Mutation of PTPN11 (Encoding SHP-2) Promotes MEK Activation and Malignant Progression in Neurofibromin-Deficient Cells in a Manner Sensitive to BRAP Mutation

SIMPLE SUMMARY: Germline mutations of NF1 cause neurofibromatosis type 1 (NF1), which is characterized by multiple benign peripheral nerve sheath tumors known as neurofibromas. In some individuals with NF1, plexiform neurofibromas can give rise to malignant peripheral nerve sheath tumors. Here, we applied genomic DNA sequencing to NF1-derived tumors and identified additional genetic alterations in PTPN11 (encoding Src homology region 2 domain-containing phosphatase-2 (SHP)-2) and BRAP associated with NF1 tumor malignancy. We found that the forced expression of the mutant form of SHP-2 activated the protein kinase MEK and increased tumorigenic activity in NF1 cells, and that these effects were attenuated by the forced expression of the mutant form of BRCA1-associated protein (BRAP). This suppressive action of mutant BRAP was not apparent in NF1-intact cells. Our data indicate that the combination of NF1 mutation and PTPN11 mutation drives the malignancy of NF1 cells and that SHP-2 inhibition by BRAP is a potential therapeutic strategy for NF1-associated malignant tumors. ABSTRACT: Germline mutations of NF1 cause neurofibromatosis type 1 (NF1) through the activation of the RAS signaling pathway, and some NF1 patients develop malignant peripheral nerve sheath tumors (MPNSTs). Here, we established subclones of the human NF1-MPNST cell line sNF96.2 that manifest increased tumorigenic activity and increased phosphorylation of the protein kinases MEK and Akt relative to the parental cells. Genomic DNA sequencing identified 14 additional heterozygous mutations within the coding regions of 13 cancer- and other disease-related genes in these subclones. One of these genes, PTPN11, encodes SHP-2, and the forced expression of the identified G503V mutant of SHP-2 increased both tumorigenic activity and MEK phosphorylation in parental sNF96.2 cells, suggesting that the combination of PTPN11 and NF1 mutations induces the pathological activation of the RAS pathway. These effects of SHP-2 (G503V) were inhibited by the coexpression of the G370A mutant of BRAP, which was also detected in the highly malignant subclones, and this inhibition was accompanied by the calpain-dependent cleavage of SHP-2 (G503V). The cleavage of SHP-2 (G503V) and suppression of MEK phosphorylation mediated by BRAP (G370A) were not detected in NF1-intact (HeLa) cells. Tumor promotion by SHP-2 (G503V) and its suppression by BRAP (G370A) may serve as a basis for the development of new treatment strategies for NF1.