical Nurse Specialist for Neurofibromatosis for Complex NF1 Service

Guy’s and St Thomas’ NHS Foundation Trust.

4. Nadira Bullock -  Patient Centred Solutions Lead, Alexion, AstraZeneca Rare Disease

5.   Judith Eelloo -  RGN, RSCN, MPhil

Lead Nurse Complex NF1 Service

Genomic Medicine

Manchester University NHS Foundation Trust

Background: Neurofibromatosis Type 1 (NF1) is an autosomal dominant genetic condition with a birth incidence of 1 in 2500 to 1 in 3000 ¹ and can lead to the development of benign tumours called plexiform neurofibromas in 20-50% of patients.^3-7 Selumetinib, an oral MEK inhibitor, has been approved for treating symptomatic and inoperable plexiform neurofibromas in children aged 3-18 in the UK.²

To support patients and carers, a comprehensive treatment guide was developed and distributed across two national NF centres in the UK. 

Method: The guide was developed by two clinical nurse specialists from both National UK NF1 centres and reviewed by both patient advocacy groups with the support and funding of Alexion, AstraZeneca Rare Disease.   The guide aims to ensure NF1 patients are supported in their treatment journey and to aid communication between healthcare providers including the National NF1 Centre, paediatric oncology shared care units (POSCUs) and local paediatric care. The guide is comprehensive but needs to be clear and include picture guidance to support children and those with learning difficulties. Coloured tabbed sections covered various aspects, such as how to take Selumetinib, daily oral and skin care, possible side effects, upcoming medical appointments and support groups. Feedback was collected after 3 months from nine different patients or carers, eight of whom were already undergoing Selumetinib treatment.  The age of the patients was not recorded.  

Results: A scale of 1-5 was used.  High feedback scores, indicated that the guide was user-friendly (8/9 – scoring 5/5 or 4/5), easy to navigate (8/9 – scoring 5/5 or 4/5) and provided reassurance (8/9 – scoring 5/5 or 4/5), with visual aids enhancing comprehension. Users found the sections on daily care routines and what to expect particularly helpful. The guide has facilitated communication with schools and extended family members, supporting a wider support network for the patient. Additionally, it assisted in managing side effects and keeping track of appointments. One patient valued the importance of the treatment guide for NF1 by comparing it to the family-held oncology records used in cancer care. One respondent suggested integration of a section on mental health will allow users to document their emotional well-being.

Conclusion: The patient and carer guide for Selumetinib treatment enhances understanding and management of the treatment process and has proven to be a valuable resource.  Future developments will explore the feasibility of converting the guide into a mobile application, providing a more accessible and interactive platform. 

References 

1. Rosalie E Ferner, Susan M Huson, Nick Thomas, Celia Moss, Harry Willshaw, D Gareth Evans, Meena Upadhyaya, Richard Towers, Michael Gleeson, Christine Steiger, Amanda Kirby - Guidelines for the diagnosis and management of individuals with neurofibromatosis 1 - JMG Unlocked 2007
2. NICE – National Institute for Health and Care Excellence - Selumetinib for treating symptomatic and inoperable plexiform neurofibromas associated with type 1 neurofibromatosis in children aged 3 and over. Highly specialised technologies guidance Reference number: Highly Specialised Technologies guidance HST20 Published: 05 May 2022
3. Evans DG, Baser ME, McGaughran J, Sharif S, Howard E, Moran A. Malignant peripheral nerve sheath tumours in neuro- fibromatosis 1. J Med Genet 2002; 3 9: 3 11-4. 
4. Kim A, Gillespie A, Dombi E, et al. Characteristics of children enrolled in treat- ment trials for NF1-related plexiform neu- rofibromas. Neurology 2009; 7 3: 1 273-9. 
5. Korf BR. Plexiform neurofibromas. Am J Med Genet 1999; 8 9: 3 1-7. 
6. Mautner VF, Asuagbor FA, Dombi E, et al. Assessment of benign tumor burden by whole-body MRI in patients with neuro- fibromatosis 1. Neuro Oncol 2008; 1 0: 5 938
7. Nguyen R, Kluwe L, Fuensterer C, Kentsch M, Friedrich RE, Mautner VF. Plexiform neurofibromas in children with neurofibromatosis type 1: frequency and associated clinical deficits. J Pediatr 2011; 159 (4 ): 6 52-5 .e2. 

{{Plexiform neurofibroma}}

Sadali Wanniarachchi¹, Jannath Begum-Ali², Paola Piza Martinez³, Jonathan Green¹, Mark H Johnson⁴ Emily J.H. Jones², Shruti Garg¹  &  the STAARS and EDEN Teams

1 Division of Psychology & Mental Health, The University of Manchester

2 Centre for Brain & Cognitive Development, Birkbeck University of London

3 School of Biosciences, University of Cardiff 

4 Department of Psychology, University of Cambridge, Cambridge, UK.

Background: Neurofibromatosis type 1 (NF1) is a monogenic neurodevelopmental condition associated with a higher prevalence of attention deficit hyperactivity disorder (ADHD). Emerging research suggests ADHD in NF1 differs from idiopathic ADHD. Identifying early differences may clarify neurodevelopmental pathways and inform personalised treatments. This study examined early motor activity in infants with NF1 compared to those with a family history of idiopathic ADHD. The primary aim was to assess whether early motor activity differences emerge among NF1, elevated-likelihood ADHD (EL-ADHD), and typically developing (TD) infants. It was hypothesised that group differences will emerge between the first and second year of life when activity-related differences become apparent.

Methods: Data were collected as part of the Early Development in Neurofibromatosis Type 1 (EDEN) project, a longitudinal study on early functional development in NF1. Infants at 10 and 14 months across three cohorts (NF1, EL-ADHD, TD) were assessed using accelerometery, behavioural video coding, and standardised developmental scales. Group differences were analysed via ANOVA and Kruskal-Wallis tests in the statistical software R.

Results: Accelerometery data showed no group differences at 10 months. However, by 14 months, motion entropy differed between NF1 and EL-ADHD cohorts (p<0.05), whilst motion intensity differed between NF1 and both EL-ADHD and TD groups (p<0.05). Total activity levels did not differ across groups. Likewise, video coding analyses revealed that at both timepoints, time spent in stillness and motion were similar across all three groups. However, NF1 infants spent significantly less time in locomotion than TD (p=0.0287) and EL-ADHD infants (p=0.0412) and more time in non-locomotor positions than TD infants (p=0.0489). By 14 months, time spent in non-locomotor positions, and locomotion was comparable across groups. However, NF1 infants performed more simple locomotion (p=0.000227) and less complex locomotion (p=0.0174) than EL-ADHD infants, although no differences were observed when compared to TD infants. 

Conclusion: Our findings indicate that infants with NF1 show early motor differences at as early as 10 months of age. Total activity levels were similar across groups, but NF1 infants displayed differences in movement complexity, engaging in more simple locomotion and less in complex locomotion compared to EL-ADHD infants. The similar activity levels between NF1 and EL-ADHD infants may reflect emerging ADHD traits in NF1. These findings highlight the potential for early motor differences as markers for neurodevelopmental outcomes and underscore the need for targeted early interventions in NF1 to support motor development.

Funding Disclosure: Sadali Wanniarachchi was funded by the University of Manchester, Nerve Tumours UK, and the Medical Research Council for their PhD. The study from which data were used was supported by Action for Medical Research (GN2385), Rosetrees Trust (A2213), and the Medical Research Council (MR/K021389/1). Additional support was provided by MQ (MQ14PP_83), the EU-AIMS and AIMS-2-TRIALS programmes, which received funding from the Innovative Medicines Initiative (IMI) Joint Undertaking (Grant Nos. 115300 and 777394), the European Union’s FP7 and Horizon 2020 programmes, as well as in-kind contributions from the European Federation of Pharmaceutical Industries and Associations (EFPIA). Further funding came from Autism Speaks, Autistica, and SFARI.

{{Cognition / Behavioral / Learning}}

Lombardi, Francina¹ ; Salvador, Hector²; Parra, Alba²; Blanco, Ignacio²; Juan Luis Becerra²;Ramos, Federico²
 1-Child Neurologist, Neuropediatrics Department, FLENI, Buenos Aires, Argentina
 2-Neuro-oncology Department, Hospital Sant Joan de Déu, Barcelona, España

Introduction: Neurofibromatosis Type 1 (NF1) is associated with significant neurocognitive and emotional challenges. Emotional intelligence (EI) refers to the ability to recognize, understand, manage, and regulate both one’s own emotions and those of others. Deficits in EI may impact social interactions, quality of life, and psychological well-being. This study aims to assess the effectiveness of different rehabilitation strategies in improving EI in NF1 patients using a structured intervention framework, contributing to the identification of personalized, evidence-based treatments.

Objectives:
 *Primary: To evaluate the impact of three different rehabilitation programs on EI and regulation in NF1 patients.
*Secondary:
 -To compare the effectiveness of interventions in improving quality of life and emotional regulation.
 -To assess changes in clinical symptoms, including pain management, following rehabilitation.
 -To determine the feasibility and acceptability of different intervention strategies.

Study Design: This is a prospective, randomized, interventional clinical study. The sample will include 60 patients diagnosed with NF1, aged 16-24 years. Intervention duration is 24 weeks. Patients will be randomized into three intervention groups. Pre- and post-intervention comparisons will be made using repeated-measures ANOVA. Qualitative feedback from participants will be analyzed to evaluate program acceptability.
Baseline and Follow-up Assessments:
 1- Baseline Evaluation: EI and regulation will be assessed using:
-Mayer-Salovey-Caruso Emotional Intelligence Test (MSCEIT) (objective measure of EI as a cognitive ability).
-Difficulties in Emotion Regulation Scale (DERS) (self-report measure of emotional regulation deficits).
-Behavior Assessment System for Children, Third Edition (BASC-3) (assesses emotional, behavioral, and social functioning in real-life contexts).
 -Quality of life and pain perception measures will also be collected.
2- The same assessments will be repeated after 24 weeks to measure intervention outcomes.

 Outcome Measures: Primary outcomes will include changes in baseline scores. Secondary outcomes will include changes in quality of life, improvement in pain perception and management, and participant adherence and satisfaction with the interventions.

Conclusion: Emotional intelligence is considered a fundamental skill for all individuals, and rehabilitating these aspects in children with NF1 could significantly impact their ability to manage emotions, improve interpersonal relationships, and cope with challenging situations. This study aims to provide critical insights into the most effective intervention strategies for improving EI and emotional regulation in NF1 patients. The findings could guide the development of targeted rehabilitation programs, enhancing social and emotional functioning in this population, leading to a positive effect on their quality of life and self-esteem.

{{Cognition / Behavioral / Learning}}

Introduction:
Neurofibromatosis type 1 (NF1) is a genetic disorder that affects multiple organ systems, with cutaneous and neurological manifestations being the most commonly recognized. However, the cardiological complications associated with NF1 often remain underdiagnosed, despite their potential to significantly impact long-term health. It is estimated that between 15% and 30% of patients with NF1 experience cardiovascular abnormalities, ranging from structural issues to functional heart dysfunctions. These manifestations, if not detected early, can substantially affect the patient's quality of life. Given the complexity of these cases, cardiological surveillance in NF1 patients is not only a medical necessity but also a clinical challenge requiring a high level of awareness, particularly among pediatric cardiologists. Furthermore, with the increasing use of novel treatments for plexiform neurofibromas, there is a growing need to recognize the potential cardiotoxic effects of these therapies. Early detection of cardiological complications can make a significant difference in preventing irreversible cardiovascular damage.

Objectives:
To assess the prevalence and types of cardiological manifestations in pediatric patients with NF1 at a high-complexity pediatric hospital.

Materials and Methods:
We conducted a retrospective review of clinical records from pediatric patients diagnosed with NF1 (ages 0-24) followed at our institution. A total of 117 patient records were reviewed, including their electrocardiograms (ECGs) and echocardiograms.

Results:
The cohort included 117 patients with a mean age of 12.26 years. Of these, 36.7% (43 patients) had no prior cardiological evaluation. Among the 74 patients who had undergone cardiological assessments, 16.2% (12 patients) presented with cardiovascular abnormalities. Six patients had abnormal echocardiograms: 3 with mitral valve insufficiency due to prolapsed valves, 1 with multiple ventricular fibromas, 1 with a ventricular septal defect (VSD), and 1 with a combination of an atrial septal defect (ASD), VSD, and patent ductus arteriosus (PDA). Five patients showed ECG abnormalities: 2 with short PR intervals, 2 with supraventricular tachycardia, and 1 with first-degree atrioventricular (AV) block. One patient was diagnosed with hypertension.

Conclusion:
Cardiological monitoring is crucial for all pediatric patients with NF1, as a significant proportion (15-30%) exhibit cardiovascular manifestations that can be identified and managed if detected early. Pediatric cardiologists should be well-versed in the potential cardiovascular issues associated with NF1 to enable timely diagnosis and intervention. Additionally, with the rise of novel therapies for plexiform neurofibromas, it is essential to be aware of the potential cardiotoxic effects of these treatments. Early identification of these complications can help mitigate the need to discontinue critical therapies, ensuring better outcomes and quality of life for NF1 patients.

{{Other}}

Ciavarelli P.,MD, PhD¹; Sarotto L., MD¹; Abudi V. MD²; Valieri M., MD³; Altzul M., MD³; Di Fonso G., MD¹: ; Sockolovsky, M., MD, PhD¹.

1. 
   
2. Clinicas Hospital, University of Buenos Aires, Argentina. 
   • Garraham Children Hospital, Buenos Aires, Argentina. 
   • Italian Hospital of Buenos Aires. Argentina.
3. 
   

Introduction

Neurofibromatosis (NF) is a genetic disorder that affects the development of neural crest derived tissues, including ocular structures in the eye, such as the sclera, choroid, ciliary body and part of the cornea and predispose to refractive defects such as myopia. 

There are publications about the prevalence of myopia in patients with NF1, which is higher than the prevalence of myopia in the general population. Myopia has an overall prevalence of approximately 30%. 

The aim of this work is to describe the prevalence of myopia and astigmatism in a series of patients with NF1 and compare it with the general population.

Materials and Methods

A retrospective and cross-sectional study of patients with NF1, treated at the Clinicas Hospital, the Italian Hospital, and Garraham Hospital was performed. The clinical and demographic characteristics of patients seen between January 1, 2024 and January 31, 2025 were collected from medical records. The sample was described with measures of central tendency and dispersion for continuous numerical variables and percentage for categorical variables. To compare the prevalence of myopia in the sample studied and the estimated prevalence in the general population, a one-way test of proportions was performed. A p value of less than 0.05 was considered statistically significant. R software was used for the statistical analysis.

Results

The study included a total of 123 participants, with a mean age of 31.2 years (SD = 16.9). Of the total sample, 56.9% (n = 70) were women and 43.1% (n = 53) were men. Of the participants, 98.4% (n = 121) were of Argentine nationality, while 1.6% (n = 2) were foreigners. Participants were recruited from three medical centers: 32.5% (n = 40) from Garraham, 30.9% (n = 38) from HCJSM, and 36.6% (n = 45) from Hospital Italiano. The prevalence of myopia was 49.6% (n = 61), with a right eye involvement of 41.5% (n = 51), and 47.2% (n = 58) in the left eye. When comparing with the general population, a statistically significant difference was observed X-squared = 21.563, p-value = <0.001. Regarding astigmatism, 52.8% (n = 65) presented astigmatism.

Discussion

In this study, a significantly higher prevalence of myopia and astigmatism was observed in patients with neurofibromatosis type 1 (NF1) compared to the general population, further research is needed to clarify this relationship.

Conclusion: These results suggest that refractive defects are more common in NF1, could be an additional feature?

{{Other}}

Abstract

Neurofibromatosis Type I (NF1) frequently leads to the development of cutaneous neurofibromas (CNFs), which are benign yet debilitating tumors that currently lack effective therapeutic options and urgently need novel treatment strategies. Oncolytic viruses (OVs), a promising cancer immunotherapist class, have successfully targeted various malignancies, while their effectiveness in treating immunologically "cold" tumors like CNFs remains poorly understood. In this study, we investigated the therapeutic potential of OVs for CNFs by employing single-nucleus and single-cell RNA sequencing to analyze tumor samples before and after OV treatment. Our findings revealed post-treatment alterations in the tumor microenvironment (TME), including increased infiltration of T cells, B cells, and macrophages, alongside reduced Schwann cells (SCs), the primary tumorigenic cells in neurofibromas. We found that OVs directly induced apoptosis in SCs and inhibited angiogenesis by disrupting VEGF signaling. Furthermore, OV therapy transformed the TME from an immunologically "cold" to a "hot" state, enhancing antitumor immune responses. These findings highlight the multifaceted efficacy of OVs in treating CNFs and suggest their broader potential in modulating tumor-immune interactions.

Graphic Abstract

{{BIOINFORMATICS AND COMPUTATIONAL OMICS}}

Kathryn M. Lemberg MD PhD¹, Lior Levy BS¹, Susan Aja PhD¹, D. Wade Clapp MD², and Miriam Bornhorst MD³

¹Johns Hopkins University School of Medicine, ²Indiana University School of Medicine, ³Ann and Robert Lurie Children’s Hospital at Northwestern University

Funding: Congressionally Directed Medical Research Program Neurofibromatosis New Investigator Award (PI: Bornhorst, co-I: Lemberg.)

Disclosures: No relevant disclosures

Title: Metabolic phenotyping of the plexiform neurofibroma model Periostin-Cre+;NF1^flox/flox demonstrates lower weights and increased energy expenditure compared to non-NF1 controls 

Purpose: Children and adults with neurofibromatosis type I (NF1) who develop nervous system tumors such as gliomas and plexiform neurofibromas (pNF), tend towards lower body weights than general population norms [1-4], even when consuming calorie dense diets [5]. The reason for this observation is not well understood, and there are limited published studies of the interactions of energy metabolism and tumor growth in animal models of NF1 [6-9]. The aim of this work was to compare the metabolic phenotype of mice with loss of NF1 that develop pNF to litter-matched control mice. 

Methods: We compared male Periostin-Cre+;NF ^flox/flox mice [10, 11] with litter-matched Cre- mice (n=9-11/arm) beginning at 1 month of life. Mice were fed standard chow. We collected weekly weights, monthly QNMR body compositions, and performed indirect calorimetry at 3 months, which is reported onset of peripheral nerve hyperplasia in this model. VO₂ and VCO₂ were used to calculate respiratory exchange ratio (RER) and energy expenditure (EE). Glucose and insulin tolerance testing was also performed. 

Results: Periostin-Cre+;NF1^flox/flox (Cre+) mice have lower body weights than Periostin-Cre-;NF1^flox/flox (Cre-) controls (21.36+ 1.36 versus 22.91 + 1.53 g; p = 0.03) at 3 months of life. Fat mass was significantly lower in Cre+ mice at 4 months (2.89 + 0.88 versus 4.22 + 1.19 g; p=0.01). RER was lower in Cre+ mice (0.92 + 0.02 versus 0.96 + 0.03; p<0.01) while energy expenditure per kg of body weight was ~20% higher in Cre+ mice (19.29 + 1.32 versus 15.87 + 1.53 kcal/kg; p<0.0001). Food intake was slightly higher in Cre+ mice (11.39 kcal vs 10.32 kcal, p = 0.024), while no differences in activity were observed between the groups. Cre+ mice had lower peak glucose than controls (174.44 + 17.68 versus 207.55 + 36.62 mg/dl, p = 0.02). 

Conclusions: Similar to what has been observed in non-tumor forming models and in people with NF1, lower weights, fat mass, and higher EE were observed in the Cre+ mice compared to Cre- controls. Lower RER and lower peak blood glucose suggest differences in carbohydrate and fat metabolism in NF1 versus control animals. The study remains ongoing to collect data through at least 6 months of life. Future directions include evaluating hormone differences between the groups, assessing the metabolic response of Cre+ mice to high fat diet, and analyzing metabolomic profiles of tissues, including serum, peripheral nerve and liver, collected at study termination. 

References: 

1.       Lemberg, K.M., et al., Anthropometric measurements of children with neurofibromatosis type I: impact of plexiform neurofibroma volume and treatment. Pediatr Res, 2024.

2.       Koga, M., Y. Yoshida, and S. Imafuku, Nutritional, muscular and metabolic characteristics in patients with neurofibromatosis type 1. J Dermatol, 2016. 43(7): p. 799-803.

3.       Ly, K.I. and J.O. Blakeley, The Diagnosis and Management of Neurofibromatosis Type 1. Med Clin North Am, 2019. 103(6): p. 1035-1054.

4.       Clementi, M., et al., Neurofibromatosis type 1 growth charts. Am J Med Genet, 1999. 87(4): p. 317-23.

5.       de Souza, M.L., et al., Nutrient intake in neurofibromatosis type 1: A cross-sectional study. Nutrition, 2015. 31(6): p. 858-62.

6.       Botero, V., et al., Neurofibromin regulates metabolic rate via neuronal mechanisms in Drosophila. Nat Commun, 2021. 12(1): p. 4285.

7.       Summers, M.A., et al., Developmental dosing with a MEK inhibitor (PD0325901) rescues myopathic features of the muscle-specific but not limb-specific Nf1 knockout mouse. Mol Genet Metab, 2018. 123(4): p. 518-525.

8.       Tritz, R., et al., Nf1 heterozygous mice recapitulate the anthropometric and metabolic features of human neurofibromatosis type 1. Transl Res, 2021. 228: p. 52-63.

9.       Chan, A., et al., Maternal obesogenic diet operates at the tumor cell of origin to increase incidence and decrease latency of neurofibromatosis type 1 optic pathway glioma. Neuro Oncol, 2024. 26(12): p. 2339-2351.

10.     Fisher, M.J., et al., Cabozantinib for neurofibromatosis type 1-related plexiform neurofibromas: a phase 2 trial. Nat Med, 2021. 27(1): p. 165-173.

11.     Flint, A.C., et al., Combined CDK4/6 and ERK1/2 Inhibition Enhances Antitumor Activity in NF1-Associated Plexiform Neurofibroma. Clin Cancer Res, 2023. 29(17): p. 3438-3456.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Haley M. Hardin, MS¹; Ethan W. Hass, BS¹; Robert Allaway, PhD²; Helen Morrison, PhD^3,4; Cristina Fernandez-Valle, PhD¹

¹Burnett School of Biomedical Sciences, University of Central Florida, Orlando, FL, United States of America

²Sage Bionetworks, Seattle, WA, United States of America

³Leibniz Institute on Aging, Fritz Lipmann Institute, Jena, Germany 

⁴Faculty of Biological Sciences, Friedrich-Schiller University, Jena, Germany

Purpose: NF2-related schwannomatosis (NF2) predisposes individuals to benign schwannomas, meningiomas, and ependymomas. Schwann cells (SCs) lacking merlin have multiple dysregulated signaling cascades activated by receptor tyrosine kinases and cell adhesion molecules, including integrins and cadherins, which contribute to schwannoma development and growth. A current theory on the pathogenesis of NF2-schwannomas posits that they arise after a nerve injury from SCs that assumed a “repair state” phenotype but cannot re-differentiate due to loss of functional merlin. ERK’s persistent high activity, together with deregulated YAP activity caused by merlin loss, is associated with SC hyperplasia following nerve injury in an NF2 conditional mouse model. Previous research by this lab evaluated MEK inhibitors for NF2 treatment. In merlin-deficient human SCs (MD-HSCs) treatment with the MEK inhibitor trametinib induced a G0/G1 cell cycle arrest as well as a morphological transformation; they shift from small spindle shaped cells to large, flattened cells.

Methods: We conducted immunofluorescent staining, western blots, and qPCR on MD-HSCs treated with trametinib to evaluate transcription factors regulating the repair and myelination phenotype of SCs. We also analyzed the transcriptome and proteome analysis of isogenic wild-type and MD-HSCs treated with trametinib for differential regulation of genes characterizing the SC myelination or repair state. 

Results: Our results show that merlin loss in our human schwannoma model cells increases expression of genes, including several AP-1 transcription factors, that support the repair phenotype. Transcriptome and proteome analysis of MD-HSCs treated with trametinib for 24 hours shows upregulation of genes involved in extracellular matrix receptor interactions and cell adhesion. MD-HSCs also significantly downregulated transcription of SOX2 and YAP1 following trametinib treatment. Immunofluorescent staining showed increased stress fiber and focal adhesion formation as well as decreased nuclear localization of c-Jun and increased nuclear Egr2/Krox20 in MD-HSCs treated with trametinib, which is consistent with a pro-myelinating SC phenotype.

Conclusion: We demonstrate that our human schwannoma model cells reflect the repair phenotype observed in NF2-null mouse models and human vestibular schwannomas, and that treatment with a MEK-inhibitor monotherapy in MD-HSCs induces a shift from a repair towards a pro-myelinating phenotype.

Funding: This project was funded by a grant to Cristina Fernandez-Valle from DOD (W81XWH-21-1-0228), and Haley Hardin is funded by CTF as a young investigator awardee (2022-01-002).

Disclosures: No relevant financial disclosures.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Mashu Futagawa, PhD, Department of Clinical Genomic Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Japan

Full List of Authors: 

Tetsuya Okazaki, Eiji Nakada, Chika Fukano, Risa Osumi, Fumino Kato, Yusaku Urakawa, Hideki Yamamoto, Toshifumi Ozaki, Akira Hirasawa

Funding: 

This research was supported by Japan Agency for Medical Research and Development (AMED) under Grant Number JP23bm1423027.

Disclosure: The authors declare that they have no competing interests.

Purpose: Previous studies reported that patients with neurofibromatosis type 1 (NF1) develop neoplasms at a significantly higher rate than the general population (Uusitalo E et al., 2016, Landry JP et al., 2021). However, few studies have evaluated the risk of cancer in Japanese NF1 patients. This study retrospectively investigated the risk of cancer development in patients with NF1 pathogenic variant (PV) confirmed by genetic testing.

Methods: We reviewed the history of malignant tumors in 52 participants from our Mid-West Japan Hereditary Tumor Cohort Study (https://cgm.hsc.okayama-u.ac.jp/en/cohort/) who were genetically confirmed to harbor NF1 PV. In addition, we analyzed whether the risk of malignant tumors was associated with the type or location of NF1 PVs.

Results: The mean age at the initial visit was 36 ± 21.4 years (range: 2–76 years), with 55.8% of the subjects being female. A history of malignancy was identified in 34.6% (18/52) of the patients. Among adults (over 18 years), 43.6% (17/39) had a history of malignancy, including malignant peripheral nerve sheath tumors (MPNST) in 7.7% (3/39) and breast cancer in 20% (5/25) of women. The mean age at breast cancer diagnosis among adult women (54.8 years). Notably, a trend toward a higher risk of malignant tumors was observed in patients harboring missense variants.

Conclusions: The mean age at diagnosis of breast cancer in carriers of the NF1 PV was higher than the commonly reported age of onset of 30-50 years for breast cancer in this patient population. This finding suggests that breast cancer surveillance should be extended beyond the age of 50 years in this population. Our study suggests that Japanese patients with NF1 have a high risk of developing malignant tumors. Further large prospective studies are needed to validate these findings and to explore additional risk factors for malignancy in NF1.

{{Other}}

Tina Thomas, PhD; Caitlyn J. Cap, PsyD; Meredith J. Goyette, MA; Megan Knauss, MSW, LGSW; Rebecca Levitt, BA; Emily J. Carlson, PhD; Jarrod J. Sotos, PhD; Karin S. Walsh, PsyD

(All authors are affiliated with Children’s National Hospital)

Research Category: Clinical research: cognition/behavioral/learning

Funding: District of Columbia Intellectual and Developmental Disabilities Research Center (DC-IDDRC) Award U54HD090257 by NICHD (PI: V. Gallo).

Disclosures: The authors have no conflicts of interest to disclose. 

Title: Adaptive Functioning in NF1: The Influence of Cognition, Executive Functioning, and Social Determinants of Health in Children and Young Adults

Purpose: Adaptive functioning (AF) is a crucial set of skills necessary for building independence and preparing for transition to adulthood. These abilities are often underdeveloped in individuals with Neurofibromatosis Type 1 (NF1), for whom transition planning is of vital importance given the range of medical, cognitive, and psychosocial challenges they face. We aimed to understand the impact of executive functioning (EF), intelligence (IQ), and social determinants of health (SDOH) on AF in the NF1 population, as an important step in developing interventions to support transition readiness. We hypothesized that IQ, EF, and SDOH would be associated with AF.

Methods: We extracted data from a large retrospective clinical neurocognitive dataset and medical record review. Inclusion criteria were NF1 patients who were clinically administered an adaptive measure. Neurocognitive data included IQ (Weschler scales), EF (Delis-Kaplan Executive Function System [D-KEFS] and parent-proxy reported Brief Rating Inventory of Executive Function [BRIEF]), and AF data (parent-proxy reported Adaptive Behavior Assessment System [ABAS]). The Child Opportunity Index 3.0 (COI) was extracted as a measure of SDOH. Impairment was classified as 1.5 SD below the mean. Statistical methods included parametric and non-parametric correlations and multiple regressions.

Results: Participants (N=62) were 12.34 years of age (SD=3.66, Range=8-23) and 47.7% female. Average IQ was 80.95 (SD=16.21, Range=47-115) with 36.9% classified as impaired. Adaptive data had a mean of 83.52 (SD=17.43, Range=50-120) with 29.2% classified as impaired. AF was significantly correlated with parent-reported EF (N=62, r =-0.68, p<0.001) and DKEFS Verbal Fluency Switching (N=32, r=0.42, p=0.016), but not IQ or COI. In regression, parent-reported EF strongly predicted AF (β=-0.57, p<0.001), although DKEFS Switching did not (β=0.27, p=0.070). When subdomains of parent-reported EF were examined in regression, cognitive regulation strongly predicted AF (β=-0.73, p=0.001) but emotional and behavioral regulation did not. When EF predictors were regressed on AF subdomains, cognitive regulation remained most predictive: conceptual (β=-0.64, p=0.007), social (β=-0.49, p=0.044), and practical (β=-0.61, p=0.007). 

Conclusion: Consistent with previous literature, EF, specifically cognitive regulation, significantly predicted AF in our sample of children and young adults with NF1. IQ and COI were not related to AF. Small sample size was a limitation of this study. Future research may consider further analysis of medical risk factors as well as individual and family factors on AF. Overall, the current findings suggest that interventions targeted towards “real world” application of EF factors, particularly cognitive regulation skills, may help to facilitate AF development in the NF1 population. 

{{Cognition / Behavioral / Learning}}

Purpose: Nerve sheath tumors of the vocal cord are extremely rare. We describe the case of a Neurofibromatosis 1 (NF1) presenting with hoarseness and underlying vocal cord neurofibroma. 

Methods and Results: A 41-year-old patient with NF1 presented with one year of hoarse voice. He described a sudden change in his voice that was noticed during the day while speaking. The impairment remained constant since onset and was without pain, but speech required more effort. No dysphagia was noted. He had no history of smoking. Videostroboscopy was performed in laryngology clinic, demonstrating fullness of the right true fold with relatively normal appearing epithelium and reduction in vibratory capacity. These findings were suggestive of a subepithelial cyst. The patient underwent direct microlaryngoscopy with microflap excision. Rather than a cyst, a subepithelial fusiform mass of right true vocal fold was found, with dense anterior and posterior attachments was visualized. It was dissected free while preserving the vocal ligament and much of the superficial lamina propria and epithelium as possible. Pathology showed coexpression of S100 protein and CD34 in the neoplastic cell population with approximately equal frequency supporting the diagnosis of neurofibroma.  Following surgery, the patient reported a significant improvement in vocal quality and decreased strain producing sound. His Voice Handicap Index-10 Score improved from 32 to 20 (< 11 considered normal). Follow up video stroboscopy showed mild erythema, stiffness and a whitish patch of the right vocal cord, consistent with early stage of healing. In contrast to the preoperative findings, the vocal fold closure was complete and not impaired by the mass. Literature review shows that this is a rare presentation and will be described in detail.

Conclusion: Neurofibroma of the vocal cord is a rare presentation in patients with NF1. Patients presenting with voice changes should be referred for laryngeal evaluation.

Fig 1: Preoperative image from videostroboscopy showing fullness of the midportion of the right true vocal fold

Fig 2: View of right true vocal fold with decrease in the fullness, but expected inflammation for this point postoperatively

Figure 3: Intraoperative photo showing fusiform submucosal mass lesion involving much of the membranous vocal fold on the right

Figure 4: intraoperative view at conclusion of procedure showing a smooth vocal fold edge, preserved epithelium other than the incision itself, but greater erythema than is typical for these procedures in the non-neoplastic setting.

{{Other}}

Jarrod J. Sotos, PhD; Rebecca Levitt, BA, Emily J. Carlson, PhD; Meredith J. Goyette, MA; Christina Thomas, PhD; Caitlyn J. Cap, PsyD; Megan Knauss MSW, LGSW; Karin S Walsh, PsyD 

(all authors are affiliated with Children’s National Hospital)

Funding: Lambert Family Foundation, Gilbert Family Foundation 

Disclosures: None 

Title: Not-So-Sweet Dreams: Sleep and Emotion Regulation Problems in Children with NF1 

Purpose: Children with Neurofibromatosis Type 1 (NF1) face well-documented cognitive and attentional challenges and are also at heightened risk for internalizing difficulties (e.g., anxiety, depression, somatization), externalizing behaviors (e.g., impulsivity, aggression, oppositionality), and emotion regulation deficits. A recent caregiver survey conducted by this group highlighted significant sleep disturbances (e.g., trouble falling asleep, frequent nighttime awakenings) and impairments (e.g., daytime sleepiness, attention difficulties, irritability) in children with NF1. Despite these findings, research on sleep in NF1 and its connection to emotion regulation and internalizing/externalizing (I/E) behaviors remains limited. This study expands on our previous work by investigating the relationship between sleep difficulties and emotion regulation in children with NF1 using norm-referenced parent questionnaires.

Methods: Caregivers of 31 children with NF1 (ages 6-16) provided ratings of their child’s emotion regulation and I/E behaviors using the Behavior Rating Inventory of Executive Function - Second Edition (BRIEF-2) and the Behavior Assessment System for Children – Third Edition (BASC-3). Sleep disturbance and sleep impairment were measured using parent-proxy Patient Reported Outcomes Measurement Information System (PROMIS) measures. Additional sleep factors were assessed using the Child and Adolescent Sleep Checklist (CASC). Data were analyzed using Pearson correlations and independent samples t-tests.

Results: Participants (M_age = 10.83 years, SD = 2.95) averaged 8.78 hours of sleep on weeknights (SD = 1.51), with 39% (n = 12) reporting sleep disturbance and impairment, respectively. Those with sleep difficulties slept 1 to 1.5 hours less per night than their peers. Sleep disturbance and impairment were significantly associated with greater emotion regulation difficulties, internalizing behaviors, and externalizing behaviors. T-tests indicated that children with sleep disturbance and impairment exhibited more pronounced emotion regulation challenges and higher levels of internalizing symptoms, along with lower adaptability, social skills, leadership, daily living skills, and communication abilities compared to those without sleep difficulties. However, no significant differences emerged between groups in externalizing behaviors. Exploratory analyses of specific internalizing behaviors revealed that those with reported sleep disturbance had reported greater somatization symptoms, and participants with sleep impairment reported greater challenges with mood.

Conclusions: This study highlights the significant impact of sleep disturbance and impairment on emotion and behavior functioning in children with NF1. Those with disrupted sleep show greater emotion dysregulation and increased internalizing symptoms, particularly somatization and mood issues. Clinicians should assess sleep routinely and implement targeted interventions to improve sleep quality, reducing emotional distress and enhancing well-being.

{{Cognition / Behavioral / Learning}}

Idrisova Zhannat¹, Zhumadullaev Bahram², Zhienkulova Gulzhanat³, Rakhimbekova Farida¹

¹Kazakh National Medical University named after S.D. Asfendiyarov, Department of Nervous Diseases, Scientific Research Institute of Fundamental Medicine named after B. Atchabarov

²Scientific Research Institute of Pediatrics and Pediatric Surgery, Department of Tumors (No. 2) 

³Oncology outpatient department, "University Medical Center"

Funding - Genetic research was carried out within the framework of the Grant of the Ministry of Education and Science of the Republic of Kazakhstan AP19676226 "Study of genetic markers and environmental factors in phacomatoses and neurogenic tumors" for 2023-2025

No Disclosures

Purpose: Clinical case study of neurofibromatosis phenotype spectrum in identical twins with NF1 diagnosis  

Methods: Acquisition of clinical data of NF1 (neurofibromatosis 1) patients with observations, magnetic resonance imagining (MRI), specialty consultations, medical-genetic testing. 

Results: Both boys, 11 years old, presented with multiple café au lait spots from 1 to 3 cm, up to 10 spots in in the axillary area. Diagnosis was verified by genetic testing – nonsense mutation in NF1 gene chr17:31206297C>T, c.1318C>T, p.Arg440Ter in both twins. 

First twin had small plexiform neurofibroma above m. sternocleidomastoideus on the left, below the corner of the lower jaw on the left, measuring 1.5 by 1 cm, bulging 0.5 cm out the skin surface. Second twin had from 2019 (age of 6 years old) a large neurofibroma in the left inguinal-iliac region on the left, emanating from the roots and plexus at the level of S1-S5, dimensions 7 cm by 4 cm and by 2 cm; according to MRI in 2024 in STIR regime the tumor had sized 10cm by 9 cm by 4.5 cm. In 2022, first biopsy confirmed NF1, in 2024, the tumor started to enlarge (see Figure 1) with volume 13 cm by 10 cm and by 6cm, additionally the boy started complaining the pain appeared during walking and at rest. Same year, biopsy of pelvic tumor masses at the Korean clinic was performed. Biopsy results: increase cellularity marked, nuclear pleomorphism marked, necrosis present, mitosis 2/10 HPF; suggestive of malignant peripheral nerve sheath tumor. Patient underwent 4 courses of chemotherapy, tumor decreased in volume, pain almost gone, treatment continues.

Conclusion: Identical twins with the same mutation in NF1 gene have a significant difference in the clinical picture (phenotype): one child has classic Neurofibromatosis type 1 with multiple (>10) coffee-with-milk spots and one small plexiform neurofibroma, and the other twin has a malignant tumor of the membranes of peripheral nerves of the inguinal-iliac region.

Figure 1. Massive pelvic neurofibroma of 11 years old boy (one from identical twin pair). 

{{Malignant peripheral nerve sheath tumors (MPNST)}}

Arjun Yogaratnam, BS,¹ Jayla Melvin, MLIS,² Omololu Omoteso, BS,³ Staci Martin, PhD,² Daniel Wikstrom, BA,² Paige Little, BS,² Brigit Rweikiza, BA,² Anne Dufek, MS, CPNP,² Brian Stewart, MA,⁴ Joshua Roberts, PhD,¹ Michael Fisher, MD,³ Chelsea Kotch, MD, MSCE,³ Carlos Romo M.D.,³ Pamela L. Wolters Ph.D.²

¹Johns Hopkins University, ²National Cancer Institute, ³Children’s Hospital of Philadelphia, ⁴Metricwire, Inc.

Funding: This research was supported in part by the Neurofibromatosis Therapeutic Acceleration Program (NTAP) and the Intramural Research Program of the National Institutes of Health.

Disclosures: PLW and SM received funding from NTAP.

Title: Validation of novel NF1-specific patient-reported outcome measures to assess pain related to plexiform neurofibromas for clinical trials: Preliminary data analysis

Purpose: Patient-reported outcome (PRO) measures are used to evaluate the effects of treatments on symptoms and function. To date, there are no validated PRO measures for pain related to plexiform neurofibromas (pNF), which are needed for clinical trials and critical for drug approval. We previously developed PRO measures assessing pNF-related pain intensity and interference in daily life from extensive qualitative research. The current study examines the psychometric properties of these two novel surveys in individuals with neurofibromatosis type 1 (NF1) and pNFs to support validation. 

Methods: Individuals with NF1-pNF, ages ≥ 8 years, with pNF-related pain, and a consistent analgesic regimen (none or stable >3 months) or MEK therapy (none or >12 months), are eligible for either in-person or remote enrollment at 3 NF centers (target N=120). At enrollment, participants complete a demographic form and PROMIS Anxiety and Depression measures. During a practice session, they download the Catalyst Metricwire mobile app, and they review and complete the two new PRO measures: 1) PAin INtensity Scale for pNF (PAINS-pNF) assessing spike and chronic tumor pain intensity of a target pNF (0=no tumor pain to 10=worst tumor pain) and 2) the Pain Interference Index for pNF (PII-pNF) assessing tumor pain interference (0=none to 6=completely) as well as validated measures of pain intensity (eCAS; 0-100) and pain interference (PROMIS-PI; T-scores, Mean=50, SD=10). During the study, participants complete the PAINS-pNF and eCAS nightly for two consecutive weeks and the PROMIS-PI and additional measures at days 7 and 14. Data analyses were conducted using non-parametric statistics.

