Neurofibromatosis type 1: a multidisciplinary approach to care

Introduction Neurofibromatosis type 1 (NF1) is one of the most common hereditary tumor syndromes. The average incidence of NF1 in the world is 1:3000 of the population. The characteristic signs of the disease are neurofibromas and café-au-lait macules on the skin. 60 % of patients with NF1 develop specific skeletal anomalies: scoliosis, chest deformity, pseudarthrosis, requiring surgical treatment and long-term rehabilitation. It is necessary to develop prognostic criteria for the development of severe skeletal anomalies in NF1 and take early measures to prevent their progression. Congenital pseudarthrosis of the tibia is diagnosed in 5 % of children with NF1, accounting for 80 % of all cases of this pathology in the general population. Spinal scoliosis is detected in 60 %, osteoporosis in 50 %, chest deformity in 37.6 %, microgenia in 53 %, increased head circumference in 25 %, sphenoid wing dysplasia in 12 %, facial asymmetry in 10 % of patients with NF1. The aim of the review is to focus on the pathogenesis of skeletal anomalies development in NF1 that result in disorders of the musculoskeletal system in NF1 in order to take early measures for the prevention and treatment of the disease. Materials and method The review is based on numerous studies found in the databases: PubMed, Scopus, Web of Science, published mainly over the past 5 years. The suitable studies were searched by keywords and their combinations «neurofibromatosis type 1» with the words «skeletal abnormalities», «musculoskeletal system», «pseudarthrosis», «scoliosis», «pathogenesis», «deformation», «treatment», «frequency», «prevalence», «genotype-phenotype correlation», « modifier genes». Results and discussion The pathogenesis of skeletal anomalies is due to both the loss of heterozygosity of the NF1 gene in pseudoarthrosis and the effect of neurofibromin deficiency on the development of connective tissue. Currently, the only effective drugs for the treatment of tumor syndrome in NF1 are inhibitors of mitogen-activated kinase (MEK), which suppress the increased activity of Ras oncogenes. A promising issue is the study of the effect of MEK inhibitors on the progression of skeletal anomalies in patients with NF1 in the treatment of tumor syndrome. Therefore, dynamic observation by an orthopedic surgeon with an objective assessment of the observed changes is of great importance in the management of patients. It is necessary to widely introduce molecular genetics methods for confirming the diagnosis of NF1 in the clinic in cases of a combination of skeletal anomalies with individual signs of the disease, since the manifestations of NF1 are steadily progressing with age, even in the presence of erased and atypical forms of the disease. Since the analysis of scientific literature has shown the possible influence of modifier genes on the pathogenesis of NF1, the search for mutations in these genes is promising. Conclusion Most patients with NF1 develop orthopedic pathology, which is associated with the role of the NF1 gene in the development of connective tissue. The increased mutability of this gene causes the loss of heterozygosity in the development of congenital pseudoarthrosis of the tibia. At the same time, NF1 driver mutations are detected in 10 % of sporadic malignant neoplasms. Therefore, the role of somatic mutations in the NF1 gene in the development of skeletal anomalies in the general population is probable. The methods of NF1 therapy that are under investigation may become the basis for the complex treatment of oncological and orthopedic patients.

Understanding the biological pathways critical for common neurofibromatosis type 1 (NF1) peripheral nerve tumours is essential, as there is a lack of tumour biomarkers, prognostic factors and therapeutics. We used gene expression profiling to define transcriptional changes between primary normal Schwann cells (n = 10), NF1-derived primary benign neurofibroma Schwann cells (NFSCs) (n = 22), malignant peripheral nerve sheath tumour (MPNST) cell lines (n = 13), benign neurofibromas (NF) (n = 26) and MPNST (n = 6). Dermal and plexiform NFs were indistinguishable. A prominent theme in the analysis was aberrant differentiation. NFs repressed gene programs normally active in Schwann cell precursors and immature Schwann cells. MPNST signatures strongly differed; genes up-regulated in sarcomas were significantly enriched for genes activated in neural crest cells. We validated the differential expression of 82 genes including the neural crest transcription factor SOX9 and SOX9 predicted targets. SOX9 immunoreactivity was robust in NF and MPSNT tissue sections and targeting SOX9 – strongly expressed in NF1-related tumours – caused MPNST cell death. SOX9 is a biomarker of NF and MPNST, and possibly a therapeutic target in NF1.

