Immortalization of human normal and NF1 neurofibroma Schwann cells

Abstract

Neurofibromas, which are benign Schwann cell tumors, are the hallmark feature in the autosomal dominant condition neurofibromatosis 1 (NF1) and are associated with biallelic loss of NF1 gene function. There is a need for effective therapies for neurofibromas, particularly the larger, plexiform neurofibromas. Tissue culture is an important tool for research. However, it is difficult to derive enriched human Schwann cell cultures, and most enter replicative senescence after 6-10 passages, impeding cell-based research in NF1. Through exogenous expression of human telomerase reverse transcriptase and murine cyclin-dependent kinase (mCdk4), normal (NF1 wild-type), neurofibroma-derived Schwann cells heterozygous for NF1 mutation, and neurofibroma-derived Schwann cells homozygous for NF1 mutation were immortalized, including some matched samples from the same NF1 patient. Initial experiments employed retroviral vectors, while subsequent work utilized lentiviral vectors carrying these genes because of improved efficiency. Expression of both transgenes was required for immortalization. Molecular and immunohistochemical analysis indicated that these cell lines are of Schwann cell lineage and have a range of phenotypes, many of which are consistent with their primary cultures. This is the first report of immortalization and detailed characterization of multiple human NF1 normal nerve and neurofibroma-derived Schwann cell lines, which will be highly useful research tools to study NF1 and other Schwann tumor biology and conditions.

EZH1/2 function mostly within canonical PRC2 and exhibit proliferation-dependent redundancy that shapes mutational signatures in cancer

“…Considering the lack of evidence for PRC2-independent role of EZH2 that would explain why it is not found to be mutated in MPNST, we investigated the alternative hypothesis that the absence of mutations affecting EZH2 might reflect compensation by EZH1. We inactivated EZH1 and EZH2 separately or in combination in the ipNF05.5 cell line derived from a plexiform neurofibroma (29), corresponding to the tumor type from which MPNSTs arise. The resulting mutant cells were compared with SUZ12-mutant cells derived from the same cell line (GSE118186, ref.…”

Regulation of localization and function of the transcriptional co-activator YAP by angiomotin

“…hSC2λ cells were obtained from the laboratory of Dr. Margret Wallace (Li et al, 2016). All cell lines were authenticated by short tandem repeat (STR) DNA profiling (DDC Medical).…”

Malignant Peripheral Nerve Sheath Tumors: From Epigenome to Bedside

“…In addition, Li and colleagues developed a series of immortalized SC lines from healthy individuals and from NF-1 patients. Furthermore, NF1-null and heterozygous immortalized cell lines were established from the same individual, providing isogenic cells for study (155). The availability of immortalized SC lines will expedite the process of understanding genetic and epigenetic aberrations that result from loss of NF1, and those that occur during MPNST evolution.…”

“…Crucially, potential interventions must be considered within the context of an NF1 heterozygous individual, because therapies that target hyperactive Ras signaling could exert deleterious effects in individuals with germline NF1 haploinsufficiency. NF1 heterozygous immortalized SC lines will be helpful in studying this narrowed therapeutic window (155).…”