Results: To date, 55 participants enrolled with 53 completing the study (2 withdrew after consent); mean age=33 years; range=8-69; 47% male, 62% White, and 91% not Hispanic. For the new measures, internal consistency was excellent (Cronbach’s alpha: PAINS-pNF=0.91, PII-pNF=0.96) with all item-total correlations >0.7. Spearman correlation coefficients indicated excellent construct validity for the two PAINS-pNF items of spike and chronic tumor pain intensity with the eCAS tumor pain item (r_s=0.90, r_s=0.80, respectively, both p<.001) and for the PII-pNF total score with the PROMIS-PI (r_s=0.75, p<.001). Known groups validity was demonstrated by significantly different median PII-pNF scores between NF1 symptom severity groups (p=0.011; mild=0.33 vs. moderate-severe=1.29) and higher median PAINS-pNF spike pain intensity in participants taking analgesics compared to those who do not (p=0.004; pain meds=5.0 vs. no pain meds=1.0). Median ratings of spike and chronic pain also were significantly different between descriptive pain categories (e.g., none, mild, moderate, severe; p<.001). Higher PROMIS Anxiety and Depression T-scores were correlated with greater PII-pNF total scores (r_s=0.33, p=.017; r_s=0.30, p=.029, respectively) suggesting a relationship between social-emotional function and pain interference in daily life, but not with PAINS-pNF spike or chronic tumor pain intensity.

Conclusion: The new NF1-specific PRO measures assessing pNF-related spike and chronic pain intensity and pain interference in daily life show excellent reliability and validity based on preliminary data. These measures are being used as secondary endpoints in a phase III MEK inhibitor trial that will determine their sensitivity to assess change with treatment. Recruitment for individuals with NF1-pNF <21 years and >40 years for the current study is ongoing to achieve target enrollment and complete the validation process.

{{Other}}

Lalitha Gopalan¹, Ph.D 

¹Division of Experimental Hematology and Cancer Biology, Cancer & Blood Diseases Institute Cincinnati Children's Hospital Medical Center, 3333 Burnet Ave., Cincinnati, OH 45229, USA.  

Abstract 

Neurofibromatosis type 1 (NF1) patients are prone to developing plexiform neurofibromas (PNFs). Previous work showed that the NF1 protein neurofibromin can bind p97/valosin-containing protein (VCP) in rat brain (Wang et al, 2011). We found VCP protein increased expression in both mouse and human PNFs versus wild type mouse dorsal root ganglia or normal human nerves. Co-immunoprecipitation confirmed that VCP binds to NF1 in purified mouse Schwann cells and tumor lysates. In vivo treatment with CB-5083, a p97/VCP inhibitor that causes poly-ubiquitinated protein accumulation and proteotoxic stress, inhibited cell proliferation, increased apoptosis, and reduced PNF volume in the DhhCre;Nf1fl/fl mouse model. However, some mice were resistant to CB-5083 mediated tumor shrinkage. To investigate mechanisms that underly this resistance, we developed a CB-5083-resistant cell line by exposing an NF1 knockout immortalized human Schwann cell line to stepwise increasing concentrations of CB-5083 (0.1 µM to 6.5 µM). RNA sequencing on the resistant cells identified 805 differentially expressed genes (vs non-resistant, 3-fold change, p<0.05), with TGF-beta signaling predicted as the top pathway. We confirmed the overexpression of TGFβ1, TGFβ2, TGFβR1, TGFβR2, and SMAD7 genes by qRT-PCR, and Western blotting confirmed the overexpression of TGFβR1 and TGFβR2 proteins. Pre-clinical testing combining CB-5083 with a TGFβ inhibitor will be carried out.

(Supported by NIH-R01 NS097233 to JW). 

Author list: Lalitha Gopalan¹, Ashley Hall¹, Liang Hu¹, Mi-Ok Kim², Eva Dombi³, Nancy Ratner^1,4, and Jianqiang Wu^1,4.

¹Division of Experimental Hematology and Cancer Biology, Cancer & Blood Diseases Institute Cincinnati Children's Hospital Medical Center, 3333 Burnet Ave., Cincinnati, OH 45229, USA. ²Department of Epidemiology and Biostatistics, UCSF, 1450 3rd Street, San Francisco, CA 94143, USA. ³Pediatric Oncology Branch, National Cancer Institute, Bethesda, MD 20892, USA. ⁴Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, OH, 45267 USA.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Ethan W Hass, BS¹; Lenna Huelbes, BS¹; Sofia A Oliveira, BS¹; Alexander W Sutton¹; Haley Hardin, MS¹; Anna Nagel, PhD¹; John Ragheb, MD²; Mislen Bauer, MD^2,3; Cristina Fernandez-Valle, PhD¹

¹University of Central Florida College of Medicine, Orlando FL

²Nicklaus Children’s Hospital, Miami FL

³Kidz Medical Services, Miami FL

Purpose: Patients with NF2-related schwannomatosis (NF2-SWN) develop bilateral vestibular schwannomas as well as paraspinal schwannomas. Our studies sought to confirm results of high-throughput drug screens conducted on primary schwannoma cells isolated from a debulked paraspinal schwannoma resected from a severe pediatric patient.

Methods: Previous time lapse drug sensitivity screens identified three pharmacologics out of 20 tested that produced a 50% decrease in well confluence compared to DMSO controls at 100nM over 96 hours. These were: the dual PI3K/HDAC inhibitor CUDC-907, the Src family kinase inhibitor dasatinib, and the combination of a MEK inhibitor, trametinib with a bromodomain and extraterminal protein (BET) inhibitor, BMS-986158. Follow-up flow cytometry studies evaluated the ability of each treatment to cause cell cycle arrest (EdU incorporation) or cell death (violet ratiometric assays). Drug target modulation was validated using automated capillary-based immunoblotting.

Results: Trametinib and dasatinib both promoted a G1 cell cycle arrest, whereas BMS-986158 promoted a G2 cell cycle arrest. CUDC-907 and the combination of trametinib with BMS-986158 were the only treatments that promoted apoptotic cell death. Trametinib and BMS-986158 were ineffective alone. By 6 hours of treatment, dasatinib completely inhibited FAK phosphorylation at tyrosine 576 (Y576) required for catalytic activity and caused proliferation arrest. Brigatinib, an ALK inhibitor that also inhibits FAK, did not reduce proliferation in the drug screens at the doses tested; however, it effectively reduced total FAK levels by 30% with detectable Y576 phosphorylation at 6 hours of treatment. FAK Y576 phosphorylation by SRC promotes its activity. Dasatinib dramatically increased total SRC; phosphorylation of tyrosine 527, a site of negative regulation by CSK, was absent. This is consistent with a cellular compensation mechanism against SRC inhibition.

Conclusion: Of the 20 therapeutics tested on primary paraspinal schwannoma cells, we identified three effective candidates. Dasatinib was the most cytostatic drug tested and caused SRC-dependent inhibition of FAK. Comparatively, the FAK inhibitor in clinical trials for NF2-SWN, brigatinib reduced total FAK levels but failed to arrest proliferation. CUDC-907 and trametinib in combination with BMS-986158 both promoted cell death. This data supports the use of limited patient samples for mechanistic drug studies and personalized medicine to advance the standard of care for NF2-SWN.

Funding: All experiments were funded by the University of Central Florida.

Disclosures: No relevant financial disclosures.

{{THERAPEUTICS}}

Eva Dombi¹, Jonathan Samet², Jenny P. Garzon², Carolyn R. Raski², Maria P. Silva² Michael Sawin², Robert Listernick², Carlos E. Prada²

¹Pediatric Oncology Branch, National Cancer Institute, Bethesda, MD, ²Ann & Robert H. Lurie Children’s Hospital of Chicago, Chicago, IL.

Funding: This study is funded by the National Institute of Neurological Disorders and Stroke (NINDS) Award Number: 1R61NS122094-01 and supported in part by the Intramural Research Program of the NIH/NCI.

Disclosures: None

Title: Developmental Abnormality of the Sacroiliac (SI) Joint is an Underreported Complication of Neurofibromatosis Type 1 (NF1)

Purpose: To present a possible new NF1-related developmental abnormality in children and young adults with NF1 involving the SI joint.

Methods: We report three participants out of 112 enrolled, with SI joint involvement noted on whole-body magnetic resonance imaging (WB-MRI). These individuals are part of a cohort of 112 participants in a longitudinal NF1 study (NCT05238909) at Ann & Robert H. Lurie Children’s Hospital of Chicago in collaboration with the National Institutes of Health. A detailed review of clinical history and imaging was conducted.

Results: Participants 1-3, all female, were 12, 17, and 32 years, respectively, at time of WB-MRI. For each, there was an underlying internal plexiform neurofibroma (PN) adjacent to the SI joint. All three were asymptomatic, without history of joint dislocations or pain. Participant 1: The first WB-MRI for this individual showed osseous dysplasia of the right SI joint, including widening of the fibrofatty portion. Follow-up WB-MRI a year later showed osseous dysplasia of the right SI joint with an intervening linear soft tissue abnormality consistent with infiltrative intra-articular PN. The tumor volume was too small to perform volumetrics. Participant 2: The first WB-MRI revealed dysplasia of the right posterior iliac crest and right hemisacrum with abnormal widening of the right S1 neural foramen. T2 hyperintense signal was noted in the right SI joint region, extending into the neural foramen, consistent with a focal diffuse PN. The tumor volume was too small to perform volumetrics. Participant 3: The first WB-MRI revealed a widened and dysplastic appearance of the left SI joint in the superior aspect. T2 hyperintense signal within the fibrofatty portion of the joint was identified, extending into the subcutaneous tissues. A year later, her second WB-MRI showed a PN with reticular tissue extending from the left SI joint into the surrounding posterior soft tissues. The tumor volume measured 61.7 mL. 

Conclusion: Our findings show an underreported developmental abnormality of the SI joint in NF1. MRI characteristics include diffuse, non-mass-like PN tissue infiltrating and widening the SI joint. There is an associated osseous dysplasia of the bones of the SI joint. PN was identified in three asymptomatic individuals, demonstrating the value of WB-MRI.

Figure 1. (A, B) Axial and coronal STIR images show an infiltrative linear plexiform neurofibroma (blue arrows) in the left sacroiliac joint. There is widening of the joint and thinned dysplastic appearance of the posterior iliac crest. (C) Axial STIR image of the pelvis demonstrates a plexiform neurofibroma infiltrating the right sacroiliac joint characterized by linear T2 hyperintense signal that widens the joint (long blue arrow). The adjacent right posterior iliac crest is thinner than the unaffected left side consistent with dysplasia of the bone (short arrow). (D) Axial STIR image of the pelvis demonstrates a linear plexiform neurofibroma infiltrating and widening the right sacroiliac joint (blue arrow).

{{Bone abnormalities}}

Anne Dufek, CPNP, MS¹, Eva Dombi, MD¹, Andrea M Gross, MD¹, Arjun Kanuri, MD, MBA², Hemaxi Patel, RN, BSN³, Alexia Torres, BS, MPH³, Andrea Baldwin, CPNP, MSN³, Kara Heisey, BS³, Brigitte C Widemann, MD¹

Disclosures: No relevant financial relationships

Affiliations:

¹National Cancer Institute, Pediatric Oncology Branch, Bethesda, MD

²Center for Plastic Surgery, Chevy Chase, MD

³Clinical Research Directorate, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, MD, USA

Title: Successful resection of selumetinib resistant superficial facial plexiform neurofibromas in two pediatric patients with Neurofibromatosis type 1

Purpose: Neurofibromatosis type 1 is a tumor predisposition syndrome characterized by peripheral nerve sheath tumors, including histologically benign plexiform neurofibromas (PN). PN occur in several variants including typical fascicular/multi-nodular tumors, diffuse appearing tumors without clear internal structure and distinct nodular lesions. Preliminary analyses suggest that response to MEK inhibitor (MEKi) treatment may vary between these PN types. For example, distinct nodular lesions appear to respond less frequently than typical appearing PN and may grow or develop despite ongoing therapy. Here, we describe two participants in the phase I/II trial of the MEKi selumetinib, who underwent successful surgical resection of a skin infiltrating superficial PN, that demonstrated growth during selumetinib treatment, while shrinkage was maintained in the underlying, deeper target PN.

Methods: Two participants in the Phase I/II trial of selumetinib for children with NF1 and inoperable PN (SPRINT) (NCT01362803) are presented here. Information was extracted from the medical record and study database.

Results: Participant 1 was 6 years old when starting selumetinib treatment for a right facial submandibular PN.  Her target tumor was classified as typical multi-nodular/fascicular PN, localized predominantly in the deep facial structures but extending into subcutaneous/cutaneous layers, and interdigitating with surrounding tissues. Approximately 1 year into selumetinib treatment, a new area of diffuse superficial tumor developed over the right mandible adjacent to the target PN. She remained on treatment for approximately 7 years at which time her target PN volume was -32.1% from baseline while the superficial PN continued to grow. She underwent plastic surgery to remove the superficial lesion, which was a PN with diffuse intra- and extra-neural components; there were no surgical complications.  She continues on selumetinib post-operatively and has not had any regrowth of the superficial tumor in 43 months of observation.  

Participant 2 was 7 years old at the time of starting selumetinib treatment for her right facial and periorbital PN, which had similar structural features to the PN of participant 1. At baseline, there was a small area of superficial PN visible on imaging over the right mandible adjacent to the target PN which shrank slightly during the first year of selumetinib treatment, but then began growing markedly over the next 6 years while her target PN volume was -48.8% from baseline. Due to worsening appearance concerns, the growing superficial portion of the tumor was resected by plastic surgery, with pathology showing cutaneous-subcutaneous neurofibroma with a small intraneural neurofibroma component; there were no surgical complications. She continues on selumetinib post-operatively and has not had any regrowth of the superficial tumor in 12 months of observation.

Conclusions: We present here two children treated with selumetinib for facial PN who had partial response in their deep, multi-nodular target PN, but experienced some growth in their diffuse superficial skin infiltrating PN while on selumetinib treatment. In both cases, the superficial tumors were safely resected by plastic surgery with good cosmetic outcome and no regrowth has been observed. These cases suggest that superficial and diffuse PN may respond differently to MEKi therapy than typical multi-nodular/fascicular PN. In these cases, plastic surgery may be a treatment option to safely improve appearance and selumetinib treatment may prevent regrowth. Additional prospective evaluations will be needed to determine if structural PN differences influence response to MEKi treatment and to further assess the role of surgery on superficial PN. 

{{Plexiform neurofibroma}}

Nicole M. Brossier, MD, PhD

Washington University in St. Louis

Disclosures: None.

Abstract: Children with the cancer predisposition syndrome Neurofibromatosis Type 1 (NF1) have ~15-20% risk of developing low-grade gliomas of the optic pathway (NF1-OPG). Recent studies suggest that nonrandom factors, such as the specific germline NF1 mutation and environmental exposures, may increase this risk. One potential factor is maternal obesity, as our laboratory has recently demonstrated that maternal obesogenic diet increased Nf1-OPG penetrance and accelerated tumor formation in murine models, consistent with data from the general population showing that children of obese mothers develop several types of tumors at increased rates. Previous research has also shown that an obesogenic diet triggers low-grade inflammation, which can promote tumor formation in other model systems. Based on these observations, we hypothesized that maternal obesogenic exposure would modulate the risk of Nf1-OPG formation through increased inflammatory signaling. Methods: We exposed both dams and offspring to an obesogenic high-fat, high-sucrose diet (Ob) to mimic typical U.S. dietary conditions, with control diet (CD)-exposed animals serving as a comparison. At 6 weeks, we analyzed serum and optic nerves from both groups. Results: Immunohistochemistry revealed increased microglial and T-cell infiltration in the optic nerves of Ob-exposed NF1-OPG mice, suggesting a potential increase in inflammatory stimuli. However, there were no significant differences in circulating inflammatory cytokine levels between the two groups, as measured by cytokine array and confirmatory ELISA. Similarly, quantitative PCR analysis of optic nerves did not show upregulation of inflammatory cytokines in the tumor microenvironment. RNA sequencing of optic nerves from Ob- and CD-exposed mice also revealed no evidence of increased inflammatory signaling pathways. Instead, Ob-exposed nerves exhibited upregulation of pathways related to neuronal signaling and brain development. Conclusion: Our findings suggest that maternal obesogenic exposure does not increase the risk of NF1-OPG development through heightened inflammatory cytokine pathways. However, the observed alterations in neuronal signaling pathways raise the possibility that crosstalk between these pathways and tumor cells may contribute to tumor formation.

Full Author List: Gemma Martin, BS; Sabiha Bano, BS; Vasavi Koneru, MS; Manish Adhikari, PhD; Reagan Winegar; Nicole M. Brossier, MD, PhD

Funding: Neurofibromatosis Therapeutic Acceleration Program (NTAP; 2005106568 to NMB) and St. Louis Hospital Children’s Foundation (DR2019688 to NMB).

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Maria Luisa Garau¹, Kimia Hashemi², Sara Zanchetti¹, Monica Forzan¹, Francesca Boaretto¹, Alessandra Friso¹, Elisabetta Zanoletti³, Katharina Wimmer² and Eva Trevisson^1,4

¹ Clinical Genetics Unit, Dept of Women’s and Children’s Health, University of Padova. Italy

² Human genetics department, Medical University Innsbruck, Austria

³ Dept of Neurosciences, University of Padova. Italy 

⁴ Institute of Pediatric Research IRP, Fondazione Città della Speranza, Padova, Italy 

Disclosure: The authors declare no conflict of interest

Funding: none. 

Background: NF2-related Schwannomatosis (NF2-SCH) is an autosomal dominant condition predisposing to the development of schwannomas and multiple meningiomas. The detection of bilateral vestibular nerve schwannomas is considered enough to establish a clinical diagnosis of NF2-related Schwannomatosis. NF2-SCH caused by ring chromosome 22 is a rare entity and patients usually present a more complex phenotype. 

Methods: We report a case of a 37 years old male who was referred to our center following diagnosis of schwannoma of the left VIII cranial nerve and magnetic resonance [WK1] [G2] anomaly compatible with bilateral formation. The patient's brother and mother presented the same clinical and radiological picture. In addition, the patient spermiogram revealed azoospermia. We performed next generation sequencing of a schwannomatosis gene panel, NF2 MLPA, cytogentic analysis, fluorescence in situ hybridization, array analysis, and PacBio long-read sequencing (LRS).

Results: Cytogenetic analysis uncovered the presence of ring chromosome 22 and a translocation t(1;22)(p36.3;q13.1) while no underling NF2 PV was identified by NGS and MLPA. Further characterization by FISH analysis and LRS uncovered the exact translocation breakpoint on chromosome 22 and 1 and showed that this translocation and the ring 22 formation led to constitutional ~104kb  deletion on chromosome 22. Molecular characterization of DNA from the left VIII cranial nerve schwannoma showed loss of the entire chromosome 22 except for the 22q13.1-13.33 region translocated to chromosome 1.

Conclusions: Azoospermia and peripheral nervous system tumors are described in patients with complex rearrangements involving chromosome 22. However, application of chromosomal analysis is rarely reported in cases of familial NF2-SCH. Our case underlines the importance of considering cytogenetic analysis in case of vertical transmission mother-to-offspring of a purely NF2 phenotype and shows the potential of PacBio LRS to resolve the exact nature of complex rearrangements.

 [WK1]Spell out abreviation

 [G2]Metterei direttamente bilateral…

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Varun Arunachalam Chandran¹, Nils Muhlert², Stavros Stivaros³, Grace Vassallo⁴, Caroline Lea-Carnall², Shruti Garg¹

1.) Division of Psychology and Mental Health, Faculty of biology, medicine and health, University of Manchester, United Kingdom

2.) Division of Psychology, Communication & Human Neurosciences, Faculty of biology, medicine and health, University of Manchester, United Kingdom

3.) Division of Informatics, Imaging & Data Sciences, Faculty of biology, medicine and health, University of Manchester, United Kingdom

4.) Manchester Centre for Genomic Medicine, Manchester University NHS Foundation Trust, Oxford Road, Manchester, United Kingdom

It has been hypothesised that children with NF1 show an aberrant brain-behaviour relationship linked to Autism Spectrum Conditions (30%) and Attention Deficit Hyperactivity Disorder (50%) (Garg et al., 2013). Previous studies have shown T2-white matter hyperintensities (in the thalamus and basal ganglia) and widespread white matter microstructure differences (diffusion MRI) in NF1 suggesting aberrant myelination (Karlsgodt et al., 2012, Feldmann et al., 2010). However, no studies have investigated quantitative myelin in NF1 yet. Objectives: (i) Compare quantitative myelin levels across the grey and white matter at whole brain and lobar level between NF1 and neurotypicals, and determine (ii) Relationship between estimated myelin levels and working memory (WM) performance in children with NF1.

Participants include forty-eight children and adolescents with NF1 (age: 11-18 years; gender: 25 males and 23 females). T1-weighted and T2-weighted structural images were acquired using a Philips 3T MRI scanner. One hundred and sixty-eight (age and gender-matched) control structural MRI images were collected from the Human Connectome Project. Visuospatial n- back tasks were used to ascertain WM performance. The image preprocessing pipeline includes bias-field correction, coregistration, calculation of T1W/T2W maps, segmentation, spatial normalisation and calculation of GM/WM metrics. ANCOVA and Pearson’s partial correlation was used to test the whole brain and lobar level GM/WM myelin in the case-control and correlation analysis in NF1 respectively, while the effects of age and gender were controlled. In addition, age and sex-related myelin differences were tested between NF1 and controls.

Our results showed significantly reduced grey matter/white matter at whole brain level (F=28.165, p<0.001), and the frontal (F=6.906, p<0.009), temporal (F=27.548, p<0.001), parietal (F=7.440, p<0.007) and occipital (F=28.906, p<0.001) lobes in NF1 relative to neurotypical controls. Stratified age- and sex-related myelin differences were found significantly low in NF1 than the controls at whole brain level. However, no significant correlation was found between myelin and WM performance either at whole brain or lobar level in NF1.

Our findings show that quantitative GM/WM myelin differences are consistent with previous animal model studies in NF1 (López-Juárez et al., 2017). Reduced GM/WM myelin in the whole brain and the frontal, temporal, parietal and occipital lobes suggest NF1 gene mutation may affect oligodendrocytes producing myelin in NF1. Low quantitative myelin may manifest as a consequence of loss of neurofibromin in NF1 leading to reduction in mature oligodendrocytes/decompaction of myelin (Mayes et al., 2013). T1W/T2W ratio is a useful neuroimaging biomarker to identify quantitative myelin differences in NF1.

Keywords: Neurofibromatosis-1; Myelin; Grey/White matter; T1W/T2W ratio; Working memory performance.

References:

Garg, S., Green, J., Leadbitter, K., Emsley, R., Lehtonen, A., Evans, D.G. and Huson, S.M., 2013. Neurofibromatosis type 1 and autism spectrum disorder. Pediatrics, 132(6), pp.e1642-e1648.

Feldmann, R., Schuierer, G., Wessel, A., Neveling, N., & Weglage, J. (2010). Development of MRI T2 hyperintensities and cognitive functioning in patients with neurofibromatosis type 1. Acta Paediatrica, 99(11), 1657-1660.

Karlsgodt, K.H., Rosser, T., Lutkenhoff, E.S., Cannon, T.D., Silva, A. and Bearden, C.E., 2012. Alterations in white matter microstructure in neurofibromatosis-1.

López-Juárez, A., Titus, H.E., Silbak, S.H., Pressler, J.W., Rizvi, T.A., Bogard, M., Bennett, M.R., Ciraolo, G., Williams, M.T., Vorhees, C.V. and Ratner, N., 2017. Oligodendrocyte Nf1 controls aberrant notch activation and regulates myelin structure and behavior. Cell reports, 19(3), pp.545-557.

Mayes, D.A., Rizvi, T.A., Titus-Mitchell, H., Oberst, R., Ciraolo, G.M., Vorhees, C.V., Robinson, A.P., Miller, S.D., Cancelas, J.A., Stemmer-Rachamimov, A.O. and Ratner, N., 2013. Nf1 loss and Ras hyperactivation in oligodendrocytes induce NOS-driven defects in myelin and vasculature. Cell reports, 4(6), pp.1197-1212.

Funding and support:

This study is funded by National Institute for Health and Care Research-Biomedical Research Centre (NIHR-BRC), Manchester under the Mental Health theme. In addition, the abstract submission is supported by Dr Shruti Garg from the University of Manchester and data collection from the study team members of the Synaptic Plasticity in Neurodevelopment (SPIN) Lab.

Ethical Requirements:

This study was conducted after receiving approval from the NHS research ethics committee.

Declaration:

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this abstract.

Abstract submission categories (options): 

1.1.7      Biomarkers

2.2          Cognition/Behavioural/Learning

{{Cognition / Behavioral / Learning}}

Purpose: Neurofibromatosis type 1 (NF1) is an uncommon genetic disorder associated with plexiform neurofibromas (PNs). Therapeutic options have historically been limited. However, recent clinical data have shown expanded and improved outcomes with the use of MEK inhibitors. The purpose of this study was to assess the educational impact of a continuing medical education (CME) activity on the multidisciplinary (MDT) clinicians’ understanding of ongoing data and application to care.  

Methods:  The educational intervention consisted of a text-based CME/CE-certified expert interview. Educational impact was measured with a pre-/post-education assessment including multiple-choice knowledge questions. Data from relevant learners who completed pre- and/or post-education assessments were included. Relative changes in percentage of correct responses were used to measure improvement in knowledge. A McNemar test assessed significant levels of changes reported, with P values < .05 considered statistically significant. The activity launched Nov. 21, 2024. Data were collected through Feb. 2025.

Results: Improvements were seen after education across clinician groups by learning theme, with statistical improvements observed throughout for neurologists, pediatricians, and surgeons. Nurses did not see significant gains in knowledge.  

Conclusions: The study demonstrates the overall success of an online CME activity in enhancing MDT members’ understanding of the impact of evolving clinical evidence in the care of patients with NF1-associated PNs. Despite improvements, it also underscores the critical remaining gaps collectively and by specialty. Additional educational activities and analyses are warranted to further clarify the link between education and patient care improvements, particularly as data continue to shape standards of care.

{{Plexiform neurofibroma}}

Amelia Khan, BSc (Hons), mCSP; Mandy Myers MSc BSc (Hons)/RGN, RSCN, SCPHN; Katrina Kettle BSc (Hons/RN); Jamie Gibson, BSc (Hons), FHEA, mCSP.  Guys and St Thomas’ NHS Foundation Trust, UK

Disclosure: Alexion AstreZeneca Rare Diseases sponsorship to attent CTF Conference for Mandy Myers

Purpose:

Functional Neurological Disorders (FND) are one of the most common diagnoses made in Neurology clinics (1). FND refers to symptoms of altered motor or sensory function with no organic cause, resulting in symptoms such as limb weakness, numbness, pain, and seizures (2). Recent studies have shown that the incidence of FND within paediatrics is estimated at 1.3 to 6.0 per 100,000, and 10% of all FND diagnosis are made alongside a pre-existing neurological condition (2). Whilst consensus on how to rehabilitate adults with FND has been documented (3), there is no guidance on how to rehabilitate paediatric patients with additional conditions such as Neurofibromatosis Type 1 (NF1).

Designing, testing and implementation of nascent treatment approaches are often blurred by insufficient reporting of treatment theory (4), that is, how active ingredients of a treatment lead to change in aspects of a patients’ function (5). The rehabilitation treatment specification system (RTSS) provides a countermeasure to this by challenging the clinician to specify treatment theory through its common language and tripartite structure (6). We sought to utilise the RTSS to aid reasoning and specification of the treatment theory of a rehabilitation programme for a paediatric NF1 patient living with FND.

Methods: 

Case report presentation of the physical therapy rehabilitation of a 17-year-old female with NF1. She presented with medical complications including an extensive optic pathway glioma treated with chemotherapy, shunted hydrocephalus and post-operative right-sided cerebral haemorrhage. Her presenting issues included a five-year history of right sided functional weakness resulting in wheelchair mobility, with a background of anorexia and an unexpected family bereavement. The RTSS was applied a priori to reason and specify treatment theory of the rehabilitation programme. 

Results: 

Treatment components included: improving muscular endurance via hypertrophic change and increasing muscle unit activation; and improving selective trunk control by using distraction ingredients to override functional pathways and aid in neuroplastic change. Distraction ingredients included therapeutic finger painting, popping bubbles and catching objects out of her base of support. All task ingredients encouraged trunk flexion, side-flexion and rotation; tasks which she was unable to perform without distraction ingredients. Over several sessions she was able to progress through her ingredient’s difficulty, leading to increased postural control. Volitional components encouraged independent performance of mindfulness. 

Conclusion: 

This case report demonstrates a specified rehabilitation for FND in an NF1 paediatric patient, through use of the RTSS, a reasoned rehabilitation intervention for this population is available for replication, modification and testing. 

References:

1. Stone, J., Carson, A., Duncan, R., Roberts, R., Warlow, C., Hibberd, C., Coleman, R., Cull, R., Murray, G., Pelosi, A., Cavanagh, J., Matthews, K., Goldbeck, R., Smyth, R., Walker, J., & Sharpe, M. (2010). Who is referred to neurology clinics?--the diagnoses made in 3781 new patients. Clinical neurology and neurosurgery, 112(9), 747–751. https://doi.org/10.1016/j.clineuro.2010.05.011
2. Yong, K., Chin, R. F. M., Shetty, J., Hogg, K., Burgess, K., Lindsay, M., McLellan, A., Stone, J., KamathTallur, K., & Edinburgh Paediatric FND Study Group (2023). Functional neurological disorder in children and young people: Incidence, clinical features, and prognosis. Developmental medicine and child neurology, 65(9), 1238–1246. https://doi.org/10.1111/dmcn.15538
3. Nielsen, G., Stone, J., Matthews, A., Brown, M., Sparkes, C., Farmer, R., Masterton, L., Duncan, L., Winters, A., Daniell, L., Lumsden, C., Carson, A., David, A. S., & Edwards, M. (2015). Physiotherapy for functional motor disorders: a consensus recommendation. Journal of neurology, neurosurgery, and psychiatry, 86(10), 1113–1119. https://doi.org/10.1136/jnnp-2014-309255
4. Dijkers M. P. (2019). An End to the Black Box of Rehabilitation?. Archives of physical medicine and rehabilitation, 100(1), 144–145. https://doi.org/10.1016/j.apmr.2018.09.108
5. Whyte J. (2014). Contributions of treatment theory and enablement theory to rehabilitation research and practice. Archives of physical medicine and rehabilitation, 95(1 Suppl), S17–23.e2. https://doi.org/10.1016/j.apmr.2013.02.029
6. Hart, T., Dijkers, M. P., Whyte, J., Turkstra, L. S., Zanca, J. M., Packel, A., Van Stan, J. H., Ferraro, M., & Chen, C. (2019). A Theory-Driven System for the Specification of Rehabilitation Treatments. Archives of physical medicine and rehabilitation, 100(1), 172–180. https://doi.org/10.1016/j.apmr.2018.09.109

{{Cognition / Behavioral / Learning}}

Chong, Wai Chin^1,2, Subramaniam, Bavani^1,2, Roy, Debasish^1,2, Mizoguchi, Sean^1,2, Yadavilli, Sridevi^1,2, Bornhorst, Miriam³, Packer, Roger¹, Nazarian, Javad⁴.

1. Brain Tumor Institute, Center for Cancer and Immunology Research, Children’s National Hospital, Washington, District of Columbia, United States of America.
2. Center for Genetic Medicine, Children’s National Research and Innovation Campus, Children’s National Hospital, Washington, District of Columbia, United States of America. 
3. Department of Pediatric Hematology-Oncology, Ann & Robert H Lurie Children’s Hospital of Chicago, Illinois, United States of America.  
4. Department of Oncology, Children’s Research Center, University Children’s Hospital Zurich, Zurich, Switzerland.   

Background: Neurofibromatosis type 1 (Nf1) is a common autosomal dominant disorder that is caused by mutations in neurofibromin 1 (NF1) gene and affecting 1 in 3000 children worldwide. Nf1 patients have higher risk of developing different tumors, including glioma. Importantly, comprehensive genomic analysis revealed high frequency of cyclin-dependent kinase inhibitor 2A (CDKN2A) inactivation in malignant NF1 mutant glioma as compared to its benign counterpart, suggesting its role in driving malignant transformation in NF1 mutant glioma. However, little is known about this CDKN2A dependent malignant transformation, due to lack of appropriate preclinical glioma models for comprehensive profiling. To address this knowledge gap, we utilized CRISPR editing platform to generate isogenic lines with different combinations of NF1 and CDKN2A mutation, aiming to investigate the role of CDKN2a in malignant transformation of NF1 mutant glioma.

Method: We utilized CRISPR technology to knock out NF1 and CDKN2A genes in NF1 and CDKN2A wildtype RES186 glioma cell. The cell growth and migration potential of the generated isogenic lines were assessed and compared using cell proliferation and Boyden chamber invasiveness assays. The mRNA-seq was performed on these isogenic lines to compare the global transcriptomic changes and enriched molecular events. Further, these isogenic lines were mcherry-luciferase labelled and engrafted in the NSG immunodeficient murine models to evaluate their proliferation and invasiveness potential under in-vivo condition.      

Results: Our findings revealed that RES186 NF1^KO has similar growth and invasive rate as its parental line. Conversely, RES186 CDKN2A^KO possessed both increased growth and invasive rate. Strikingly, the RES186 NF1^KO CDKN2A^KO has the highest growth and invasive rate as compared to its parental and other isogenic lines. The global transcriptomic analysis further revealed additional enrichment in collagen degradation process solely in RES186 NF1^KO CDKN2A^KO cells. These results highlight the interplay of NF1 and CDKN2a in regulating the growth and invasiveness dynamics of glioma cells.

Conclusion: In conclusion, our preliminary study suggests that the concurrent loss of NF1 and CDKN2a enhanced growth and invasiveness in gliomas. The established CRISPR isogenic model will provide a platform to explore the collaborative role of these genetic alterations in tumor transformation.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Bavani Subramaniam, PhD^1,2, Wai Chin Chong, PhD^1,2, Surajit Bhattacharya, PhD², Gary Cunnningham, PhD², Roger Packer, MD^1,2, Miriam Bornhorst, MD³, Javad Nazarian, PhD⁴.

¹Brain Tumor Institute, Center for Cancer and Immunology Research. Children’s National Hospital, Washington, DC, USA. 

²Center for Genetic Medicine Research, Children’s National Hospital, Washington, DC, USA. 

³Northwestern University Feinberg School of Medicine, Ann & Robert H. Lurie Children’s Hospital of Chicago, IL, USA.

⁴Department of Oncology, Children's Research Center, University Children's Hospital Zurich, Zurich, Switzerland 

Neurofibromatosis type 1 (NF1) is a tumor predisposition syndrome that is characterized by hyperactivation of the RAS/MAPK signaling pathway. Our analysis of NF1-associated high grade gliomas revealed that 57% of these tumors harbor homozygous deletions in chromosome 9p21.3, encompassing the tumor suppressor genes CDKN2A, CDKN2B, and MTAP. Methylthioadenosine phosphorylase (MTAP) is critical for the methionine salvage pathway and its loss leads to increased vulnerability to PRMT5 inhibition. PRMT5 methylation activity is also associated with multiple kinases in the RAS/MAPK signaling pathway, warranting further investigation on a combined PRMT5 and RAS/MAPK inhibition therapy.

Cell lines (n= 4) with loss-of-function mutations in the NF1, CDKN2A/B, and MTAP genes and CRIPSR-Cas9 MTAP-knockout (MTAP-KO) cells were first validated for MTA accumulation using liquid chromatography-mass spectrometry (LC-MS). Cell viability assays were then performed using a brain-penetrant, MTA-cooperative PRMT5 inhibitor, TNG908. Specific inhibition of PRMT5 was confirmed by immunoblotting for symmetric dimethylarginine (SDMA) expression and genes involved in the RAS/MAPK signaling pathway. Additionally, the impact of TNG908 on cell cycle progression and apoptosis was assessed using flow cytometry. Combination of TNG908 with MEK inhibitors was also assessed.

MTAP-deleted and MTAP-KO cells showed increased MTA accumulation compared to wild-type MTAP (MTAP-WT) cells, resulting in significant sensitivity towards TNG908. PRMT5 inhibition was confirmed by reduced SDMA expression in MTAP-KO cells but not in MTAP-WT cells, indicating TNG908’s MTA-dependent inhibition activity. PRMT5 inhibition also significantly increased (p<0.05) the proportion of cells in the G2/M phase of the cell cycle and enhanced apoptotic activity (p<0.05). Combinatorial inhibition using TNG908 and Binimetinib demonstrated significant synergy (ZIP score= 14.36) in MTAP-KO cells and additive activity in TM31 cells (ZIP score= 0.272). 

These findings highlight PRMT5 inhibition as a promising therapeutic strategy for MTAP-deleted NF1 HGGs. Moreover, the combination of TNG908 and Binimetinib shows strong potential and warrants further investigation in vivo.

{{DRUG DISCOVERY AND DEVELOPMENT}}

Kyungmin Ji, George J. Schwenkel,  Yong Xu, Zhenggang Zhang, and Michael Chopp

Departments of Neurology, Henry Ford Health, Detroit, MI 48202, USA;  Biomedical Engineering, Electrical and Computer Engineering, and Pharmacology, Wayne State University, Detroit, MI 48202, USA. * Correspondence, kji1@hfhs.org

1) Funding: This work is supported by DoD USAMRAA Neurofibromatosis Research Program-New Investigator Award (W81XWH2210564) to Kyungmin Ji. 

2) Disclosures: None

3) Purpose: Neurofibromatosis Type 1 (NF1) plexiform neurofibroma (hereafter called pNF1) is present in ~50% of NF1 patients, and is a complex tumor composed of abnormal Schwann cells and cells of the surrounding tumor microenvironment (TME). Fibroblasts, the most abundant cell types within the pNF1 TME, are implicated in tumor formation and regulate extracellular matrix (ECM) stiffness by producing excessive amounts of collagen. Using 3D pNF1 culture models, we previously demonstrated that fibroblasts have a critical role in tumor growth and invasiveness through their secretome. However, it is poorly understood how fibroblasts affect a long-term drug response of pNF1. Delineating molecular mechanisms of drug resistance is crucial to designing new therapies to improve therapeutic outcome in pNF1. In the present study, we will investigate how fibroblasts affect drug resistance of pNF1 via their secretome and/or their modulation of ECM stiffness.

4) Methods: As a pre-clinical model to evaluate drug resistance of pNF1 tumor cells, we used our unique 3D/4D (3D + real-time) cultures of human pNF1 cells in the context of their TME, i.e., fibroblasts in the present study, at a ratio of 1 tumor: ½ or ¼ fibroblasts grown in our patented culture chips (TAME chips; Patent #: US 10,227,556 B2). Our 3D/4D culture model supports growth of the 3D cultures, live-cell confocal imaging in real-time (4D) and non-invasive, high-content analysis and drug testing over extended long-term culture periods. Two immortalized human pNF1 cell lines (ipNF95.11b C and ipNF05.5; Nf1^-/-) and patient-derived primary fibroblasts (Nf1^+/-) are used for 3D cultures. FDA-approved selumetinib and mirdametinib are used for testing drug resistance of pNF1. We employed: 1) 3D parallel cocultures of pNF1 tumor cells and fibroblasts in which the two cell types share media without direct contact to test effects of fibroblast-derived secretome, and 2) two kinds of 3D ECM hydrogels, naturally derived and synthetic ones, to test effects of different ECM stiffness produced mainly by fibroblasts. We used live-cell imaging and 3D quantitative analysis to evaluate drug responses in pNF1 tumor cells. 