Neurofibromatosis type 1 (NF1) is a genetic disorder with a wide range of manifestations and severity. Currently, the few available NF1 treatments target specific manifestations, with no available therapies targeted to correct the underlying driver of all NF1 manifestations. Evidence supports that haploinsufficiency in NF1 caused by a decreased amount of wild-type (WT) neurofibromin in all Nf1+/- cells directly causes or facilitates a range of NF1 manifestations. Consequently, NF1 haploinsufficiency correction therapy (NF1-HCT) represents a potentially effective approach to treat some NF1 manifestations. NF1-HCT would normalize the level of WT neurofibromin in all NF1-haploinsufficient cells, including those integral to the NF1 phenotype such as Schwann cells (SCs), melanocytes, neurons, bone cells, and cells of the tumor microenvironment. This would correct altered cellular signaling pathways and, in turn, restore normal function to cells with a retained WT allele. NF1-HCT will not restore WT neurofibromin in NF1-/- cells; however, by restoring function in the surrounding Nf1+/- microenvironment cells, NF1-HCT is predicted to have a beneficial effect on NF1-/- cells. NF1-HCT is expected to have a clinical effect in some NF1 manifestations, as follows: (i) prevention, or delay of onset, of potential manifestations; and (ii) reversal, or halting/slowing progression, of established manifestations. This review describes the rationale for NF1-HCT, including specific NF1 considerations (e.g., NF1 clinical phenotype, neurofibromin function/regulation, NF1 mutational spectrum, genotype-phenotype correlation, and the impact of haploinsufficiency in NF1), HCT in other haploinsufficient diseases, potential NF1-HCT drug treatment strategies, and the potential advantages/challenges of NF1-HCT.

1. Mustafa Tekin, MD* 2. Joann N. Bodurtha, MD, MPH† 3. Vincent M. Riccardi, MD‡ 1. 2. *Clinical Genetics Fellow. 3. 4. †Associate Professor of Human Genetics, Pediatrics, Obstetrics and Gynecology, Virginia Commonwealth University/Medical College of Virginia Hospitals, Richmond, VA. 5. ‡President, The Neurofibromatosis Institute, La Crescenta, CA. Objectives After completing this article, readers should be able to: 1. Define cafe au lait spots typical of neurofibromatosis type 1 (NF1) and describe their frequency and variability in the normal population. 2. List three or more genetic disorders other than NF1 that are associated with cafe au lait spots. 3. Summarize three or more clinical manifestations and molecular bases of NF1 and NF2. 4. List the diagnostic criteria for NF1. 5. Summarize clinical findings of genetic disorders other than NF1 associated with cafe au lait spots. Every pediatrician faces the challenge of deciding if a patient who has cafe au lait (CAL) spots has an underlying genetic condition. CAL spots typical of neurofibromatosis type 1 (NF1) are discrete, round or oval, uniformly hyperpigmented skin patches. Their color varies from light to dark brown, and the border may be smooth or irregular. They usually are smaller in newborns, enlarge as children get older, and are less prominent in adults. The histologic basis of CAL spots is increased melanin content, with the presence of giant melanosomes in both melanocytes and basal keratinocytes and no melanocytic proliferation. The giant melanosomes in CAL spots are not unique to NF1; they can be seen in unaffected skin of adults who have NF1 and occasionally in normal skin of healthy individuals. Therefore, the presence of giant melanosomes is not helpful for diagnosing NF1. The frequency and number of CAL spots vary in the general population according to ethnic background and age. Sometimes otherwise healthy children who have red hair and often are of Irish or Welsh background have multiple areas of patchy hyperpigmentation. Similarly, multiple patchy areas of hyperpigmentation can occur in healthy children who have mixed ethnic backgrounds in which two parents have very different skin colors. CAL spots were noted in 0.3% of Caucasians and 18% of African-Americans …

