Genomic Analysis and Biochemical Investigation Reveal Mechanisms of Schwann Cell Transformation and Treatment Resistance in Malignant Peripheral Nerve Sheath Tumors (MPNSTs).

Abstract

PURPOSE/OBJECTIVE(S)\nIn contrast to benign Schwann cell derived tumors such as neurofibromas and schwannomas, malignant peripheral nerve sheath tumors (MPNSTs) metastasize, and have poor outcomes. MPNSTs are associated with loss of NF1, a tumor suppressor that inhibits Ras signaling, and loss of Polycomb Repressive Complex 2 (PRC2) which resulting in epigenetic dysregulation. Translationally, MPNSTs are resistant to Ras/MEK inhibition with selumetinib and radiation therapy yet the mechanisms underlying how PRC2 loss cooperates with Ras misactivation to drive transformation and treatment resistance remains unclear. Here, we performed DNA methylation profiling and RNA-sequencing of Schwann cell tumors and cell lines as well as investigated in vitro responses to radiotherapy and selumetinib.\n\n\nMATERIALS/METHODS\nWe identified 119 Schwann cell tumor resection specimens with sufficient tissue for DNA methylation profiling (66 schwannoma, 13 neurofibroma, 40 MPNSTs) and RNA-sequencing (23 schwannoma, 9 neurofibroma, 9 MPNSTs). To study cell signaling and treatment responses, we utilized primary human cell lines generated from NF1 intact peripheral nerve, NF1 mutant neurofibromas, and MPNSTs (n\u202f=\u202f6). We assessed Ras signal transduction by immunoblot/immunoprecipitation for downstream effectors and performed cell viability assays following selumetinib or radiotherapy.\n\n\nRESULTS\nPrincipal component analysis (PCA) of DNA methylation and RNA-sequencing data demonstrated a gradient of epigenetic signatures across Schwann cell tumors, with schwannomas comprising a distinct cluster from neurofibromas and MPNST. Moreover, differential DNA methylation and RNA-sequencing analyses both revealed enrichment for PRC2 target genes and repression of Schwann cell differentiation genes in MPNSTs. RNA-sequencing of MPNST versus neurofibroma cell lines demonstrated a PRC2 mediated signature enriched for cell differentiation genes and negative feedback regulators of Ras signaling, suggesting PRC2 dysregulation mediates de-differentiation and increased Ras pathway output in MPNSTs. In contrast to neurofibroma cells, MPNST cells were deficient in PRC2 and NF1 while exhibiting increased basal active Ras-GTP levels insensitive to serum starvation. PRC2 deficient MPNST cell lines were more resistant to selumetinib and radiation therapy when compared to NF1 deficient neurofibroma cells.\n\n\nCONCLUSION\nPRC2 loss causes epigenetic dysregulation that may mediate Schwann cell de-differentiation programs and altered Ras signaling in MPNSTs compared to neurofibromas. Translationally, these epigenetic changes may contribute to increased resistance to targeted therapy and radiation. The mechanisms underlying transformation and treatment resistance upon PRC2 loss are under further investigation.