5) Results: We observed a greater viability of pNF1 tumor cells in parallel cocultures with fibroblasts than in pNF1 monocultures treated with selumetinib or mirdametinib. Moreover, pNF1 tumor cells grown in 3D ECM hydrogel with higher stiffness (7 kPa) are more resistant to the drug treatment compared to ones in 3D ECM hydrogel with lower stiffness (1.5 kPa). The pNF1 tumor cells cultured with fibroblast-derived conditioned media or grown in 3D ECM hydrogel with higher stiffness displayed more increased level of p-glycoprotein, a major mechanism of drug resistance induced by actively pumping drugs out of cells. 

6) Conclusions: Our results demonstrate that fibroblasts significantly increase drug resistance of tumor cells via their secretome, and the modulation of ECM stiffness and p-glycoprotein is a potential mechanism underlying fibroblast-mediated drug resistance in pNF1. The results of this study may lead to improvement of the therapeutic outcome of pNF1 patients by identifying means to ameliorate drug resistance of pNF1.

TUMOR BIOLOGY AND DISEASE MECHANISM

• Authors: Full name, academic credentials and institutional affiliation for all authors.

Jared Zhao, B.S., Johns Hopkins University School of Medicine.

Bryan Ward, M.D., Johns Hopkins University School of Medicine.

George Liu, M.D., Johns Hopkins University School of Medicine.

Charles C. Della Santina, Ph.D., M.D., Johns Hopkins University School of Medicine.

• Funding: Indicate the granting agency and mechanism whose fiscal support made the research possible.

No funding

• Disclosures: Any relevant financial relationships for author(s). If chosen for CME accreditation, additional disclosure documentation may be required.

No disclosures relevant to this case.

• Purpose: Statement of why the study was conducted.

This case report aims to present a facial nerve schwannoma manifesting as unilateral hearing loss, discuss the erosion of facial nerve schwannoma into the inner ear, and how this erosion influences the decision-making process regarding surgical intervention.

• Methods: Brief description of how the research was conducted.

This case report was compiled by thoroughly reviewing the patient’s medical chart—including clinical notes, imaging studies, and operative reports—and synthesizing these findings with current peer-reviewed literature on facial nerve schwannomas.

• Results: Summary of key findings, to include text and/or images/tables/graphs.

A young adult male presented with a 10-month history of intermittent left ear fullness and hearing loss, which began acutely after a presumed episode of acute otitis media. Audiometry indicated left-sided conductive hearing loss, yet all cranial nerves were functioning normally. Both otomicroscopy and CT scans suggested a middle ear mass involving the ossicles, initially raising suspicion for cholesteatoma. However, MRI revealed that the lesion was more likely a facial nerve schwannoma, with CT imaging revealing the tumor extending from the mastoid region to the sinus tympani, eroding into the horizontal semicircular canal, and encompassing the stapes. Surgical decompression of the facial nerve was performed, and a biopsy confirmed a schwannoma. Genetic testing (for NF2, SMARCB1, and LZTR1) was negative. A follow-up spine MRI detected a 1 mm peripheral nerve sheath tumor in the cauda equina, meeting the criteria for schwannomatosis-NOS (not otherwise specified). Postoperatively, the patient developed mild lower facial weakness and tinnitus in the left ear. Despite stable tumor size on imaging, conductive hearing loss persisted due to the involvement of the ossicular chain. There is also a recognized risk of future sensorineural hearing loss should the tumor continue to grow.

• Conclusion: Interpretation of results and the implications/significance of the study.

This case underscores the importance of thorough imaging and vigilant monitoring to distinguish facial nerve schwannomas from common middle ear pathologies, as bony erosion can result in conductive or sensorineural hearing and vestibular loss. Detecting a small peripheral nerve sheath tumor in the cauda equina highlights broader syndromic considerations and the need for long-term surveillance.

Schwannoma

Julia R. Plank Ph.D.¹, Elveda Gozdas Ph.D.¹, Jennifer Bruno Ph.D.¹, Chloe A. McGhee B.A.¹, Hua Wu Ph.D.², Mira M. Raman M.S.¹, Manish Saggar Ph.D. ¹, Tamar Green M.D.¹

¹Division of Interdisciplinary Brain Sciences, Department of Psychiatry and Behavioral Sciences, 1520 Page Mill Road, Palo Alto, CA 94304, USA 

²Center for Cognitive and Neurobiological Imaging, Stanford University, Stanford, CA 94305, USA 

Purpose: Preclinical models of neurofibromatosis type-1 (NF1) show aberrations in myelin content, however, there is a need for in vivo techniques that can detect myelin alterations in clinical populations (1,2). We investigated quantitative T1 mapping (QT1) as a clinically feasible tool for measuring myelin content in children with NF1. Methods: Fifteen children with NF1 (M_age=9.59, SD_age=2.24; 8 male) and 23 typical developing (TD; M_age=10.2, SD_age=3.48; 11 male) peers participated in this prospective study. We collected structural, multi-shell diffusion-weighted, and QT1 MRI data on a GE Premier 3T scanner. QT1 maps were produced using open-source Python code. The macromolecular tissue volume (MTV), a proxy for myelin concentration, was extracted from 39 white matter tracts using TRActs Constrained by UnderLying Anatomy within FreeSurfer (3). We extracted R1 values from 360 brain regions per subject based on the Human Connectome Project multi-model parcellation (4). Previous work shows the R1 (1/T1) and MTV measurements generated by QT1 are reliable measures of myelin content (5,6). We compared MTV in the 39 white matter tracts and cortical R1 in 360 regions between-groups using analyses of covariance, including age and sex as covariates in R 4.4.1 (R Core Team, 2024). Results: Thirty-six of 39 tracts showed significantly increased MTV in NF1 relative to TD (p_FDR <.05), indicating elevated white matter myelin in NF1 (Figure 1). The largest effect sizes were found in the genu of the corpus callosum (p_FDR<.001, Cohen’s d=1.40), the right (p_FDR<.001, d=1.40) and left cingulum bundle-dorsal (p_FDR<.001, d=1.37), and the left uncinate fasciculus (p_FDR<.001, d=1.37). We did not find any significant differences in cortical R1 between NF1 and TD, suggesting similar levels of cortical myelin content in both groups. Conclusion: Our findings suggest increased white matter myelin content in NF1 relative to TD. Mouse models of NF1 shows proliferation of oligodendrocyte precursor cells (7). We expect that the affected oligodendrocyte lineage may contribute to the elevated myelin observed in this work, however, further studies are needed to uncover the exact mechanism. The significance of these findings warrants further investigation with a larger sample size and greater statistical power. Myelin is often linked to cognitive functioning, thus future studies should investigate relationships between myelin content and cognition in NF1 (8).  Our results and the short scan time required (approximately 3 minutes for a QT1 acquisition) suggest QT1 may be a promising technique for clinical translation.

Funding: This project was supported by grants: Contract grant sponsor: National Institute of Child Health and Human Development; Contract grant number: 123752K23 and R01HD108684 to T.G; The Stephen Bechtel Endowed Faculty Scholar in Pediatric Translational Medicine, Stanford Maternal & Child Health Research Institute to T.G. Contract grant sponsor: Neurofibromatosis Therapeutic Acceleration Program (NTAP) at the John Hopkins University School of Medicine to T.G. Its contents are solely the responsibility of the authors and do not necessarily represent the official views of The Johns Hopkins University School of Medicine. The funding sources had no role in the study design, collection, analysis, and interpretation of the data.

References

1. López-Juárez A, Titus HE, Silbak SH, Pressler JW, Rizvi TA, Bogard M, et al. Oligodendrocyte Nf1 controls aberrant Notch activation and regulates myelin structure and behavior. Cell Rep. 2017 Apr 18;19(3):545–57.
2. de Blank P, Nishiyama A, López-Juárez A. A new era for myelin research in Neurofibromatosis type 1. Glia. 2023 Dec;71(12):2701–19. 
3. Yendiki A, Panneck P, Srinivasan P, Stevens A, Zöllei L, Augustinack J, et al. Automated probabilistic reconstruction of white-matter pathways in health and disease using an atlas of the underlying anatomy. Front Neuroinform. 2011 Oct 14;5:23.
4. Glasser MF, Coalson TS, Robinson EC, Hacker CD, Harwell J, Yacoub E, et al. A multi-modal parcellation of human cerebral cortex. Nature. 2016 Aug 11;536(7615):171–8.
5. Mezer A, Yeatman JD, Stikov N, Kay KN, Cho N-J, Dougherty RF, et al. Quantifying the local tissue volume and composition in individual brains with magnetic resonance imaging. Nat Med. 2013 Dec;19(12):1667–72.
6. Yeatman JD, Wandell BA, Mezer AA. Lifespan maturation and degeneration of human brain white matter. Nat Commun [Internet]. 2014; Available from: https://www.nature.com/articles/ncomms5932
7. Bennett MR, Rizvi TA, Karyala S, McKinnon RD, Ratner N. Aberrant growth and differentiation of oligodendrocyte progenitors in neurofibromatosis type 1 mutants. J Neurosci. 2003 Aug 6;23(18):7207–17.
8. Buyanova IS, Arsalidou M. Cerebral White Matter Myelination and Relations to Age, Gender, and Cognition: A Selective Review. Front Hum Neurosci. 2021 Jul 6;15:662031.

Other

PRESENTING AUTHOR: J. Elliott Robinson

FULL AUTHOR LIST: Zoe Cappel, Ronald Waclaw, PhD, Gregory Schwartz, PhD; and J. Elliott Robinson, MD, PhD

Rasopathy Program, Division of Experimental Hematology and Cancer Biology, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH, 45229, USA

Department of Pediatrics, University of Cincinnati, Cincinnati, OH, 45229, USA

Department of Ophthalmology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA

DISCLOSURES: None

ABSTRACT

Neurofibromatosis type 1 (NF1) is an autosomal dominant disorder caused by loss-of-function mutations in the NF1 gene that encodes the neurofibromin protein. Neurofibromin loss leads to increased signaling in the Ras-mitogen activated protein kinase (MAPK) pathway (Ras-Raf-MEK-ERK) that is critical for nervous system development and function. The neurocognitive symptoms of neurofibromatosis type 1 (NF1) include impaired executive functioning, autistic features, speech and language delays, and sensory processing abnormalities. NF1 patients also have a high incidence of attention deficit/hyperactivity disorder (ADHD), which is associated with diminished ability to suppress distractive stimuli and increased sensitivity to aversive sensory cues. Here, we show that mice modeling NF1 (Nf1+/- mice) are hypersensitive to aversive visual stimuli, such as threatening looming discs that simulate predator approach from above. This phenotype was also observed in a novel mouse model of MAPK1-related Rasopathy that contains a patient-derived ERK2 gain-of-function mutation (Mapk1A174V/+ mice) but not in mice with increased mTOR signaling (Tsc2+/- mice). Conditional expression of an activated MEK1 mutant in cortical excitatory neurons and glia was sufficient to enhance threat reactivity, whereas MAPK activation in the midbrain, which plays a critical role in innate behavioral responses to visual threats, or in forebrain interneurons was not. Our findings were not caused by changes in retinal sensitivity to light given that Nf1+/- mice exhibited no differences in baseline or light-evoked retinal ganglion cell (RGC) firing in intact retinal preparations ex vivo or in physiological assays that test RGC function in vivo. Taken together, our results suggest that increased MAPK activation in the Nf1-deficient cerebral cortex alters top-down control of subcortical sensory processing centers, which, in turn, enhances visual threat reactivity. Currently, we are employing circuit- and cell-type specific Nf1 knockout models and calcium imaging techniques to refine our understanding of the effects of neurofibromin loss and MAPK pathway activation on attentional and sensory function.

FUNDING: This work was directly supported by NINDS R01NS126108, a SFARI Bridge to Independence Award, and a Cincinnati Children’s Trustee Grant Award to JER. 

TUMOR BIOLOGY AND DISEASE MECHANISM

Emily White, MDPhD Candidate, Department of Pediatrics, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, IN.

Full List of Authors: Emily E. White, Marisa D. Ciesielski, Brooke E. Hickey, Hannah N. Leffew, Dana K. Mitchell, Henry E. Mang, Melissa L. Fishel, Laura M. Snell, Eric A. Albright, Andrew E. Horvai, D. Wade Clapp, Steven D. Rhodes

Disclosures: SDR has served as advisor and independent contractor for SpringWorks Therapeutics and Practice Point Communications for activities unrelated to these studies. 

Purpose: Atypical neurofibromas exhibit increased signatures of immune cell surveillance and T cell infiltration, but the role of T cells in governing malignant transformation of MPNST precursor lesions remains unclear. We hypothesized that effector CD4+ and/or cytotoxic CD8+ T cells are critical in preventing malignant transformation of plexiform and atypical neurofibroma precursors. The goal of this work was to address a critical gap in our understanding of how T cell subsets influence the behavior of MPNST and their precursor lesions.

Methods: Single cell RNA sequencing (scRNAseq) with sub-type anchor correction for alignment in Seurat (STACAS) was utilized to integrate multiple scRNAseq datasets and characterize the functional states of infiltrating T cells in human NF1-PNSTs. Effector cytotoxic, helper, regulatory, and exhausted CD4+ and CD8+ T cell subsets were quantified in native tumor tissue by multiplexed immunohistochemistry (IHC). Finally, preclinical mouse models that recapitulate MPNST development from neurofibroma precursors were used to evaluate the impact of CD4+ and CD8+ T cell depletion on tumor progression.

Results: We observed changes in single cell transcriptional programs of infiltrating T cells in human PNSTs with a shift from naïve, effector, and cytotoxic CD4+ and CD8+ T cells in PNF to regulatory (Treg) and exhausted states in MPNST. Furthermore, multi-chromogenic IHC of human PNST showed robust T cell infiltration in atypical neurofibroma with a predominance of CD4+ T helper 1 (Th1) cells versus T cell exclusion with Treg predominance in MPNST. In parallel studies, the ratio of CD4⁺FOXP3⁺ Tregs to total CD4⁺ T cells increased as MPNSTs progressed in immunocompetent C57BL/6 mice engrafted with syngeneic Nf1^+/-;Trp53^+/- MNF463A cells. To assess the functional contribution of T cells in restraining transformation and malignant outgrowth, immunocompetent syngeneic recipient mice were orthotopically engrafted with primary Nf1-Cdkn2a^-/- Schwann cell precursors or MNF463A cells and treated with monoclonal antibodies against CD4, CD8, and the combination versus an IgG2b isotype control (n=8/group). Kaplan-Meier analysis showed that while depletion of CD4+ T cells alone had minimal impact on tumor progression, CD8+ depletion and combined CD4+/CD8+ T cell depletion profoundly accelerated the latency and penetrance of MPNST (p<0.0001, log-rank test).

Conclusions: Interactions between CD4+ and CD8+ T cell subsets are critical in governing malignant transformation of NF1-PNSTs.  Changes in T cell phenotypes across the neurofibroma to MPNST continuum may inform future diagnostic and therapeutic strategies for the early detection and prevention/treatment of malignancy.

Funding: This work was supported by K08NS128266-03 (NIH/NINDS) to SDR and T32CA272370-02 (NIH/NCI) to DWC/MRK.

TUMOR BIOLOGY AND DISEASE MECHANISM

Zhichao Wang^1,2*, Qingfeng Li^1,2, Mingde Liao³, Hao Wu⁴, Xuewen Xu⁵, Guangshuai Li⁶ 

^*Corresponding author: Zhichao Wang, Email address: shmuwzc@163.com

¹Neurofibromatosis Type 1 Center and Laboratory for Neurofibromatosis Type 1 Research, Shanghai Ninth People’s Hospital, Shanghai JiaoTong University School of Medicine.

²Department of Plastic and Reconstructive Surgery, Shanghai Ninth People’s Hospital, Shanghai JiaoTong University School of Medicine.

³The First Affiliated Hospital of Guangxi Medical University.

⁴Xuanwu Hospital, Capital Medical University.

⁵West China Hospital, Sichuan University

⁶The First Affiliated Hospital of Zhengzhou University

Funding: AstraZeneca China

Disclosure: This study was funded by an independent study research grant from Alexion, AstraZeneca Rare Disease. Alexion AstraZeneca Rare Disease reviewed the publication, without influencing the opinions of the authors, to ensure medical and scientific accuracy, and the protection of intellectual property. Authors retained control and final authority over publication content and decisions, including the choice of journal.

Background

Neurofibromatosis type I (NF1) affects about 1 in every 3000 people worldwide. 30-50% NF1 patients develop plexiform neurofibromatosis (PN) which grow rapidly in early childhood and can cause severe complications. This systemic disease imposes a heavy psychosomatic and financial burden on patients and their caregivers. 

Methods

We performed a retrospective analysis of the NF1 patients visited 5 hospitals between Jan 2019 and May 2024. The prevalence, disease characteristics, progression free survival (PFS) and treatment pattern of NF1-PN were assessed.

Results

Totally 2277 NF1 patients were enrolled.

Table 1. Prevalence of NF1 manifestations

51.08% (1163/2277) of NF1 patients developed PN. The median age of patients at PN diagnosis was 14 years old. Head & neck (51.5%) was the most common locations of PN. 55.03% (640/1163) of patients with PN had PN related complications and 26.41% (169/640) of them had more than 1 complication. The most common complications are disfigurement (63.44%), functional abnormality (31.88%) and pain (30.57%). 

22.79% (265/1163) patients received surgery. The mean surgery times was 1.14. 27.69% (72/260) patients had PN complete resected, 28.46% (74/260) patients had PN subtotal resected and 43.85% (114/260) had PN partial resected. 5.67%（66/1163）patients had drug therapy and selumetinib was the mostly used drug(93.9%, 62/66). 56.66% (659/1163) patients had follow-up and 32.56%（380/1163）patients only had follow-up without any treatment. The median PFS of 327 patients with PN evaluated by doctor was 23.36 months (95%CI 22.11~25.36). 173 patients were evaluated as progression, 47.40% of progressions were not realized by patients. 107 patients who received surgery were evaluated by doctors after surgery. The median after-surgery PFS was 23.33 months (95%CI 16.03~27.50). Patients who received complete resection had significant longer PFS than patients who received partial resection (47.44 months vs 23.36 months, HR 0.50, 95%CI 0.30-0.82, P=0.006).

Conclusion 

The retrospective analysis shows PN can be found in 51.08% NF1 patients and can cause severe disease burden. PN is a progressive disease. Early diagnosis, early treatment and regular follow-up are important to NF1-PN. Only a small proportion of PN can be fully resected and partially surgery can’t prolong the PFS of PN. With drug treatment approved for PN, inoperable PN needs new treatment plan.

Authors: Cheng-Jiang Wei, MD^{1, 2}; Zhi-Chao Wang, MD, MPH^{1, 2}; Jun Yang, MD, PhD^{1, 2}; Qing-Feng Li, MD, PhD^{1, 2}

Affiliations: 

1. Neurofibromatosis Type 1 Center and Laboratory for Neurofibromatosis Type 1 Research, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, China.

2. Department of Plastic and Reconstructive Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China.

Funding: 

National Natural Science Foundation of China (82472579; 82102344; 82172228)

Disclosures: 

The authors declared no conflict of interest. 

Abstract：

Purpose: Early differentiation between malignant peripheral nerve sheath tumors (MPNSTs) and plexiform neurofibromas (PNFs) is essential for improving the prognosis of patients with neurofibromatosis type 1 (NF1). Radiological imaging provides a noninvasive method for MPNST screening; however, current manual identification requires substantial expertise from radiologists and is time-consuming. Deep learning models have shown promise in differentiating between benign and malignant lesions. However, traditional deep learning methods face challenges in this context due to interference from the heterogeneity of whole-body backgrounds and limited data availability. This study aimed to develop highly accurate MRI-based deep-learning models for the early automated screening of MPNSTs against complex whole-body background (Figure 1).

Methods: In this study, we collected a Chinese seven-center cohort to address the challenge of limited data, comprising 347 subjects (251 PNF, 96 MPNST) with multiparametric MRI data. A total of 3150 T2-weighted MRI images with typical tumor lesions were selected (2141 PNF, 1009 MPNST). Furthermore, our one-step model incorporated eight normal tissue/organ labels to provide contextual information, offering a solution for tumors embedded in complex backgrounds. To address privacy concerns, we employed a lightweight deep neural network suitable for deployment in hospital settings.

Results: This model demonstrated excellent performance in simultaneous area detection and differential diagnosis (accuracy: 85.71%, threshold: 85%) while requiring less computational power compared to other classical U-Net- and ResNet-based models. Additionally, compared to another one-step model trained exclusively on PNF/MPNST labels, incorporating information from surrounding normal organs/tissues improved diagnostic accuracy from 61.90% to 79.37% for MPNST and from 46.67% to 70% for PNF (Figure 2).

Conclusion: This advancement could represent a significant milestone in the early clinical screening of MPNST and provide a novel approach for the ‘fully automated’ identification of whole-body background tumors, such as metastatic cancers.

Figure 1. A flowchart of the study process of our MRI-based model development

Figure 1 legend. We developed automatic segmentation and classification deep-learning AI models for the differential diagnosis of MPNST from benign PNF. In the training process, we introduced the surrounding information to augment their performance. The Figure 1 was created with Biorender.com.

Figure 2. The results classified by the one-step whole-body tumor identification deep learning model

Figure 2 legend. (a) The confusion matrix of the training process. (b) The correct prediction percentages in the test set. An image might include several tumor lesions, and the percentage was calculated both along the number of images and tumor lesions at threshold values as 85%/90%. (c) Examples of the PNF images in the test set classified by the model. (d) The confusion matrix of the results of independent test set 2. The model showed an accuracy of 84.75% and a false negative rate of 15.25% in this independent test set. (e) Model performance comparison between the deep learning model trained on PNF/MPNST labels only and one that includes surrounding normal organ/tissue information. The model trained with information from surrounding normal organs/tissues enhanced diagnostic accuracy from 61.90% to 79.37% for MPNST and from 46.67% to 70% for PNF, affirming our hypothesis.

Plexiform neurofibroma

Authors: Saihu Mao, MD¹, Song Li, MD¹, Zezhang Zhu, MD¹

1 Division of Spine Surgery, Department of Orthopedic Surgery, Nanjing Drum Tower Hospital, The Affiliated Hospital of Nanjing University Medical School, Zhongshan Road 321, Nanjing 210008, China

Corresponding author: Zezhang Zhu, E-mail: zhuzezhang@126.com

Disclosures: The authors declare that they have no competing interests.

Title: Convexity coronal imbalance in dystrophic scoliosis secondary to type I neurofibromatosis: classification and its importance for surgical decision-making 

Purpose: Trunk shift causing coronal imbalance (CI) is frequently encountered in unstable dystrophic scoliosis (DS) secondary to Type I Neurofibromatosis (NF-1). Surgical rebalance of coronal malalignment is challenging with high occurrence of iatrogenic aggravation. To develop a comprehensive classification of convex coronal spinal malalignment and propose a treatment algorithm for this specific condition.

Methods: A review of NF-1CI cases where different types of convex CI, rebalancing strategies, as well as the outcomes were identified and analyzed.

Results: Two main convex CI patterns were defined: thoracic CI (type 1) and thoracolumbar/lumbar CI (type 2), and were further subtyped by the compensatory behavior of the upper hemi-curve (straight or curved morphology). The incidence of post-op persistent CI ≥3cm was 0.0% and 63.6% for Type1 and Type 2 groups, respectively. Quantitative analysis revealed that the over correction of the compensatory upper hemi-curve and insufficient correction of the lower hemi-curve playing the role of imbalance driver were more significant in the imbalanced group (△Upper Arc Translation/|△Lower Arc Translation|: 31.8±34.4% vs. 109.6±60.0%, p=0.008; △Upper Arc Inclination/|△Lower Arc Inclination|: 33.5±37.3% vs. 89.8±36.6%, p=0.012). A surgical rebalancing algorithm was proposed to treat each subtype.

Conclusion: This new classification of convex CI emphasized that coronal rebalance should be obtained with cooperative correction that the lower hemi-curve should be aggressively corrected while the morphology of pre-op upper hemi-curve determined whether it played the role of fine-tuning or radical adjustment for coronal realignment. This is particularly true for type 2 NF-1CI patients with poor distal screw purchase and utmost avoidance of pelvic fixation.

Key words: NF1; Convexity coronal imbalance; dystrophic scoliosis; surgical decision-making

{{Bone abnormalities}}

Authors: Dan Liu, Ph.D., Florida State University, USA; Xiaoli Zong, Ph.D., Army Medical University, China; Pamela L. Wolters, Ph.D., National Cancer Institute, USA; Karin S. Walsh, Psy.D., Children’s National Hospital, USA; Jennifer Janusz, Psy.D., Children’s Hospital Colorado, USA; Bonita P. Klein-Tasman, Ph.D., University of Wisconsin–Milwaukee, USA; Stephanie M. Morris, M.D., Kennedy Krieger Institute, USA; Carrie E. Bearden, Ph.D., University of California, Los Angeles, USA; Jonathan M. Payne, Psy.D., Murdoch Children's Research Institute, Australia; Natalie Pride, Ph.D., Children's Hospital at Westmead, Australia; Peter L. Stavinoha, Ph.D., University of Texas MD Anderson Cancer Center, USA; Yang Hou, Ph.D., Florida State University, USA

Funding: (a) Congressionally Directed Medical Research Programs, Neurofibromatosis Research Program [W81XWH2110504]; (b) Center for Cancer Research, National Cancer Institute, Intramural Research Program; (c) Florida State University Faculty Startup Funding; and (d) University of Kentucky Faculty Startup Funding. 

Disclosures: The authors declare no financial conflict of interest.

Title: Age-varying associations between executive function and internalizing/externalizing behaviors in children with neurofibromatosis type 1: Integrative analyses of data from nine institutions

Purpose: Children with neurofibromatosis type 1 (NF1) frequently encounter cognitive and behavioral difficulties, such as challenges with executive function and both internalizing and externalizing behaviors. The severity of these issues can change over time. While executive function is known to relate to internalizing and externalizing behaviors differently at different ages in typically developing children, the pattern of these associations in children with NF1 is unclear. This study aims to explore these associations across ages to deepen our understanding and inform the development of age-appropriate interventions.

Methods: Individual-level data from 1,049 children with NF1, ages 3–18 years, across nine institutions in the U.S. and Australia were combined using integrative data analysis. Executive function, including inhibitory control, flexibility, emotional control, working memory, and planning/organization, was assessed with parent-rated Behavior Rating Inventory of Executive Function (BRIEF). Internalizing and externalizing symptoms were measured using the Child Behavior Checklist and the Behavior Assessment System for Children. Time-varying effect modeling (TVEM), a nonparametric method, was used to examine the associations between executive function and internalizing and externalizing symptoms across different ages, while controlling for sex. TVEM estimates these associations as continuous functions of age, providing curvilinear intercept and slope estimates with 95% confidence intervals (CIs). Significant associations are indicated by 95% CIs that do not include zero, and significant age-related differences by non-overlapping 95% CIs at specific age points.

Results: Elevated BRIEF scores, indicating greater impairment in all five domains, were consistently associated with greater internalizing and externalizing symptoms from ages 3 to 18 years. The magnitude of some associations varied significantly across different ages. Notably, the association between emotional control and internalizing symptoms (Figure 1) increased over time, becoming significantly stronger in middle to late adolescence (ages 16-18) compared to early childhood (ages 4-7). In contrast, the association between inhibitory control and externalizing symptoms (Figure 2) was strongest in early childhood (ages 5-8) and significantly decreased during early and middle adolescence (ages 12-16).   

Conclusion: The findings indicate that impaired executive function may contribute to persistent internalizing and externalizing problems among children with NF1 from early childhood through late adolescence. Interventions targeting different executive function domains could help mitigate these emotional and behavioral issues, with their effectiveness potentially varying by developmental stage. For example, early childhood interventions that focus on improving inhibitory control may be particularly effective in reducing externalizing symptoms, while interventions aimed at enhancing emotional control may be especially beneficial for addressing internalizing problems during adolescence.

Cognition / Behavioral / Learning

Authors: Michael J. Lippincott1, Alberto Mendoza-Valderrey PhD2, Jenna Tomkinson1, Huyen Thi Lam Nguyen2, Cameron Mattson1, Dave Bunten1, Michelle Mattson-Hoss PhD3, Herb Sarnoff3, Stephanie J. Bouley PhD4, James A. Walker PhD4, Sara Gosline PhD5, Alice Soragni PhD2, Gregory P. Way PhD1

¹Department of Biomedical Informatics, University of Colorado School of Medicine. Aurora, CO. ²Department of Orthopedic Surgery, David Geffen School of Medicine, University of California Los Angeles, CA. ³Infixion Bioscience, Inc., San Diego, CA. ⁴Center for Genomic Medicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA. ⁵Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland, WA.

Funding: A portion of this work was funded by the Gilbert Family Foundation NGMI Award #923014.

Disclosures: MMH and HS are employees of iNFixion Bioscience. GPW is on the scientific advisory board of iNFixion. AS is a founder and owner of Icona BioDx.

Title: High-content microscopy for characterizing and predicting drug response in NF1^-/- Schwann cell cultures and NF1 patient-derived tumor organoids.

Purpose: We are extending the foundation of our drug screening program for NF1 to quantify high-content microscopy readouts of cell phenotypes.

Methods: We applied the high-content, multiplex fluorescence microscopy assay called Cell Painting¹ in two-dimensional Schwann cell cultures (Figure 1). We collected Cell Painting images of two isogenic Schwann cell lines, one of wildtype genotype (NF1^+/+) and one of NF1 null genotype (NF1^-/-). Separately, we also optimized a Cell Painting assay in three-dimensional NF1 patient-derived organoids.² This is the first time this assay has been applied in this setting. We and others have shown that Cell Painting quantifies cell health phenotypes³, compound mechanism of action⁴, toxicity⁵, cancer cell resistance⁶, and provides new information not captured by molecular readouts like gene expression.⁷ We developed a 3D image analysis pipeline to segment individual cells, perform quality control, and quantify cell-type specific, high-content features from drug screening NF1 patient-derived organoids.

Results: In 2D, we showed proof-of-concept that pairing high-content microscopy with machine learning can distinguish NF1 genotypes in Schwann cells.⁸ Our machine learning model predicted NF1 genotype in a never-before-seen holdout test set of Schwann cells over 2x better than random (Figure 2). In 3D, our image analysis pipeline successfully segments and processes high-content, single-cell morphology features from organoids (Figure 3).  We screened dozens of drugs in different NF1 patient cutaneous neurofibroma samples and quantified the phenotypic impact of drugs with different targets and mechanisms of action. All data analysis code is publicly available on GitHub to accelerate community efforts.

Conclusion: The human eye can only detect large differences in fluorescence microscopy images. Instead, high-performance computing and machine learning are required to distinguish subtle but important patterns. 2D-based microscopy methods are amenable for extremely high-throughput drug screening and can identify therapeutic agents that make NF1 patient cells look healthy.⁹ Our 3D-based microscopy methods can prioritize drugs in a physiologically relevant environment specific to individual NF1 patients. Combined, our phenotypic drug screening platform may help to discover new and personalized therapeutic agents for NF1 patients while avoiding drug toxicity pitfalls and side-stepping high attrition rates in traditional drug discovery pipelines. 

References:

1. Bray, M.-A. et al. Cell Painting, a high-content image-based assay for morphological profiling using multiplexed fluorescent dyes. Nat. Protoc. 11, 1757–1774 (2016).

2. Thi Lam Nguyen, H. et al. A platform for rapid patient-derived cutaneous neurofibroma organoid establishment and screening. Cell Rep Meth. Volume 4, Issue 5 (2024).

3. Way, G. P. et al. Predicting cell health phenotypes using image-based morphology profiling. Mol. Biol. Cell 32, 995–1005 (2021).

4. Wong, D. R., Logan, D. J., Hariharan, S., Stanton, R. & Kiruluta, A. Deep representation learning determines drug mechanism of action from Cell Painting images. bioRxiv (2022) doi:10.1101/2022.11.15.516561.

5. Nyffeler, J. et al. Bioactivity screening of environmental chemicals using imaging-based high-throughput phenotypic profiling. Toxicol. Appl. Pharmacol. 389, 114876 (2020).

6. Kelley, M. E. et al. High-content microscopy reveals a morphological signature of bortezomib resistance. Elife 12, (2023).

7. Way, G. P. et al. Morphology and gene expression profiling provide complementary information for mapping cell state. Cell Syst 13, 911–923.e9 (2022).

8. Tomkinson, J. et al. High-content microscopy and machine learning characterize a cell morphology signature of NF1 genotype in Schwann cells. BioRxiv, 10.1101/2024.09.11.612546 (2024)

9. Frost, M. et al. Rationale for haploinsufficiency correction therapy in neurofibromatosis type 1. J. Transl. Genet. Genom. 6, 403–428 (2022).

{{DRUG DISCOVERY AND DEVELOPMENT}}

High-Throughput Screening Identifies Polymeric Nanoparticles for Delivery of Full Length Human  NF1 gene to Schwann Cells

Funding: This work is supported by a grant from the Gilbert Family Foundation Gene Therapy Initiative.

Purpose

Polymeric nanoparticles (PNPs) are modular delivery systems that can be optimized for targeted therapeutic delivery. Advances in rational design, including high-throughput screening and computational approaches, have shifted nanoparticle development from trial-and-error methods to structured, data-driven strategies. This has significantly improved preclinical discovery pipelines for nanomaterials. Here, we employ a high-throughput design-build-test-learn platform to enhance the delivery efficiency of PNPs for the treatment of Neurofibromatosis Type 1 (NF1), a genetic disorder caused by the loss of functional neurofibromin. NF1 gene replacement therapy holds immense clinical potential but is hindered by challenges in delivering the large 8.5 kbp NF1 gene using viral vectors. To overcome this, we utilized cationic PNPs for encapsulating and delivering the full-length human NF1 plasmid (EF1a-NF1-2A-EGFP, ~20 kbp) to human Schwann cells (SCs).

Methods and Results

Designing the perfect PNP is complex due to vast chemical diversity, biological system intricacies, and current modeling limitations. To address this, we used our High-Throughput Synthesis, Characterization, and Assessment of Nanoparticles (HIT SCAN™) platform to systematically screen over 500 different PNPs and identify the most promising “hit” candidates. The polymers were synthesized via reversible addition-fragmentation chain-transfer (RAFT) polymerization and self-assembled into PNPs. Our approach consisted of two rounds of screening.

In the first round, we screened ~450 first-generation (G1) PNPs derived from six selected monomers to assess plasmid loading, transfection efficiency (TE), and cytotoxicity in immortalized human NF1-null SCs (hTERT NF1-/- ipNF97.4). Select PNPs with specific monomer combinations emerged as promising candidates for plasmid encapsulation and delivery. Twenty hit PNPs demonstrated partial neurofibromin restoration in SCs. In vivo testing confirmed gene expression in mice and rat cortex, but G1 PNPs exhibited insufficient TE to fully restore neurofibromin function.

In the second round, a machine learning (ML)-guided approach was used to refine the design space. An ML model, trained on successful G1 candidates, identified key structural and compositional features, leading to a second-generation (G2) of 96 PNPs. G2 PNPs exhibited a 5x increase in TE in SCs, from ~4% in G1 to ~20% in G2. Functional assays assessing biodistribution, delivery efficacy, and neurofibromin restoration in vivo are ongoing.

Conclusion

By combining our HIT SCAN™ platform with data-driven PNP evolution, we identified novel PNPs capable of efficiently delivering the large NF1 gene to SCs. The 5x improvement in TE in G2 PNPs underscores the potential of high-throughput screening for accelerating nanomedicine development for genetic disorders like NF1.

{{THERAPEUTICS}}

Epilepsy in a cohort of pediatric patients with Neurofibromatosis type 1 in a tertiary center in Argentina.

Objectives: To describe the electroclinical characteristics of epilepsy in a pediatric series with NF1 and compare them with those reported in the literature. 

To analyze electroclinical and radiological patterns and therapeutic response.

Materials and methods: We performed a retrospective descriptive analysis of patients with NF1 evaluated by Neurology at Garrahan Hospital during the last 10 years. We included children from 1 month to 17 years old with positive clinical criteria of Neurofibromatosis type 1 (NF1) according to the latest guidelines of the National Institute of Health of the United States (NIH) and who had associated epilepsy. We defined epilepsy according to the ILAE 2022 criteria. We classified epileptic seizures according to ILAE 2017. Two authors analyzed electroencephalograms (EEGs). We excluded any patient with paroxysmal non-epileptic episodes or symptomatic seizures.

Results: 203 records were reviewed, of which 22 (10.8%) were reported with seizures, 2 had presented acute symptomatic generalized seizures and were therefore excluded, 20 (9.8%) presented epilepsy, 10 (50%) were focal epilepsies, 8 (40%) generalized and 2 (5%) epileptic encephalopathy of difficult control.

Of the focal seizures, 5 had pathological electroencephalogram and only presented structural epilepsy due to a high-grade glial lesion.  

Of the generalized epilepsies, 8 had EEG without foci or paroxysms, only 2 had lesions, one for a Moyas Moya syndrome and the other for a neuronal migration disorder.

As for treatments, 15 controlled seizures with one drug, 2 with two, 2 with four and 1 with five antiepileptic drugs.

Conclusions: Our series confirms that children with NF1 are more predisposed to epilepsy than the rest of the child population, 10.8%. We also evidenced that we did not register significant percentage differences between focal and generalized epilepsies. And that focal epilepsies one patient had a structural lesion. In turn, we recorded that most of these patients controlled their epileptic seizures with an antiepileptic drug and only two had refractory epileptic encephalopathy. 

{{Other}}

High Mobility Group A2 (HMGA2) drives plexiform neurofibroma growth and malignant transformation

Authors: Brooke E Hickey¹, Marissa D. Ciesielski¹, Emily E. White², Hannah N. Leffew¹, Alyssa A. Flint¹, Henry E. Mang¹, Dana K. Mitchell¹, Steven P. Angus^1,3,4, D. Wade Clapp^1,2,4, and Steven D. Rhodes^1,2,4,5

1Department of Pediatrics, Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN, USA

2Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis, IN, USA 

3Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, IN, USA 

4Indiana University Simon Comprehensive Cancer Center, Indiana University School of Medicine, Indianapolis, IN, USA 

5Division of Pediatric Hematology/Oncology/Stem Cell Transplant, Indiana University School of Medicine, Indianapolis, IN, USA 

CONFLICT OF INTEREST STATEMENT

SDR has served as advisor and independent contractor for SpringWorks Therapeutics and Practice Point Communications for activities unrelated to these studies. 

FUNDING

This abstract/presentation was supported by funding from the Neurofibromatosis Therapeutic Acceleration Program (NTAP) at the Johns Hopkins University School of Medicine. Its contents are solely the responsibility of the authors and do not necessarily represent the official views of The Johns Hopkins University School of Medicine.

ABSTRACT

Purpose: Malignant peripheral nerve sheath tumors (MPNSTs) are the leading cause of mortality in neurofibromatosis type 1 (NF1) and often arise from pre-existing plexiform (PN) and atypical neurofibroma (ANF). While genetic disruption CDKN2A and PRC2 complex members are frequently observed in ANF and MPNST, higher order transcriptional mechanisms governing the trajectories of these precursor lesions remain largely uncharted. High Mobility Group A2 (HMGA2) is a non-histone chromatin binding protein that modulates stem-like gene expression programs by altering chromatin structure. While silenced postnatally, we found that HMGA2 is aberrantly re-expressed in MPNST, leading us to hypothesize that HMGA2 drives MPNST progression by reactivating embryonic growth programs in Schwann cell precursors. 

Methods: We utilized genetically engineered mouse models with conditional Nf1, Ink4a/Arf, and Hmga2 alterations driven by PostnCre, as well as CRISPR-Cas9 editing of Hmga2 in primary Schwann cell precursors (SCPs) and human MPNST cell lines. RNA sequencing identified Let7-HMGA2 axis disruption and key HMGA2-dependent target genes which we validated by qRT-PCR, western blot, and immunohistochemical staining (IHC). Orthotopic xenograft studies assessed in vivo tumorigenesis. 

Results: RNAseq revealed a 7.6 log2 fold increase in Hmga2 expression in MPNST versus precursor lesions (padj <0.001), confirmed by western blot and IHC across mouse and human neurofibromas and MPNST. This was accompanied by downregulation of Let7 miRNAs, which physiologically suppress HMGA2. While conditional knockout (KO) of Hmga2 in Nf1f/f;Hmga2f/f;PostnCre+ mice did not impact PN genesis, ectopic HMGA2 overexpression in Nf1flox/+;Hmga2Δ/+;PostnCre+ mice resulted in the development of massive facial PN and impaired survival (p < 0.0001, log-rank test). CRISPR-Cas9 knockout of Hmga2 in primary Nf1-Ink4a/Arf  -/- SCPs delayed MPNST outgrowth (n=2 clones, 8 mice/group) and completely abolished tumor growth in human JH-2-002 MPNST cells (n=3 clones, 8 mice/group) in orthotopic xenografts. Contrastingly, Hmga2 KO in Nf1/Ink4a/Arf +/− SCPs did not alter MPNST-free survival, suggesting a role for Ink4a/Arf gene dose in modulating the impact of HMGA2 on malignant growth. Compensatory upregulation of Hox-family genes in RNAseq analysis of KO-tumors suggests an adaptive resistance mechanism that merits further exploration. 