Neurofibromatosis type 1 (NF1) is a rare genetic disorder, characterized by the development of benign and malignant nerve tumors. Although all individuals with NF1 harbor genetic alterations in the same gene, the clinical manifestations of NF1 are extremely heterogeneous even among individuals who carry identical genetic defects. In order to deepen the understanding of phenotypic manifestations in NF1, we comprehensively characterized the prevalence of 18 phenotypic traits in 2051 adults with NF1 from the Children’s Tumor Foundation’s NF1 registry. We further investigated the coassociation of traits and found positive correlations between spinal neurofibromas and pain, spinal neurofibromas and scoliosis, spinal neurofibromas and optic gliomas, and optic gliomas and sphenoid wing dysplasia. Furthermore, with increasing numbers of cutaneous neurofibromas, the odds ratio of malignant peripheral nerve sheath tumor increased. Phenotypic clustering revealed 6 phenotypic patient cluster subtypes: mild, freckling predominant, neurofibroma predominant, skeletal predominant, late-onset neural severe, and early-onset neural severe, highlighting potential phenotypic subtypes within NF1. Together, our results support potential shared molecular pathogenesis for certain clinical manifestations and illustrate the utility of disease registries for understanding rare diseases.

To characterize the adaptive behavior profile of children with neurofibromatosis type 1 (NF1) and determine its relationship to neuropsychological functioning and non-neoplastic T2-weighted hyperintense brain lesions on brain magnetic resonance imaging (MRI). In this cross-sectional study, we retrospectively reviewed neuropsychological reports from 104 children with NF1 (56 males, 48 females; mean age 10y 4mo; standard deviation [SD] 3y 4mo; range 3y 5mo-17y 6mo), and extracted data from a range of cognitive and behavioral measures, including the Adaptive Behavior Assessment System (ABAS). Brain MRI was retrospectively reviewed in 42 individuals. Adaptive Behavior Assessment System scores were continuously distributed and pathologically shifted by 0.79 to 1.26SD across Conceptual, Social, and Practical domains, and 46.5% of individuals had a composite score in the borderline or impaired range. Impairment in adaptive functioning was correlated with deficits in executive function (r=-9.543, p<0.001), externalizing problems (r=-0.366, p<0.001), and attention (r=-9.467, p=0.001). Cluster analysis revealed three distinct phenotypic subgroups, one of which exhibited normal cognitive ability, but impaired adaptive functioning, with persistent deficits in executive function, behavioral problems, and attention-deficit/hyperactivity disorder symptomatology. There was no relationship between ABAS scores and the number or location of unidentified bright objects. Adaptive functioning deficits are common among children with NF1 and are associated with impairment in other cognitive/behavioral domains, independent of general cognitive ability. Deficits in adaptive behavior are common in children with neurofibromatosis type 1 (NF1). Poor adaptive functioning is associated with impairments in executive function, externalizing behaviors, and attention, regardless of cognitive ability. The presence or location of unidentified bright objects do not predict adaptive behavior skills in children with NF1.

