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Supplementary Figure 1 from Glomus Tumors in Neurofibromatosis Type 1: Genetic, Functional, and Clinical Evidence of a Novel Association

Neurofibromatosis type 1 (NF1) is a common disorder that arises secondary to mutations in the tumor suppressor gene NF1. Glomus tumors are small, benign but painful tumors that originate from the glomus body, a thermoregulatory shunt concentrated in the fingers and toes. We report 11 individuals with NF1 who harbored 20 glomus tumors of the fingers and 1 in the toe; 5 individuals had multiple glomus tumors. We hypothesized that biallelic inactivation of NF1 underlies the pathogenesis of these tumors. In 12 NF1-associated glomus tumors, we used cell culture and laser capture microdissection to isolate DNA. We also analyzed two sporadic (not NF1-associated) glomus tumors. Genetic analysis showed germ line and somatic NF1 mutations in seven tumors. RAS mitogen-activated protein kinase hyperactivation was observed in cultured NF1−/− glomus cells, reflecting a lack of inhibition of the pathway by functional neurofibromin, the protein product of NF1. No abnormalities in NF1 or RAS mitogen-activated protein kinase activation were found in sporadic glomus tumors. By comparative genomic hybridization, we observed amplification of the 3′-end of CRTAC1 and a deletion of the 5′-end of WASF1 in two NF1-associated glomus tumors. For the first time, we show that loss of neurofibromin function is crucial in the pathogenesis of glomus tumors in NF1. Glomus tumors of the fingers or toes should be considered as part of the tumor spectrum of NF1. [Cancer Res 2009;69(18):7393–401]

Supplementary Figure Legend from Glomus Tumors in Neurofibromatosis Type 1: Genetic, Functional, and Clinical Evidence of a Novel Association

Supplementary Table 1 from Glomus Tumors in Neurofibromatosis Type 1: Genetic, Functional, and Clinical Evidence of a Novel Association

Supplementary Table 2 from Glomus Tumors in Neurofibromatosis Type 1: Genetic, Functional, and Clinical Evidence of a Novel Association

Little is known about the influence that environmental stressors may have on genome-wide methylation patterns, and to what extent epigenetics may be involved in environmental stress response. Yet, studies of methylation patterns under stress could provide crucial insights on stress response and toxicity pathways. Here, we focus on genome-wide methylation patterns in the microcrustacean Daphnia magna, a model organism in ecotoxicology and risk assessment, exposed to the toxic cyanobacterium Microcystis aeruginosa. Bisulfite sequencing of exposed and control animals highlighted differential methylation patterns in Daphnia upon exposure to Microcystis primarily in exonic regions. These patterns are enriched for serine/threonine amino acid codons and genes related to protein synthesis, transport and degradation. Furthermore, we observed that genes with differential methylation corresponded well with genes susceptible to alternative splicing in response to Microcystis stress. Overall, our results suggest a complex mechanistic response in Daphnia characterized by interactions between DNA methylation and gene regulation mechanisms. These results underscore that DNA methylation is modulated by environmental stress and can also be an integral part of the toxicity response in our study species. ispartof: Environmental Science & Technology vol:51 issue:2 pages:924-931 ispartof: location:United States status: published

Neurofibromatosis type 1 (NF1) is a common, autosomal dominant, tumor-predisposition syndrome that arises secondary to mutations in NF1. Glomus tumors are painful benign tumors that originate from the glomus body in the fingers and toes due to biallelic inactivation of NF1. We karyotyped cultures from four previously reported and one new glomus tumor and hybridized tumor (and matching germline) DNA on Illumina HumanOmni1-Quad SNP arrays (∼1 × 10(6) SNPs). Two tumors displayed evidence of copy-neutral loss of heterozygosity of chromosome arm 17q not observed in the germline sample, consistent with a mitotic recombination event. One of these two tumors, NF1-G12, featured extreme polyploidy (near-tetraploidy, near-hexaploidy, or near-septaploidy) across all chromosomes. In the remaining four tumors, there were few cytogenetic abnormalities observed, and copy-number analysis was consistent with diploidy in all chromosomes. This is the first study of glomus tumors cytogenetics, to our knowledge, and the first to report biallelic inactivation of NF1 secondary to mitotic recombination of chromosome arm 17q in multiple NF1-associated glomus tumors. We have observed mitotic recombination in 22% of molecularly characterized NF1-associated glomus tumors, suggesting that it is a not uncommon mechanism in the reduction to homozygosity of the NF1 germline mutation in these tumors. In tumor NF1-G12, we hypothesize that mitotic recombination also "unmasked" (reduced to homozygosity) a hypomorphic germline allele in a gene on chromosome arm 17q associated with chromosomal instability, resulting in the extreme polyploidy. © 2012 Wiley Periodicals, Inc. ispartof: Genes, Chromosomes & Cancer vol:51 issue:5 pages:429-437 ispartof: location:United States status: published