Conclusions: While dispensable for PN initiation, ectopic overexpression of HMGA2 in SCPs drives PN progression and promotes malignant outgrowth in vivo in a subset of MPNST cell lines and primary SCPs – dependent on Ink4a/Arf gene dose. Future studies will characterize compensatory mechanisms that enable some tumor cells to bypass HMGA2 disruption and identify therapeutic vulnerabilities in HMGA2-addicted MPNSTs to inform clinical translation. 

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Title: Long-term hematologic effects of selumetinib treatment in children with inoperable plexiform neurofibromas 

Authors: Alan H. Siegel1, Andrea Baldwin2, Kara Heisey2, Kailey Herrera2, Anne Dufek3, Hemaxi Patel2, AeRang Kim1, Miriam Bornhorst4, Brian D. Weiss5, Peter de Blank6, Michael J. Fisher7, Brigitte C Widemann3, Andrea M Gross, MD3

Affiliations:

1Children’s National Hospital, Washington, DC; 2Clinical Research Directorate, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, MD; 3National Cancer Institute, Pediatric Oncology Branch, Bethesda, MD; 4Lurie Children’s Hospital, Chicago, IL, 5Riley Children’s Hospital, Indianapolis, IN; 6Cincinnati Children’s Hospital Medical Center, Cincinnati, OH; 7Children’s Hospital of Philadelphia, Philadelphia PA

Funding: This research is supported by the Intramural Research Program of the NCI

Disclosures: No relevant financial relationships 

Purpose: Selumetinib, a mitogen-activated protein kinase kinase (MEK) inhibitor, is the first FDA-approved treatment for children with neurofibromatosis type 1 (NF1)-associated inoperable plexiform neurofibromas. In the phase 1/2 clinical trial of selumetinib for children with inoperable plexiform neurofibromas (SPRINT, NCT01362803), treatment was associated with mild and reversible leukopenia (14%), neutropenia (22%), anemia (36%), and thrombocytopenia (12%). To maintain a durable tumor response, prolonged treatment is necessary; therefore, it is important to understand any potential long-term adverse effects. This study characterizes hematologic changes associated with long-term use of selumetinib in pediatric patients.

Methods: Data was obtained from the study database for the SPRINT trial. Comprehensive blood count (CBC) with differentials were collected per protocol at baseline and before cycles 2, 3, 4, 5, 7, 9, 11, 13, 17, 21, 25, then every 6 cycles (1 cycle = 28 days). Patient enrollment began in 2011 (Phase 1) and 2015 (Phase 2) and data was collected through February 7, 2025 and included all CBCs obtained for the duration of treatment including periods of drug hold. Parameters evaluated included white blood cell count (WBC, k/µL), hemoglobin (Hgb, g/dL), platelets (Plts, k/µL) and absolute neutrophil count (ANC, cells/µL). Simple linear regressions and descriptive statistics were calculated with GraphPad Prism version 10.4.1. 

Results: There were 98 participants (59 male, median age at enrollment 10.6 years, range 3-18.5) treated for a median 67.1 cycles per person (range 2.7 to 143.5) with a total 2597 CBCs obtained (median 24 per participant, range 3 to 78).  Across all CBCs, the WBC, ANC, Hgb, and Plts were within the normal range 91%, 88%, 77% and 91% of the time, respectively.  The minimum values for these cell lines at any timepoint were WBC 2.02 K/uL, ANC 690 cells/µL, Hgb 6.9 g/dL, and Plts 95 k/µL. No participants required a dose reduction or drug hold for a hematologic adverse event.  There was no statistically significant change over time for 73% (n=72), 87% (n=85), 59% (n=58) and 74% (n=73) of participants for WBC, ANC, Hgb and Plts, respectively. For those with a statistically significant change over time (p<0.05), the median slopes were -0.0266 (range -0.1333 to 0.0624) for WBC, -0.0149 (range -0.0729 to 0.2743) for ANC, 0.0133 (range -0.5793 to 0.0777) for Hgb, and -0.3802 (range -5.352 to 18.03) for Plts. 

Conclusion: The data reported here is the longest follow-up to date for pediatric patients with NF1 on selumetinib. No evidence of clinically meaningful changes in hematologic parameters was observed.  Though small statistically significant changes were seen in some participants, there was no clear trend (positive or negative) in any parameter.  Hgb had the highest proportion of patients with a significant trend (41%) however it was a positive trend and not consistent with previously reported anemia. Notably, in all instances where the minimum values were below normal, the decreases were transient and often associated with intercurrent illness. These results are preliminary and additional analyses are ongoing.

{{Plexiform neurofibroma}}

Authors: Sarah Mohabeer1, Frank McCormick PhD FRS DSc1,2, Matthew J. Sale PhD 1,*, Harish N. Vasudevan MD PhD3,4,*

Funding: CTF Gene Therapy Initiative

Disclosures: N/A

Title: A suite of NF1 reconstitution systems reveal GAP activity achieves transient but not long-term efficacy in MPNST cells

Word Count: 374/400 words

Purpose: Neurofibromatosis type-1 (NF-1) is a monogenic syndrome caused by germline mutation of the NF1 gene, making the possibility of gene therapy conceptually attractive. However, NF1 reconstitution efforts have been hampered by effective expression systems and both the optimal cargo and cellular context for NF1 gene therapy remain unclear. Here, we report the development of multiple NF1 expression systems and their effects in malignant peripheral nerve sheath tumor (MPNST) models. 

Methods: We optimized transient and stable expression systems for three NF1 constructs: full length NF1 (NF1full), the catalytically inactive arginine finger mutant (NF1R1276P), and the minimum GAP-related domain (NF1GRD-CaaX) membrane anchored via the HRAS prenylation signal (CaaX). Expression systems were validated in HEK293T cells followed by functional and biochemical analysis in two NF1 null MPNST cell lines, ST88-14 and JH-2-002, to investigate the effects of NF1 reconstitution. 

Results: ALFA tagged transient NF1FL and NF1GRD-CaaX fully blocked downstream phospho-MEK (pMEK) and phospho-ERK (pERK) activation in HEK293T but not MPSNT cells. Stable lentiviral doxycycline inducible expression revealed the NF1GRD-CaaX was superior to NF1FL in blocking both pERK activation and decreased cell growth in MPSNT cells. In contrast, the NF1R1276P showed minimal effect on both signaling and cell growth. However, continued NF1FL or NF1GRD-CaaX expression led to MPNST cells silencing expression through genetic mutation or deletion of these constructs while NF1R1276P expression was maintained, underscoring the selective pressure of long term catalytically competent NF1 expression. This occurs at least in part due to the stability of the NF1 protein following doxycycline induction in MPNST cells. To overcome this limitation, we developed and validated an E. coli dihydrofolate reductase degron system to translationally regulate NF1 protein expression, providing a more dynamic expression window. Finally, we also designed a dual NF1 and tamoxifen inducible CRAF system to circumvent NF1 silencing through downstream RAF activation. 

Conclusion: Transient and short-term stable reconstitution of catalytically competent NF1 blocks RAS signaling and cell growth in MPNST cells, but long-term reconstitution is overcome through NF1 mutation. Moreover, our data suggests the dominant function of NF1 is through its Ras GAP catalytic activity, as the NF1R1276P did not affect biochemical or functional outputs and was not subject to the same selective pressure as NF1FL or NF1GRD, motivating the development of alternate systems for NF1 reconstitution

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Authors: Diane C. Zelman, PhD and Erica B. Leif, M.A.; Alliant International University – San Francisco Bay Area, California; Robert Gore, PhD;  Adena Regional Medical Center, Chillicothe, Ohio

Funding: This research was funded by a contract number 2023-04-001 from the Children’s Tumor Foundation awarded to Diane Zelman, PhD. 

Disclosures: The authors have no relevant financial or non-financial disclosures

Title: Quantifying Self-Care among Adults with Neurofibromatosis I: Validation of a New Disease-Specific Measure

Purpose: Neurofibromatosis 1 (NF1) requires complex self-care, including symptom management, identifying healthcare, and addressing psychological health. The Self-Care of Chronic Illness Inventory (SC-CII) is a widely-used “generic” measure of self-care among people with chronic illness that quantifies monitoring, management, and maintenance of chronic illness, and self-care confidence (Riegel et al., 2012). Riegel and colleagues have subsequently validated several disease-specific versions of the SC-CII for self-care behaviors relating to specific health conditions (e.g., hypertension, diabetes, and heart disease). There exists no self-care measure specific to NF1.  The research objective was to create a disease-specific patient-reported outcome measure of self-care in people with NF1. 

Methods: Participants were 317 adults with NF 1 recruited from the Neurofibromatosis Registry (age 18-80, 71.3% female, 86.4% Caucasian, 33.5% college-educated, 54.6% with Worst Pain > 4 on a 1-10 scale) who completed measures online. An earlier study (Leif & Zelman, 2024) conducted interviews with 22 individuals with NF1 to establish a compendium of self-care behaviors, and based on these findings, self-care theory and the generic SCI-II,  sixty-eight candidate items for the new self-care measure were generated.  Each item described a form of self-care scored on a 1-5 Likert frequency scale ranging from Never/Rarely to Very Often. Participants also completed the 20-item generic SC-CII (Riegel et al., 2018) and measures of pain, sleep, quality of medical care and quality of life.  

Results: In several iterations of Confirmatory Factor Analysis (CFA), items were dropped at each step to improve fit statistics to establish a set of domains corresponding to important self-care constructs. The final 22-item solution comprised five subscales (Medical Monitoring, Self-Monitoring, Engaging Social Support, Stress Management, and Self-Maintenance) and two four-item pain self-care subscales for those with pain (Personal Pain Care and Pain Medication/Professional Interventions).  The new disease-specific NF1 measure better met criteria for CFA relative to the generic SCI-II, and showed stronger relationships with key outcomes such as anxiety, depression, sleep, pain, autonomy and relatedness with the healthcare team. 

Conclusion:  This research was the first study to quantify disease self-care in NF1 and is one of few studies to investigate pain management as an aspect of self-care in NF1 (e.g., Buono et al.,2018). This is a promising measure for quantifying self-care that can also stimulate dialogues in healthcare settings about care needs. Subsequent research will continue to establish the suitability of this measure for patients and clinicians. 

{{Cognition / Behavioral / Learning}}

Authors: Nikita Bagaev, MD (Schepens Eye Research Institute, Massachusetts Eye and Ear, Harvard Medical School)

Jonathan Soucy, PhD (Schepens Eye Research Institute, Massachusetts Eye and Ear, Harvard Medical School)

Aubin Mutschler (Schepens Eye Research Institute, Massachusetts Eye and Ear)

Petr Baranov, MD, PhD (Schepens Eye Research Institute, Massachusetts Eye and Ear, Harvard Medical School)

Funding: Gilbert Family Foundation, 533492

Disclosure: This research was supported by Gilbert Family Foundation - 533492. The authors declare no conflicts of interest. The IMARIS software was provided by Neurobiology Imaging Facility of Harvard Medical School for research purposes. The authors confirm that the research was conducted ethically, adhering to all relevant guidelines and regulations for animal studies. All animal procedures were approved by the Institutional Animal Care and Use Committee (IACUC) of Schepens Eye Research Institute. The authors have no financial or personal relationships with other individuals or organizations that could inappropriately influence their work

Title: Morphometric analysis of microglia in the retina and optic nerve

Purpose: Microglia promote many diseases including some clinical manifestations of Neurofibromatosis type 1 (NF1). Particularly visual impairments seen in NF1 patients can be result from damage of retinal ganglion cells (RGC) by activated microglia. This study performs morphometric analysis of microglia in the retina and optic nerve of mice with a mutation in the Neurofibromin 1 (Nf1) gene to investigate reactive microglia in NF1.

Methods: Retinal whole-mounts and optic nerves from Nf1 fl/fl mice were stained with ionized calcium binding adaptor molecule 1 (Iba1), a pan-myeloid marker, and 4′,6-diamidino-2-phenylindole (DAPI) for cell nuclei. To increase the image resolution we incubated tissues in the clearing solution and put them on the concave microscope slides mounting with clearing media. Then confocal 3D images were acquired using the z-stack function of Olympus Fluoview FV3000 microscope (160 μm depth, 2 μm steps, ×20 magnification). IMARIS software (Version 10.0.1) was used to reconstruct the microglial surface and Filament Tracer function was applied to reconstruct the processes and spines of microglia cells. Vantage function was utilized for quantitative analysis and classification of the microglia based on cell area and processes length. 

Results: This study successfully characterized microglia morphology and cell states (homeostatic, reactive, hyper-ramified, rod-shaped) in the retina and optic nerve of Nf1 fl/fl mice. The pipeline included tissue clearing, high-resolution image acquisition, and artificial intelligence-powered image analysis. This allowed for detailed description of microglia parameters, precise cell classification, and extraction of numerous cell features (cell area, soma size, process number, process length, Sholl analysis). 

Conclusions: This pipeline provides a framework for high-throughput image analysis in the retina and optic nerve, particularly relevant for NF1 research. This protocol may be adapted for other image analysis applications.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Authors: Abbott Catesby Pinney (College of William & Mary), Aidan Robinson (Stevens Institute of Technology)

Funding: N/A

Disclosures: N/A

Title: A Landscape Analysis of Machine Learning Models for Pain Monitoring and Prediction

Purpose: Pain is an inherently subjective experience that complicates effective diagnosis and treatment, particularly in patients with schwannomatosis, where pain is often severe and underreported. Advances in machine learning and continuous physiological monitoring now offer promising avenues for objective pain assessment. This project conducts a scoping review of existing pain prediction models—examining machine learning models trained on physiological data—to identify research gaps and promising methodologies.

Methods: A comprehensive search of publications in the PubMed database was conducted using variations of query terms “machine learning,” “artificial intelligence,” “deep learning,” “pain,” “analgesic,” “prediction,” “classification,” and “estimation.” 26 publications that introduce at least one machine learning model capable of predicting instantaneous or long-term pain were included in the survey. Information such as the model architecture (e.g., SVMs, CNNs, RNNs), its training dataset (e.g., UNBC-McMaster Shoulder Pain Expression Archive Database, Infant COPE database), and the modality of the training data (heart rate, skin conductance, EEG, EMG), was extracted from each included paper to identify trends and gaps in pain prediction models. 

Results: Within our survey, we have identified two main types of pain monitoring machine learning models: instantaneous and long-term. We found that these models usually fall into another two categories, being pain intensity prediction and pain/no pain classification. We have identified functional magnetic resonance imaging (fMRI) and electroencephalogram (EEG) data as being the leading two data modalities used to identify pain signatures. Finally, we found that the majority of the models in the survey use original datasets which are unique to the studies in which the models are introduced.

Conclusion: Our review identified several trends in pain monitoring machine learning models. The labelling of most models into either pain intensity prediction or pain/no pain classification provides a useful framework for discussion of autonomous pain monitoring techniques. The recognition of a trend towards using neurological signatures for monitoring highlights both the potential of EEG and fMRI as well as the relative lack of other data modalities which might be more convenient and practical for those living with chronic or recurring acute pain. Finally, the identified lack of a common dataset for training, validation, and testing among the models gathered in this survey limits the feasibility of comparing their reported accuracies and other metrics. This emphasizes the need for a large-scale, accessible pain recognition dataset for more objective comparison between pain recognition models.

{{THERAPEUTICS}}

Authors: Jeremy Jacobson1, Yannick Mahlich1, Chang In Moon1,2,3, M. Ryan Weil4, Sara JC Gosline1,5

1.    Pacific Northwest National Laboratory, Seattle, WA, 

2.    Lester and Sue Smith Breast Center and Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. 

3.    Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030, USA. 

4.    National Cancer Institute Frederick National Laboratory, Frederick, MD 

5.    Oregon Health and Sciences University, Portland, OR

Funding: Department of Defense NFRP

Disclosures: None

Title: CoderData: a Python package and collection of benchmark datasets to enable development of machine learning models for drug sensitivity in NF1 organoids

Purpose: While numerous machine learning models have been published that utilize omics data for drug sensitivity prediction1, these methods often require larger, more diverse datasets than are currently available for NF1. Enhancing drug response prediction is vital for the NF community, which currently has only has two approved NF1 therapies2,3. The purpose of CoderData is to provide a series of harmonized datasets4-15—including cell lines, ex vivo, organoid, and patient derived xenograft (PDX) data—to empower both machine learning and deep learning analytical approaches in inferring drug response in NF1.

Methods: We developed the Cancer Omics and Drug Experiment Response Data (CoderData) Python package that allows users to download, reformat, and perform balanced splits creating data ready for model integration. Our automated build pipeline 1) automates the collection of drug structure and cancer data from multiple sources including the NF Data Portal16, 2) harmonizes drug curves, omics measurements, and mutation variant nomenclature, and 3) publishes the data on FigShare with programmatic retrieval using our Python package. We plan to benchmark drug response models using multiple cancer model types to attain better predictions than those based solely on PDXs.

Results: To date we have curated molecular measurements (genomics, transcriptomics, proteomics, copy number) from approximately 4300 unique samples across multiple model systems such as two-dimensional cell lines (1280) with multiple measurements, organoid models (388), patient-derived xenografts (25), ex vivo (1022) and tumor data (1561). This collection includes data from patient-derived NF1 tumors (malignant peripheral nerve sheath tumors and cutaneous neurofibromas). Additionally, we assembled drug response data for roughly 55,000 unique drugs across these samples, and further integration of patient-derived organoids (86) is underway. 

The package is available at https://pnnl-compbio.github.io/coderdata/ .

Conclusions: The integration of harmonized data from multiple cancer model systems, particularly organoid data, may advance the power of machine learning and standard statistical models to predict drug sensitivity in rare NF1 tumors. The CoderData package not only facilitates training and benchmarking of these models through an accessible Python package but also supports cross-application between cell line/tumor data and organoid systems through data harmonization. This cross-model integration enhances the potential for robust, generalizable predictions and positions CoderData as the only benchmark dataset to combine cell line, ex vivo, and NF1 organoid data—fostering collaboration and further contributions from the research community17,18.

References: 1. Partin et al., Frontiers in Medicine 2023. 2. Gross et al., The New England Journal of Medicine, 2020. 3. Moertel et al., Journal of Clinical Oncology 2024. 4. Barretina et al., Nature 2012. 5. Haverty et al., Nature 2016. 6. Garnett et al., Nature 2012. 7. Corsello et al., Nature Cancer 2020. 8. Seashore-Ludlow et al., Cancer Discovery 2015. 9. Mpindi et al., Nature 2016. 10. Shoemaker, Nature Reviews Cancer 2006. 11. Basu et al., Cell 2013. 12. Lindgren et al., Journal of Proteome Research 2021. 13. Bottomly et al., Cancer Cell 2022. 14. Pino et al., Cell Reports Medicine 2024. 15. Dehner et al., JCI Insight 2021. 16. Allaway et al., Scientific Data 2019. 17. Larsson et al., Neuro-oncology 2023. 18. Nguyen et al., Cell Reports Methods 2024.

{{BIOINFORMATICS AND COMPUTATIONAL OMICS}}

Isam A. Naber, MD, Marco Giovannini, MD, PhD, Jeremie Vitte, PhD.

Department of Head and Neck Surgery, David Geffen School of Medicine at UCLA, University of California Los Angeles, Los Angeles, CA, 90095, USA.

Funding: This research is funded and supported by the Department of Defense CDMRP NFRP (W81XWH1810622)ADD GRANT NUMBER to JV, and Children Tumor’s Foundation Young Investigator Award (2024-01-004) to IAN.

Disclosures: The authors report no disclosures. 

Title: Proteome analysis of inner ear fluids in NF2-SWN mouse models with hearing loss

Purpose: Clinical data suggests that hearing loss in NF2-related schwannomatosis (NF2-SWN) patients does not correlate directly with vestibular schwannoma (VS) size or growth rate. This challenges the assumption that hearing loss results solely from tumor compression and suggests other contributing factors. High-resolution FLAIR MRI has detected increased cochlear signal and elevated intralabyrinthine protein in 94% of ears in patients with hearing loss, indicating a possible contribution to the pathogenesis. This study aims to investigate the underlying mechanisms of hearing loss in NF2-SWN-associated vestibular schwannomas using NF2 mouse models, potentially identifying prognostic biomarkers.

Methods: This longitudinal study will use two NF2-SWN mouse models, presenting with different mechanisms of progressive hearing loss (P0-NF2Δ2-3 and Periostin-Cre;Nf2flox/flox) at 3, 6, 9 and 12 month timepoints. We are performing ABR/DPOAE to monitor their hearing levels, dissecting cochleae to evaluate the extent of proteinaceous precipitates in the cochlear fluids with H&E staining. We are preparing sensory epithelium whole mounts to assess the number of cochlear hair cells using Phalloidin/Hoechst staining. Then, we will extract protein precipitates from the cochleae and analyze them with mass spectrometry for identification of potential biomarkers.

Results: Whole mounts of sensory epithelium from control mice were microdissected and stained using rhodamine phalloidin with optimized dilutions. Imaging was conducted using a confocal microscope and maximum projections will beere used for hair cells counting. Control mice were used to compare the quality of H&E-stained cochlear sections after paraffin or OCT embedding, to optimize their use for laser microdissection of protein precipitates, while maintaining the morphology of the cochleae. 

Conclusion: The results of this study will allow us to dissect the pathophysiology of hearing loss in NF2-SWN and sporadic VS patients and might reveal promising prognostic biomarkers of hearing loss for informed clinical management.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Development of a genomics-based nanopore sequencing test for the differential diagnostics of MPNSTs and ANNUBPs.

Presenting author: Bernat Gel

Germans Trias i Pujol Research Institute (IGTP)

Malignant peripheral nerve sheath tumors (MPNST) are soft tissue sarcomas with bad prognosis and the leading cause of Neurofibromatosis Type 1 (NF1)-related mortality. In the NF1 context, MPNSTs usually arise from preexisting benign plexiform neurofibromas (pNF) through a premalignant state called atypical neurofibromatous neoplasm with unknown biological potential (ANNUBP), although it is currently not possible to determine if a given ANNUBP will ever progress into an MPNST. Surgery is currently the only curative treatment for these tumors. However, differentiating between these tumors radiologically and histologically, especially in the transition phases, can be difficult. 

This project aims to develop a new molecular test for the differential diagnosis of ANNUBPs and MPNSTs, including the detection of ANNUBPs in the early steps of MPNST transformation, based on the recurrent alterations found in MPNSTsz. The test is based on nanopore sequencing and takes advantage of the selective sequencing capabilities these sequencers have to perform in-silico enrichment and their ability to determine nucleotide sequence and DNA methylation simultaneously.

We performed a detailed genomic characterization of a set of MPNSTs using WGS, RNA-seq and methylation arrays and identified recurrent alterations seen on subgroups of MPNSTs at the genetic, genomic (copy-number, loss of heterozygosity, and chromosomal rearrangements) and epigenomic level. We designed an enrichment panel targeting those regions and are currently testing its performance in surgical specimens and minimally invasive biopsies. We are also developing a custom data analysis pipeline to process the generated data, extract all key features and produce a reasoned and data-backed classification of the samples. 

In conclusion, we are developing a new genomics-based tool for better ANNUBP and MPNST diagnostics, with a potential impact on patient’s management, survival and quality of life.

Full author list: Miriam Magallón-Lorenz1,2, Alba Hernández-Gallego3, Gustavo Tapia3, Helena Mazuelas2, Itziar Uriarte-Arrazola2, Judit Farrés-Casas2, Sara Ortega-Bertran4, Edgar Creus-Bachiller4, Juana Fernández-Rodríguez5,6, Lopez-Gutierrez JC7, Alicia Castañeda8, Maria Saigí9, Marc Cucurull9, Ignacio Blanco10, Claudia Valverde11,12, Cleofé Romagosa13, Héctor Salvador8, Conxi Lázaro4,6, Meritxell Carrió2, Eduard Serra2,6, Bernat Gel1,2,14

1)    Translational Genomics and Bioinformatics Group, Germans Trias I Pujol Research Institute, Can Ruti Campuc, Badalona, Barcelona, 08916, Spain

2)    Hereditary Cancer Group, Germans Trias i Pujol Research Institute (IGTP); Can Ruti Campus, Badalona, Barcelona, 08916; Spain

3)    Department of Pathology, Germans Trias i Pujol University Hospital (HUGTiP), Can Ruti Campus, Badalona, Barcelona, 08916; Spain

4)    Hereditary Cancer Program, Catalan Institute of Oncology (ICO-IDIBELL), L'Hospitalet de Llobregat, Barcelona, 08098; Spain

5)    Mouse Lab, IDIBELL, Hospitalet de Llobregat, Barcelona, Spain

6)    Centro de Investigacion Biomedica en Red de Cancer (CIBERONC), Madrid, Spain

7)    Department of Pediatric Surgery, University Hospital La Paz, Madrid, Spain

8)    Pediatric Cancer Center Barcelona, Sant Joan de Déu Barcelona Children’s Hospital, 08950, Barcelona, Spain

9)    Catalan Institute of Oncology (ICO), Germans Trias i Pujol Research Institute (IGTP)

10)    Clinical Genomics Unit, Germans Trias i Pujol University Hospital (HUGTiP), Can Ruti Campus, Badalona, Barcelona, 08916; Spain

11)    Department of Medical Oncology, Hospital Universitari Vall d’Hebron, Barcelona, Spain.

12)    Vall d’Hebron Institute of Oncology, Barcelona, Spain.

13)    Department of Pathology, Hospital Vall d’Hebron, Barcelona, Spain

14)    Departament de Fonaments Clínics, Universitat de Barcelona, 08036, Barcelona, Spain

Funding: This work has been supported by the Children’s Tumor Foundation (CTF) with a Clinical Research Award (CTF-2023-10-001). It has also received support from Instituto de Salud Carlos III National Health Institute funded by FEDER funds - a way to build Europe -[PI20/00228; PI23/00583; PI23/00422] and Fundación Proyecto Neurofibromatosis. We would like to thank the constant support of Fundación Proyecto Neurofibromatosis and the AANF and AcNefi patient associations.

{{Malignant peripheral nerve sheath tumors (MPNST)}}

Cryptanoside A, a cardiac glycoside epoxide as a potential treatment for NF2-SWN related tumors

Halder, Sushanta, PhD

Nationwide Children's Hospital

Cardiac glycosides, such as digoxin and digitalis, are Na⁺/K⁺-ATPase (NKA) antagonists used to treat heart failure and exhibit anti-tumor effects. Cryptanoside A is a naturally occurring cardiac glycoside epoxide isolated from the plant Cryptolepis dubia (Burm.f.). Unlike digoxin and digitalis, cryptanoside A has distinct structural variations, contributing to a comparatively lower binding affinity to NKA and potentially lower side effects. It has also been shown to be cytotoxic to colon, breast, and ovarian cancer, and melanoma cell lines. We aim to evaluate the growth inhibitory and anti-tumor activities of cryptanoside A in neurofibromatosis type 2-related schwannomatosis (NF2-SWN) tumors and its mechanism of action. We extracted cryptanoside A from Cryptolepis dubia. Its purity was confirmed via high-performance liquid chromatography and mass spectroscopy, and its structural identity verified using nuclear magnetic resonance. Cryptanoside A effectively inhibited the growth of NF2-deficient benign meningioma Ben-Men-1 and AG-NF2-Men cells and malignant meningioma KT21-MG1 cell lines with the IC₅₀ values of 101, 104, and 131 nM, respectively. Similarly, schwannoma cells were vulnerable to growth inhibition by cryptanoside A. Treatment of AG-NF2-Men cells with cryptanoside A at 1x IC₅₀ led to both G₁ and G₂/M arrest, and at 2x and 5x IC₅₀ led to G₂M arrest after one-day treatment. Incucyte Caspase-3/7 Green Apoptosis Assay demonstrated that cryptanoside A induces caspase cleavage, indicating apoptosis. Western blots confirmed PARP cleavage in cryptanoside A-treated cells. Additionally, cryptanoside A increased the levels of the phosphorylated stress activated protein kinase p-p38^SAPK and the DNA damage response marker γH2A.X. Experiments are in progress to evaluate the anti-tumor activity of cryptanoside A in the orthotopic AG-NF2-Men xenograft model. Also, we are performing transcriptomic analysis to examine the signaling pathways affected by cryptanoside A. Our preclinical findings support further development of cryptanoside A as a potential treatment for NF2-SWN associated tumors. (Supported by CancerFree KIDS)

Author list: Sushanta Halder¹, Janet L Oblinger¹, Yulin Ren², A Douglas Kinghorn², Long-Sheng Chang¹

¹Center for Childhood Cancer, Abigail Wexner Research Institute at Nationwide Children’s Hospital and Department of Pediatrics, The Ohio State University College of Medicine; ²Division of Medicinal Chemistry and Pharmacognosy, The Ohio State University College of Pharmacy

{{DRUG DISCOVERY AND DEVELOPMENT}}

Association between Neurofibromatosis Type 1 and Mixed Pheochromocytoma: A Case Report of a Pediatric Patient

Lombardi, Francina; Palomar, Nicolas; Cardozo, Agustin; Achilli, Gabriela; Diez, Blanca.
 FLENI, Buenos Aires, Argentina.

Introduction:
Neurofibromatosis type 1 (NF1) is a genetic disorder characterized by an increased risk of developing tumors, including pheochromocytomas. These tumors, rare in the general population, have a higher incidence in individuals with NF1. Pheochromocytomas are neuroendocrine tumors that affect the adrenal glands and are responsible for excessive production of catecholamines. In some cases, dopamine-secreting pheochromocytomas are present and are known as mixed pheochromocytomas. This variant refers to pheochromocytomas that present morphological features of both typical pheochromocytomas and neuroblastomas. This variant is particularly rare in pediatric patients with NF1, which poses significant diagnostic and therapeutic challenges. Identification of this type of pheochromocytoma is crucial due to the clinical implications in the management of hypertension and other associated complications.

Objectives:
The objective of this study is to describe the clinical, diagnostic, and therapeutic characteristics in a pediatric patient with NF1 who developed a mixed pheochromocytoma, in order to enhance the understanding of this rare association and optimize its management.

Materials and Methods:
A retrospective, observational study was conducted on a 13-year-old pediatric patient diagnosed with NF1 and mixed pheochromocytoma. Data were collected regarding clinical presentation, diagnostic studies, surgical interventions, and treatments.

Results:
A 12-year-old male patient, diagnosed with NF1, began with recurrent, progressively severe headaches. Upon admission to our institution, records showed elevated blood pressure, prompting a brain and spinal MRI, which revealed a nodular mass in the left adrenal gland, initially presumed to be a neurofibroma. Urinary catecholamine tests were requested, the results were 9.5 mg/24 hours (NV < 5.2 mg).
 Scintigraphy showed a left adrenal mass suggestive of neuroblastoma. The patient underwent laparoscopic left adrenalectomy with tumor resection. The biopsy confirmed adrenal neoplasm composed of pheochromocytoma/ganglioneuroblastoma. No further treatment was required, and the patient remained asymptomatic, with no cardiovascular compromise, normalizing blood pressure readings. Follow-up imaging was unremarkable.

Conclusion:
Mixed pheochromocytomas in pediatric patients with NF1 are an extremely rare entity, with an estimated incidence in the literature of approximately 0.1% to 5.7% of patients with NF1. These tumors should be considered in the differential diagnosis of children with unexplained hypertension. Early diagnosis and surgical intervention are crucial for the successful management of these patients. Given the risk of recurrence, continuous long-term follow-up is recommended to detect potential recurrences and manage associated complications.
 Management of pediatric patients with NF1 and mixed pheochromocytoma requires a multidisciplinary approach to ensure timely diagnosis and appropriate treatment.

{{Other}}

Neurological manifestations in neurofibromatosis type 1: experience of a tertiary center with a cohort of 203 Argentine children.

INTRODUCTION: NF1 is an autosomal dominant genetic disorder with expression variability, it is due to mutation in the neurofibromin gene. It can be associated with epilepsy, neurodevelopmental disorders and cerebrovascular disease.

OBJECTIVES: To describe the most prevalent neurological manifestations in a series of pediatric patients with NF1 seen at Hospital Garrahan in Argentina in the last 10 years.

POPULATION AND METHODS: We performed a descriptive, retrospective analysis of the medical records of patients with clinical criteria for NF1 with an age range of zero to 15 years, evaluated by neurology at Hospital Garrahan in the last 10 years. 

RESULTS: We registered 203 patients, 99 females with a mean age of 3 years. 133 patients reported neurodevelopmental disorders with learning difficulties in 54%, psychopedagogy was the most indicated therapy (85%). 22 (11.3%) presented focal and generalized epilepsy not always associated with structural lesions, 12 of them (54%) with pathological electroencephalogram. The most indicated drug was clobazam. 83 patients 22 (11.3%) presented focal and generalized epilepsy not always associated with structural lesions, 12 of them (54%) with pathological electroencephalogram. The most indicated drug was clobazam.83 patients (50%) had hamartomatous lesions in the cerebellum, 2 patients reported Moya Moya syndrome.

CONCLUSION: Our study shows that neurodevelopmental disorders are frequent in this population, among them the most diagnosed was learning disorder. Epilepsy is more common than in the general population, and focal seizures are more frequent, requiring prolonged antiepileptic drugs but with good control. In agreement with previous series, bright lesions in cerebellum were frequently observed.

{{Other}}

Extracellular Vesicles derived from heterogeneous Nf2−/− Schwann cells present 

distinct proteomic signatures.

Sara Veiga, Ph.D. Krantz Family Center for Cancer Research, Massachusetts General Hospital, Harvard Medical School, Boston, MA. 

Schwannomas are surprisingly heterogeneous and develop in complex microenvironments with a plethora of different cell types, such as immune cells.  Macrophages are abundant in many schwannomas, but it is not known how they are recruited to and interact with the tumors. Understanding how tumor heterogeneity arises and how different cells contribute to this is crucial to improve clinical responses. Extracellular vesicles (EVs) are a known means of communication between cells and, in the central nervous system, EVs play a significant role in glial tumor development. They carry diverse cargo consisting of proteins, nucleic acids, enzymes and lipids, which contributes to their diverse roles in disease. The contributions of EVs to schwannoma development have not been extensively studied, and not much is known about their effects in disease context. In this study we are investigating how schwannoma intrinsic heterogeneity influences EV cargo and how EVs influence macrophage recruitment.

For this study we used an in vitro model that captures intrinsic heterogeneity seen in schwannoma. Nf2-deficient Schwann cells cultured two different conditions adopt distinct phenotypic states that mirror intrinsic heterogeneity. EVs were isolated from the cell conditioned media of both conditions and characterized by nanoparticle tracking analysis (NTA) to assess size and Western blot to identify classic EV markers. A proteome profiler assay was used to identify the presence/absence of 111 different cytokines in EVs from the two different conditions. Transwell assay was used to explore the role of EVs in bone marrow derived macrophage recruitment. 

Our data shows that EVs isolated from the two different Nf2-deficient Schwann cell populations present a similar size, around 130nm. Furthermore, nutrient deprivation resulted in a different loading of Alix, but not HSP70 into the EVs. These classic EV markers detected after isolation also confirm the presence of a pure EV population. Importantly, proteome profiler analysis suggests that the two different Nf2-deficient Schwann cell populations produce EVs with distinct cytokine cargoes. Several of the identified analytes such as CCL2, CCL3 and CCL5 are known macrophage chemoattractants, suggesting that intrinsic heterogeneity within schwannomas may differentially impact macrophage recruitment. These data shed light onto the roles of EVs in schwannoma, and onto how heterogeneity can affect vesicular cargo. Our ongoing work aims to define how these changes impact macrophage recruitment into the tumors.

Sara Veiga1,2, Christine Chiasson-MacKenzie1,3, Emily Wright1,3, Jeremie Vitte4, Marco Giovannini4, Andrea McClatchey1,3*, Shannon Stott1,2,5*

1 Krantz Family Center for Cancer Research, Massachusetts General Hospital, Charlestown, MA. 2 Department of Surgery, Harvard Medical School, Boston, MA. 3 Department of Pathology, Massachusetts General Hospital, Harvard Medical School, Boston, MA. 4 Department of Head and Neck Surgery, David Geffen School of Medicine at UCLA and Jonsson Comprehensive Cancer Center (JCCC), University of California Los Angeles, Los Angeles, CA. 5 Center for Engineering in Medicine, Massachusetts General Hospital, Boston, MA. *Corresponding authors.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Authors: Madilyn R Stahl, BS1,2; Garrett M Draper, BS1; Duncan J Clarke, PhD2; David A Largaespada, PhD1,2

1Department of Pediatrics, Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota, USA

2Department of Genetics, Cell Biology and Development, University of Minnesota, Twin Cities, Minneapolis, Minnesota, USA

Funding: NINDS R01NS115438, NCI R01NS086219, Pre-Clinical Research Award Neurofibromatosis Research Initiative (NFRI) through Boston Children’s Hospital (GENFD0001769008), Children's Tumor Foundation Drug Discovery Initiative Award and Synodos for NF1 Award, Zachary Bartz NF Research FundAmerican Cancer Society Research Professor Award (RP-17-216-06-COUN)

Disclosures: D.A.L. is the co-founder and co-owner of several biotechnology companies, including NeoClone Biotechnologies, Inc., Discovery Genomics, Inc. (recently acquired by Immusoft, Inc.), B-MoGen Biotechnologies, Inc. (recently acquired by Biotechne Corporation), and Luminary Therapeutics, Inc. The business of all these companies is unrelated to the contents of this research. All other authors have no disclosures.

Title: PRC2 Loss Drives Genome Instability in Malignant Peripheral Nerve Sheath Tumors

Purpose: Malignant Peripheral Nerve Sheath Tumors (MPNSTs) have been characterized with recurrent chromosomal imbalances, including gains, losses, and rearrangements. In most cases a component of the Polycomb Repressive Complex 2 (PRC2) is lost, which results in a global loss of the repressive mark H3K27me3. MPNSTs that have loss of the PRC2 also present with a higher number of chromosomal abnormalities, suggesting a link between genome instability and the PRC2. Topoisomerase IIα (Topo II) is located on recurrent gain site chromosome 17q, and the resulting upregulation is linked to poor survival in MPNST patients. Topo II is an essential enzyme that maintains genome stability by deconcatenating sister chromatids during mitosis to release tension from supercoils and untangle knots in DNA. The ability of Topo II to efficiently deconcatenate DNA relies on its chromatin tethering domain and that domain’s interaction with methylated histones, including H3K27me3. The twin purposes of this study were to determine if loss of the PRC2 and H3K27me3 can cause mitotic defects in human Schwann linage cells and if drugs that target the mitotic apparatus are selectively toxic to PRC2-deficient cells. In this way, this study may inform therapeutic opportunities for treating PRC2-deficient MPNSTs and suggest MPNST-specific mechanisms for the acquisition of genome instability.

Methods: We utilized a human induced pluripotent stem cell (iPSC)-derived model system, iPSC-Schwann cell precursors (iPSC-SCPs) to understand how MPNSTs acquire genome instability over time with loss of PRC2. This model includes loss of NF1, CDKN2A/CDKN2B, and SUZ12. To visualize errors in mitosis, including ultra-fine bridges (UFBs) and aberrant chromosome segregation, we used fluorescence microscopy. To determine if PRC2-deficient cells are more sensitive to drugs targeting mitosis, we conducted a drug screen in immortalized NF1-deficient Schwann cells. To assess how PRC2 loss contributes to tumorigenesis, we are analyzing anchorage independent growth in vitro. After tumor formation, we will determine if PRC2 loss leads to more genome instability via low-pass whole genome sequencing to determine copy number alterations and structural variants. 

Results: Our studies revealed that key indicators of genomic instability, UFB formation and aberrant segregation, are increased in NF1- and CDKN2A/CDKN2B-deficient iPSC-SCPs treated with an inhibitor of PRC2. We have also shown that immortalized NF1-deficient Schwann cells with PRC2 loss have a heightened sensitivity to mitosis-targeted drugs, including Haspin Kinase and Topo II inhibitors, revealing the potential for therapeutic exploitation.