Novel Mouse Model Developed to Show Transformative Link, Results Validated in Human TumorsResearchers at UCLA's Jonsson Comprehensive Cancer Center showed for the first time that the loss or decreased expression of the tumor suppressor gene PTEN plays a central role in the malignant transformation of benign nerve tumors called neurofibromas into a malignant and extremely deadly form of sarcoma.The work, a collaboration between the Institute for Molecular Medicine, the molecular and medical pharmacology department and the cancer center's Sarcoma Program, could lead to the development of new therapies that target the cell signaling pathway regulated by PTEN. A novel mouse model of neurofibromatosis type 1 (NF1) developed at UCLA first illustrated the importance of PTEN tumor suppressor in malignant transformation and this finding was validated in human malignant peripheral nerve sheath tumors (MPNST), the deadly sarcomas.The study is being published in the early online edition of the peer-reviewed journal Proceedings of the National Academy of Sciences."The loss of expression of PTEN in the human sarcomas we studied mirrored the loss of PTEN in mice, and we anticipate being able to target this pathway abnormality for the development of new methods of diagnosis and treatment" said Dr. Fritz Eilber, director of the Sarcoma Program and an assistant professor of surgical oncology. "Within the sarcoma world, malignant peripheral nerve sheath tumors are one of the most lethal sub-types, so this is a significant finding and may lead to new and more effective treatments."NF1 is one of the most common genetically inherited disorders, with an incidence of about 1 in every 2,500 births, said, Dr. Hong Wu, associate director of the molecular medicine institute, a Jonsson Cancer Center researcher and senior author of the study."Patients with NF1 have an about 10 percent lifetime risk of developing this lethal sarcoma sub-type," Wu said.The study also showed that Positron Emission Tomography (PET) scanning with the glucose analogue FDG - both in the mice and in humans - is a highly accurate way to distinguish between the benign tumors and the malignant ones, indicating that this non-invasive imaging technology is valuable in assessing therapeutic response to new treatments.Wu created the mouse model with two of her graduate students, Caroline Gregorian and Jonathan Nakashima, co-first authors of this paper. It was created by altering two cell signaling pathways that are commonly activated in peripheral and central nervous system cancers, the RAS/RAF/MAPK & PTEN/P13K/AKT pathways, known to regulate cell proliferation, survival and differentiation."When we began to generate mouse models to mimic different human cancers, we usually did gene-based analysis to see the relevance of a specific gene in the development of the cancer," Wu said. "But we realize that sometimes targeting the cell signaling pathways that organize and instruct cells to function, both for normal functions of our body and also in abnormal ways in disease, are more important and informative than the individual gene"The mouse model developed benign neurofibromas, but then progressed to the deadly sub-type of sarcoma. The neurofibromas had half the normal levels of PTEN and the sarcomas had a complete loss of PTEN. Since PTEN is an important factor in suppressing cells from becoming malignant, this could provide an explanation for the sequence of the normal cells transforming into benign neurofibromas that could then transform into cancer.Wondering if this was also the case in people, Dr. Wu collaborated with Eilber and pathologist Dr. Sarah Dry, director of the Institute of Molecular Medicine's Pathway Pathology Center, and a multidisciplinary team of physician-scientists to determine if people with this sarcoma sub-type also had little or no PTEN."This type of collaboration is the hallmark of the work at the Jonsson Cancer Center and molecular medicine institute - translating discoveries in a basic science lab into discoveries in patients," Wu said.Currently, there are no effective treatments to prevent the benign NF1 tumors from transforming into cancer. The genetically engineered mouse model will be used to screen drugs that may be able to target the signaling pathway regulated by PTEN, to block signals that instruct the cells to change from a benign state to a malignant one, providing treatment options for patients with the deadly form of cancer."I think these findings will help us provide a better diagnosis that can determine if the neurofibroma is becoming a malignant tumor or not," Eilber said. "But more importantly, the loss of the PTEN and its associated signaling pathways gives us targets for therapy and it may lay the foundation for treatment in other sarcomas as well."Also involved in the research were Dr. Paul Mischel, Dr. Simin Liu, Dr. Phioanh Leia Nghiemphu, Dr. Greg Lawson, Dr. Michael Sofroniew and Dr. Michael Phelps, director of the molecular medicine institute and creator of the PET scannerThe study was funded by the United States Department of Health and Human Services, the National Cancer Institute, the National Institutes of Health, UCLA's Jonsson Comprehensive Cancer Center, the American Cancer Society, the Brain Tumor Society, the Henry Singleton Brain Cancer Research Program and the James S. McDonnell Foundation.UCLA's Jonsson Comprehensive Cancer Center has more than 240 researchers and clinicians engaged in disease research, prevention, detection, control, treatment and education. One of the nation's largest comprehensive cancer centers, the Jonsson center is dedicated to promoting research and translating basic science into leading-edge clinical studies. In July 2009, the Jonsson Cancer Center was named among the top 12 cancer centers nationwide by U.S. News & World Report, a ranking it has held for 10 consecutive years. For more information on the Jonsson Cancer Center, visit our website at http://www.cancer.ucla.edu.Kim IrwinUCLA's Jonsson Comprehensive Cancer Center(310) 206-2805

