PP17. RHO GTPASE SIGNALLING IN CANCER CELL INVASION AND ANGIOGENESIS IN BRAIN TUMOURS

Abstract

Listen

Translate article

AbstractINTRODUCTION: Rho GTPases are molecular switches that cycle between GDP-bound inactive and GTP-bound active states. They transduce signals from the extracellular environment and downstream of oncogenes to control the actin cytoskeleton, and cellular functions including cell motility, cell cycle progression, polarity, malignant transformation, invasion and metastasis. Activation of Rho proteins is controlled by guanine nucleotide exchange factors (GEFs), and GTPase activating proteins (GAPs). Rho GEFs and GAPs are frequently overexpressed in tumours and they are tractable targets for cancer therapy. Rho proteins Rac1 and Cdc42 are deregulated in GBM. Our study is focused on understanding the role of Rac GEF DOCK4 and Cdc42 GEF DOCK9 [Abraham S, Scarcia M, ..Mavria G (2015) Nat Commun. 6: 7286] in the tumour cell and stroma cell compartments of glioblastomas. METHOD: i) The role of DOCK4 and DOCK9 in GBM invasion was analysed using organotypic spheroid assays, the U251 model and patient-derived models of GBM ii) DOCK4 expression was analysed in GBM patient samples and related to expression of stem cell marker nestin. iii) Blood vessel types were characterised in GBM patient samples and related to histological parameters and patient outcome following treatments. iii) The role of DOCK4 was investigated in the vasculature of heterozygous Dock4 null mice and syngeneic GBM tumour models implanted orthotopically. RESULTS: i) There was reduced outgrowth of GBM cells in spheroid assays with DOCK4 or DOCK9 knockdown. A complex of DOCK4 with DOCK9 was identified by immunoprecipitation in U251 cells. ii) Paired DOCK4 and nestin scores of immunostaining from 19 GBM patient samples showed DOCK4 scores correlated with nestin scores using two-tailed Spearmans Rho. Oncomine microarray data shows significant increase of DOCK4 levels in cultured GBM cells isolated from tumours (n=25) compared to neural stem cells (p=8.16 x 104). iii) Hyperproliferative blood vessels in GBM show larger diameter compared to normal blood vessels and increased number of glomeruloid structures was observed in recurrences iv) Heterozygous Dock4 genetic deletion results in reduction of blood vessel diameter. CONCLUSIONS: Tumour cell invasion into adjacent normal brain and vascular proliferation are pathologic hallmarks of GBM. Our results show that DOCK4 may be implicated in both vascular growth and glioblastoma invasion. Delineating further the signalling network in which DOCK4 participates may shed considerable light in these processes and identify new targets for treatments.