Conclusion: This study will deepen our understanding of the biological consequences of PRC2 loss in MPNSTs and establish a rationale for the use of drugs targeting the mitotic apparatus in PRC2 deficient tumors. We are integrating mechanistic studies with preclinical therapeutic validation to understand the acquisition of genome instability in these tumors and inform new treatment opportunities.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Authors: Kavita Y Sarin, MD PhD1, Kexin Huang PhD2, Vandhana Krishnan3, Jeffrey Sagun BS1, Ekshika Patel BS1, Xing Hu PhD1, Katya Vera BS1, Peter Caroline MS1, Jasmine Mehta BS1, Jeanie Ramos BS1,  Hanqi Caroline Yao BS1, Audris Chiang MD1, Michelle Lin BS1, Said Chosro Farschtsch MD4, Frank D. Buono, PhD7, Frank McCormick PhD6, Jaishri O. Blakeley MD7, Carlos G. Romo MD7

1Department of Dermatology, Stanford University 2Department of Computer Science, Stanford University 3gRED Computational Sciences, Genentech Inc  4 Department of Neurology, University Medical Center Hamburg - Eppendorf 5Department of Psychiatry, Yale University  6Dept. of Cellular and Molecular Pharmacology, UCSF, 7Department of Neurology, Johns Hopkins University

Funding: The Neurofibromatosis Therapeutic Acceleration Program  Disclosures: Kavita Sarin is a scientific advisor for NFlection Therapeutics. Carlos Romo is a consultant for Alexion Pharmaceuticals and SpringWorks Therapeutics. Jaishri Blakeley is a consultant for Alexion Pharmaceuticals Therapeutics and SpringWorks Therapeutics.

 Integrated GWAS with a functional genomics knowledge graph (KGWAS) identifies six novel loci associated with cutaneous neurofibroma development. 

Introduction. Cutaneous neurofibromas (cNFs) are benign skin tumors that develop in the majority of individuals with Neurofibromatosis Type 1 (NF1), often leading to significant physical and psychological morbidity. Despite their prevalence and impact, there is considerable variability in the number of cNFs that develop across affected individuals, posing a major challenge to understanding and treating these burdensome lesions. Although over 3,000 pathogenic variants in the NF1 gene have been identified, few have been linked to cNF number, suggesting that genetic modifiers outside the NF1 locus likely play a critical role in influencing individual susceptibility to tumor development.

Purpose. This study aims to identify genetic modifiers of cNF development by leveraging a large, crowd-sourced dataset that integrates genetic, photographic, and clinical information from individuals with NF1.

Methods.  A cohort of 607 participants with NF1 aged > 40 contributed photographs covering seven body regions, completed clinical surveys, and provided saliva samples for whole-genome sequencing. Participants were stratified into four groups based on cNF number as measured by the investigators from submitted photographs: 1-10, 11–100, 101–500, and more than 500 tumors. We applied Knowledge Graph GWAS (KGWAS), a novel geometric deep learning framework that integrates genome-wide association summary statistics with a large-scale functional genomics knowledge graph. This method enhances detection power in small-cohort GWAS by assessing the association strength of genetic variants through aggregated biological evidence from interacting molecular elements within the graph.

Results. KGWAS identified six suggestive loci associated with increasing cNF number at a significance threshold of P < 5 × 10⁻⁷. These novel SNPs occur within nerve and brain tissue-specific regulatory elements and implicate loci involved in neuronal cell survival, epithelial-mesenchymal transition, and axonal guidance and polarity. These loci may represent novel genetic modifiers influencing cNF formation in individuals with NF1.  

Conclusions. Potential genetic modifiers of cNF number in adults with NF1 have been identified, offering new insights into the biological pathways driving tumor development and potential targets for future therapeutic intervention

{{Cutaneous and subcutaneous neurofibroma}}

Authors: Devon ­Gunter BA^{1, 2}, Patricia Richey MD^1,2,3, Margaret Funk MS^1,2, Jamie Hu MD^1,2, Linh Pham BS^1,2, Fernanda H. Sakamoto MD PhD^1,2, Scott Plotkin MD PhD⁴, Justin Jordan MD MPH⁴, Joshua M. Roberts PhD⁵, William Farinelli BS^1,2, Lilit Garibyan MD PhD^1,2, Jaishri Blakeley MD⁵, R. Rox Anderson MD^1,2

1. 1.    Wellman Center for Photomedicine, Massachusetts General Hospital, Boston, MA

   2.    Department of Dermatology, Harvard Medical School, Boston, MA

   3.    Department of Dermatology, Boston University, Boston, MA

   4.    Department of Neurology, Massachusetts General Hospital, Boston, MA

   5.    Department of Neurology, Johns Hopkins University, Baltimore, MD

Funding: The Neurofibromatosis Therapeutic Acceleration Program (NTAP) at Johns Hopkins University.

Disclosures: No relevant financial relationships or conflicts of interest.

Title: Dosimetry for Treatment of Cutaneous Neurofibromas (cNFs) by Surfactant Injection

Purpose: We previously found that a single injection of two FDA-approved surfactants (deoxycholate and polidocanol) is effective in reducing cutaneous neurofibromas (cNFs) both in-vivo and ex-vivo^1,2. The purpose of this clinical study was to examine dose-response of these agents. 

Methods: 20 adults (ages 26-74; average age 48±16; 5M/15F) with small (2-8mm) cNFs were recruited in an Institutional Board Review-approved study. Tumors amenable to treatment were selected, then randomized into either 1) deoxycholate or  2) polidocanol groups. The individual lesions within these groups were then randomized to treatment or control (minimum of 6 cNFs per condition, with no upper limit for treated cNF). Deoxycholate-treated tumors received intratumoral injection of 10mg/mL deoxycholate (Kybella^R, Allergan-AbbVie); polidocanol-treated tumors received intratumoral injection of 1% polidocanol (Asclera^R, Merz). Control lesions received no treatment and were observed only for comparison. Each treatment had 6 escalating doses, determined by a fixed ratio of tumor volume to injected surfactant volume. Tumor diameter was used to approximate tumor volume. The 6 doses used were: 0.50x, 0.75x, 1.00x, 1.25x, 1.50x, or 1.75x the volume of the corresponding tumor. Topical anesthetic (5% lidocaine/prilocaine) was applied for 40 minutes before treatment. The following outcome measures were assessed at baseline, 3 months, 6 months, and 12 months post-treatment: 1) tumor height and volume (via 3D Cherry Imaging^R); 2) pain score (0-10); 3) clinician and patient assessment of tumor clearance; 4) and side effects (by CTCAE v5 scale), including inflammation, pigmentation, and ulceration. Biopsies were obtained from 13 participants 3 months post-treatment. 18 of 20 subjects completed all assessments, and 396 cNFs total (107 deoxycholate-treated; 102 polidocanol-treated; 187 untreated controls) were measured over all timepoints.

Results: No adverse events > grade 2 occurred. Treatment with both surfactants showed a significant (p<0.05) decrease in tumor volume. Tumor reduction increased with the volume injected, for both agents (Figure 2).   

Conclusion: A single injection of either polidocanol or deoxycholate into NF1-associated cNFs causes significant, dose-dependent reduction of tumor volume and height, as shown. Injected volumes up to 1.75x the tumor volume are well-tolerated. Injection causes temporary inflammation and mild pain (deoxycholate: 3.2±1.6, polidocanol: 3.3±1.8). Local pigmentation changes occurred in a minority of subjects, which persisted in only one. These local-injection treatments for cNF are well-tolerated and promising.  

References:

1. Funk, M, et al. Preliminary study using deoxycholic acid and polidocanol injections for the treatment of cutaneous neurofibromas. Presented at: Children’s Tumor Foundation NF Conference; June 24-27, 2024; Scottsdale, AZ.
2. Richey P, Funk M, Sakamoto F, et al. Noninvasive treatment of cutaneous neurofibromas (cNFs): Results of a randomized prospective, direct comparison of four methods. J Am Acad Dermatol. Published online December 10, 2023. doi:10.1016/j.jaad.2023.11.058

{{Cutaneous and subcutaneous neurofibroma}}

Understanding the role of apelin-mediated angiogenesis in NF2-associated tumors.

Srirupa Bhattacharyya1 (PhD), Roberta L. Beauchamp1 (BA), Vijaya Ramesh1(PhD)

Department of Neurology and Center for Genomic Medicine, Massachusetts General Hospital

Funding: Children’s Tumor Foundation Young Investigator Award (to S.B), National Institutes of Health R01 NS113854 (to V.R)

Disclosure of relevant financial relationships: The authors have no relevant financial relationships to disclose.

Purpose: NF2-associated meningiomas and schwannomas are highly vascular tumors, and while VEGF inhibition with Bevacizumab has benefited some NF2-related schwannomas, a majority of NF2-associated meningiomas often remain nonresponsive suggesting alternative angiogenic mechanisms. This study aimed to investigate the role of the angiogenic peptide apelin and its regulation by the mTORC1 pathway in NF2-null meningiomas as a potential strategy to overcome resistance to Bevacizumab for meningiomas.

Methods: We quantified APLN expression using qRT-PCR and the presence of its receptor (APLNR/APJ) by immunoblotting in several NF2-deficient meningioma lines. The effect of exogenous apelin on downstream signaling was examined in two immortalized NF2-null meningioma lines by assessing the phosphorylation status of key substrates. Given that mTORC1 signaling is known to regulate the expression of various angiogenic factors and considering that NF2 loss leads to aberrant activation of the mTORC1 pathway, we investigated the role of mTORC1 in regulating apelin expression. Additionally, we developed a 3D in vitro meningioma spheroid model co-cultured with human umbilical vein endothelial cells (HUVECs) to mimic vascularization and evaluated endothelial sprouting under basal conditions, and the angiogenic impact of apelin stimulation. Finally, we explored the role of mTORC1 by treating the angiogenesis model with RMC-6272.

Results: Increased expression of APLN in meningioma lines was confirmed using qRT-PCR, and immunoblotting validated the presence of APLNR/APJ receptor in all cell lines tested. Stimulation with exogenous apelin revealed activation of several downstream APJ signaling substrates that are shown to play a major role in angiogenesis and proliferation. Notably, in the 3D meningioma co-culture model, endothelial cells showed formation of robust primitive tube-like structures, reminiscent of a crude vessel-like appearance and interestingly basal conditions also sustained angiogenic sprouting, highlighting the importance of meningioma cell derived factors in angiogenesis. There was also an increase in both the total number of endpoints and the total number of junctions upon stimulation with apelin indicating a potent angiogenic effect of apelin. Treatment with the mTORC1 inhibitor RMC-6272 led to down-regulation of apelin expression, along with a significant reduction of sprouting angiogenesis suggesting potential involvement of the mTORC1 pathway in regulating angiogenesis of NF2- deficient meningiomas.

Conclusion: Our study establishes the importance of the angiogenic peptide apelin in NF2-associated tumors. Moreover, our results demonstrate, for the first time that meningioma spheroids alone are sufficient to initiate angiogenesis in the absence of external growth factors and suggest a role for mTORC1 signaling in meningioma vascularization.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Application of Mek inhibitor in patients with NF

Introduction: 

NF is classified into three main types: NF1, NF2, and schwannomatosis. NF1, the most prevalent form, is associated with mutations in the NF1 gene and often presents with cutaneous neurofibromas and plexiform neurofibromas. NF2, caused by mutations in the NF2 gene, typically involves bilateral vestibular schwannomas. Despite advances in surgical techniques, the treatment of NF remains challenging. Recent therapeutic developments have introduced MEK inhibitors as a promising treatment for plexiform neurofibromas in NF1. 

Case Presentation: We present 3 patients with NF using Mek inhibitor profiles, 2 NF1 and 1 NF2. 

Case 1: 14 year old boy diagnosed with NF1. Four years ago a right popliteal fossa swelling was found, which grew gradually, affecting joint mobility and limping with pain. MRI suggested neurofibromatosis（Fig.1）. After 1 week of treatment with Mek inhibitors, he developed a rash on the face and chest, posterior chest, and a rash on the trunk with pruritus. Dermatology consultation suggested the possibility of facial thrush and the trunk rash was a possible drug rash. After 4 months of continued medication, the tumor began to soften and the pain was reduced.

Case2: A 10 year old boy was diagnosed with NF1. His mother had a history of NF1. 3 years ago the child presented with a subcutaneous mass on the right side of the neck, which gradually increased in size and was often associated with pain. MRI suggested plexiform neurofibromatosis (Fig.2). After 4 months of treatment with Mek inhibitor, the texture of the right neck mass became softer. The pain disappeared.

Case3:A 19-year-old male underwent 3 surgeries for intracranial multiple meningiomas and intravertebral ventricular meningiomas. Molecular testing suggested NF2 p.Tyr150Ter mutation, and MRI showed multiple enhancing nodules in the bilateral internal auditory canal and Meckel's cavity, which led to the diagnosis of NF 2 (Fig.3). The patient voluntarily requested to take Mek inhibitor, and after 8 months of medication, the patient complained of hearing improvement without other side effects or complications, and is still under further follow-up.

These inhibitors have shown efficacy in reducing tumor volume and improving patient outcomes. However, the long-term efficacy and safety of MEK inhibitors in NF1 patients remain under investigation. Additionally, the potential application of MEK inhibitors in NF2 is still unexplored, warranting further research to determine their effectiveness in this context. This highlights the need for continued investigation into targeted therapies to improve the management of NF.

Fig.1

MRI of Case 1: right knee joint subcutaneous and soft tissues multiple abnormal enhancement foci, combined with the history of the disease, consider neurofibromatosis.

Fig.2

MRI of Case 2 showed multiple irregular nodules on the right side of the cervical musculature, under the sternocleidomastoid muscle, and in the right axilla. The tumor extends along the intervertebral foramina into the spinal canal, the right intervertebral space is enlarged. Combined with the medical and family history, plexiform neurofibromatosis was considered. 

Fig.3 

MRI of Case 3 showed multiple enhancing nodules in the bilateral internal auditory canal and Meckel's cavity, which led to the diagnosis of neurofibromatosis type 2 (NF2).

{{Plexiform neurofibroma}}

Identification of different Malignant Peripheral Nerve Sheath Tumor types in NF1 patients

Presenting author: Eduard Serra, PhD

Germans Trias i Pujol Research Institute (IGTP)

Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive soft-tissue sarcomas appearing either sporadically or in the Neurofibromatosis type 1 (NF1) context. MPNSTs tend to metastasize and have a poor prognosis. The diagnosis of MPNSTs can be challenging and heterogeneity exists, since different tumor entities may share overlapping histological characteristics. Our working model on MPNST development is as follows: MPNST initiates progression through the loss of key tumor suppressor genes (TSG) (NF1, CDKN2A, SUZ12/EED, TP53) in the cell of origin. Then, a general genomic rearrangement occurs, that needs to be compatible with cell viability for tumor progression. The resulting cell identity, with its expression and functional properties, will define MPNST histology and clinical behavior. The present project aimed to clarify the genomic characteristics of MPNSTs, evaluate their utility for understanding MPNST heterogeneity and identify any potential biological and clinical value.

We performed a comprehensive genomic analysis in an initial set of 20 clinically diagnosed MPNSTs that helped us identify recurrent genetic and genomic features characteristic of MPNSTs: TSG inactivation signature, low mutation burden, absence of activating mutations, global LOH status and copy-neutral (CN) LOH regions and specific copy number genomic profile. These genomic characteristics were confirmed using 50 additional MPNSTs from Cortes-Ciriano et al. (2023) we completely re-analyzed. With the extended set of MPNSTs and considering  these genomic features, we identified three distinct types of MPNSTs present in NF1 patients, mainly defined by their combination of inactivated TSGs.

The first group of MPNSTs bears the inactivation of NF1, CDKN2A and PRC2, exhibits a conventional histology and loss of H3K27 trimethylation. A second group of MPNSTs bears the inactivation of NF1, TP53 and PRC2, and associates with conventional histology with heterologous elements, loss of H3K27 trimethylation and, commonly, a complete 2n genome in LOH. The third group is composed of tumors found only in NF1 patients, bearing the inactivation of NF1 and CDKN2A, highly rearranged genome, with no activating mutations, and associates with a conventional MPNST histology, maintaining the H3K27 trimethylation. Finally, we identified a highly heterogeneous group of tumors, diagnosed initially as MPNSTs, with different TSG inactivation signatures, with activating mutations and diverse histological presentations and methylation patterns. This group needs further characterization but is probably composed of many tumors not being MPNSTs.

We will present the potential biological and clinical implications of this classification of NF1-associated MPNSTs. 

Full author list:

Miriam Magallón-Lorenz^1,2, Juana Fernández-Rodríguez^3,4, Helena Mazuelas², Itziar Uriarte-Arrazola², Sara Ortega-Bertran⁵, Edgar Creus-Bachiller⁵, Aleix Mendez¹⁶, Eva Rodríguez⁶, Mariona Suñol⁶, Carlota Rovira⁶, Raquel Arnau⁶, Juan Carlos Lopez-Gutierrez⁷, Alicia Castañeda⁸, Isabel Granada¹⁶, Alba Hernandez⁹, Gustavo Tapia⁹, Maria Saigí¹⁰, Marc Cucurull¹⁰, Ignacio Blanco¹¹, Claudia Valverde^12,13, Cleofé Romagosa¹⁴, Héctor Salvador⁸, Conxi Lázaro^4,5, Meritxell Carrió², Eduard Serra^2,4,*, Bernat Gel^1,2,15,*

1. Translational Genomics and Bioinformatics Group, Germans Trias I Pujol Research Institute, Can Ruti Campus, Badalona, Barcelona, 08916, Spain
2. Hereditary Cancer Group, Germans Trias i Pujol Research Institute (IGTP); Can Ruti Campus, Badalona, Barcelona, 08916; Spain
3. Mouse Lab, IDIBELL, Hospitalet de Llobregat, Barcelona, Spain
4. Centro de Investigacion Biomedica en Red de Cancer (CIBERONC), Madrid, Spain
5. Hereditary Cancer Program, Catalan Institute of Oncology (ICO-IDIBELL), L'Hospitalet de Llobregat, Barcelona, 08098; Spain
6. Pathology Department, Sant Joan de Déu Barcelona Children’s Hospital, 08950 Barcelona, Spain
7. Department of Pediatric Surgery, University Hospital La Paz, Madrid, Spain
8. Pediatric Cancer Center Barcelona, Sant Joan de Déu Barcelona Children’s Hospital, 08950, Barcelona, Spain
9. Department of Pathology, Germans Trias i Pujol University Hospital (HUGTiP), Can Ruti Campus, Badalona, Barcelona, 08916; Spain
10. Catalan Institute of Oncology (ICO), Germans Trias i Pujol Research Institute (IGTP)
11. Clinical Genomics Unit, Germans Trias i Pujol University Hospital (HUGTiP), Can Ruti Campus, Badalona, Barcelona, 08916; Spain
12. Department of Medical Oncology, Hospital Universitari Vall d’Hebron, Barcelona, Spain.
13. Vall d’Hebron Institute of Oncology, Barcelona, Spain.
14. Department of Pathology, Hospital Vall d’Hebron, Barcelona, Spain
15. Departament de Fonaments Clínics, Universitat de Barcelona, 08036, Barcelona, Spain
16. Hematology Department, Hospital Germans Trias i Pujol, Institut Català d'Oncologia, Josep Carreras Leukaemia Research Institute, Barcelona, Spain

* Co-last authors

Funding: This work has been supported by the Instituto de Salud Carlos III National Health Institute funded by FEDER funds—a way to build Europe—[PI20/00228; PI23/00583; PI23/00422]; Fundació La Marató de TV3 (51/C/2019). The work has also supported by Fundación Proyecto Neurofibromatosis, and the Generalitat of Catalonia (2021 SGR 00967).

References: Cortes-Ciriano, Isidro et al. “Genomic Patterns of Malignant Peripheral Nerve Sheath Tumor (MPNST) Evolution Correlate with Clinical Outcome and Are Detectable in Cell-Free DNA.” Cancer discovery vol. 13,3 (2023): 654-671. doi:10.1158/2159-8290.CD-22-0786

{{BIOINFORMATICS AND COMPUTATIONAL OMICS}}

Authors: Song Li, MD¹, Yong Qiu, MD¹, Zezhang Zhu, MD¹, Saihu Mao, MD¹

1 Division of Spine Surgery, Department of Orthopedic Surgery, Nanjing Drum Tower Hospital, The Affiliated Hospital of Nanjing University Medical School, Zhongshan Road 321, Nanjing 210008, China

Corresponding author: Saihu Mao, Siemens_636@163.com

Disclosures: The authors declare that they have no competing interests.

Title: Surgical treatment of dystrophic kyphoscoliosis secondary to neurofibromatosis type 1: is three-column osteotomy necessary?

Purpose: To evaluate whether the patients with thoracic kyphosis secondary to neurofibromatosis type 1 (DKS-NF1) must undergo three-column osteotomy during deformity correction surgery.

Methods: 84 patients with DKS-NF1 were retrospectively analyzed, and the average age was 17.7±6.9 years. There were 50 cases with single curve, 18 cases with double curves, and 16 cases with triple curves; kyphosis was found in 42 cases in the thoracic area, 31 cases in the thoracolumbar area, and 11 cases in the lumbar area. According to whether the patient has undergone three column osteotomy, they were divided into two groups: None three column osteotomy group (N-3CO) and three column osteotomy group (3CO). The preoperative, postoperative and follow-up imaging parameters of the two groups were measured and analyzed to evaluate the surgical effect.

Results: 74 patients were divided into the N-3CO group and 3CO consisted of 10 patients. The age of patients in the N-3CO group was significantly younger than that in the 3CO group (15.8 ± 4.8 years vs. 29.4 ± 10.2 years, P<0.001), and the proportion of preoperative traction in this group was significantly higher than that in the 3CO group (26/74 vs. 0, P=0.027). The patients with kyphosis in the 3CO group was mainly located in the thoracolumbar and lumbar area, significantly higher than that in the N-3CO group (10/10 vs. 32/74, P=0.003). The magnitude of kyphosis in the two groups were 73.8±20.9° and 63.1±21.4° before surgery, respectively (P=0.136). After surgery, they were corrected to 43.1±20.9° and 21.1±22.8°, respectively (P=0.003), with correction rates of 43.7±19.6% and 84.1±78.7%, respectively (P<0.001). At the last follow-up, they were maintained at 46.5±20.9° and 24.6±25.5°, respectively (P=0.003). The Cobb angle of the main curve was corrected from preoperative 83.0±29.0° and 66.3±17.7° (P=0.081) to postoperative 50.6±20.8° and 40.8±15.6° (P=0.155), with correction rates of 38.3±16.6% and 39.3±12.7% (P<0.849), respectively. At the last follow-up, they were maintained at 52.3±20.5° and 43.1±18.2°, respectively (P=0.185). The proportion of multi-rod system application and screw density in 3CO group were significantly higher than that in N-3CO group (8/10 vs. 20/74, P=0.002; 72.0±11.3% vs. 61.4±14.6%, P=0.033). The incidence of complications in the two groups was 12.2% (N-3CO group, 9/74) and 20% (3CO group, 2/10), respectively, with no statistically significant difference (P=0.613).

Conclusion: Three column osteotomy is mainly used to treat adult kyphosis in DKS-NF1 patients. While the none-three-column-osteotomy methods were mainly applied in young patients. Posterior approach with posterior column osteotomy was the mostly used plan, while anterior supplementary fusion was also an important option. For severe cases, preoperative Halo gravity traction can be used as a routine strategy. This can basically meet the treatment needs of young patients with DKS-NF1, and has good feasibility and safety as well as ideal deformity correction.

Key Words: NF1; dystrophic kyphoscoliosis; three-column osteotomy

{{Bone abnormalities}}

Authors: Song Li, MD¹, Yong Qiu, MD¹, Zezhang Zhu, MD¹, Saihu Mao, MD¹

1 Division of Spine Surgery, Department of Orthopedic Surgery, Nanjing Drum Tower Hospital, The Affiliated Hospital of Nanjing University Medical School, Zhongshan Road 321, Nanjing 210008, China

Corresponding author: Saihu Mao, Siemens_636@163.com

Disclosures: The authors declare that they have no competing interests.

Title: The incidence, mechanism, and clinical outcomes of postoperative coronal imbalance in dystrophic lumbar scoliosis secondary to Neurofibromatosis Type 1

Purpose: To investigate the incidence, mechanism, and clinical outcomes of postoperative coronal imbalance in dystrophic lumbar scoliosis secondary to Neurofibromatosis Type 1.

Methods: A retrospective analysis was conducted on the data of 33 patients with dystrophic lumbar scoliosis secondary to Neurofibromatosis Type 1 (DLS-NF1) (with the apex located at L1 or below) who underwent surgical correction in our hospital from January 2007 to January 2018. There were 21 males and 12 females, with an average age of 14.5 ± 3.2 years and an average follow-up time of 34.0 ± 26.3 months. 11 cases were fused with distal internal fixation to L4 (i.e., the number of compensatory intervertebral discs at the distal end was 2), and 22 cases were fused to L5 (15 cases) or below (5 cases) (i.e., the number of compensatory intervertebral discs at the distal end was 1 or none). According to coronal balance (CB), patients with scoliosis are classified into three types: Type A (balanced state: CB<30mm); Type B (concave imbalance: CB≥30mm and C7PL located on the concave side of the main curve); Type C (convex imbalance: CB≥30mm and C7PL located on the convex side of the main curve).

Results: Satisfactory correction of lumbar scoliosis and kyphosis were obtained after surgery. CB increased from an average of 22.4±17.5mm before surgery to 27.6±10.9mm after surgery. There were 18 Type A patients, 0 Type B patients, and 15 Type C patients before surgery. After surgery, there were 17 Type A patients, 0 Type B patients, and 16 Type C patients. 16 cases (48.5%, 16/33) had coronal imbalance after surgery, of which 5 cases (27.8%, 5/18) had preoperative Type A conversion to postoperative Type C, and 11 cases (73.3%, 11/15) had preoperative Type C conversion to postoperative Type C (5/18 vs. 11/15, p=0.015). In addition, the proportion of postoperative Type C patients with a distal compensatory disc count of 1 or none was significantly higher than that of Type A patients (17/20, vs. 5/13, p=0.009). All patients with postoperative coronal imbalance showed improvement in decompensation 3-6 months after surgery.

Conclusion: The incidence of postoperative coronal imbalance is higher in DLS-NF1 patients (48.5%), which may be mainly due to preoperative convex coronal imbalance and fewer postoperative distal compensatory discs. However, all postoperative coronal imbalance patients can improve during follow-up.

Key Words: NF1; dystrophic lumbar scoliosis; coronal imbalance

{{Bone abnormalities}}

Authors: Song Li, MD1, Yong Qiu, MD1, Zezhang Zhu, MD1, Saihu Mao, MD1

1 Division of Spine Surgery, Department of Orthopedic Surgery, Nanjing Drum Tower Hospital, The Affiliated Hospital of Nanjing University Medical School, Zhongshan Road 321, Nanjing 210008, China

Corresponding author: Saihu Mao, Siemens_636@163.com

Disclosures: The authors declare that they have no competing interests.

Title: Identification of key genes and pathways of decreased bone mineral density in NF1 patients with dystrophic scoliosis by microarray and integrated gene network analysis

Purpose: To explore the differential genetic expression profile, Gene Ontology terms, and Kyoto Encyclopedia of Genes and Genomes pathways in human trabecular bone (HTB)-derived cells of dystrophic scoliosis secondary to neurobromatosis type 1 (DS-NF1) and compare these to normal controls. 

Methods: Bone mineral density (BMD) was measured and compared between DS-NF1 patients and normal controls. Then, microarray analysis was used to identify differentially expressed genes (DEG) of HTBs between DS-NF1 patients and normal controls. Functional and pathway enrichment analysis was performed based on GO and KEGG pathway databases. Then, STRING database, Cytoscape and MCODE were used to construct the protein-protein interaction (PPI) network and screen hub genes. Further, the hub genes and gene clusters determined by module analysis were analyzed by pathway enrichment. Six potential key genes were selected for reverse transcription polymerase chain reaction (RT-PCR). 

Results: The BMD of lumbar spine, femoral neck and total femur in DS-NF1 patients significantly decreased. Bioinformatics analysis showed that there were 401 previously unrecognized DEGs (238 up-regulated genes and 163 down regulated genes) in HTBs of DS-NF1 patients, which were mainly enriched in immune response, type I interferon signaling, TNF signaling pathway and RIG-I-like receptor signal pathway. Five hub genes such as STAT1, OASL, IFIH1, IRF7 and MX1 were identified by PPI network. These genes were mainly enriched in Jak-STAT and RIG-I-like receptor signaling pathway. PPI network analysis also found an independent dysregulated protein cluster containing CCL2, CXCL1, CXCL3, CX3CL1, TLR1 and CXCL12. The former five genes were significantly down-regulated, while CXCL12 was significantly up-regulated. All these genes are closely related to immune response, suggesting that immune response may play an important role in the pathogenesis of DS-NF1 with decreased bone mass.

Conclusion: In this study, STAT1, OASL, IFIH1, IRF7 and MX1 were identified as the key genes of DS-NF1-related bone mass reduction. Immune response may play a key role in this process, and CXCL12 mediated osteogenesis may play a protective role in the reduction of BMD. This requires further research to clarify the pathogenesis of osteopenia in DS-NF1.

Key words. NF1, bone mineral density, osteopenia, immune response

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Title: Discovery of bicyclic peptides that engage wild-type KRAS and block downstream signaling in NF1^(-/-) Schwann cells.

Authors:  Sandeep Lohan^a,b, Bipasana Shakya^a,b, Jian Liu^c, Aslamuzzaman Kazi^a, Grant Davis^c, Said Sebti^a, Deeann Wallis^c, Matthew Hartman^a,b

Addresses:

^aDepartment of Chemistry, Virginia Commonwealth University, 1001 W Main St Richmond, VA 23284-2006, USA

^bMassey Comprehensive Cancer Center, Virginia Commonwealth University, 410 College St. Richmond, VA 23219, USA.

^c Department of Genetics, University of Alabama at Birmingham, Birmingham, AL 35294, USA

Abstract:

Overactivation of Ras is the primary basis for neurofibromatosis type 1 (NF1) disease.  Despite intense drug discovery efforts, Ras proteins have proven to be difficult targets for small molecule drug discovery.  Therapies for NF1 have; therefore, resorted to targeting kinases involved in downstream signaling which are more “druggable.”  Macrocyclic peptides as a therapeutic modality are attracting increasing attention due to their ability to bind to targets intractable to small molecules; however, lack of cell permeability is an ongoing challenge.  To find peptides that engage KRAS in cells, we used mRNA display to generate bicyclic libraries, with one cycle containing a short, cyclic cell penetrating peptide to enable cellular entry.  After 7 rounds of in vitro selection, we identified putative hits from next generation sequencing (NGS) data.  After scale up synthesis, surface plasmon resonance (SPR) and biolayer interferometry (BLI) data revealed Kds as low as 100-200 nM against the GDP-bound and GcP-bound conformation of WT KRAS. Treatment of IPN 97.4 NF-1^(-/-) Schwann cells with our peptides at micromolar concentrations led to a reduction of phospho-ERK in a time and dose-dependent manner.  Similar effects were observed on cancer cell lines with aberrant KRAS signaling.   Together, our data highlights the power of mRNA display to find macrocyclic peptides with dual properties of cell penetration and target binding, and provides leads for future drug discovery for molecules that directly engage Ras proteins for the treatment of NF1.

{{DRUG DISCOVERY AND DEVELOPMENT}}

Authors:

Roope A. Kallionpää, PhD, University of Turku, Finland

Milla Lehto, BSc, University of Turku, Finland

Elina Uusitalo, PhD, University of Turku, Finland

Roosa E. Kallionpää, PhD, Auria Biobank, University of Turku and Turku University Hospital, Finland

Pauliina Kronqvist, MD, PhD, University of Turku and Turku University Hospital, Finland

Sirkku Peltonen, MD, PhD, University of Helsinki and Helsinki University Hospital, Finland; University of Turku and Turku University Hospital, Finland; University of Gothenburg and Sahlgrenska University Hospital, Sweden

Juha Peltonen, MD, PhD, University of Turku, Finland

Funding: Children’s Tumor Foundation Young Investigator Award (Award ID: 2023-01-006; doi: https://doi.org/10.48105/CTF.CTF-2023-01-006.pc.gr.172004), Cancer Foundation Finland, Turku University Hospital, Helsinki University Hospital.

Disclosures: The authors declare no conflicts of interest.

Increased myxoid stroma in NF1-associated breast cancer

Purpose: Women with neurofibromatosis 1 (NF1) are at an increased risk for breast cancer, and the prognosis of NF1-associated breast cancer is often poor. Data from neurofibromas and model systems have shown that NF1 can have major effects on tumor microenvironment. We now aimed to characterize histological features of NF1-associated breast cancer.

Methods: A set 26 breast cancers from women with NF1 was analyzed. A frequency-matched control cohort of 75 breast cancers from non-NF1 women was retrieved from Auria Biobank (Turku, Finland). The matching of the control breast cancers accounted for age at diagnosis (<40, 40-50, ≥50 years), estrogen receptor status, HER2 status, tumor grade and histological subtype. A pathology specialist, blinded to the NF1 status of the samples, scored hematoxylin & eosin sections for myxoid stroma and tumor budding.

Results: Myxoid stroma was observed significantly more frequently in NF1-related breast cancers (69%) than in control breast cancers (40%), yielding an odds ratio (OR) of 3.33 (95% CI 1.20-10.1, P=0.013). Among the breast cancers with myxoid stroma, the proportion of myxoid stroma was higher in NF1-related breast cancers than in the control breast cancers (P=0.025). Especially breast cancers diagnosed after 50 years of age showed increased rate of myxoid stroma in NF1 versus the controls (OR 4.03, 95% CI 1.06-17.5, P=0.023). Tumor budding was detected in 50% of the NF1-related breast cancers and 37% of the control breast cancers, yet the difference was not statistically significant (OR 1.67, 95% CI 0.62-4.54, P=0.354). Upon further stratification, many NF1-related breast cancers showed single tumor budding complexes (NF1 42%, control 15%), and abundant budding was more common in the control breast cancers (NF1 7.7%, control 23%).

Conclusion: NF1 affects the stromal composition of breast cancer, which may contribute to the development and growth of NF1-associated breast cancer.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Single-Center Retrospective Case Series on LZTR1 Germline Mutations in Patients Without  Established Diagnosis of LZTR1-Related Schwannomatosis 

Navarro, Laura, BS¹; Guevara, Gabriel, BS, MS¹; Maga, Tara, PhD, MS, CGC¹; Balay, Lara, Tara, MSed, MS, CGC¹; Maraka, Stefania, MD¹

University of Illinois at Chicago¹

Introduction

Germline loss-of-function mutations in the leucine zipper-like transcription regulator 1 (LZTR1) gene are associated with the development of non-neurofibromatosis type 2 (NF2) related schwannomatosis (non NF2-SWN). Diagnostic criteria for LZTR1-related schwannomatosis (LZTR1-SWN) include the presence of schwannomas on imaging and a confirmed LZTR1 mutation. Schwannoma typically develop between 20-40 years of age. This case series presents three patients with incidental findings of LZTR1 germline mutations without confirmed schwannomas and thereby they do not meet diagnostic criteria for LZTR1-related schwannomatosis at this time.

Methods

We conducted a single-center retrospective study at the University of Illinois at Chicago, reviewing patients who underwent genetic testing between 2022-2024 and were found to have LZTR1 mutation without established diagnosis of LZTR1-SWN. Data collected included clinical features, imaging results, genetic and oncologic history, family history, age, and gender.

Results

Three patients with confirmed LZTR1 mutations were included. Patient 1 is a 51-year-old female with a history of invasive ductal carcinoma of the breast who underwent genetic screening due to her cancer history. She presented with bilateral tinnitus and thigh pain, but no focal neurologic deficits. Her family history included renal cell carcinoma and breast cancer. Neuroimaging, including MRI spine and MRI internal auditory canal (IAC) showed no schwannomas. Patient 2 is a 32-year-old female with papillary thyroid carcinoma, screened for genetic mutation due to a family history of ovarian, breast, and cervical cancer. She presented with bilateral tinnitus and balance problems but had no focal neurologic deficits. MRI of the spine and IAC were negative for schwannomas. Patient 3 is a 33-year-old female with a history of breast sarcoma and Cowden syndrome, who underwent genetic screening due to her cancer history. There were no focal neurologic deficits or pertinent positive symptoms at presentation. MRI of the abdomen revealed a T2 hyperintense liver lesion, and a PET scan identified hilar lymphadenopathy, which was biopsied and found to be benign.

Conclusion

This case series presents three patients with LZTR1 germline mutation, which were incidentally found on a screening test due to personal or family history of cancer. Given their young ages, they may develop schwannomas in the future, potentially meeting diagnostic criteria for LZTR1-SWN. Further research is needed to explore the potential association between LZTR1 germline mutation, LZTR1-SWN, extraneural malignancies and other cancer predisposition syndromes. 

{{Schwannoma}}

Authors: Dan Liu, Ph.D., Florida State University, USA; Pamela L. Wolters, Ph.D., National Cancer Institute, USA; Bonita P., Klein-Tasman, University of Wisconsin–Milwaukee, USA; Karin S. Walsh, Psy.D., Children’s National Hospital, USA; Jonathan M. Payne, Psy.D., Murdoch Children's Research Institute, Australia; Natalie Pride, Ph.D., Children's Hospital at Westmead, Australia; Stephanie M. Morris, M.D., Kennedy Krieger Institute, USA; Yang Hou, Ph.D., Forida State University, USA

Funding: (a) Congressionally Directed Medical Research Programs, Neurofibromatosis Research Program [W81XWH2110504]; (b) Center for Cancer Research, National Cancer Institute, Intramural Research Program; (c) Florida State University Faculty Startup Funding; and (d) University of Kentucky Faculty Startup Funding. 

Disclosures: The authors declare no financial conflict of interest.

Title: Age-varying associations between ADHD symptoms and internalizing/externalizing behaviors in children with neurofibromatosis type 1: Integrative analyses of data from six institutions

Purpose: Children with neurofibromatosis type 1 (NF1) experience higher rates of Attention-Deficit/Hyperactivity Disorder (ADHD) and internalizing and externalizing behaviors compared to their typically developing peers. The severity of these symptoms flucatuates across different ages; however, the age-varying associations between ADHD symptoms and internalizing or externalizing behaviors in children with NF1 remain poorly understood. This study aims to investigate these associations across various ages to inform the development of age-appropriate interventions.

Methods: We integrated individual-level data from 685 children with NF1 (ages 3–18 years) across six institutions in the United States and Austrania, using integrative data analysis procedures. ADHD symptoms (inattention, hyperactivity/impulsivity, and combined ADHD) were assessed through parent-reported Conners Rating Scales and ADHD Rating Scales. Internalizing and externalizing symptoms were measured using the Child Behavior Checklist and the Behavior Assessment System for Children. Time-Varying Effect Modeling (TVEM), a nonparametric method, was used to examine the associations between ADHD symptoms and internalizing and externalizing symptoms across ages, while controlling for sex. TVEM estimates these associations as continuous functions of age, providing curvilinear intercept and slope estimates with 95% confidence intervals (CIs). Significant associations are indicated by 95% CIs that do not include zero, and significant age-related differences by non-overlapping 95% CIs at specific age points.

Results: Elevated inattention, hyperactivity/impulsivity, and combined ADHD scores were consistently correlated with elevated internalizing and externalizing symptoms from ages 3 to 17 (see Figure 1 for an example). The strength of these correlations remained stable across ages, with one exception. Specifically, the correlation between hyperactivity/impulsivity and externalizing symptoms was most pronounced during early childhood (ages 5–8), diminished through late childhood, reached its lowest point during early adolescence (ages 13–14), and then stabilized in middle adolescence (Figure 2).

Conclusion: Findings are consistent with existing research on the associations between ADHD symptoms and internalizing/externalizing behaviors in the general population. The results indicate that children with NF1 who exhibit elevated internalizing and externalizing symptoms should be continuously monitored for ADHD symptoms from early childhood through middle adolescence. Interventions targeting these symptoms may be more effective if they concurrently address ADHD symptoms, with particular attention to developmental stages. For example, early childhood interventions focusing on hyperactivity/impulsivity may be particularly beneficial for mitigating externalizing symptoms, in contrast to interventions in early adolescence.