Simple SummaryNeurofibromatosis type 1 (NF1) is a genetic disorder caused by pathogenic variants in the NF1 tumor suppressor gene. In 5–10% of NF1 patients, a large heterozygous deletion of the whole NF1 gene is identified, leading to the commonly called “NF1 microdeletion syndrome”. NF1-deleted patients were previously reported to develop a particularly severe form of the disease with frequent cognitive impairment and an increased risk of benign and malignant tumors. Here, we performed a comprehensive clinical assessment of the largest NF1-deleted cohort to date, including 126 NF1 patients with a deletion of the NF1 gene. This work provides new insights into the specific phenotype associated with NF1 deletions and may contribute to improve the follow-up care of NF1 patients. Complete deletion of the NF1 gene is identified in 5–10% of patients with neurofibromatosis type 1 (NF1). Several studies have previously described particularly severe forms of the disease in NF1 patients with deletion of the NF1 locus, but comprehensive descriptions of large cohorts are still missing to fully characterize this contiguous gene syndrome. NF1-deleted patients were enrolled and phenotypically characterized with a standardized questionnaire between 2005 and 2020 from a large French NF1 cohort. Statistical analyses for main NF1-associated symptoms were performed versus an NF1 reference population. A deletion of the NF1 gene was detected in 4% (139/3479) of molecularly confirmed NF1 index cases. The median age of the group at clinical investigations was 21 years old. A comprehensive clinical assessment showed that 93% (116/126) of NF1-deleted patients fulfilled the NIH criteria for NF1. More than half had café-au-lait spots, skinfold freckling, Lisch nodules, neurofibromas, neurological abnormalities, and cognitive impairment or learning disabilities. Comparison with previously described “classic” NF1 cohorts showed a significantly higher proportion of symptomatic spinal neurofibromas, dysmorphism, learning disabilities, malignancies, and skeletal and cardiovascular abnormalities in the NF1-deleted group. We described the largest NF1-deleted cohort to date and clarified the more severe phenotype observed in these patients.

Nf1 heterozygous mice recapitulate the anthropometric and metabolic features of human neurofibromatosis type 1

Investigating therapeutic nonsense suppression in a neurofibromatosis mouse model

