Neurofibromin Deficiency and Extracellular Matrix Cooperate to Increase Transforming Potential through FAK-Dependent Signaling

Abstract

Plexiform neurofibromas (Pnfs) are benign peripheral nerve sheath tumors that are major features of the human genetic syndrome, neurofibromatosis type 1 (NF1). Pnfs are derived from Schwann cells (SCs) undergoing loss of heterozygosity (LOH) at the NF1 locus in an NF1+/− milieu and thus are variably lacking in the key Ras-controlling protein, neurofibromin (Nfn). As these SCs are embedded in a dense desmoplastic milieu of stromal cells and abnormal extracellular matrix (ECM), cell–cell cooperativity (CCC) and the molecular microenvironment play essential roles in Pnf progression towards a malignant peripheral nerve sheath tumor (MPNST). The complexity of Pnf biology makes treatment challenging. The only approved drug, the MEK inhibitor Selumetinib, displays a variable and partial therapeutic response. Here, we explored ECM contributions to the growth of cells lacking Nfn. In a 3D in vitro culture, NF1 loss sensitizes cells to signals from a Pnf-mimicking ECM through focal adhesion kinase (FAK) hyperactivation. This hyperactivation correlated with phosphorylation of the downstream effectors, Src, ERK, and AKT, and with colony formation. Expression of the GAP-related domain of Nfn only partially decreased activation of this signaling pathway and only slowed down 3D colony growth of cells lacking Nfn. However, combinatorial treatment with both the FAK inhibitor Defactinib (VS-6063) and Selumetinib (AZD6244) fully suppressed colony growth. These observations pave the way for a new combined therapeutic strategy simultaneously interfering with both intracellular signals and the interplay between the various tumor cells and the ECM.

Neurofibromin Deficiency Causes Epidermal Growth Factor Receptor Upregulation through the Activation of Ras/ERK/SP1 Signaling Pathway in Neurofibromatosis Type 1-Associated Malignant Peripheral Nerve Sheet Tumor

“…These results can explain how neurofibromin regulates EGFR expression; reduction of neurofibromin levels induces the activation of the Ras/ERK signaling pathway, leading to an increase in SP1 phosphorylation and binding to the EGFR gene promoter region and subsequently promoting EGFR transcription. Downregulation of neurofibromin expression levels has been suggested to be involved in the resistance to anticancer drugs caused by hyperactivation of Ras/ERK signaling in NF1-associated MPNST cells [38], clear cell kidney cancer cells [39], mouse embryonic fibroblasts, and mouse neurofibroma-associated Schwann cells [40]. EGFR dysregulation contributes to MPNST transformation by promoting the proliferation of Schwann cells in NF1 and induces drug resistance in MPNST [41].…”

Integrated genomic analysis of NF1-associated peripheral nerve sheath tumors: an updated biorepository dataset

“…To further characterize the MPNST and PNF cohorts, we evaluated the expression of genes previously implicated in MPNST biology [29]. We found that ERBB3, S100B, CNP, PMP22, PDGFB, and TGFBR2 were significantly downregulated in MPNST compared to PNF, while CCNB2, TWIST1, COL6A3, PTK7, GAS1, and TGFB2 were significantly upregulated in MPNST (Figure 4C,D) (|fold change| > 1.2 with adjusted P < 0.05), consistent with previous reports [29,30].…”

Neurofibromatosis Type 1 Gene Alterations Define Specific Features of a Subset of Glioblastomas

“…Neurofibromin regulates the dynamic reorganisation and turnover of actin filaments through its interacting partners such as Ras-related C3 botulinum toxic substrate 1 (Rac1) [ 114 , 115 ], Lim kinases (LIMK1/2) [ 77 , 84 , 114 , 116 ], syndecan-2 [ 117 ], and focal adhesion kinase (FAK) [ 118 , 119 , 120 ] among others. Neurofibromin binding to syndecan-2 induces actin polymerisation and filopodia formation in dendrites [ 117 ].…”

“…Neurofibromin binding to syndecan-2 induces actin polymerisation and filopodia formation in dendrites [ 117 ]. Along the same line, neurofibromin interaction with FAK regulates cell migration [ 120 ]. Given its role as a modulator of cytoskeletal and focal adhesion as well as a negative regulator of RAS signalling, mutations, or loss of NF1 results in disruption of the extracellular matrix and induction of EMT.…”