Figure 1. Age-varying associations between inattention 

and internalizing symptoms. Significant associations are indicated by 95% confidence intervals (CIs) that do not include zero. Significant age differences are indicated by non-overlapping 95% CIs between specific age points. 

Figure 2. Age-varying associations between hyperactivity/impulsivity and externalizing symptoms. Significant associations are indicated by 95% confidence intervals (CIs) that do not include zero. Significant age differences are indicated by non-overlapping 95% CIs between specific age points. 

{{Cognition / Behavioral / Learning}}

Bae-Hoon Kim^{2, #}, Yeon-Ho Chung^{2, *}, Soyoung Park^{1, *}, Tae-Gyun Woo², Minju Kim², Bum-Joon Park^{1,2, #} 

¹ Department of Molecular Biology, College of Natural Science, Pusan National University, Busan, Republic of Korea. ² Rare Disease R&D Center, PRG S&T Co., Ltd, Busan, Republic of Korea. ^*, ^# These authors contributed equally to this work

Funding: This work was supported by National Research Foundation of Korea (NRF) grants funded by the Korean government (MSIT) (RS-2024-00399681, RS-2024-00339289 and RS-2024-00442484).

Title: Targeted Therapeutic Potential of PRG-N-01 in NF2-Related Schwannomatosis: Preclinical Evaluation

Purpose

Neurofibromatosis type 2-related schwannomatosis (NF2-SWN) is a genetic disorder characterized by the formation of multiple benign tumors within the nervous system, primarily intracranial and spinal schwannomas. NF2-SWN arises due to pathogenic mutations in the NF2 gene, leading to dysregulated tumor suppressor activity. Given the substantial surgical risks and high recurrence rates associated with multifocal tumors, the development of effective targeted therapies is imperative. However, to date, no approved pharmacological interventions exist.

Methods

To elucidate the molecular mechanism of action of PRG-N-01, a novel therapeutic candidate for NF2-SWN, we performed direct binding assays and mass spectrometry-based protein interaction studies. In vitro anti-proliferative effects of PRG-N-01 were assessed using primary Schwann cells derived from an NF2 mouse model and patient-derived schwannoma cells. In vivo therapeutic efficacy and prophylactic potential were evaluated through both intraperitoneal and oral administration in the NF2 mouse model (Postn-Cre; Nf2^f/f). Furthermore, RNA sequencing of dorsal root ganglia (DRG) was conducted to determine transcriptomic alterations following treatment. Pharmacological properties, including absorption, distribution, metabolism, excretion (ADME), pharmacokinetics, pharmacodynamics, and toxicology, were systematically characterized through preclinical investigations.

Results

PRG-N-01 was identified as a selective binder to the N-terminal kinase domain of TGFβ receptor type 1 (TβR1), disrupting its interaction with Raf kinase inhibitory protein (RKIP) and thereby stabilizing RKIP expression. In vivo administration of PRG-N-01 effectively suppressed schwannoma growth in the DRG, while early oral treatment demonstrated prophylactic benefits in NF2-SWN progression. Gene expression profiling revealed that PRG-N-01 downregulated tumor-promoting pathways while promoting transcriptional programs associated with Schwann cell differentiation and normal cellular metabolism. Preclinical evaluations confirmed that PRG-N-01 exhibits favorable drug-like properties, supporting its potential as a viable therapeutic agent.

Conclusions

Our findings provide compelling preclinical evidence supporting PRG-N-01 as a promising targeted therapy for NF2-SWN. These results establish a strong rationale for advancing PRG-N-01 into clinical trials, underscoring its potential as a novel therapeutic strategy for NF2-SWN patients.

THERAPEUTICS

Authors: Tingting Zheng^1,2†, Beiyao Zhu^1,2†, Zhichao Wang^1,2*, Qingfeng Li^1,2*

Affiliations:

¹ Department of Plastic and Reconstructive Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine; Shanghai, 200011, China.

² Neurofibromatosis Type 1 Center and Laboratory for Neurofibromatosis Type 1 Research, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine; Shanghai, 200011, China.

^†These authors contributed equally to this work. Email: srlala1022@sjtu.edu.cn (T.Z.); beiyao.zhu@sjtu.edu.cn (B.Z.); shsmuliujun@163.com (J.L.).

*Corresponding author. Email: dr.liqingfeng@shsmu.edu.cn (Q.L.); shmuwzc@163.com (Z.W.). Telephone: 13301990666 (Q.L.); 13816382311 (Z.W.).

Disclosures: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Funding: National Natural Science Foundation of China (82102344, 82172228).

Title: Gene therapy strategies and prospects for neurofibromatosis type 1

Purpose: To summarize the gene therapy strategies for neurofibromatosis type 1 (NF1) and related research progress. 

Methods: The recent literature on gene therapy for NF1 at home and abroad was reviewed. The structure and function of the NF1 gene and its mutations were analyzed, and the current status as well as future prospects of the transgenic therapy and gene editing strategies were summarized. 

Results: NF1 is an autosomal dominantly inherited tumor predisposition syndrome caused by mutations in the NF1 tumor suppressor gene, which impair the function of the neurofibromin and lead to the disease. It has complex clinical manifestations and is not yet curable. Gene therapy strategies for NF1 are still in the research and development stage. Existing studies on the transgenic therapy for NF1 have mainly focused on the construction and expression of the GTPase-activating protein-related domain in cells that lack of functional neurofibromin, confirming the feasibility of the transgenic therapy for NF1. Future research may focus on split adeno-associated virus (AAV) gene delivery, oversized AAV gene delivery and the development of new vectors for targeted delivery of full-length NF1 cDNA. In addition, the gene editing tools of the new generation have great potential to treat monogenic genetic diseases such as NF1, but need to be further validated in terms of efficiency and safety.

Conclusion: Gene therapy, including both the transgenic therapy and gene editing, is expected to become an important new therapeutic approach for NF1 patients.

Xin Zhang^*,MD    Xiaojun Yuan^#,MD

 Xinhua Hospital affiliated to Shanghai JiaoTong University School of Medicine, Shanghai, China;

Abstract

Purpose

Selumetinib, a MEK inhibitor, has been approved in China for treating symptomatic, unresectable plexiform neurofibromas (PN) in children aged 3 years and older with neurofibromatosis type 1 (NF1). This Phase I trial (CTR20200357) aims to evaluate the long-term efficacy and safety of Selumetinib in Chinese pediatric patients with NF1-associated PN, presenting updated findings from follow-up assessments conducted over a period of up to 4 years.

Methods

From December 2020 to February 2025, 16 pediatric patients with symptomatic/progressive PN received Selumetinib (25 mg/m² BID) until disease progression or intolerable adverse events (AEs). Efficacy endpoints included PN volume reduction (REiNS criteria), café-au-lait spots, health-related quality of life (HRQoL), pain, and physical function. Safety assessments included AEs and AEs of special interest (AESIs). MRI and clinical evaluations were performed every 4 cycles for the first 24 cycles, and every 6 cycles thereafter, with 14 patients completing over 42 cycles (median follow-up: 44 cycles).

Results

The cohort comprised 16 pediatric patients (median age: 11 years; range: 4-16). Patients received a median of 44 cycles of Selumetinib (range: 20-53). One patient discontinued treatment due to disease progression after the 20th cycle. At the initial data cutoff (DCO) (August 15, 2023), the overall response rate (ORR) was 81.3% (95% CI: 54.4%, 96.0%), with no complete responses (CR), 13 confirmed partial responses (cPR), 2 unconfirmed partial responses (uPR), and 1 stable disease (SD). Independent central review (ICR) reported an ORR of 62.5% (95% CI: 35.4%, 84.8%). All patients exhibited a reduction in target PN volume compared to baseline, with a mean reduction of -42.30% (±15.95%) by investigator assessment and -29.56% (±22.62%) by ICR. At the final DCO (February 28, 2025), 14 patients (87.5%) had a median volume reduction of 45.6% (range: 20.1%-61.2%). Clinical assessments indicated significant improvements in pain management, physical function, HRQoL, and symptom severity. Treatment-related AEs were reported by all patients, predominantly fever (38%) and rash (50%), all graded as 1 or 2 severity per CTCAE 5.0.

Conclusions

Selumetinib demonstrates durable efficacy, with a median response duration of 3.5 years, along with sustained functional benefits and a favorable safety profile in pediatric patients with NF1-associated PN. These findings support its long-term use with proactive management of AEs. Discrepancies between investigator assessments and independent central review highlight potential bias in subjective imaging evaluations, emphasizing the need for emerging AI imaging technologies to enhance objectivity in clinical trial assessments.

Plexiform neurofibroma

Authors: Sarah Morrow^1,2,3, Shelley A.H. Dixon¹, Christopher Davis¹, Colin Beach^1,3, Christine A. Berryhill¹, Emily Zhang¹, Christine A. Pratilas⁴, D. Wade Clapp¹, Steven D. Rhodes¹, Steven P. Angus^1,2,3.

¹Department of Pediatrics, IU School of Medicine, Indianapolis, IN

²Department of Pharmacology and Toxicology, IU School of Medicine, Indianapolis, IN

³Simon Comprehensive Cancer Center, IU School of Medicine, Indianapolis, IN

⁴Johns Hopkins University, School of Medicine, Baltimore, MD

Funding: This work was supported by the Young Investigator Award from the Children’s Tumor Foundation (SM), a Career Enhancement Program Award from the DHART SPORE (SPA), a Team JOEY Award from the Heroes Foundation (SPA), a New Investigator Award from the DOD NFRP (NF2000038) (SPA), and generous support from the Riley Children’s Foundation (DWC, SDR, SPA).

Title: Characterizing the role of ZNF423 in Neurofibromatosis Type 1 (NF1)-related malignant peripheral nerve sheath tumors (MPNST).

Purpose: Recent work in our lab utilized transcriptomics to analyze two SUZ12-deficient human MPNST cell lines with restored PRC2 activity, which revealed downregulation of ZNF423, a lineage-specific transcription factor. These results were validated in murine tumor cells-of-origin isolated from GEMMs of plexiform neurofibroma (PNF) and MPNST, which revealed ZNF423 was significantly upregulated in MPNST compared to both wild-type controls and PNF, suggesting a potential role for this protein in MPNST tumorigenesis. Our goal is to identify potential therapeutic targets for NF1-related MPNST by elucidating ZNF423-dependent signaling mechanisms that may influence MPNST lineage, identity and proliferation.

Methods: Human MPNST cell line models were employed for RNA interference, RNA sequencing, immunoblotting, proteomic analysis of the kinome, cell-based assays, and in vivo implantation.

Results: Short-interfering RNA (siRNA) depletion of ZNF423 from four human MPNST cell lines altered MPNST cell morphology, and significantly decreased cell proliferation and viability compared to controls. EdU/DAPI staining further confirmed these effects, showing a marked decrease in DNA synthesis and hence impaired cell cycle progression and proliferation in ZNF423 knockdown cells. To establish the biological mechanism underlying these phenotypic effects, we performed RNA sequencing, which revealed significant transcriptional changes, with overlapping gene expression patterns amongst the four cell lines. Shared significantly downregulated genes were enriched for pathways involved in proliferation, migration and development. In contrast, shared upregulated genes were enriched for pathways involved in UV response, DNA damage checkpoint activation and cell cycle arrest, suggesting a shift toward stress-induced inhibition of tumor growth. To assess the effects of ZNF423 knockdown in vivo, we generated a human MPNST cell line with stable ZNF423 suppression using shRNA, which was implanted into the sciatic nerve of NRG mice. Tumor growth was measured over time revealing that ZNF423 knockdown led to marked reduction in tumor size compared to controls, and individual tumor growth curves show a consistent slower progression in the ZNF423 knockdown mice. 

Conclusion: These studies suggest that ZNF423 regulates key pathways that maintain MPNST survival, as depletion reduces MPNST cell viability and interferes with tumor growth in vivo. Ongoing studies will further delineate ZNF423-dependent signaling pathways using ChIP-seq, affinity purification-mass spectrometry (AP-MS), and multiplex inhibitor bead mass spectrometry (MIB/MS) kinome profiling to reveal druggable targets within these pathways. 

TUMOR BIOLOGY AND DISEASE MECHANISM

ZHANG Xin ,MD  GUAN Wenbin ,MD  

Author Affiliations: Department of Pediatric Hematology/Oncology, Xinhua Hospital, Shanghai Jiaotong University, School of Medicine, Kongjiang Road 1665, Shanghai, 200092, PR China.

Abstract

Purpose: This case study highlights the diagnosis and management of a malignant triton tumor (MTT) in a 3-year-old girl with neurofibromatosis type I (NF1). The case underscores the importance of accurate diagnosis and treatment strategies for pediatric patients with complex tumor presentations associated with NF1.

Methods: The patient was referred to the pediatric oncology department due to a recurrent mass in her left wrist. Initial imaging studies, including ultrasound and MRI, revealed a soft tissue mass consistent with a neurofibroma. A repeat excision was performed to evaluate the tumor’s nature, followed by pathological analysis and genetic testing. Immunohistochemical staining was conducted to assess tumor markers. The patient subsequently received six cycles of Ifosfamide/Doxorubicin chemotherapy. After chemotherapy, a second surgical excision was performed, and her response was monitored through PET/CT scans.

Results: Genetic sequencing identified a germline mutation in the NF1 gene, consistent with the patient’s diagnosis of neurofibromatosis type I (NF1). Additionally, somatic mutations were detected within the tumor tissue, including a heterozygous mutation in NF1 and deletions in CDKN2A/B. Pathological examination of the excised mass revealed features characteristic of MTT, with positive immunohistochemical markers such as Desmin, MyoG, and MyoD1, while P16, SOX10, S100, and H3K27me3 were negative. Following chemotherapy, pathological analysis of the second surgical excision confirmed the presence of active tumor cells, indicating persistent disease. PET/CT scans demonstrated reduced metabolic activity surrounding the tumor site, suggesting a partial response to treatment, with no evidence of systemic metastasis.

Conclusions: The case exemplifies the challenging nature of diagnosing MPNST and its subtypes in pediatric patients, particularly in the context of NF1. MTT, a rare and aggressive form of MPNST, requires a multifaceted treatment approach, including surgical resection and chemotherapy. The detection of active tumor cells after chemotherapy highlights the need for continued vigilance in monitoring and treating residual disease. Despite high recurrence rates and poor prognosis, this patient exhibited no tumor growth or metastasis one year post-diagnosis, suggesting that early surgical intervention combined with targeted therapy may enhance outcomes. Further research into the efficacy of MEK inhibitors and combination therapies is warranted to improve management strategies for NF1-associated tumors. This case emphasizes the need for comprehensive genetic analysis and molecular profiling to facilitate tailored treatment options for pediatric sarcomas with ambiguous clinical presentations.

Malignant peripheral nerve sheath tumors (MPNST)

Ravindran, R.¹, Pundavela, J.¹, Rizvi, TA.¹, Volz, A.¹, Kedei, N.², Largaespada, DA.³, Shern, J.⁴, Ratner N.¹

¹ Division of Experimental Hematology and Cancer Biology, Cincinnati Children’s Hospital Medical Center 3333 Burnet Ave., Cincinnati, OH, 45229, USA. ² Collaborative Protein Technology Resource, OSTP, Center for Cancer Research, NCI, NIH, Bethesda, MD 20892, USA. ³ Departments of Pediatrics and Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, Minnesota. ⁴ Pediatric Oncology Branch, Center for Cancer Research, National Institutes of Health, Bethesda, MD.

Schwann cells are the cells of origin in plexiform neurofibroma (PNF), and inflammation plays an essential role in PNF formation and growth. Tumor formation occurs long after Nf1 loss in mouse models of the disease, suggesting that Schwann cells undergo secondary changes during tumor formation.  Prior single cell RNA-Sequencing implicated NF-κB expression as upregulated late in tumor formation, and thus such a potential second signal.  In human and mouse neurofibromas, p-65 was nuclear (active) in many neurofibroma cells, at least some of which were Schwann cells (Kershner et al., 2022).

Purpose: Identify the tumor Schwann cells in PNF and investigate the role of the NF-κB pathway in Schwann cell reprogramming after Nf1 loss.

Methods: A multiplex antibody panel was developed for the analysis of the PNF microenvironment and to identify the tumor Schwann cells. In vitro assays and RNA-Seq were utilized to determine the effect of NF-κB pathway modulation on Nf1 mutant Schwann cells. In vivo pathway modulation was also studied in a mouse model.

Results: Multiplex imaging revealed that tumor Schwann cells, but not pre-tumor Schwann cells in the same mice, express the cell surface markers CD44 and CD49f. We used these markers to isolate the tumor cell population by flow cytometry and characterized isolated tumor cells (versus non-tumor Schwann cells) by RNA-Seq.  We also cultured Nf1 mutant Schwann cells and showed that they upregulate CD44+ CD49f+ expression when exposed to of a variety of stressors, including prolonged serum depletion, Poly I:C (inflammatory agent), and certain cytokines (ie., Il1b, Tnfa), or when infected with activated IKK2, which activates the NF-κB pathway. As these cells increased expression of these surface proteins, they also showed nuclear (active) p65.  Preliminary results from ongoing in-vivo experiments show that low dose treatment of DhhCre; Nf1 fl/fl mice with the NF-κB pathway inhibitor BAY 11-7082 slightly reduced tumor volume, and higher-dose treatment reduced CD44+ CD49f+ tumor cells.

Conclusion: An inflammatory microenvironment is formed with activation of the inflammation associated NF-κB pathway in Schwann cells, resulting in an upregulation of the CD44+ and CD49f+ marker expression in the Schwann cells, and an overall reprogramming of the Schwann cells within the neurofibroma. A two-step process to PNF formation is proposed, with Nf1 loss in Schwann cells being the first step and the formation of an inflammatory microenvironment via activation of the NF-κB pathway and Schwann cell reprogramming as the second step.

TUMOR BIOLOGY AND DISEASE MECHANISM

Authors: Saihu Mao, MD¹, Song Li, MD¹, Zezhang Zhu, MD¹

1 Division of Spine Surgery, Department of Orthopedic Surgery, Nanjing Drum Tower Hospital, The Affiliated Hospital of Nanjing University Medical School, Zhongshan Road 321, Nanjing 210008, China

Corresponding author: Zezhang Zhu, E-mail: zhuzezhang@126.com

Disclosures: The authors declare that they have no competing interests.

Title: Convexity coronal imbalance in dystrophic scoliosis secondary to type I neurofibromatosis: classification and its importance for surgical decision-making 

Purpose: Trunk shift causing coronal imbalance (CI) is frequently encountered in unstable dystrophic scoliosis (DS) secondary to Type I Neurofibromatosis (NF-1). Surgical rebalance of coronal malalignment is challenging with high occurrence of iatrogenic aggravation. To develop a comprehensive classification of convex coronal spinal malalignment and propose a treatment algorithm for this specific condition.

Methods: A review of NF-1CI cases where different types of convex CI, rebalancing strategies, as well as the outcomes were identified and analyzed.

Results: Two main convex CI patterns were defined: thoracic CI (type 1) and thoracolumbar/lumbar CI (type 2), and were further subtyped by the compensatory behavior of the upper hemi-curve (straight or curved morphology). The incidence of post-op persistent CI ≥3cm was 0.0% and 63.6% for Type1 and Type 2 groups, respectively. Quantitative analysis revealed that the over correction of the compensatory upper hemi-curve and insufficient correction of the lower hemi-curve playing the role of imbalance driver were more significant in the imbalanced group (△Upper Arc Translation/|△Lower Arc Translation|: 31.8±34.4% vs. 109.6±60.0%, p=0.008; △Upper Arc Inclination/|△Lower Arc Inclination|: 33.5±37.3% vs. 89.8±36.6%, p=0.012). A surgical rebalancing algorithm was proposed to treat each subtype.

Conclusion: This new classification of convex CI emphasized that coronal rebalance should be obtained with cooperative correction that the lower hemi-curve should be aggressively corrected while the morphology of pre-op upper hemi-curve determined whether it played the role of fine-tuning or radical adjustment for coronal realignment. This is particularly true for type 2 NF-1CI patients with poor distal screw purchase and utmost avoidance of pelvic fixation.

Key words: NF1; Convexity coronal imbalance; dystrophic scoliosis; surgical decision-making

Authors: Jennifer L. Faulkner, MS, CRS and Erika Santos Horta, MD (University of Arkansas for Medical Sciences)

Funding: This work was supported by the Children’s Tumor Foundation.

Disclosures: Dr Erika Santos Horta is the recipient of the “Adult NF Clinic Program Building Pilot Project."

Dr Erika Santos Horta has also worked as a member of advisory board for SpringWorks Therapeutics and Alexion Pharmaceuticals. 

Title: A Comparison of Two Electronic Data Capture (EDC) Platforms to Create an NF Registry

Purpose: Compare the utility of maintaining a neurofibromatosis registry in both REDCap and OpenClinica platforms.

Methods: A patient registry was created to help manage UAMS’ adult neurofibromatosis clinic. REDCap (Vanderbilt University, USA) was the first choice due to its ease of use and ubiquity. After curating data elements, Epic Business Intelligence for UAMS was consulted to examine the feasibility of extracting data. Diagnoses of NF1, NF2, and/or Schwannomatosis were used to determine the registry population. ICD-11 codes were used to identify the presence of complications and/or tests, and treatment history was gathered using pharmacy and service orders. This information plus patient encounters were gathered quarterly. After receiving extracted Epic data, they were transformed for upload into REDCap. The data were edited and expanded via manual entry by the physician accessing patient charts 

After months of using REDCap, it was decided to build a registry that enabled varying numbers of rows, representing encounters, that could be added as needed to a grid. This led to OpenClinica being chosen as an alternative EDC platform. In addition to housing the same data elements as REDCap, OpenClinica also enabled this addition of rows, allowing comparison of different encounters on a single page.

Results: Of the 76 data elements selected, 61 were able to be easily extracted and imported into REDCap, and with somewhat more difficulty, OpenClinica. REDCap can clone projects which proved useful in creating “snapshots” of each project before cloning and adding newly extracted data. Both are user friendly, but in different ways. OpenClinica can add rows to a grid, facilitating following a patient through different services. REDCap has useful status icons and the capability to build graphics.  

Conclusion: Both EDC platforms are useful for creating and maintaining patients’ registries. REDCap offers a smooth pipeline for data import and export while OpenClinica offers the ability to increase the number of data elements captured on a given page at will.

{{Other}}

Authors: Phioanh L. Nghiemphu, MD^1,2, Conrad L. Cordova, MSN³, Abby Crites, BJ,⁴ Alyssa Bowling, PharmD,⁵ Xiaoqin Yang, PhD⁶, and Theresa Dettling, MS^5,*

*Presenting author

Affiliations: ¹UCLA Neuro-Oncology Program, David Geffen School of Medicine, University of California, Los Angeles, CA, USA; ²Department of Neurology, David Geffen School of Medicine, University of California, Los Angeles, CA, USA; ³Patient author; ⁴IQVIA, Durham, NC, USA; ⁵Alexion, AstraZeneca Rare Disease, Boston, MA, USA; ⁶Merck & Co., Inc., Rahway, NJ, USA.

Funding: Alexion, AstraZeneca Rare Disease and Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway, NJ, USA.

Disclosures: Phioanh L. Nghiemphu has received grants or contracts from Chimerix, Bristol Myers Squibb, Novartis, Erasca, Takeda, National Comprehensive Cancer Network, Department of Defense, NI. Phioanh L. Nghiemphu has also received payment/honoraria from Alexion and Springworks, has participated on a Data Monitoring Board for California Institute for Regenerative Medicine, and has had an unpaid leadership/fiduciary role in other board, society, committee, or advocacy group for the Society of Neuro-oncology and American Academy of Neurology. Theresa Dettling and Alyssa Bowling are employees of, and own stocks in, Alexion, AstraZeneca Rare Disease. Xiaoqin Yang is an employee of Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway, NJ, USA and owns stocks in Merck & Co., Inc., Rahway, NJ, USA. Abby Crites is an employee of IQVIA. 

Adult Patient and Caregiver Perspectives on the Impact of Neurofibromatosis Type 1-Associated Plexiform Neurofibroma (NF1-PN): Insights from a US Qualitative Survey

Purpose: This study aimed to improve understanding of the disease burden, healthcare experiences, and unmet needs of adults with NF1-PN in the USA, from the perspective of patients and caregivers.

Methods: Individual, 45-minute, double-blind telephone interviews were conducted in the USA (July 27–August 4, 2023). Participants (≥18 years) had NF1-PN or were caregivers of adults with NF1-PN. All participants were required to be involved in NF1-PN treatment decisions. Caregivers who participated were not associated with patients who participated. Interviewers followed discussion guides, which contained questions covering patient background and NF1-PN diagnosis, treatment journey, relationship with healthcare providers, and unmet needs. All participants were recruited through physician referrals, patient organizations, and patient databases.

Results: The study included 11 adult patients with NF1-PN and two caregivers of adult patients with NF1-PN. Overall, 62% of patients were aged ≥35 years. Most patients (85%; n=11/13) had been diagnosed in childhood. Additionally, patients had multiple other NF1-associated conditions, including pain-related disorders (n=10), psychiatric disorders (n=7), and chronic migraine (n=6). NF1-PN affected most aspects of patients’ lives, including overall quality of life, mental health, personal relationships, and participation in school/work. Of patients diagnosed in childhood, 82% (n=9/11) had been transitioned to an adult practitioner. Transition to adult care occurred at 18/19 years for most patients and was typically driven by the pediatric care team (n=6/9). The transition process was uncomplicated for most patients (56%; n=5/9); the remainder experienced challenges, including finding an appropriate provider (n=3/9). Most patients (77%; n=10/13) reported receiving routine healthcare annually; patients with lapses in care (n=3/13) provided reasons including lack of local physicians (n=1/13), lack of healthcare insurance (n=2/13), and the perception that there is nothing that can treat/cure NF1-PN (n=3/13). NF1-PN medication management for adults included pain medications over-the-counter (n=9) and prescription (n=3), vitamin supplements (n=1), and skin creams/oils (n=2). When asked what could be improved in the management of NF1-PN, patients and caregivers cited more informed care and disease state information (n=7) and improved treatments for NF1-PN (n=13), including medications that would slow/stop PN growth (n=6), reduce PN size (n=8), improve pain management (n=5), and were covered by insurance (n=2).

Conclusion: This US-based study of adults with NF1-PN and caregivers demonstrated the high disease burden on patients despite variability in manifestations and highlighted the need for more informed care and improved treatment options.

{{Plexiform neurofibroma}}

Authors: Phioanh L. Nghiemphu, MD,^1,2,*, Abby Crites, BJ³, Alyssa Bowling, PharmD⁴, Xiaoqin Yang, PhD⁵, and Theresa Dettling, MS⁴

*Presenting author

Affiliations: ¹UCLA Neuro-Oncology Program, David Geffen School of Medicine, University of California, Los Angeles, CA, USA; ²Department of Neurology, David Geffen School of Medicine, University of California, Los Angeles, CA, USA; ³IQVIA, Durham, NC, USA; ⁴Alexion, AstraZeneca Rare Disease, Boston, MA, USA; ⁵Merck & Co., Inc., Rahway, NJ, USA.

Funding: Alexion, AstraZeneca Rare Disease and Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway, NJ, USA.

Disclosures: Phioanh L. Nghiemphu has received grants or contracts from Chimerix, Bristol Myers Squibb, Novartis, Erasca, Takeda, National Comprehensive Cancer Network, Department of Defense, NI. Phioanh L. Nghiemphu has also received payment/honoraria from Alexion and Springworks Therapeutics, has participated on a Data Monitoring Board for California Institute for Regenerative Medicine, and has had an unpaid leadership/fiduciary role in other board, society, committee, or advocacy group for the Society for Neuro-Oncology and American Academy of Neurology. Abby Crites is an employee of IQVIA. Xiaoqin Yang is an employee of Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway, NJ, USA and owns stocks in Merck & Co., Inc., Rahway, NJ, USA. Theresa Dettling and Alyssa Bowling are employees of, and own stocks in, Alexion, AstraZeneca Rare Disease. 

Healthcare Providers’ (HCPs’) Perspectives on the Management of Adults With Neurofibromatosis Type 1 (NF1) and Plexiform Neurofibroma (PN): Insights From a US Survey

Purpose: The aim of this study was to better understand experiences, approaches, and challenges in treating and managing adults with NF1-PN from the perspective of US physician HCPs.

Methods: Quantitative data were collected through 25-minute online surveys (July 27–August 23, 2023). HCPs were asked for their perspectives on unmet needs in the diagnosis, management, and treatment of adults with NF1-PN. A subset of participants participated in quali-quant interviews to provide additional context to their responses. HCPs must have been in practice for 3–35 years, spent ≥50% of their time in direct patient care, and managed ≥100 patients in the last 12 months for any condition, including ≥3 adults (aged ≥18 years) with NF1-PN.

Results: Overall, 41 HCPs completed the survey; 4/41 participated in quali-quant interviews. HCPs reported the most common manifestation in patients prior to NF1-PN diagnosis as neurofibromas (66% adults). The most burdensome manifestations reported for adult patients were brain stem gliomas (80%), visual impairment (78%), neuropathic pain (76%), other (non-neuropathic) pain (73%), and bladder/bowel dysfunction (68%). HCPs generally reported follow-up appointments with most adults every 2–3 months (68%). Physical exam of the skin (92%) and diagnostic imaging (79%) were the most common PN monitoring methods. The most common treatments for adult PN-related issues included physiotherapy (76%) and pain medication (90%). Types of pain medication included pregabalin (76%), gabapentin (73%), analgesics (66%), and non-steroidal anti-inflammatory drugs (61%). While selumetinib was FDA approved for the treatment of pediatric patients with symptomatic, inoperable NF1-PN during the study period, 46% of physicians reported use of selumetinib in adult patients and 50% of physicians reported use in pediatric patients. Over two-thirds of HCPs highlighted both the limited and lack of effective treatment options as the greatest challenges in the management of adult NF1-PN. With regard to continuity of care, HCPs reported that 17% of patients fall out of care when transitioning from pediatric to adult care and attribute this to lack of transition guidelines/standards as a key challenge.

Conclusion: From the perspectives of HCPs, adult patients with NF1-PN experience a considerable number of issues, including high rates of reported pain. Additionally, HCPs generally reported that adults have a high overall burden of disease, face many challenges in their transition from pediatric to adult care, and could benefit from more effective treatment options to manage manifestations of NF1-PN.

{{Plexiform neurofibroma}}

Authors: Jadwiga N. Bilchak, PhD, Kyla Mace, PhD, Kevin Wu, Matthew S. Kayser, MD PhD

Institution: University of Pennsylvania

Funding: CTF-2023-01-004, DOD-NFRP W81XWH2010206

Abstract Title: Connecting sleep and sensory deficits in a Drosophila model of NF1

Disclosure of relevant financial relationships: None

Background: Neurofibromatosis type 1 (NF1) is mainly characterized by benign tumors throughout the nervous system. However, individuals with NF1 also experience cognitive and social deficits, disrupted circadian rhythms, poor sleep quality, and altered sensory perception. Although these non-tumor manifestations affect up to 80% of the NF1 population, it is largely unknown how they arise, and whether they influence one another. Interestingly, these traits are highly conserved across species, including in Drosophila models of NF1.  Here, we leverage this powerful animal model to examine the relationship between sensory integration and sleep disruption in NF1.

Methods and Results: Sensory input can either disrupt sleep or can be harnessed to enhance sleep quality. Gentle vibration, for example, improves sleep in rodents and humans, with recent studies indicating similar effects in Drosophila. Here, we used a vibration-induced sleep (VIS) paradigm to investigate how sleep is influenced by sensory input a fly model of NF1. Adult fly sleep was recorded at baseline for 24 hours, followed immediately by 24 hours of exposure to gentle, continuous vibration. Nf1 flies exhibited reduced and fragmented sleep at baseline, consistent with published work. Additionally, while control flies showed VIS, Nf1 flies failed to increase sleep in response to vibration, suggesting deficits in the sleep-sensory connection.

 To determine if VIS or baseline sleep deficits are due to excessive sensory input, we reduced mechanosensory signals by surgically removing antenna, where most mechanosensory neurons reside. This manipulation reduced VIS in controls, as expected, but failed to normalize baseline sleep or VIS in Nf1 flies (Fig.1). This suggested that absence of VIS in NF1 does not result from hypersensitivity to vibration. We then used genetic approaches to directly activate mechanosensory neurons in vivo. In controls, we found increased sleep similar to VIS, but in Nf1 flies there was no effect.

To decouple potential sensitivity differences from sleep, we exposed flies to an hour of vibration while they were awake. Control flies responded with an initial spike in activity during the first 5 minutes of vibration. Surprisingly, NF1 flies showed a similar response, regardless of vibration intensity. Furthermore, mechanosensory neurons in Nf1 flies appear morphologically normal. 

Conclusion: Together, these findings suggest that mechanosensory neurons in Nf1 flies function normally during wake, but their sensory input is not effectively integrated to influence sleep. This work sheds light on the relationship between sensory integration and sleep in NF1, pointing towards novel treatment strategies for disrupted sleep. 

DRUG DISCOVERY AND DEVELOPMENT

Authors

Isha Mol, MS¹; Timothy Bell, MHA²; Yannan Hu, PhD¹; Abraham J. Langseth, PhD²; Michael D. Weber, PharmD²; and Hoora Moradian, PhD³

¹Cytel, Rotterdam, Netherlands; 

²SpringWorks Therapeutics, Inc, Stamford, CT, US; 

³Cytel, Vancouver, Canada.

ABSTRACT

Purpose: Mirdametinib is the first FDA-approved MEK1/2 inhibitor for both adult and pediatric patients 2 years of age and older with NF1 who have symptomatic PN not amenable to complete resection. We report efficacy and safety outcomes utilizing ITC in children with NF1-PN treated with mirdametinib vs selumetinib.

Methods: Unanchored matching-adjusted indirect comparison (MAIC) and simulated treatment comparison (STC) analyses evaluated outcomes of mirdametinib (ReNeu; NCT03962543) vs selumetinib (SPRINT; NCT01362803), where feasible.

Results: Mean best percent reduction from baseline in target PN volume by blinded independent central review (BICR) was significantly greater with mirdametinib vs selumetinib (MAIC: -36.0% vs -22.8%; STC: -36.2% vs -22.8%). The mean best percent reduction from baseline difference (95% CI) in the MAIC was -13.2% (-22.4%, -4.1%; P=.005; effective sample size [ESS]=54); in the STC was -13.4% (-22.2%, -4.4%; P=.004; N=54). In the MAIC and/or STC, all-grade treatment-related adverse event (TRAE) rates were significantly lower with mirdametinib vs selumetinib for dermatitis acneiform, diarrhea, nausea, vomiting, fatigue, blood creatinine phosphokinase increase, dry skin, pruritus, constipation, abdominal pain, stomatitis, hair color change, headache, and neutrophil count decrease (P<.05). Rate of dose reductions due to TRAEs was significantly lower with mirdametinib vs selumetinib (MAIC: 12% vs 28%, odds ratio [OR]=.355, P=.048; STC: 11% vs 28%, OR=.309, P=.028). Differences in confirmed objective response rate by BICR (MAIC: 51.3% vs 44%, ESS=21; STC: 55.5% vs 44%; N=56) and rates of all-grade paronychia (MAIC/STC: 30% vs 42%), white blood cell count decrease (MAIC: 6% vs 14%; STC: 5% vs 14%), alopecia (MAIC/STC: 13% vs 8%), and asymptomatic ejection fraction decrease (MAIC: 20% vs 14%; STC: 18% vs 14%) were not significantly different between mirdametinib vs selumetinib. Study limitations include possible confounding of unmeasured treatment effect modifiers.

Conclusions: Mirdametinib demonstrated a significantly greater mean best percent reduction from baseline in target PN volume, lower rate of dose reductions due to TRAEs, and lower rates of most TRAEs vs selumetinib in children with NF1-PN. None of the analyses conducted significantly favored selumetinib.

Supported by: SpringWorks Therapeutics, Inc.

DISCLOSURES

TB, MW, and AJL: Employees, stock ownership, SpringWorks Therapeutics, Inc.

IM, YH, and HM: Employees of Cytel, who received consulting fees from SpringWorks Therapeutics, Inc, to conduct the analyses and for medical writing support.

Plexiform neurofibroma

Authors: Nathan K. Leclair¹, Mattia Brugiolo¹, Ryan Englander¹, Kanish Mirchia², Melike Pekmezci², Harish N. Vasudevan³*, Olga Anczukow¹*

Affiliations:¹The Jackson Laboratory for Genomic Medicine, Farmington, CT; ²Pathology Division, University of California San Francisco, San Francisco, CA; ³Departments of Radiation Oncology and Neurological Surgery, University of California San Francisco, San Francisco, CA.

^*corresponding authors

Funding:

Disclosures: None

Purpose: Neurofibromatosis type 1 (NF-1) is a cancer predisposition syndrome associated with the development of benign plexiform neurofibromas (pNFs) which can transform into malignant peripheral nervous sheath tumors (MPNSTs). While the genomic and epigenetic underpinnings of this transformation have been described, changes in alternative RNA-splicing (AS) during malignant transformation are not well understood. AS is frequently dysregulated in human tumors, promoting the expression of pro-tumorigenic mRNA isoforms, and represent an untapped repertoire for novel therapeutics. Here, we define the AS landscape during pNF to MPNST transformation in patient samples and cell lines, and identify targetable RNA splicing alterations. 

Methods: Bulk short-read RNA-sequencing (RNA-seq) from human pNFs and MPNSTs across three cohorts (UCSF, UoT, GeM) was analyzed using rMATS to quantify AS and DESeq for gene expression. Reverse-transcription polymerase chain reaction (RT-PCR) using RNA from pNF and MPNST-derived cell lines was used to validate AS splicing differences between tumor types. Finally, splice-switching antisense oligonucleotides (ASOs) were designed to modulate AS events. 

Results: Using samples from UCSF as a discovery cohort, we identified 660 AS events significantly differently spliced during transformation from pNFs to MPNSTs. Of these 233 AS events (35%) were also detected in the University of Toronto cohort and 299 (45%) in the GeM cohort. These AS events were enriched for genes associated with mitotic spindle and epithelial-mesenchymal transition signatures, including for example, increased inclusion of an alternative cassette exon in Fibronectin (FN1) in MPNSTs compared to pNFs, which regulates FN1 protein localization and function. We further identified 13 RNA splicing regulators differentially expressed between MPNSTs and pNFs, including MBNL1, PTBP1, IGF2BP1 which have known roles in cancer. Finally, we demonstrate that splice-switching ASOs that promote FN1 exon skipping decrease proliferation, migration, and invasion of MPNST cell lines. We also show that ASOs targeting AS events in known RAS/MAPK regulators HRAS, KRAS, RRAS, or RASA3 decrease proliferation of MPNST cells and synergize with the MEK inhibitor selumetinib.

Conclusion: We identify multiple AS events in MPNSTs and provide proof-of-concept for targeting of AS with ASOs as single agents and to synergize with MEK inhibitors, the only FDA approved therapy for pNFs. While our work reveals that AS represents an actionable target in NF-1, further research is needed to comprehensively map the AS landscape in NF-1 patients using long-read and single cell approaches, and to test AS-targeting therapeutic approaches in preclinical models.

TUMOR BIOLOGY AND DISEASE MECHANISM

AUTHORS :, Danny Bergeron¹, Camille Plante² and Jean-Philippe Brosseau^2,3,4

AFFILIATIONS :

Université de Sherbrooke, Faculté de médecine et des sciences de la santé, Département de Microbiologie et Infectiologie 2. Département de Biochimie et Génomique Fonctionnelle; 3. Centre de recherche du centre hospitalier universitaire de Sherbrooke (CRCHUS); 4. Institut de recherche sur le cancer de l`université de Sherbrooke (IRCUS)

FUNDING : This research was supported by an operating grant from the US DoD CDMRP-NFRP-NIA (NF230030)

DISCLOSURES : The authors declare no conflict of interest.

PURPOSE : The main objective is to determine the landscape of alternative splicing change in human neurofibroma fibroblasts 

METHOD : We performed RNAseq in 3 normal skin fibroblasts and 3 cutaneous neurofibroma fibroblasts. We performed differential percent of splicing index (PSI) analysis to identify alternative splicing events associated with neurofibroma fibroblasts. We also performed Gene ontology analysis (DAVID bioinformatics) to get insights into their functional role. We performed DESeq2 analysis to measure the change in splicing factor expression.