Neurofibromatosis type 1 (NF1) is an autosomal dominant genetic syndrome caused by a mutation in or deletion of the NF1 gene. Mutations in the NF1 gene lead to the production of a nonfunctional version of neurofibromin that cannot regulate cell growth and division. As a result, tumors such as neurofibromas can form along nerves throughout the body. The NF1 phenotype is highly penetrant and occurs in 1 in 3,000 to 4,000 people worldwide. Individuals with an altered NF1 gene are at an increased risk of developing benign and/or malignant tumors. NF1 has been shown to negatively regulate Ras activity. Ras-driven cancer cells have also been known to alter glucose and glutamine metabolism. In the present study, we evaluated the role played by NF1 status in determining the sensitivity of sarcoma and melanoma cell lines to glutaminase inhibition and its effect on Ras activity. We tested a panel of soft tissue sarcoma and melanoma cell lines that were either wild-type, null or mutant for NF1. Results from our in vitro proliferation assay showed that compared to wild-type NF1 (STS26T, LS141) cell lines, NF1 mutant (MPNST) and NF1 null (ST88 and MeWo) cell lines showed greater sensitivity to inhibition of proliferation by glutaminase inhibitors CB-839 and BPTES. Western blot analysis showed induction of apoptosis and down regulation of mTORC1 targets such as phospho-S6K and phospho-S6 by glutaminase inhibitors only in NF1 null and NF1 mutant but not wild-type NF1 cell lines. Gene silencing experiments showed that siRNA mediated knockdown of NF1 sensitizes LS141and STS26T cell lines to glutaminase inhibition. Conversely, overexpression of wild-type NF1 GRD (GAP related domain) in MeWo cell line resulted in decreased sensitivity to glutaminase inhibition when tested in a cell proliferation assay, thus, confirming the role played by NF1. Previous reports have shown that mutation or deletion of NF1 results in activation of Ras. In order to test the effect of glutaminase inhibition on Ras activity, we carried out Ras-GTP pull down assay following the treatment with glutaminase inhibitors in NF1 null and wild-type NF1 cell lines. Our results showed that glutaminase inhibition leads to down regulation of activated RAS in NF1 null but not wild-type NF1 cells. SiRNA mediated knockdown of NF1 followed by glutaminase inhibition in wild-type NF1 cell line (LS141) resulted in decreased Ras activity, further confirming our hypothesis. Results from patient derived MPNST tumor xenograft model showed a significant suppression of tumor volume when tumors were treated with glutaminase inhibitor compared to vehicle control. Taken together, our data strongly indicates that NF1 status determines the sensitivity of sarcoma and melanoma cell lines to glutaminase inhibition. Further research is warranted to explore glutaminase inhibition as potential therapy for patients with NF1 loss and/or mutation. Citation Format: Tahir N. Sheikh, Parag P. Patwardhan, Gary K. Schwartz. Neurofibromatosis type 1 (NF1) status determines sensitivity of soft tissue sarcoma and melanoma cell lines to glutaminase inhibitors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 19.

e22550 Background: Neurofibromatosis Type 1 (NF1) is a rare hereditary condition that predisposes to the development of a variety of tumors. Variability in disease manifestation, prevalence and severity, in combination with a low prevalence complicates the efficient conduct of clinical trials in NF1. NF1-associated tumors mainly but not exclusively affect the nervous system as well as the skin, and a percentage of those tumors exhibit a significantly increased risk of malignant transformation. There is an urgent need to identify effective treatments for NF1-associated tumors. The development of innovative trial designs that accelerate the ability to assess new investigational agents is essential. The European Patient-Centric Clinical Trial Platforms (EU-PEARL) is an Innovative Medicines Initiative (IMI) project, aiming to create platform trials for diseases with high unmet medical need. As part of EU-PEARL, we designed a basket-platform trial in NF1 that could serve as a prototype for accelerating drug studies in rare cancer predisposition syndromes. Methods: The trial was designed by NF1 experts, a dedicated statistician and a trial designer. Subprotocols were written for previously prioritized manifestations including plexiform neurofibroma, cutaneous neurofibroma, low-grade glioma, and optic pathway glioma. Collaboration with NF1 experts from the USA was sought to ensure alignment with trials for NF1 in the USA. Results: To optimally learn from a small number of potential participants, patients should be able to participate in both an observational and a treatment phase. The observational phase will serve as longitudinal natural history study, providing data that can be used as a comparator for the treatment arms. To enter the treatment phase, patients must meet additional eligibility criteria. Patients will be randomized to a sequence of available drugs, rather than one single drug. This may allow for the addition of newly identified drugs during the course of the trial. If a drug concept fails or unacceptable toxicity arises, patients may re-enter the observational phase or be re-randomized to a different treatment arm if eligible. Drug-specific eligibility criteria and endpoints are listed separately in Intervention-Specific-Appendices (ISAs), allowing flexibility and adaptability that is needed for a highly variable and progressive rare disorder like NF1. This trial design allows optimal learning from a limited number of patients. Conclusions: We designed a basket-platform trial for four manifestations of NF1. This trial design addresses challenges that may be encountered when designing a clinical trial for NF1. This trial will be the future of clinical trials for NF1 in Europe. Its design could serve as a prototype for other rare diseases: it enhances the chances of finding beneficial treatments by optimizing patient inclusion and invigorating international collaborations.