Neurofibromatosis type 1 (NF1) is a common disorder that arises secondary to mutations in the tumor suppressor gene NF1. Glomus tumors are small, benign but painful tumors that originate from the glomus body, a thermoregulatory shunt concentrated in the fingers and toes. We report 11 individuals with NF1 who harbored 20 glomus tumors of the fingers and 1 in the toe; 5 individuals had multiple glomus tumors. We hypothesized that biallelic inactivation of NF1 underlies the pathogenesis of these tumors. In 12 NF1-associated glomus tumors, we used cell culture and laser capture microdissection to isolate DNA. We also analyzed two sporadic (not NF1-associated) glomus tumors. Genetic analysis showed germ line and somatic NF1 mutations in seven tumors. RAS mitogen-activated protein kinase hyperactivation was observed in cultured NF1−/− glomus cells, reflecting a lack of inhibition of the pathway by functional neurofibromin, the protein product of NF1. No abnormalities in NF1 or RAS mitogen-activated protein kinase activation were found in sporadic glomus tumors. By comparative genomic hybridization, we observed amplification of the 3′-end of CRTAC1 and a deletion of the 5′-end of WASF1 in two NF1-associated glomus tumors. For the first time, we show that loss of neurofibromin function is crucial in the pathogenesis of glomus tumors in NF1. Glomus tumors of the fingers or toes should be considered as part of the tumor spectrum of NF1. [Cancer Res 2009;69(18):7393–401]

The polycomb repressive complex 2 (PRC2) exerts oncogenic effects in many tumour types1. However, loss-of-function mutations in PRC2 components occur in a subset of haematopoietic malignancies, suggesting that this complex plays a dichotomous and poorly understood role in cancer2, 3. Here we provide genomic, cellular, and mouse modelling data demonstrating that the polycomb group gene SUZ12 functions as tumour suppressor in PNS tumours, high-grade gliomas and melanomas by cooperating with mutations in NF1. NF1 encodes a Ras GTPase-activating protein (RasGAP) and its loss drives cancer by activating Ras4. We show that SUZ12 loss potentiates the effects of NF1 mutations by amplifying Ras-driven transcription through effects on chromatin. Importantly, however, SUZ12 inactivation also triggers an epigenetic switch that sensitizes these cancers to bromodomain inhibitors. Collectively, these studies not only reveal an unexpected connection between the PRC2 complex, NF1 and Ras, but also identify a promising epigenetic-based therapeutic strategy that may be exploited for a variety of cancers.

The identification of cancer drivers is a major goal of current cancer research. Finding driver genes within large chromosomal events is especially challenging because such alterations encompass many genes. Previously, we demonstrated that zebrafish malignant peripheral nerve sheath tumors (MPNSTs) are highly aneuploid, much like human tumors. In this study, we examined 147 zebrafish MPNSTs by massively parallel sequencing and identified both large and focal copy number alterations (CNAs). Given the low degree of conserved synteny between fish and mammals, we reasoned that comparative analyses of CNAs from fish versus human MPNSTs would enable elimination of a large proportion of passenger mutations, especially on large CNAs. We established a list of orthologous genes between human and zebrafish, which includes approximately two-thirds of human protein-coding genes. For the subset of these genes found in human MPNST CNAs, only one quarter of their orthologues were co-gained or co-lost in zebrafish, dramatically narrowing the list of candidate cancer drivers for both focal and large CNAs. We conclude that zebrafish-human comparative analysis represents a powerful, and broadly applicable, tool to enrich for evolutionarily conserved cancer drivers., Kathy and Curt Marble Cancer Research Fund, Arthur C. Merrill, National Institutes of Health (U.S.) (Grant CA106416), National Institutes of Health (U.S.) (Grant ROI RR020833), National Institutes of Health (U.S.) (Grant 1F32GM095213-01)

Legius syndrome presents as an autosomal dominant condition characterized by cafe-au-lait macules with or without freckling and sometimes a Noonan-like appearance and/ or learning difficulties. It is caused by germline loss-of-function SPRED1 mutations and is a member of the RAS-MAPK pathway syndromes. Most mutations result in a truncated protein and only a few inactivating missense mutations have been reported. Since only a limited number of patients has been reported up until now, the full clinical and mutational spectrum is still unknown. We report mutation data and clinical details in fourteen new families with Legius syndrome. Six novel germline mutations are described. The Trp31Cys mutation is a new pathogenic SPRED1 missense mutation. Clinical details in the 14 families confirmed the absence of neurofibromas, and Lisch nodules, and the absence of a high prevalence of central nervous system tumors. We report white matter T2 hyperintensities on brain MRI scans in 2 patients and a potential association between postaxial polydactyly and Legius syndrome. (C) 2010 Wiley-Liss, Inc.