RESULTS : We identified 68 alternative splicing events associated with neurofibroma fibroblasts. Gene ontology analysis indicated that this splicing isoform signature is associated with cytoplasm intracellular location. We also identified 19 splicing factors with a significant change in gene expression between normal and neurofibroma fibroblasts. Overall, our preliminary results suggest a regulated alternative splicing program where a set of neurofibroma fibroblast splicing factors control the expression of specific neurofibroma fibroblast isoforms to contribute to the function of neurofibroma fibroblast phenotype ultimately.

CONCLUSION : We determined the landscape of alternative splicing change in human neurofibroma fibroblasts, setting the first step toward understanding the functional contribution of alternative splicing to neurofibroma fibroblast biology. 

TUMOR BIOLOGY AND DISEASE MECHANISM

Rupa Siva and Chunling Yi

Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, Washington DC, USA

Neurofibromatosis Type 2 (NF2) mutant tumors exhibit increased oxidative stress, making them vulnerable to ferroptosis, a regulated form of cell death driven by lipid peroxidation. Nuclear factor erythroid 2-related factor 2 (Nrf2), encoded by the Nfe2l2 gene, is a master regulator of antioxidant responses that suppresses ferroptosis. On the other hand, the YAP/TAZ transcriptional co-activators, partnering with the TEAD family of transcription factors, play a critical role in promoting NF2 tumor growth. This study investigates the functional relationships between Nrf2 and YAP/TAZ/TEAD during NF2 schwannoma development and explores therapeutic potentials of inducing ferroptosis as a therapeutic strategy for treatment of NF2. Genetically engineered murine models of Postn-Cre-driven Nf2 knockout (Nf2KO) and Nf2:Nfe2l2 double knockout (DKO) were first established to determine the effects of Nrf2 co-deletion on NF2 tumorigenesis. Nf2KO and DKO mice exhibited similar rates of dorsal root ganglion enlargement and overall survival, and both developed spontaneous schwannomas that expressed mature Schwann cell markers p75NTR and Sox10. These results demonstrate that Nrf2 is dispensable for NF2 tumorigenesis. To interrogate how Nrf2 inactivation affects the sensitivity of NF2 schwannoma cells to ferroptosis, murine and human Nrf2 knockout schwannoma cell lines were generated. Compared to their wild type counterparts, both murine and human schwannoma cells lacking Nrf2 showed increased sensitivity to known ferroptosis inducers, which corresponded to a lack of induction of canonical Nrf2 target genes such as HO-1 and NQO1. Intriguingly, clinical YAP/TAZ/TEAD inhibitors phenocopied the effects of Nrf2 deletion in sensitizing NF2 schwannomas to ferroptosis inducers, suggesting a synergistic effect of inducing oxidative stress and suppressing YAP/TAZ/TEAD transcriptional activity. Taken together, our findings provide a mechanistic basis and highlight the therapeutic potential of co-targeting the Nrf2-mediated redox balance and YAP/TAZ signaling in inducing ferroptotic death in NF2-associated tumors.

Funding: This research is supported by the DOD Neurofibromatosis Research Program (W81XWH-19-1-0537) and the Children’s Tumor Foundation Drug Discovery Initiative Award.

Disclosures: The authors declare no competing interests.

TUMOR BIOLOGY AND DISEASE MECHANISM

1. Authors: Full name, academic credentials, and institutional affiliation for all authors.
   1. Dr Abinaya Seenivasan BA DCH DNB MRCPCH Trainee¹
   2. Dr Vivian Tang MBChB MRCP FRCR Consultant Paediatric Neuroradiologist ^1,5
   3. Prof Stavros Stivaros BSc (Hons) MBChB FRCR PhD Consultant Paediatric Neuroradiologist ^1,5
   4. Dr Chris Duff BM BCh MA FRCSEd FRCS(Plast) Plastic Surgeon ^1,5
   5. Prof John Paul Kilday MBChB FRCPCH PhD Paediatric oncologist ^1,5
   6. Prof Ian Kamly MBChB, FRCS(SN) MD Paediatric neurosurgeon ^1,5
   7. Mr Dimitros Varthalis MD FRCS Paediatric neurosurgeon ^1,5
   8. Dr Emma Burkitt-Wright MBChB, MPhil, MRCP, PhD consultant clinical genetics ^2,5
   9. Helen Young MB, ChB, PhD, MRCP(UK) consultant dermatologist ^3,5
   10. David Mowatt BDS FDSRCS MBBS FRCS (Plast) MD consultant plastic surgeon ^4,5
   11. Alexander TJ Lee PhD MBChB MRCP BSc (Hons) PGDip, consultant medical oncologist ^4,5
   12. Judith Eelloo clinical nurse specialist ^2,5
   13. Eileen Hupton NF1 specialist nurse^2,5
   14. Bibitha Saji Specialist nurse ^2,5
   15. Carol Irving BSc (Hons), RN Child, Specialist Practitioner Child Health^2,5
   16. Caroline Newby NF1 specialist nurse ^2,5
   17. Natalie Hayes MDT co-ordinator ^2,5
   18. Dr Grace Vassallo, MD DCH MRCPCH consultant Paediatric Neurologist ^1,5
   19. Dr Sean Booth consultant paediatric radiologist ^1,5
2. Affiliations: (1) Royal Manchester Children’s Hospital, Manchester Foundation Trust, UK, (2) St Mary’s Hospital, Manchester Foundation Trust, UK, (3) Salford Royal NHS Foundation Trust, Manchester, UK, (4) The Christie NHS Foundation Trust, Manchester, UK, (5) Complex NF1 Service, St Mary’s Hospital, Manchester, UK

2. Funding: nil

3. Disclosures: Dr. Grace Vassallo is a medical advisor for Alexion.

4. Title: Outcomes of paediatric patients with Neurofibromatosis 1 (NF1) treated with MEK inhibitors in the north of England 

5. Purpose: In 2009, National Health Services (NHS) England established two Highly Specialised Services (HSS) for Complex NF1 in Manchester and London. In May 2022, NICE approved Selumetinib for the treatment of symptomatic inoperable plexiform Neurofibromas in children from 3-18 years. A national pathway and shared care agreement with paediatric oncologists was published in 2023. We analysed the outcomes of patients on MEK inhibitors over 8 years in northern England. 

6. Methods: Retrospective notes from eligible children discussed in the national MEK inhibitor MDT as per National pathway for England (2017-2024) were assessed for consideration of treatment with MEK inhibitors. The main indicators for treatment were pain, disfigurement, and threat to function.

7. Results: Out of 65 eligible children discussed in MDT, 26 did not meet the full approval requirements. 8 children were enrolled in trial and 3 were on a waiting list for a trail. Their data is not included here. Since 2017, 28 children (17 males, 11 females) aged 1 to 17.1 years (average 8.5 years) have been treated with MEK inhibitors. Main indicators included disfigurement (5), pain (10), threat to function (13). All children approved for disfigurement had craniofacial plexiform. Threat to function included airway compromise (3) and swallowing and visual disturbances (1 each). 8 children had symptomatic compression-Spinal cord with 3 requiring additional decompression surgery. Side effects were reported in 21 patients: skin rashes and paronychia (17), loose stools and vomiting (2), and fatigue (1). Adjustments made for side effects included drug breaks and dose reductions, but treatments were not stopped for any of these patients.

8. Conclusion: None of the children stopped MEK inhibitors because of lack of efficacy or tolerability. Pain is a significant indicator and many of these children experienced pain relief even at lower doses of MEK inhibitors which meant that they had less side effects. Threat to function remains the main indicator for treatment with symptomatic cervical cord compression accounting for 53 % of cases. Children requiring decompression in infancy have a very difficult spinal stability course with morbidity. This group of children will need better exploration of their pathway of care.

Other

Zixin Zhang, MD ¹, Yi Ji, MD, PhD ¹

Affiliations: ¹Division of Oncology, Department of Pediatric Surgery, West China Hospital of Sichuan University, Chengdu, 610041, China

Purpose: Currently, there is no research clarifying whether drugs used to treat neurofibromatosis type 1 (NF1) affect the growth, development, and behavior of children. Therefore, the aim of this study was to explore the growth and developmental toxicity, as well as the effects on behavior and the heart, of selumetinib and binimetinib using zebrafish embryo models.

Methods: Embryos at 3 hours post fertilization (hpf) were exposed to selumetinib and binimetinib (0.5, 1, 2, 4, 8 and 16 μM) until 120 hpf (before the feeding stage). During this period, the survival and morphology of embryos were observed and recorded every 24 hours, and specific observation parameters would be assessed at specific developmental time points. As for the development of different systems and the heart, the corresponding transgenic zebrafish model were chosen, such as Tg (fli1a:EGFP), Tg (hb9:GFP), Tg (huc:GFP), Tg (mpeg:EGFP), Tg (mpx:EGFP) and Tg(myl7:GFP). Moreover, a deeper analysis of cardiac toxicity was conducted by performing apoptosis staining to observe the growth and development of myocardial cells. At the gene level, the expression of genes related to cardiac function and apoptosis was assessed. At the protein level, the expression of proteins related to myocardial enzyme profiles was measured.

Results: The results indicated that selumetinib showed slightly higher toxicity than binimetinib in terms of survival, deformity, locomotor behavior, and cardiac toxicity. The lethal concentration for both drugs was around 32 μM. At 16 μM, selumetinib significantly increased the malformation rate, while binimetinib showed minimal changes. Selumetinib also induced venous sinus congestion at 16 μM, which was not observed with binimetinib. In locomotor behavior, under light stimulation, selumetinib at 16 μM significantly decreased swimming distance and speed under light stimulation, while binimetinib had little effect; Under sound stimulation, both drugs increased swimming speed, with a more pronounced effect at higher concentrations. Cardiac toxicity results showed that at 32 μM, selumetinib significantly reduced heart rate and blood flow velocity, increased pericardial edema and AV-SV distance, and altered the expression of cardiac enzyme markers and related genes.

Conclusion: Excessively high concentrations of selumetinib significantly affected the growth and development of zebrafish embryos and caused severe cardiac toxicity, whereas binimetinib exhibited lower toxicity than Selumetinib. Therefore, in clinical practice, the dosage and blood drug concentration of selumetinib must be strictly controlled when used in children.

Funding: This work was supported by the National Natural Science Foundation of China (grant number 82473553 and 82273556), the Excellent Youth Foundation of Sichuan Province of China (grant numbers 2025NSFJQ0070), Natural Science Foundation of Sichuan Province of China (2025ZNSFSC0666, 2025ZNSFSC1525, and 2025ZNSFSC1547), the ‘0 to 1’ Project of Sichuan University (grant number 2022SCUH0033), the Med-X Center for Informatics Funding Project (YGJC004), the 1·3·5 Project for Disciplines of Excellence-Clinical Research Incubation Project of West China Hospital of Sichuan University (grant numbers 2023HXFH004), and the 1·3·5 Project for Disciplines of Excellence-Clinical Research Interdisciplinary Innovation Project of West China Hospital of Sichuan University (ZYJC21060).

Disclosures: The authors declare that there are no conflicts of interest and financial relationships.

Figure 1. Survival-related parameters after selumetinib and binimetinib intervention. (A) Survival curve of selumetinib. (B) Survival curve of binimetinib. (C) Hatching rate of selumetinib. (D) Hatching rate of binimetinib. (E) Deformity rate of selumetinib. (F) Deformity rate of binimetinib.

Figure 1. Swimming behavior analysis after selumetinib and binimetinib intervention. (A) Total swimming distance of selumetinib under light stimulation. (B) Maximum speed of selumetinib under light stimulation. (C) Total swimming distance of binimetinib under light stimulation. (D) Maximum speed of binimetinib under light stimulation. (E) Total swimming distance of selumetinib under sound stimulation. (F) Maximum speed of selumetinib under sound stimulation. (G) Total swimming distance of binimetinib under sound stimulation. (H) Maximum speed of binimetinib under sound stimulation.

DRUG DISCOVERY AND DEVELOPMENT

Authors: Ayan S. Mandal PhD^1,2,3, Jakob Seidlitz PhD^2,3,4, Margaret Gardner^2,3, Benjamin Jung PhD^2,3, Michael J. Fisher MD⁵, Aaron Alexander-Bloch MD PhD^2,3,4

1 Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA

2 Department of Child and Adolescent Psychiatry and Behavioral Science, The Children’s Hospital of Philadelphia, Philadelphia, PA, USA.

3 Lifespan Brain Institute, Children’s Hospital of Philadelphia and Penn Medicine, Philadelphia, PA, USA

4 Department of Psychiatry, University of Pennsylvania, Philadelphia, PA, USA

5 Department of Oncology, Children’s Hospital of Philadelphia, Philadelphia, PA, USA

Funding: This work was supported by the Department of Defense, NIMH R01MH134896, and by the CHOP Research Institute.

Disclosures: Drs. Seidlitz and Alexander-Bloch hold shares in and Drs. Seidlitz is a director of Centile Bioscience.

Title: Slow but sustained head growth drives macrocephaly in NF1

Purpose: NF1 is characterized by over-activation of the RAS pathway, driving a wide range of clinical manifestations including plexiform neurofibromas, low grade gliomas, cognitive issues, and macrocephaly. While macrocephaly and relatedly, megalencephaly, has been demonstrated at birth and later life in NF1, little is known about timing of head growth for these patients compared to typical development. We aimed to clarify the timing of macrocephaly in NF1 in early life which may inform the appropriate timing of interventions to address neurocognitive manifestations of the disease.

Methods: Using electronic health records from the Children’s Hospital of Philadelphia, we constructed growth charts of head circumference from age 0 to 3 years in NF1 and clinical control cohorts seen at well-child visits. Demographic information and sample sizes of available data are shown in Table 1. Using standardized (Z) scores from well-child visit growth charts, we statistically compared head circumference between NF1 patients and a left-out group of clinical controls at each well-child visit timepoint. 

Results: Consistent with prior reports, head circumference around the time of birth appears increased in NF1 compared to controls (Figure 1 A&B). However, head circumference rate-of-growth is slower for NF1 patients compared to controls over the first four months of life (Figure 1C). After that timepoint, head circumference in NF1 outpaces controls until approximately two years of age. Differences in head circumference between NF1 and controls are statistically significant at all well-child visit timepoints starting at six months (Figure 2 A&B). These patterns are recapitulated when using Center for Disease Control charts (Figure 2C). 

Conclusions: We identify a time window between four months and two years of age in which head circumference rate-of-growth is increased in NF1. Further research is needed in quantitative neuroimaging in the first years of life to determine the neurological correlates of accelerated head growth in this population and how it relates to cognitive outcomes. 

Table 1. Demographics for the NF1 and clinical control cohorts. N refers to the total number of observations in each category, of which there were many longitudinal measurements per subject.  

Figure 1. Head circumference growth charts for NF1 and clinical controls. (A) Raw head circumference data. Age is represented on a log-scale and is adjusted for gestational age at birth. PCW = Post-conceptional weeks. (B) Growth curves representing the 3^rd, 10^th, 50^th, 90^th, and 97^th percentile. (C) Rate-of-growth curves computed by calculating the first derivative of the median trajectories. For males, NF1 cases are shown in red, and well-child visits of children without NF1 are shown in blue. For females, NF1 cases are shown in blue, and well child visits of children without NF1 are shown in red.

Figure 2. Statistical comparison of head circumference differences in NF1 versus clinical control. (A) Histogram of ages at head circumference measurements for NF1 and a left-out clinical control group shows highest density of data at well-child visits. (B) Comparison of standardized scores between NF1 and left-out clinical controls at each well-child visit timepoint. P values from pairwise t-tests were adjusted for multiple comparisons using false discovery rate (FDR) correction. *** indicates P < 0.001; ** indicates P < 0.01; * indicates P < 0.05; ns indicates P > 0.05. (C) Head circumference Z scores computed using CDC growth charts at each well-child visit timepoint for NF1 subjects. 

Cognition / Behavioral / Learning

Kuangying Yang¹, Yang Lyu¹, Xiaochun Zhang¹, Dana C. Borcherding¹, Guangfeng Wang¹, Kangwen Xiao¹, Joanna K. Lempiainen², Stavriani C. Makri³, Diane E. Bender⁴, Xiyuan Zhang⁵, Jack F. Shern⁵, Christine A. Pratilas³, Benjamin A. Garcia², Angela C. Hirbe^1,*

 ^*Corresponding Author: Angela C. Hirbe, Division of Oncology, Department of Internal Medicine, Washington University School of Medicine, St. Louis, MO, USA 

¹Division of Oncology, Department of Internal Medicine, Washington University School of Medicine, St. Louis, MO, USA

²Department of Biochemistry and Molecular Biophysics, Washington University in St. Louis, St. Louis, MO, USA

³Department of Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD, USA

⁴The Bursky Center for Human Immunology and Immunotherapy Programs Immunomonitoring Laboratory, Washington University School of Medicine, St. Louis, MO, USA

⁵Pediatric Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA

Purpose: Neurofibromatosis Type 1 (NF1) is a common neurogenetic and tumor predisposition syndrome, leading to the development of benign plexiform neurofibroma (PN) that can progress to malignant peripheral nerve sheath tumors (MPNST). Loss of function of polycomb repressive complex 2 (PRC2) which methylates histone H3 on lysine 27 (H3K27me3), is a common feature of MPNST and is associated with poorer prognosis in MPNST patients. We previously demonstrated MPNST with PRC2 loss exhibit a more immune suppressive environment. To deeply understand the role of PRC2 loss in MPNST pathogenesis to inform novel treatment options for NF1-MPNST, we performed multiomic analyses to explore how PRC2 loss impacts tumor evolution and immune composition.

Methods: Paired PN and MPNST samples from 12 patients (PRC2 loss: N = 7; PRC2 retained: N = 5) were collected to investigate the molecular events and microenvironment during tumor progression. At the genomic level, we defined clonal evolution from PN to MPNST with whole exome sequencing (WES) data by a novel pipeline developed in our laboratory: Precise DNA Variant Calling (PDVC). At the transcriptomic level, we integrated bulk and single-cell RNAseq data to determine how PRC2 loss influences meta-signatures and immune cell signaling pathways through deconvolution analyses. At the epigenetic level, we performed parallel proteomic and histone analyses to determine how PRC2 loss affects histone post-translational modifications and immune responses. Finally, at the spatial proteomic level, we utilized multiplexed Hyperion Imaging Mass Cytometry (IMC) staining to construct niche-similarity networks and validate the different tumor-stroma-immune composition in PRC2-loss vs -retained MPNST as well as their precursor lesions.

Results: Diverse clonal evolution patterns including distinct types of NF1, CDKN2A, and MTAP structural variants were identified in PRC2-loss vs -retained MPNST through PDVC-WEX pipeline. Global proteomics and transcriptomics analyses indicated MPNST with PRC2 loss have poorer antitumor immune infiltration and responses compared to those with PRC2 retained. The trial study of IMC data provided a differential landscape of morphological features, spatial cellular interactions, and niche-similarity networks in PRC2-loss vs -retained MPNST. All IMC results and supplementary single-cell RNAseq data from NCI will be available before the Material Review Process of 2025 NF Conference by CTF.

Conclusion: Our findings demonstrated that MPNST with PRC2 loss are “cold tumors” with poorer antitumor immune responses and a more malignant progression path from PN, compared to those MPNST with PRC2 retained, which can potentially offer clues to develop treatments targeting the immune system for NF1-MPNST.

BIOINFORMATICS AND COMPUTATIONAL OMICS

Authors: Megan Stevens, Benjamin E. Housden

Funding: Medical Research Council

Disclosures: None

Purpose: The aim of this project is to develop effective therapies to treat tumours associated with Neurofibromatosis type 1 (NF1). NF1 affects approximately 1/3,000 live births and is associated with a variety of symptoms including the formation neurofibromas along peripheral nerves. These tumours, cause disfigurement and can disrupt nerve function leading to pain and loss of motor control. A proportion of tumours develop into malignant peripheral nerve sheath tumours (MPNSTs) that are often fatal. 

While MEK inhibitors are currently used to treat a subset of neurofibromas, these are not effective in all patients and can cause side effects. 

We previously generated a synthetic lethal interaction network for NF1, containing 54 potential drug-targets for NF1 tumours (PMID: 39129390). Initially, we focused on autophagy as a target and showed that chloroquine caused selective killing of NF1 deficient cells across a range of in vitro and in vivo models of NF1 tumours. This demonstrates the value of the network in developing new treatment options for NF1. Here, we described new results building on our synthetic lethal network to develop a synergistic drug combination with high potential as a future treatment for NF1 tumours.

Methods: To identify candidate drug-targets not conserved in Drosophila, we performed semantic similarity analysis based on the results of the synthetic lethal screens. Existing drugs were tested in Drosophila and human patient-derived cell models as well as an in vivo MPNST xenograft model. 

Results: One challenge of screening for drug targets using Drosophila cells is that human targets that are not conserved in Drosophila cannot be directly identified. To address this limitation, we performed statistical enrichment analysis to identify human-specific genes with close functional relationships with the hits from the Drosophila screens. This identified TERT as a candidate target. We then showed that a repurposed inhibitor known to affect TERT function, caused selective death of NF1 deficient patient-derived cells and MPNST-derived cells both in vitro and in vivo. Further testing revealed that this inhibitor synergises with MEK inhibition to enhance efficacy.

Conclusions: Synthetic lethal screens in Drosophila cells combined with statistical enrichment analysis is a powerful approach to identify novel treatment strategies for NF1 tumours. 

From our studies, we conclude that inhibition of the telomerase complex synergises with MEK inhibition to kill NF1-deficient tumour cells.

THERAPEUTICS

Authors: Wei Wang^1,2†, Haibo Li^1,2†, Zhichao Wang^1,2*, Qingfeng Li^1,2*

Affiliations:

¹ Department of Plastic and Reconstructive Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine; Shanghai, 200011, China.

² Neurofibromatosis Type 1 Center and Laboratory for Neurofibromatosis Type 1 Research, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine; Shanghai, 200011, China.

^†These authors contributed equally to this work. Email: dr_wangwei60@163.com(W.W.); drhaiboli@gmail.com(HB.L.).

*Corresponding author. Email: dr.liqingfeng@shsmu.edu.cn (Q.L.); shmuwzc@163.com (Z.W.).

Disclosures: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Funding: National Natural Science Foundation of China (82202470,82102344, 82172228).

Title: Multifunctional Chemoreactive Nanosonosensitizers Exert Antitumoral, Antibacterial and Wound Healing Effects on Malignant Peripheral Nerve Sheath Tumors

Purpose: Currently, surgery is the only available and effective clinical strategy for treating malignant peripheral nerve sheath tumors. However, even with complete surgical resection, poor physical health or cachexia would cause wound delay or infection. To solve this critical issue, DSF-CuO₂@DMSNs are rationally designed. 

Methods: The nanosonosensitizers are engineered to exert high performance and synergistic sonodynamic/chemodynamic/chemotherapeutic tumor treatment under the stimulation of ultrasound by producing reactive oxygen species and dithiocarbamate−copper complexes in situ in response to the tumor-specific acidic and hypoxic microenvironment. 

Results: Significant inhibition in cell viability, increment in cell apoptosis and death, and tumor cell-xenograft experiments all demonstrate the antitumoral capability of DSF-CuO₂@DMSNs in combination with ultrasound stimulation. The upregulated expression of caspase-3 and cleaved caspase-3 in both cell and resected xenograft protein lysates indicated the increment of cell apoptosis. Moreover, DSF-CuO₂@DMSNs also induce strong antibacterial effects and wound healing abilities under the activation of ultrasound both in vivo and in vitro, with histological examination results of quick wound closure, absence of bacterial infection, and new blood vessel formation. 

Conclusion: Altogether, the multifunctional DSF-CuO₂@DMSNs are highly desirable chemoreactive nanoagents that can provide a distinct postoperative therapeutic strategy for both tumor inhibition and wound recovery.

Malignant peripheral nerve sheath tumors (MPNST)

Carson Gutierrez and Kimberly Ostrow

Department of Neurology, Johns Hopkins School of Medicine

Purpose

Schwannomatosis (SWN) patients develop multiple tumors along peripheral nerves, with most experiencing significant pain. While neuropathic, nociceptive, and inflammatory pain types are reported, many describe severe pain upon palpation or light touch. Currently, surgical removal is the only effective pain relief option.

We aim to investigate the underlying mechanisms of tumor-induced pain. Tumor growth may increase pressure on nearby nerves, causing pain. Additionally, schwannoma cells secrete proinflammatory cytokines, sensitizing sensory neurons to painful stimuli both in vitro and in vivo. When injected into the glabrous skin of a mouse hind paw, conditioned medium (CM) from painful schwannomas hypersensitizes mice to mechanically induced pain demonstrated by a fourfold reduction in paw withdrawal threshold within an hour (p = 0.006), with effects lasting 48 hours (p = 0.002) in the Von Frey assay.  We hypothesize that this increase in sensitivity to light touch is linked to mechanosensitive ion channels (MSCs), which detect pressure,  stretch, and inflammation. GsMTx-4, a peptide that selectively blocks MSCs without affecting other ion channels, has shown effectiveness in reducing inflammation-induced mechanical pain and represents a promising alternative to surgery.

Methods & Results

To assess the role of MSCs in SWN CM-induced pain, we injected 10 µM GsMTx-4 into one footpad of C57Black mice alongside either control media (n=10) or painful SWN CM (n=10). One hour post-injection, Von Frey filaments were used to assess paw withdrawal threshold via the up/down method. When co-injected with CM, 10 µM GsMTx-4 prevented CM induced heightened sensitivity to light touch. GsMTx-4 had no effect on control treated mice demonstrating that mechanosensitivity is CM dependent. In a separate cohort, painful CM significantly increased hypersensitivity to light touch that lasted for 48 hours. Following this initial Von Frey testing, the mice received a 10 µM GsMTx-4 injection and underwent a second round of testing. GsMTx-4 significantly reversed the pain phenotype in these animals. Lastly, we found that SWN CM induces hyperalgesic priming and a transition to chronic pain. Mice with chronic pain treated with 10uM GsMTx-4 demonstrated a complete reversal of painful phenotype (p<0.0001).

Conclusions

Our findings demonstrate that painful SWN CM primes mice for hyperalgesia, leading to a transition from acute to chronic pain. GsMTx-4 effectively prevents and reverses acute and chronic mechanical pain induced by SWN CM. Ongoing studies aim to optimize dosage of GsMTx-4 to achieve maximum pain relief. This peptide offers a promising, minimally invasive therapeutic approach for pain in SWN patients, potentially eliminating the need for surgery.

Funding sources This work was graciously funded by The Children’s Tumor Foundation Drug Discovery Initiative Award, The Blaustein Pain Foundation and The JH Neurosurgical Pain Research Institute

{{DRUG DISCOVERY AND DEVELOPMENT}}

Pei-Yu Hung, MS, Department of Physiology, Michigan State University, East Lansing, MI

Neurofibromatosis type 1 (NF1) is a common genetic disease that predisposes approximately 50% of affected individuals to develop plexiform neurofibromas (PNFs), which can progress to highly aggressive malignant peripheral nerve sheath tumors (MPNSTs) in approximately 10% of patients. Selumetinib, a specific inhibitor of MEK1/2, is the only FDA-approved drug for NF1-associated PNFs. However, the anti-tumor effects of selumetinib are limited in MPNSTs, and the drug has dose-limiting side effects. During the formation of neurofibromatosis, a complex tumor microenvironment (TME) develops, with the infiltration of macrophages critical for growth and progression. Targeting tumor-promoting immune cells could be an alternative approach for treating or preventing the progression of NF1. The novel retinoid X receptor (RXR) agonist MSU-42011 reduces tumor growth in experimental Kras-driven lung cancers by decreasing pERK levels, reducing tumor-promoting immune cells like CD206+ macrophages and regulatory T cells, and increasing activated cytotoxic T cells.

Purpose: Given the similarities in RAS activation and immune cell infiltration in NF1 and Kras-driven lung cancer, we hypothesized that MSU-42011 can be used as an alternative approach to selumetinib and modulate immune cell populations within the NF1 TME.

Methods: We treated NF1-deficient cells and a syngeneic mouse model of MPNST with MSU-42011 and selumetinib, either alone or in combination, to evaluate their immunomodulatory and anti-tumor effects.

Results: In NF1-deficient cells, treatment with MSU-40211 or selumetinib reduced pERK protein levels, and the combination treatment enhanced this reduction. Additionally, there was a trend toward reduction in cell viability with increasing drug concentrations after the combination treatment. Moreover, conditioned media (CM) from human and mouse PNF cells increased the mRNA expression of monocyte chemoattractant CCL2 (C-C motif chemokine ligand 2) and the secretion of IL-6 and TNFα in human THP1 monocytes/macrophages and bone marrow derived macrophages (BMDM). Notably, MSU-42011 and selumetinib alone inhibited CCL2 mRNA expression in THP1 macrophages and BMDM stimulated with CM from human and mouse PNF cells, respectively, and the inhibition of CCL2 mRNA expression was greatest with the combination treatment. The combination of MSU42011 and selumetinib also significantly reduced tumor growth and reduced pERK levels, tumor-promoting CD206+ macrophages, and regulatory T cells in both a LL2 model of lung cancer driven by an activating Kras mutation and a syngeneic mouse model of MPNST.   

Conclusions: As NF1 is a complex multisystem disorder, a combination of drugs with different mechanisms will most likely be more effective than single agents. Our initial results form the justification for further preclinical evaluation in vivo. Currently, we are assessing the immunomodulatory and anti-tumor effects of MSU-42011 and selumetinib in a genetically engineered model of PNF and MPNST, with the eventual goal of translating these findings into clinical applications.

Full List of Authors: Pei-Yu Hung¹, Jessica A. Moerland², Karen T. Liby³, and Ana S. Leal³

¹Department of Physiology, Michigan State University, East Lansing, MI

²Department of Pharmacology and Toxicology, Michigan State University, East Lansing, MI

³Department of Medicine, Division of Hematology/Oncology, IU School of Medicine, Indianapolis, IN

Funding: Falk Medical Research Trust Catalyst Award

Patent applications covering the novel compounds described in this work have been applied for on behalf of Michigan State University; Karen T. Liby is a named inventor on the patent applications and a founding scientist of Akeila Bio.

{{DRUG DISCOVERY AND DEVELOPMENT}}

Juan Sebastián Botero-Meneses³, Juliana Valentina Ramos Rubio², Laura Daniela Ortiz Portela², Tatiana Camacho Carvajal ², Ana María Téllez Blanco ², José David López Báez ², and Ana-María Gómez-Carvajal¹

¹Universidad del Rosario, School of Medicine and Health Science, People, Family and Society Research Group, Bogotá, Colombia.  

²Universidad del Rosario, School of Medicine and Health Science, Bogotá, Colombia.  

³Universidad del Rosario, School of Medicine and Health Science, Neuroscience (NEUROS) Research Group, Neuroscience center Neurovitae-UR, Bogotá, Colombia.  

No funding to disclose

Objective: The purpose of this article was to conduct a systematic review of the literature pertaining social cognition in patients with Neurofibromatosis type 1 (NF1). We analyzed social cognition (SC) which encompasses a wide set of neurobiological and social processes that enable individuals to perceive, identify, and assess social events. We selected articles that evaluated empathy, social emotions, theory of mind and moral judgement. All of these, are fundamental to understand the mechanisms that underlie social functioning and has often been found to be altered in NF1 patients.

Method: We conducted a systematic review of the literature. Articles were found using two electronic databases (Scopus and PubMed) and manual searching.

The research question that guided this systematic review was addressed using a PIO (Population, Control, and Outcome) structure. We thoroughly applied criteria that allowed us to select studies which evaluated our selected outcomes in NF1 population. These included patients with NF1 and no other genetic diagnosis, the use of a control group to compare SC performance and the use of objective measurements for results.

Results: When compared with unaffected controls, individuals with NF1 (children and adults) exhibit SC dysfunction in ToM, empathetic pain perception, and social emotion recognition. No articles evaluated moral judgment.

Conclusion: Further studies are needed, and they must evaluate specific SC domains using appropriate validated tests, and assessing changes in SC in patients as they age.

Key words: Social Cognition, Empathy, Moral Judgement, Neurofibromatosis type 1, Social Emotions, Theory of Mind 

{{Cognition / Behavioral / Learning}}

Presenting Author: Cecilia Tibery (PA-C), Clinical Research Directorate (CRD), Frederick National Laboratory for Cancer Research

Introduction: People with neurofibromatosis Type 1 (NF1) and inoperable symptomatic plexiform neurofibromas (PN), may benefit from treatment with a MEK inhibitor (MEKi). An increased risk for vascular anomalies such as arteriovenous fistulas and arterial aneurysms has been described in NF1¹, however to our knowledge these have not been reported as new findings in patients on MEKi. Here we report on 2 patients with NF1 and PN who developed aneurysms while on selumetinib.

Methods: We performed a retrospective review of two NF1 patients who developed aneurysms during treatment with selumetinib on a single arm phase 2 trial (NCT02407405). Relevant patient information, including prior history of vascular anomalies, medical/surgical history, selumetinib treatment history, selected adverse events including hypertension, and interventions were extracted from the electronic medical records. Selumetinib was administered on a continuous dosing schedule at 50 mg twice daily (1 cycle=28 days). 

Results: 

Patient 1, a 30 year old with a left head and neck PN, had new neck swelling during his restaging evaluation during cycle 31 (~2.3 years) of selumetinib.  The restaging non-contrast MRI of his PN showed an internal left carotid aneurysm (1.5x1.6 cm) that was not present on MRI 6 months prior. He underwent vascular stent placement approximately 1 week after the aneurysm was discovered. Past medical history was significant for radiation therapy for a pilocytic astrocytoma at 6 years old and for a probable high grade glioma in the left cerebellar vermis at 25 years old. Patient 2, a 41 year old with a left neck and chest PN, had a restaging non-contrast MRI during cycle 67 (~5.1 years) of treatment that identified a new aymptomatic left vertebral artery aneurysm (2.5x2.9 cm) with an arteriovenous fistula that was not seen on the restaging MRI 6 months prior. He underwent a coil embolization 1 week after diagnosis. Past medical history was significant for hypertension. Both patients recovered uneventfully. 

Conclusions: People with NF1 are known to be at increased risk for developing vascular anomalies such as aneurysms¹. To our knowledge, this is the first report of the development of aneuryms in NF1 patients receiving a MEKi for inoperable PN. Both aneurysms were identified on non-contrast MRI as part of a research protocol, highlighting the need for careful review of any imaging obtained for these patients.  While the aneurysms developed on treatment with selumetinib, both patients had known   risk factors such as prior radiation therapy or hypertension, therefore a relationship between selumetinib and aneurysm was considered unlikely by the study team.  However, given the increasing use of MEKi for people with NF1, careful monitoring for the development of vascular anomalies will be important and provide additional information regarding any potential relationship.  

Full Author list 

Cecilia Tibery¹, Hari Sankaran², Eva Dombi³, Scott Lindhorst⁴, Hans Shuhaiber⁵, Alejandro Spiotta⁴, Ariana Alejandra Chacon⁴, Hasna Loulida⁴, Andrea Baldwin¹, Margaret Fagan³, Kara Heisey¹, Geraldine O’Sullivan Coyne⁶, Alice P. Chen⁶, Andrea M. Gross³, Brigitte C. Widemann³

Disclosures: None reported

Funded by the NIH intramural research program and NCI Contract No HHSN261201500003I and 75N91019D00024. 

Affiliations

¹Clinical Research Directorate, Frederick National Laboratory for Cancer Research, support to the National Cancer Institute, Center for Cancer Research, Bethesda, Maryland, USA

²Biometric Research Program, Division of Cancer Treatment and Diagnosis, NCI, NIH, Bethesda, MD 20892, USA

³Pediatric Oncology Branch, Center for Cancer Research, National Cancer Institute, NIH Bethesda, Maryland, USA

⁴Department of Neurosurgery, Medical University of South Carolina, Charleston, South Carolina, USA

⁵Department of Neurology, University of Florida, Gainesville, Florida, USA

⁶Developmental Therapeutics Clinic, Division of Cancer Treatment and Diagnosis, National Cancer Institute, Center for Cancer Research, Bethesda, Maryland, USA

References

1. G.S. Oderich, T.M. Sullivan, T.C. Bower, et al.; Vascular abnormalities in patients with neurofibromatosis syndrome type I: clinical spectrum, management, and results; 17681709J Vasc Surg, 46 (2007), pp. 475-484

Authors:

Dr Mubin Tahir, MRCPsych, Greater Manchester Mental Health NHS Foundation Trust 

Dr Grace Vassallo, MD, DCH, MRCPCH, Manchester Foundation Trust

Dr Shruti Garg, MRCPsych, PhD, University of Manchester

Dr Stavros Stivaros, FRCR, PhD, University of Manchester  

Funding:

The study was funded by the NIHR Manchester Biomedical Research Centre (NIHR203308). SG was awarded a Francis Collins Scholarship to support this study through Neurofibromatosis Therapeutic Acceleration Program (NTAP).

Disclosures:

No disclosures. 

Title:

Outcome of brain imaging in asymptomatic children with NF1

Purpose:

To analyse the outcome of MR scans of the brain undertaken on children aged 6-18 years as part of research trials.

Methods:

In 2009 NHS England commissioned two Highly Specialized Services for complex NF1. The Manchester service has undertaken two neurodevelopmental studies in children aged 6-18 years where neuroimaging was part of the research protocol and in whom they were clinically asymptomatic at enrolment into the study. The first study took place in 2019 to 2020 and the second started in 2024 and is ongoing.

Results:

The MR brain images and records of 57 children were available to us. 4 of the records were incomplete and in 8 children the imaging is still being reviewed. We have complete records on 45 children with an age range of 6-18 years and a mean / median age of 14 years. 25 are male and 20 female. Positive findings triggering further intervention were identified in 18 (40%) of children. In 2019 cohort one 14-year-old male had an asymptomatic dysembryoplastic neuroepithelial tumour (DNET). One had an asymptomatic optic pathway glioma (OPG). She has never needed treatment and has had several years of follow up with complete stability of her OPG. One child in whom the scan raised the possibility of need for cerebrospinal fluid (CSF) flow studies was lost to follow up. In 5 children nonspecific changes were seen in which it was impossible to differentiate between myelin vacuolation changes and low-grade gliomas. Their ongoing surveillance imaging continues to require yearly follow up but they remain asymptomatic. In the cohort of children 2024-2025, 3 children have asymptomatic OPGs, 3 children have slightly smaller pituitary glands and will require clinical review for growth and pubertal status and 4 children again have nonspecific changes where low grade gliomas cannot be excluded. These will all enter the clinical complex NF1 paediatric pathway in the service.

Conclusion:

The question of asymptomatic annual screening for children with NF1 continues to be raised by families and clinicians alike. Our review of neuroimaging of asymptomatic children who had a brain scan as part of a research study demonstrates that a significant proportion of these scans reveal changes that require further imaging and further clinical reviews. Reassuringly however, none of the positive findings changed the individual’s clinical outcome. Our study supports the present recommendation that MR scans should only be undertaken in children with NF1 in whom there are clinical concerns. 

Other

Soniya Chatterjee¹, Federica Tomasso², Dipak Poria ¹, Sridevi Yadavilli ¹, Roger J. Packer¹, Javad Nazarian³

¹Brain Tumor Institute, Center for Cancer and Immunology Research. Children’s National Hospital, Washington, DC 20012

² Department of Cardiology, Colorado University Anschutz Campus, Aurora CO 80045

³Department of Oncology, University Children’s Hospital Zürich, Balgrist Campus, Zürich, CH-8008, Switzerland.

schatterje@childrensnational.org, DPORIA@childrensnational.org, Federica.tomasso@cuanschutz.edu SYadavil@childrensnational.org, RPACKER@childrensnational.org, Javad.Nazarian@kispi.uzh.ch. 

Background: Neurofibromatosis type 1 (NF1) is a genetic disorder caused by NF1 gene mutations, affecting 1 in 3,000 individuals worldwide. It is characterized by cutaneous neurofibromas and an elevated tumor risk, with 15–20% of patients developing gliomas. While NF1 inactivation is linked to low-grade gliomas, the drivers of malignant transformation remain poorly understood. We hypothesized that a permissive NF1-mutant cerebral microenvironment may contribute to malignant transformation and treatment responses of gliomas. Current preclinical models, such as glioma cell lines and xenografts, fail to replicate the complex NF1 tumor microenvironment. Human induced pluripotent stem cell (hiPSC)-derived cerebral organoids provide a promising platform for modeling NF1-driven neurodevelopment and tumor biology.