BackgroundSpinal neurofibromatosis (SNF) is a related form of Neurofibromatosis type 1 (NF1) with a low incidence. Here, we report a SNF patient with NF1 (OMIM *613113) mutation in a classic NF1 family to enrich the case data.MethodsWe presented the clinical data of a 27‐year‐old female suffered from SNF. Two NF1 individuals (the mother and the brother) in the patient's family were also described. In the SNF patient, tumors in cervical were removed by surgical operation after the spinal MRI evaluation. Hematoxylin‐eosin staining and immunohistochemistry were performed to better characterize the excised tumors. NF1 exons of the patient and her NF1 families were further sequenced by the next‐generation sequencing technology.ResultsThe patient developed irregular café‐au‐lait macules, multi‐subcutaneous nodules, recurrent numbness, and weakness of both lower extremities. Multiple neurofibromas were found in the whole spine by spinal MRI. Tumor‐like cells and hyperplasia of ganglion cells were found in the excised tissue by H&E staining and immunohistochemistry, respectively. One‐year follow‐up on the SNF patient showed that after the surgery lower limb pain, numbness and convulsion were completely relieved. A common germ‐line pathogenic mutation (NM_000267.3:c.1721 + 3A>G) was found in both the SNF patient and her classic NF1 families.ConclusionA case of SNF with classic NF1 mutation in a classic NF1 family was identified for the first time, indicating that SNF may share the same gene mutation with NF1, while the different manifestation of NF1 and SNF may be related to gene modification.

Neurofibromatosis type 1 (NF1) is an autosomal dominant genetic disorder. Patients with NF1 have mono-allelic loss of the tumor suppressor gene NF1 in their germline, which predisposes them to develop a wide array of benign lesions. Intriguingly, recent sequencing efforts revealed that the NF1 gene is frequently mutated in multiple malignant tumors not typically associated with NF1 patients, suggesting that NF1 heterozygosity is refractory to at least some cancer types. In two orthogonal mouse models representing tumors associated with NF1 (neurofibroma and malignant peripheral nerve sheath tumor) and non-NF1-related tumors (papilloma and squamous cell carcinoma), we serendipitously discover that an NF1heterozygosity microenvironment accelerates the formation of benign tumors but impairs further progression to malignancy. Analysis of benign and malignant tumors commonly associated with NF1 patients, as well as those with high NF1 gene mutation frequency, reveals an antagonistic role for NF1 heterozygosity in tumor initiation and malignant transformation and helps to reconciliate the role of the NF1 gene in both NF1 and non-NF1 patient contexts. Our novel framework to study the role of NF1 in tumorigenesis integrates the concept of NF1 as a tumor predisposition gene and we have uncovered mechanisms underlying the impairment of cancer progression by tumor microenvironment cells harboring tumor suppressor genes in the haploinsufficient state, such as NF1 heterozygosity, may translate into novel therapeutic strategies.

The EVI2B gene is one of three genes embedded in intron 27b of the neurofibromatosis type 1 (NF1; M. Recklinghausen) gene, which are transcribed in the direction opposite that of the NF1 gene. The function of EVI2B and its relation to NF1 symptoms is unknown. Here, the amounts of NF1 and EVI2B mRNA were investigated in detail in cells involved in NF1 manifestations as café-au-lait macules and neurofibromas. These investigations showed that aside from the NF1 gene, EVI2B is involved in melanocyte and keratinocyte differentiation. Whereas in NF1 melanocytes from café-au-lait macules, EVI2B expression was not altered, in fibroblast-like cells derived from neurofibromas, an increased level of EVI2B mRNA was found. We investigated whether this increase was attributable to an influence of NF1 gene expression on the expression of the EVI2B gene, as suggested by the fact that the EVI2B primary transcript is antisense to the NF1 primary transcript. Investigations of cells derived from patients with different amounts of NF1 pre-mRNA showed no correlation between the amount of NF1 pre-mRNA and the increased level of EVI2B mRNA.