The polycomb repressive complex 2 (PRC2) exerts oncogenic effects in many tumor types. However, loss-of-function mutations in PRC2 components occur in a subset of hematopoietic malignancies, suggesting that this complex plays a dichotomous and poorly understood role in cancer. Our genomic, cellular, and mouse modeling data demonstrates that the polycomb group gene SUZ12 functions as tumor suppressor in PNS tumors, glioblastomas, and melanomas by cooperating with mutations in NF1. NF1 encodes a Ras GTPase-activating protein and its loss drives cancer by activating Ras. We show that SUZ12-loss potentiates the effects of NF1 mutations by amplifying Ras-driven transcription. Importantly however, SUZ12-inactivation also triggers an epigenetic switch that sensitizes these cancers to bromodomain inhibitors. Collectively, these studies not only reveal an unexpected connection between the PRC2 complex, NF1, and Ras, but also identify a promising epigenetic-based therapeutic strategy that may be exploited for a variety of cancers. Citation Format: Thomas De Raedt, Eline Beert, Eric Pasmant, Armelle Luscan, Hilde Brems, Nicolas Ortonne, Kristian Helin, Victor Mautner, Hildegard Kehrer-Sawatski, Wade Clapp, James Bradner, Meena Upadhyaya, Michel Vidaud, Eric Legius, Karen Cichowski. PRC2 loss amplifies Ras-driven transcription and sensitizes cancers to bromodomain inhibitor-based combination therapies. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-79. doi:10.1158/1538-7445.AM2014-LB-79

Patients with the familial cancer syndrome neurofibromatosis type I possess mutations in the NF1 tumor suppressor gene, which encodes a RasGAP. However a subset of patients carry a larger deletion that encompasses NF1 and the surrounding 13 genes. These patients develop hundreds of benign nervous system tumors, are at much higher risk for developing malignancies, and exhibit specific developmental defects. Therefore we hypothesized that a cooperating tumor suppressor must lie within this region. Notably, this entire region is frequently deleted in in a number of different cancers, including glioblastoma (GBM). To identify a potential cooperating tumor suppressor we analyzed a large number of NF1-associated malignancies by array CGH and detected homozygous loss of SUZ12, a member of the PRC2 repressor complex. Subsequent mutation analysis showed that SUZ12 was homozygously inactivated in a large subset of MPNSTs. Importantly, reintroduction of SUZ12 in SUZ12 deficient Malignant Peripheral Nerve Sheath tumors (MPNST) cell lines restored the function of the PRC2 complex and induced cell death. Conversely, RNAi mediated inactivation of SUZ12 significantly enhanced the tumorigenic properties of NF1-deficient GBMs. Additionally, we found that mice carrying compound heterozygous mutations in Nf1 and Suz12 developed a variety of tumor types and had a marked decrease in overall survival. As expected, knockdown of SUZ12 dramatically decreased the global H3K27 tri-methylation (repressive mark) and increased the global H3K27 acetylation (activation mark). This suggests that the pathogenic effect of SUZ12-loss is, at least in part, caused by an aberrant activation of genes due to an increase in H3K27 acetylation. Interestingly this H3K27 acetylation mark is read by BRD4, suggesting that inhibition of BRD4 might decrease the expression of critical targets. In vitro and in vivo studies indeed confirmed a hypersensitivity to BRD4 inhibition of cell lines and primary tumors lacking SUZ12. Additionally our preliminary data show an inverse correlation between the H3K27 tri-methylation level and the response to treatment with the BRD4 inhibitor JQ1, suggesting that SUZ12 levels, and thus H3K27 tri-methylation status, could be used as a biomarker. In conclusion, we showed that SUZ12 is a tumor suppressor in a number of different cancers, including MPNSTs and gliomas. These tumors are exquisitely sensitive to inhibition of BRD4. Loss of SUZ12 leads to complete ablation of the H3K27 tri-methylation mark, which could serve as a biomarker for treatment with the BRD4 inhibitor JQ1. This abstract is also presented as Poster B51. Citation Format: Thomas De Raedt, Eline Beert, Eric Pasmant, James E. Bradner, Pierre Wolkenstein, Eric Legius, Karen Cichowski. SUZ12: A novel tumor suppressor and potential biomarker for efficacy of BRD4 inhibition. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr PR15.