Purpose: To investigate the role of NF1 mutations in cerebral organoid development, glioma plasticity, and treatment response.

Methods: Cerebral organoids were generated from healthy control and NF1-mutated (NF1+/-, NF1-/-) iPSCs reprogrammed from plexiform neurofibromas. Organoids were analyzed for cytoarchitectural development at key stages using immunofluorescence assays. mCherry-labeled glioma cells TM31 and patient-derived high-grade astrocytoma with piloid features (HGAP) cells were co-cultured with cerebral organoids to assess glioma migration (qualitative imaging), proliferation (qPCR), secretome profiles (MSD), and treatment response (cell titer glow assay).

Results and Conclusions: Initial immunofluorescence analysis revealed no significant differences in neuronal development between wild-type and NF1-mutant organoids. A co-culture of our novel organoids with TM31 and patient-derived HGAP glioma cells was successfully established. Preliminary experiments indicate increased glioma migration and proliferation in NF1-mutant organoids compared to NF1 wild type controls. Image analysis further suggests localized glioma growth within specific zones of the organoid, warranting further exploration. Additionally, elevated levels of GM-CSF and IL-18—cytokines implicated in glioma growth and invasion—were detected in the secretome of NF1-mutant organoid-glioma co-cultures. These findings suggest that NF1 mutations foster a tumor-supportive microenvironment, enhancing glioma aggressiveness and potentially influencing treatment response.

To explore therapeutic strategies, we piloted a CDK4/6 inhibitor using TM31 cultures and showed drug’s efficacy in targeting tumor cell viability. Drug combination assays with MAPK/mTOR inhibitors are underway.   The most potent combinations will then be used for organoid co-cultures. 

This study highlights the utility of NF1-mutant organoids as a platform to study glioma behavior and NF1-associated tumor predisposition. Future analyses, including single-cell RNA sequencing and spatial transcriptomics, will further validate NF1's role in glioma progression. These findings pave the way for ex vivo drug testing to improve personalized therapies for NF1-driven gliomas.

{{DRUG DISCOVERY AND DEVELOPMENT}}

Objective: The aim of this study is to observe the efficacy of surgical resection and targeted drug therapy in treating plexiform neurofibromas in the neck of children.

Methods: A 3-year-old child with a progressively enlarging plexiform neurofibroma in the occipitocervical region underwent surgical resection, achieving a resection extent of over 95%. Regular imaging follow-ups were conducted postoperatively, with the follow-up period exceeding five years. Additionally, two other children with plexiform neurofibromas in the neck were treated with the targeted drug selumetinib. After starting the medication, they underwent neck MRI scans approximately every four months for follow-up.

Results: In the child who underwent surgical treatment, significant recurrence of the plexiform neurofibroma in the neck was observed at the first follow-up examination four months after surgery. Imaging studies conducted at different time points postoperatively revealed progressive enlargement of the plexiform neurofibroma in the neck. Among the two children treated with medication, the tumor in one child progressively shrank after taking the drug, while the tumor in the other child remained largely unchanged following treatment.

Conclusion：For plexiform neurofibromas in the neck, both surgical resection and targeted drug therapy have demonstrated efficacy. However, considering the rapid growth of tumors during childhood and the high likelihood of recurrence after surgery, targeted drug therapy or a combination of surgery and targeted drugs appears to be a more rational treatment approach.

Fig.1 Imaging data before and after surgery for the child who underwent surgical treatment.A: Preoperative MRI showed a large tumor in the neck.B: The MRI performed three days after surgery showed that the tumor had been largely resected.C: The tumor specimen resected during surgery.D: The MRI performed four months after surgery showed tumor recurrence.E: The MRI performed 28 months after surgery showed progressive enlargement of the tumor.F: The MRI performed 50 months after surgery showed continued enlargement of the tumor.

Fig.2 Imaging Data Before and After Targeted Drug Therapy for one Child.A: The MRI performed before the administration of the targeted drug showed a large tumor in the neck. B: The MRI performed four months after the initiation of the targeted drug therapy showed a significant reduction in the size of the neck tumor.C: The MRI performed eight months after the initiation of the targeted drug therapy showed continued shrinkage of the tumor.

Authors:
 Purva Gade¹, Marissa Howard¹, Lance Liotta¹
 ¹George Mason University, Fairfax, VA, USA

Background and Purpose: Neurofibromatosis Type 2 (NF2) is a hereditary tumor disorder caused by mutations in the NF2 gene, resulting in the loss of the tumor suppressor protein Merlin. NF2 patients develop tumors such as vestibular schwannomas and meningiomas, which cause hearing loss, tinnitus, and balance dysfunction. Current therapies are largely palliative, addressing symptoms rather than targeting tumor growth. Thus, there is a need to uncover novel mechanisms contributing to NF2 tumor progression. Cancer cells undergo metabolic reprogramming to survive oxidative stress by altering mitochondrial dynamics through fission and fusion. In healthy cells, mitophagy removes damaged mitochondria via lysosomal degradation. This process is regulated by mitochondrial proteins, including PINK. Our study identifies a novel alternative pathway secretory mitophagy by which NF2 tumor cells export damaged mitochondria via extracellular vesicles (EVs), while simultaneously exporting tumor suppressor molecules, such as Merlin. This process promotes tumor survival under oxidative stress.

Methods: Meningioma cells with NF2 mutations were treated with oxidative stress inducers combined with lysosomal inhibition to block canonical mitophagy. EVs were isolated from conditioned media and analyzed for Merlin, FIS1, and PINK1 content. Immunoprecipitation (IP) of PINK1-positive EVs was performed to detect co-exported Merlin. To investigate FIS1’s role, we conducted siRNA-mediated knockdown followed by oxidative stress treatment, then measured EV secretion, cell survival, and secretory mitophagy levels.

Results: Oxidative stress combined with lysosomal inhibition led to significantly increased EV secretion enriched for damaged mitochondria. These EVs contained Merlin, FIS1, and PINK1, indicating active export of both damaged mitochondria and tumor suppressor proteins. Immunoprecipitation confirmed Merlin co-localized with PINK1+ EVs, demonstrating tumor suppressor export. FIS1 knockdown significantly reduced secretory mitophagy and cell survival, confirming that FIS1 is critical for this adaptive process.

Conclusion: This study identifies secretory mitophagy as a previously unrecognized survival strategy employed by NF2 tumor cells. By exporting damaged mitochondria along with tumor suppressor molecules like Merlin, NF2 tumor cells enhance their survival under oxidative stress and further disrupt tumor suppressor signaling networks, promoting unchecked growth. Targeting secretory mitophagy could offer a novel therapeutic strategy to restore intracellular tumor suppressor levels and impair tumor stress adaptation mechanisms in NF2-associated tumors.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Alexandra O’Donohue^1,2, Hsien-Yin Kao^1,2, Samantha Ginn³, Gaetan Burgio⁴, Aaron Schindeler^1,2

¹   Bioengineering and Molecular Medicine Laboratory, The Children’s Hospital at Westmead and Westmead Institute for Medical Research, Westmead, NSW 2145, Australia

²    School of Chemical and Biomolecular Engineering, Faculty of Engineering, The University of Sydney, Sydney, NSW 2006, Australia

³   Gene Therapy Research Unit, Children's Medical Research Institute, Faculty of Medicine and Health, The University of Sydney and Sydney Children's Hospitals Network, Westmead, NSW 2145, Australia

⁴   The John Curtin School of Medical Research, Australian National University, Canberra, ACT 0200, Australia

Funding provided by Flicker of Hope Foundation and National Health and Medical Research Council (Australia).

Discloses: No conflicts to declare

Purpose: CRISPR base editing (CRISPR-BE) holds immense promise for rewriting single nucleotide variants (SNVs) and transforming genetic medicine. In NF2-related schwannomatosis, SNVs make up 40% of the entire pathogenic mutational burden. To demonstrate proof-of-concept for therapeutic repair, we selected a prototypical NF2 patient mutation (NF2 c.169C>T) for BE correction.

Methods: A dual-plasmid approach was used to express of fourth-generation CRISPR base editors (ABEMax and ABE8e) alongside targeted single guide RNAs (sgRNAs). A NF2 c.169C>T mutant HEK293 cell line was generated in-house by CRISPR-HDR as a resource for testing BE efficiency. Cells transfected with both plasmids were selected with puromycin and genomic DNA extracted for next generation sequencing. In parallel, a Nf2 c.169C>T mouse model was generated, which features a humanized sequence around the mutation site such that human sgRNAs would be compatible. 

Results: NGS demonstrated 76% correction of the pathogenic c.169C>T mutation with ABEMax and 90% correction with ABE8e, which could yield significant phenotypic effects if achieved in vivo. An infrequent c.167 bystander mutation was noted with both strategies, although this was predicted to lead to a less deleterious missense change. Only the heterozygous Nf2^169C>T/+ mice were viable, but akin to the Nf2 heterozygous null mouse, does not feature frequent, spontaneous tumors.

Conclusion: We have developed a base editing strategy capable of repairing the NF2 c.169C>T mutation at high efficiency in vitro. Future work will focus on in vivo delivery using a Schwann cell-targeting adeno-associated vector (AAV) to base edit Nf2^169C>T/+ Schwann cells ex vivo and in vivo.

{{THERAPEUTICS}}

Authors: Alise Blake, MS, CGC¹, Leslie Taylor, RN¹, Stacy Hines-Dowell, DNP, AGN-BC, ACGN¹, Amanda Rotenberry, FNP-BC², Roya Mostafavi, MS, CGC², Kim E. Nichols, MD¹, Eniko Pivnick, MD², Jessica Gartrell, MD¹, Melissa Perrino, MD¹

¹St. Jude Children’s Research Hospital, Oncology

²Le Bonheur Children’s Hospital

Funding: This study was funded by the American Lebanese Syrian Associated Charities.

Disclosure of relevant financial relationships: None to disclose

Background/Purpose: Neurofibromatosis Type 1 is a common genetic disorder characterized by over 2800 pathogenic variants in NF1, most with variable phenotypic presentations. However, one of the few described genotype-phenotype correlations occurs at NF1 p.Arg1809. Pinna et al. reported 14 patients with p.Arg1809 missense variants who had café au lait macules (CALMs) and Noonan-like facies, but lacked cutaneous, subcutaneous, or plexiform neurofibromas (PNs; Eur J Human Genet, 2015). Rojnueangnit et al. identified patients with p.Arg1809 missense variants are significantly more likely to have short stature or pulmonic stenosis (p<0.0001), and less likely to have cutaneous neurofibromas, optic pathway glioma (OPG), scoliosis or PNs compared to the NF1 population (p<0.05; Hum Mutat, 2015). This case report describes a patient with NF1 p.Arg1809 frameshift variant and high NF1-associated tumor burden.

Case Report: We present a 21-year-old male with maternally inherited NF1 c.5425delC; p.Arg1809AlafsTer33. Similar to the literature, he has short stature, CALMs, and Noonan-like facies including palpebral ptosis, wide neck, low set ears, and retinal pigment epithelial hypertrophy. Conversely his phenotype has been more severe with a ventricular septal defect and symptomatic OPG in infancy, scoliosis associated with multiple spinal PNs requiring surgical intervention in early childhood, then numerous PNs in late adolescence and early adulthood. At age 17, a rapidly growing thigh PN was resected revealing atypical neurofibromatous neoplasm of uncertain biologic potential (ANNUBP) with loss of CDKN2A and CDKN2B. At age 19 he developed spinal cord compression prompting a large partial resection of a C3-C4 PN followed by treatment with a MEK-inhibitor. However, treatment was discontinued after 15 cycles due to mild cardiotoxicity in the setting of a suboptimal response. Most recently a growing left forearm lesion was resected and histology confirmed ANNUBP with loss of CDKN2A. The patient continues to be followed closely due to numerous remaining nodular PNs. Family history is notable for individuals with spinal neurofibromas and Noonan-like facies, but there is no history of malignant peripheral nerve sheath tumors. 

Conclusion: This individual’s p.Arg1809 variant correlates well with the reported phenotypic overlap with Noonan syndrome. However, this frameshift variant predicts an alternate stop codon and loss of function due to premature protein truncation or nonsense-mediated mRNA decay which may explain why he does not fit the low tumor profile reported with p.Arg1809. More research is needed to understand NF1 variant impact on malignant transformation as well as response to, and toxicity from, systemic therapy with MEK inhibition.

{{Other}}

Jay Pundavela PhD¹, Samantha Anne Dinglasan BSc¹, and Nancy Ratner PhD^1,2

¹Division of Experimental Hematology and Cancer Biology, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45229, USA; ²Department of Pediatric, College of Medicine, University of Cincinnati, Cincinnati, OH 45229, USA

Funding: NIH 28840 (NR) and CancerFree Kids (JP)

Disclosure: Nothing to disclose.

Plexiform neurofibroma (PNF) is a benign peripheral nerve sheath tumor associated with Neurofibromatosis Type 1 (NF1). Beyond the presence of neoplastic Schwann cells, an inflammatory milieu enriched with immune cells, including myeloid subsets (macrophages and dendritic cells) and T lymphocytes, contributes to PNF pathogenesis. Our recent findings highlighted the critical involvement of CD8 T cells in neurofibroma tumorigenesis, after loss of Nf1 in Schwann cells (Pundavela et al., Sci. Adv., 2024). However, the impact of manipulating T cell trafficking on neurofibroma growth remained unexplored.

Aim: In this study, we investigated the modulation of T cell migration through the administration of Fingolimod—a sphingosine-1-phosphate receptor modulator approved for use in Multiple Sclerosis—on established neurofibromas in a clinically relevant preclinical mouse model (Nf1 f/f; DhhCre), which closely mimics the pathological features of NF1 in humans.

Methods: We administered Fingolimod at a daily dose of 10 mg/kg orally for 30 days to Nf1 f/f; DhhCre mice, with a parallel vehicle control group. Toxicity assessments included monitoring body weight and histological evaluations of liver, lung, and kidney tissues. Migration of lymphocytes across lymphoid organs and tumors was assessed using multiparametric flow cytometry and immunofluorescence.

Results: No significant toxicity was observed, as evidenced by stable body weights and unremarkable tissue morphology in Fingolimod-treated mice compared to controls. Importantly, Fingolimod treatment resulted in a notable reduction in tumor size. Flow cytometry analysis revealed a significant decrease in circulating T cells, with a marked reduction in CD8 T cells while CD4 T cell levels remained unchanged. Conversely, an increased frequency of CD8 T cells was noted in the lymph nodes of Fingolimod-treated mice, indicating an inhibition of CD8 T cell egress from lymphoid compartments. No alterations were detected in the CD8/CD4 T cell ratio within the spleen across both treatment groups. 

Conclusion: Our findings support a pivotal role of T cell trafficking in the growth of neurofibromas and suggest that Fingolimod may serve as an immunomodulatory agent for neurofibroma treatment. This research supports testing of therapeutic strategies that target T cell dynamics to effectively manage neurofibromas.

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Authors:  Ellen Voigt^1,2,6, Joshua J Lingo^2,6, Altay Koyas^2,6, Rebecca D Dodd, PhD^3,6, Benjamin W Darbro, PhD^4,6, Dragana Kopanja, PhD⁷, Benita S Katzenellenbogen, PhD⁸, John A Katzenellenbogen, PhD⁹, Dawn E Quelle, PhD^5,6

¹Medical Scientist Training Program, ²Cancer Biology Program, Departments of ³Internal Medicine, ⁴Pediatrics, and ⁵Neuroscience & Pharmacology, ⁶Holden Comprehensive Cancer Center, U of Iowa; ⁷Department of Biochemistry and Molecular Genetics, U of Illinois Chicago; Departments of ⁸Molecular & Integrative Physiology and ⁹Chemistry, U of Illinois Urbana-Champaign.

Funding: R01 NS119322, T32 CA078586, Adolescent Young Adult Cancer Program (U Iowa), BCRF 24-083. Disclosures: None.

Purpose: Malignant peripheral nerve sheath tumors (MPNSTs) are sarcomas that arise either spontaneously or through transformation of benign tumors, called plexiform neurofibromas (PNFs), in patients with Neurofibromatosis Type 1 (NF1). Understanding the driver mutations that cooperate to promote malignancy remains a priority in the field. One potential driver, FOXM1, is an oncogenic transcription factor that promotes malignant progression, drug resistance, and immune evasion in other cancers. There are just two published studies of FOXM1 in MPNSTs, both of which suggest it promotes the disease. We seek to understand the role of FOXM1 in these deadly tumors and hypothesize that FOXM1 is essential for MPNST pathogenesis. 

Methods: FOXM1 mRNA and protein expression were evaluated in patient matched PNFs, ANNUBPs, and MPNSTs via RNA-Seq and IHC. For in vitro studies, we generated normal human Schwann cells (NHSCs) and MPNST cell lines (S462, sNF96.2) with overexpression (OE) or knockdown (KD) of FOXM1 isoforms b and c. Cell proliferation, survival, and transformation status are being measured as is their response to historical and newly developed FOXM1 inhibitors. To directly test the in vivo role of FOXM1 in MPNST initiation and progression, de novo MPNSTs were initiated by Nf1/Ink4a/Arf editing in the sciatic nerve of Nf1^+/-, Foxm1 floxed mice relative to Nf1^+/- controls. 

Results: In patient matched tumor sets, FOXM1 mRNA was significantly elevated in MPNSTs relative to patient-matched PNF/ANNUBP precursor lesions. At the protein level, FOXM1 expression rose dramatically in a stepwise manner from normal nerve to PNFs, ANNUBPs, and MPNSTs. Increased FOXM1 expression corresponded with heightened transcriptional activation, as measured by upregulation of FOXM1 target genes in MPNSTs versus PNFs. Among its targets, upregulation of programmed death ligand 1 (PD-L1) protein correlated tightly with increased FOXM1 in patient tumors. Broad acting FOXM1 inhibitors, thiostrepton and FDI-6, effectively inhibited MPNST proliferation and induced apoptosis. Excitingly, synergistic killing of MPNST cells was obtained by combining FOXM1 inhibitors with a MEK inhibitor (mirdametinib), CDK4/6 inhibitor (palbociclib), or EGFR inhibitor (gefitinib), all of which are relevant drugs for MPNST therapy.

Conclusion: Our data suggest FOXM1 is an important driver of MPNST pathogenesis. FOXM1 expression and transcriptional activity are greatly increased in human MPNSTs compared to benign precursors from the same patients. Therapeutic inhibition of FOXM1 in vitro kills MPNST cells and effectively synergizes with several clinical drugs that antagonize MEK, CDK4/6, and EGFR. Ongoing in vitro studies will define the role of FOXM1 in Schwann cell transformation while analyses of novel Nf1^+/-, Foxm1 deficient mice will reveal the requirement of FOXM1 in NF1-associated MPNST development in vivo. To date, our evidence indicates that FOXM1 inhibition in specific combination therapies could be a new, effective strategy for treating MPNST patients.

{{THERAPEUTICS}}

Authors: Emily J Carlson, PhD; Rebecca Levitt, BA; Jarrod Sotos, PhD; Meredith J Goyette, MA; Christina Thomas, PhD; Caitlyn J Cap, PsyD; Megan Knauss, MSW, LGSW; Karin S Walsh, PsyD (all authors are affiliated with Children’s National Hospital)

Funding: Lambert Family Foundation, Gilbert Family Foundation

Disclosures: None

Purpose: Neurofibromatosis Type 1 (NF1) is associated with cognitive challenges and sleep concerns. A recent NF1 caregiver survey conducted by this group identified high levels of sleep disturbance (e.g. difficulty falling asleep) and impairment (e.g., problems caused by disrupted sleep) among children with NF1, with greater sleep concerns endorsed for children with comorbid attention-deficit/hyperactivity disorder. This study builds on these findings by examining the relationship between sleep difficulties and cognitive functioning in children with NF1 using performance-based cognitive tests and norm-based parent questionnaires. 

Methods: Thirty-two children with NF1 (ages 6-16) completed a cognitive battery evaluating attention, executive functioning (EF), and learning and memory. Measures included working memory subtests from the Wechsler Intelligence Scales, the California Verbal Learning Test (CVLT), and subtests from the NEPSY, second edition (NEPSY-II). Caregiver ratings of attention, hyperactivity, and EF were obtained using the Behavior Rating Inventory of Executive Function, Second Edition (BRIEF-2) and the Behavior Assessment System for Children, Third Edition (BASC-3). Sleep disturbance and sleep impairment were measured using parent-proxy PROMIS measures. Additional sleep factors were assessed using the Child and Adolescent Sleep Checklist (CASC). Data were analyzed using Pearson/Spearman correlations and group comparisons (t-tests and Mann-Whitney U tests). 

Results: Participants (M_age=10.28, SD=2.85) reported an average of 8.78 hours of sleep on weeknights (SD=1.5); 41% (n=13) had sleep disturbance and 38% (n=12) had sleep impairment. On average, participants with sleep disturbance and impairment had 1-1.5 less hours of sleep/night than those without. Sleep disturbance was negatively correlated with attention, EF, and both verbal and visual learning and memory performance. Sleep impairment was negatively correlated with all of these factors except for verbal learning/memory. Sleep disturbance and impairment were associated with caregiver expressed attention and EF concerns. Group comparisons revealed that participants with sleep disturbance and impairment exhibited more pronounced attention, EF, and visual learning problems than those without. Additionally, individuals with sleep impairment specifically demonstrated greater memory difficulties.

Conclusion: This study provides further evidence for the presence of sleep challenges in NF1. While children with NF1 are generally at risk for cognitive challenges, current results indicate that those with comorbid sleep problems are at even greater risk for problems with attention, EF, and learning and memory. Clinicians should monitor NF1 patients for sleep problems and be prepared to provide recommendations regarding sleep hygiene and interventions. Researchers investigating treatments aimed at improving sleep among NF1 patients should examine potential cognitive improvements associated with better sleep.

{{Cognition / Behavioral / Learning}}

Armstrong_Amy_Lack

Objective: Optimal management of malignant peripheral nerve sheath tumor (MPNST) relies on early diagnosis of malignant transformation in predisposing plexiform neurofibromas (PN) in patients with Neurofibromatosis Type 1 (NF-1).  Intratumor heterogeneity is a known occurrence in PNs where malignant transformation to MPNST can occur in part, but not all, of the underlying PN.  Efforts have been made to standardize imaging guidelines for lesions suspicious for MPNST and FDG-PET/CT is routinely performed.  Higher standardized uptake value (SUV) raises possibility for malignancy with studies using a SUV maximum (SUVmax) cut-off of 3.2-4.5, however no ideal value has been substantiated.  We assessed correlation between SUVmax and malignant pathology in NF1 patients with PN.  

Methods: We evaluated patients with NF1 enrolled on the Tissue and Blood Acquisition for Genomic Analysis and Collection of Health Information for Patients with Bone and Soft Tissue Sarcoma: The Sarcoma Oncology Group at Washington University in St. Louis who underwent FDG-PET/CT imaging.   Those patients who had subsequent biopsy or resection due to suspected malignant transformation based on FDG-PET/CT findings were reviewed.  

Results: We identified eight patients with FDG-PET/CT imaging showing a lack of correlation between high SUVmax and malignant pathology.  Median age of this group was 22 years (range 11-59 years).  Lesions with SUVmax range of 6.1-13.3 were determined to be either neurofibroma or atypical neurofibromatous neoplasm with uncertain biologic potential (ANNUBP), but not MPNST.  One patient had two lesions evaluated due to findings on FDG-PET/CT with discordant correlation between SUVmax and pathology (see below).  Furthermore, one patient’s initial biopsy of a mass with SUVmax of 7.7 revealed neurofibroma, but subsequent resection identified ANNUBP.   

Conclusions: Sampling of suspicious lesions in NF1-related PN is critical as wide resection of non-metastatic MPNST can be curative.  FDG-PET/CT is a non-invasive tool to assess for malignant transformation in patients; however, we caution that higher SUVmax can be seen in benign neurofibromas and pre-malignant ANNUBPs, the latter which does carry a risk of transformation to MPNST.  Additionally, SUVmax did not correlate with malignant pathology in the same patient.   We suggest using SUVmax to direct biopsy or resection while understanding the limitations of this modality.   In the future, we hope other non-invasive testing such as detecting circulating tumor DNA or utilization of more sensitive radiotracers can aid in diagnostic work-up for this high-risk population and studies are underway.

Figure 1. Lack of Association of SUV maximum and Malignant Pathology in Patient with Neurofibromatosis Type 1

Atypical neurofibromatous neoplasm with uncertain biologic potential (ANNUBP); Malignant peripheral nerve sheath tumor (MPNST)

{{Plexiform neurofibroma}}

Presenting Author: Alexis Murawski Stillwell^1.2

Additional Authors: Laura Lambert³, Kelley Bradley¹, Harunur Rashid¹, Hu Ke², Erik Westin¹, Min Chen², Amjad Javed², CJ Haycraft², Kai Jiao⁴, Peter Panizzi⁵, Bruce R Korf², Maria Johnson², Jeremy Foote², Deeann Wallis², Robert A Kesterson¹

Affiliations: 1. Pennington Biomedical Research Center 2. University of Alabama at Birmingham 3. Mayo Clinic 4. Augusta University 5. Auburn University

Funding: Children’s Tumor Foundation Young Investigator Award, The Giorgio Children’s Foundation for NF1, UAB Center for Precision Animal Modeling, Pennington Biomedical Research Center

Background: Animal models are critical for understanding disease and testing new therapeutics. Patients with the p.M992del mutation display a relatively mild clinical phenotype associated with café-au-lait spots but no neurofibromas, with a subgroup of patients displaying Noonan-like features. Therefore, we hypothesize that mutant p.M992del neurofibromin is hypomorphic and retains sufficient functional activity to suppress tumor formation, but not sufficient activity in all NF1 related functions.

Methods: Heterozygous mice (Nf1^M992del/+ on FVB background) were created using CRISPR/Cas9 and a repair template to introduce a 3 bp deletion in the homologous region of mouse Nf1 gene. We intercrossed heterozygous mice (Nf1^M992del/+) to assess viability of homozygous (Nf1^{M992del/M992del}) mutants, as most Nf1 mutant alleles induce early embryonic lethality when homozygous. Nf1^{M992del/M992del} animals and control littermates were assessed for growth parameters, body composition, skeletal structure (histology and X-Ray), and bone density (uCT). Histological analysis of growth plates and cranial sutures was also performed. Embryos from intercrossed heterozygous mice (Nf1^M992del/+) were analyzed between E17.5 and P0.

Results: Analysis of genotype ratios revealed perinatal lethality of Nf1^{M992del/M992del} pups (observed 101 pups born compared to 262.75 expected pups). Surviving Nf1^{M992del/M992del} mice fail to thrive and are ~50% of the size by weight of littermate controls; only 61 of the 101 observed pups survived to weaning. Mice display impaired bone organization of the sternum and craniosynostosis. Mice surviving to 6 months also display bilateral suppurative otitis media (n=2/4) and pneumonitis (n=3/4). E16.5 Nf1^{M992del/M992del} pups display double outlet right ventricle phenotype, and 6-month-old mice display mitral valve defects. Preliminary Western blots confirm the presence of mutant neurofibromin protein at levels consistent with control animals, and a significant change in pERK levels.

Conclusions: Nf1^M992del/+ mice appear to be healthy with no overt phenotype, whereas Nf1^{M992del/M992del} animals display perinatal lethality, with surviving animals severely runted. Further characterization of the skeletal and heart phenotype is ongoing. Preliminary data from backcrossing the p.M992del mutation from FVB/NJ mice to C57BL/6J strain indicates genetic background plays a role in embryonic lethality as no viable homozygous mice have been identified on the C57BL/6J background.

{{DRUG DISCOVERY AND DEVELOPMENT}}

Presenting author: Fertitta Laura, MD

Dept. of Dermatology, National Referral Center for Neurofibromatoses, Henri Mondor Hospital, Assistance Publique – Hôpitaux de Paris (AP-HP), 94010 Créteil, France.

No standardized and validated clinician-reported outcome measure exists to assess cutaneous neurofibromas (cNF) in neurofibromatosis type 1 (NF1). An international initiative recently established a core outcome set for NF1-related cNF trials, including number, size/volume, and visibility. 

The aim: To develop and validate the Nef-ASI tool designed to assess the overall severity of cNF, integrating these three outcomes.

This CERENEF study at Henri Mondor Hospital, Créteil, France followed six steps. Steps 1-2 involved surveys and working groups (N=25 healthcare professionals, HCP) to integrate cNF size, number/density, and visibility into a single score. Steps 3-6 included scoring sessions with local team members (N=9HCP, grouped by experience) and MDs (N=15) from the French rare skin disease network. Standardized 2D photographs (100cm² skin area) were used. Reliability was assessed via intraclass correlation coefficient (ICC >0.8 acceptable), assessor agreement by Bland-Altman plot, and validity by correlation with cNF severity questions.

Development Phase: Steps 1-2 defined cNF size categories (small <5mm, medium 5-9mm, large ≥10mm) with visibility-weighting factors (1, 1.5, 3) and density ranges (% skin surface: 0%, 1%, 5%, 10%, 25%, 50%, ≥75%), which were into three 21-image palettes for different skin phototypes (Fig 1). Validation Phase: Scoring sessions evaluated reliability, agreement, and feasibility: 

• Step 3: Inter-rater reliability (9 assessors, 50 photos) showed ICCs >0.8 for MDs and non-MD experts (Table 1). Bland-Altman plots indicated increased data spread in high-score cases (Fig 2).
• Step 4: Intra-rater reliability (25 rescored photos) remained high (ICCs >0.9).
• Step 5: Fifteen MDs scored 50 photos, with ICCs >0.8. NeF-ASI correlated strongly (0.81-0.88) with cNF severity questions.
• Step 6: Feasibility: 87% found NeF-ASI easy to use, with a median evaluation time of 27 seconds per image.

The NeF-ASI demonstrated strong inter- and intra-rater reliability, comparable to other dermatological CROMs like PASI. Its strong correlation with clinical scales confirms its validity. It assesses: i) topical treatments by comparing two 100cm² photos (treated vs. placebo) per patient (Target NeF-ASI) and ii) systemic treatments using three 100cm² photos (back, abdomen, thigh, as defined by Cunha et al.) to calculate a Global NeF-ASI (average of three scores) across treatment groups. Although developed in France, international experts contributed, ensuring broader applicability. 

Thus, NeF-ASI offers a cost-effective, standardized and reliable cNF severity assessment for therapeutic trials, integrating their size, number, and visibility. Its validated 2D-photograph approach enables remote assessments, supporting decentralized trial designs.

Full author list: Laura Fertitta MD presenting author, Arnaud Jannic MD, Christina Bergqvist MD, Vanessa Delique, Marie-Laure Armand, Sabine Moryousef, Salah Ferkal MD, Ouidad Zehou  MD, Houda Ayache, Philippe Le Corvoisier, Etienne Audureau, Bérénice Hebrard, Ariane Lunati-Rozie, Benoit Funalot, Kavita Y. Sarin MD, PhD, Jaishri O. Blakeley MD, Carlos G. Romo MD, Yemima Berman MD, Lu Q Le MD, PhD, Khaled Ezzedine  MD, PhD, Pierre Wolkenstein MD, PhD, and FIMARAD group

Funding: L. Fertitta is supported by the Francis S. Collins Scholars Program in Neurofibromatosis Clinical and Translational Research funded by the Neurofibromatosis Therapeutic Acceleration Program (Grant # 230115)

Figure 1: Cutaneous neurofibroma palettes constituting the Nef-ASI, corresponding to the 3 phototype groups based on the Fitzpatrick classification (1-2, 3-4, and 5-6).

Figure 2: Bland and Altman plots measuring the level of agreement between 2 assessors randomly selected in each group of 3: medical doctor (MD) experts, non-MD experts and non-experts.

Table 1: Inter and intra-rater reliability assessment of the Nef-ASI using the Intraclass Correlation Coefficient (ICC), and their 95% confidence interval (95%CI). Session 1 corresponded to the results from Step 3, and Session 2 to those from Step 4, highlighting an improvement in the tool's reproducibility with the assessors' training. ICCs range from 0 to 1: <0.5 poor, 0.5–0.74 moderate, 0.75–0.89 good and 0.9–1.0 considered excellent reliability (Koo TK, et al. J Chiropr Med. 2016).* indicates the medical doctor (MD) experts, † the non-MD experts, and ‡ the non-experts.

{{Cutaneous and subcutaneous neurofibroma}}

Authors: Takahiro Tsuchiya¹, MD; Kenta Ohara¹, MD, PhD; Satoru Miyawaki¹, MD, PhD;  Yu Teranishi¹, MD, PhD; Hiroki Hongo¹, MD, PhD; Yu Sakai¹, MD, PhD; Shogo Dofuku¹, MD, PhD; Daiichiro Ishigami¹, MD, PhD; Atsushi Okano¹, MD, PhD; Hirokazu Takami¹, MD, PhD; Masahiro Shin^{1, 2}, MD, PhD; Hirofumi Nakatomi^{1, 3}, MD, PhD; Nobuhito Saito¹ MD, PhD

¹Department of Neurosurgery, Faculty of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, Japan

²Department of Neurosurgery, Teikyo University Hospital, 2-11-1 Kaga, Itabashi-Ku, Tokyo, Japan

³Department of Neurosurgery, Faculty of Medicine, Kyorin University, 6-20-2 Shinkawa, Mitaka-shi, Tokyo, Japan

Funding: This work was supported by grants from the Japan Society for the Promotion of Science KAKENHI (grants no. 21H03041 to Dr. Saito and 23H03018 to Dr. Miyawaki, and 21J11426 to Dr. Ohara). This study was also supported by grants from the Takeda Science Foundation to Dr. Miyawaki.

Disclosures: None.

Purpose: Schwannomas are benign neoplasms arising from myelinating Schwann cells of the nerve sheath. The majority of the intracranial schwannomas are vestibular schwannomas (VS). Among non-vestibular intracranial schwannomas (non-VS), trigeminal and jugular foramen schwannomas are predominantly observed. While the loss of NF2 function plays a significant role in sporadic schwannoma tumorigenesis, a recent, large-scale study revealed the involvement of additional recurrent gene mutations, in addition to the SH3PXD2A::HTRA1 fusion gene, in sporadic schwannomas. However, the genetic landscape of non-VS remains unclear. In this study, we conducted a comprehensive genetic analysis of VS and non-VS.

Methods: Targeted panel sequencing and microsatellite analysis of 22q were performed in 51 patients with sporadic intracranial schwannomas (21 patients with VS and 30 with non-VS). The proportion of known schwannoma variants was assessed depending on the tumor origin site. Additionally, the NF2 alterations were examined to determine whether they differed between VS and non-VS.

Results: The median tumor size of non-VS was significantly larger than that of VS (38 mm, 27 mm, p < 0.01). In contrast, there were no significant differences between the VS and non-VS in median age at surgery (41.5 years vs 45.2 years, p = 0.37), female gender (42.8% vs 50.0%, p = 0.83), cystic lesions (71.4%, 73.3%, p > 0.99), postoperative stereotactic radiosurgery (14.3% vs 13.3%, p > 0.999), or regrowth after surgery (9.5% vs 16.7%, p = 0.75). Somatic NF2 mutations were frequently identified in tumor samples (25 patients, 49.0%); non-VS showed a significantly lower frequency of NF2 mutations than VS (80.9% vs 26.7%, odds ratio: 11.0 [95% confidence interval: 2.59–59.5], p < 0.001). Despite the absence of statistical significance in the frequency of 22q loss of heterozygosity (LOH) between VS and non-VS, any NF2 alterations (NF2 mutation or 22q LOH) exhibited a significant difference (95.2% vs 56.7%, p < 0.01). Truncating mutations (frameshift, nonsense, and splice-site mutations) were the most common type of mutation (22 mutations, 75.9%). The variant allele frequency of NF2 was found to be high, indicating its role as a significant contributor to tumorigenesis.

Conclusion: Our study has shown that NF2 alterations were significantly predominant in VS, and the potential involvement of factors other than NF2 inactivation in tumorigenesis, especially in non-VS. Further comprehensive molecular analyses are necessary to elucidate the mechanisms underlying these observations.

{{Schwannoma}}

INTRODUCTION: NF1 is an autosomal dominant genetic disorder with expression variability, it is due to mutation in the neurofibromin gene. It can be associated with epilepsy, neurodevelopmental disorders and cerebrovascular disease.

OBJECTIVES: To describe the most prevalent neurological manifestations in a series of pediatric patients with NF1 seen at Hospital Garrahan in Argentina in the last 10 years.

POPULATION AND METHODS: We performed a descriptive, retrospective analysis of the medical records of patients with clinical criteria for NF1 with an age range of zero to 15 years, evaluated by neurology at Hospital Garrahan in the last 10 years. 

RESULTS: We registered 203 patients, 99 females with a mean age of 3 years. 133 patients reported neurodevelopmental disorders with learning difficulties in 54%, psychopedagogy was the most indicated therapy (85%). 22 (11.3%) presented focal and generalized epilepsy not always associated with structural lesions, 12 of them (54%) with pathological electroencephalogram. The most indicated drug was clobazam. 83 patients 22 (11.3%) presented focal and generalized epilepsy not always associated with structural lesions, 12 of them (54%) with pathological electroencephalogram. The most indicated drug was clobazam.83 patients (50%) had hamartomatous lesions in the cerebellum, 2 patients reported Moya Moya syndrome.

CONCLUSION: Our study shows that neurodevelopmental disorders are frequent in this population, among them the most diagnosed was learning disorder. Epilepsy is more common than in the general population, and focal seizures are more frequent, requiring prolonged antiepileptic drugs but with good control. In agreement with previous series, bright lesions in cerebellum were frequently observed.

{{Cognition / Behavioral / Learning}}

Authors: Haydee Lara PhD*, Dhirender Ahlawat MS*

*Alexion-AstraZeneca Rare Disease Unit, Translational Sciences

Funding: NF1 Global Project Team

Disclosures: H. Lara and D. Ahlawat are AstraZeneca employees

Background & Purpose: 

The samples analyzed in this proteomics study correspond to clinical study NCT05101148 “Phase I Study to Assess the Effect of Food on the PK and Gastrointestinal Tolerability of Selumetinib in Adolescent children With Neurofibromatosis Type 1 Related Plexiform Neurofibromas”. The clinical study NCT05101148 was designed to evaluate the effect of a low-fat meal on steady state selumetinib exposure. Selumetinib was approved in the US in 2020 and in Europe in 2021; it was approved for the treatment of NF1 pediatric patients with symptomatic and inoperable PN and until recently, the drug had to be taken in a fasted state. The results of this clinical study supported the update of the label in 2024, and selumetinib can now be taken with or without food. In the proteomic analysis discussed here, we investigated the effect of selumetinib in disease-relevant pathways in this NF1 patient population, especially in the patients with skeletal manifestations. 

Methods:

We evaluated plasma samples corresponding to twenty-three patients from study NCT05101148 using aptamer-based proteomics technology. We compared protein fold changes in pre-dose plasma samples corresponding to one cycle vs. two cycles of selumetinib treatment. Similarly, we studied changes in the proteome of patients with NF1-related skeletal manifestations in the same population. Pathway enrichment analysis was also performed to assess pathways involved with the significantly dysregulated proteins. 

Results:

We found that CCL5 (RANTES) was upregulated in NF1 patient samples corresponding to cycle two of selumetinib treatment. Proteins associated with tumor growth, such as hepcidin and HMG Co-A synthase, were downregulated. When we analyzed the proteome of NF1 patients with skeletal manifestations, we found that proteins in the collagen family are upregulated. Although the role of these dysregulated collagen proteins in NF1 is unknown, other researchers have reported their relationship with other bone diseases. 

Conclusion: 

This proteomics study in NF1 patient samples treated with selumetinib demonstrates changes in proteins previously reported for this population, such as CCL5 (RANTES). The analysis also uncovered a potential link of collagen proteins dysregulation in NF1 skeletal manifestations with other bone diseases. Further validation of these findings is warranted. 

{{TUMOR BIOLOGY AND DISEASE MECHANISM}}

Authors: Emilie Everard MD^1,2, Emma De Haan², Lucie Coté¹, Ahmed Zaki El Haffaf MD³, Moujahed Labidi MD⁴, Mark Keezer MD, PhD², Philippe Major MD¹, Sarah Lapointe MD², Sébastien Perreault MD¹

1. Division of Pediatric Neurology, Department of Neurosciences, CHU Sainte-Justine, Montréal, QC, Canada