Differential effects of KRAS G-domain and hypervariable region mutants on cancer phenotypes

Identification of tumor driver genes in cancer has led to the discovery of specific allelic variants that are useful as predictive or prognostic markers. In colorectal cancer (CRC), the K-Ras GTPase is one of the most mutated oncoproteins; therefore, understanding the functional consequences of its mutations will allow us to exploit allele-specific vulnerabilities and propose potential mechanisms of resistance in cancer therapeutics. In this study, cellular phenotypic consequences of the K-Ras mutants K117I, K117N, A146T, E168K, and K172T identified in Filipino young-onset CRC (YO-CRC) patients were assessed in HCT116, Caco-2, and NIH3T3 cells. In assays assessing DNA doubling in 2D cultures and viability in 3D multicellular spheroids, only K117I, K117N, and A146T exhibited enhanced proliferation and supported a tumor-like property of increased size and viability. All mutants exhibited increased cellular migration in HCT116 and NIH3T3 but not in Caco-2 cells. None of them caused resistance to apoptosis. Compared to wild type, K117I, K117N, and A146T increased Elk1 reporter activation as well as phospho-Erk and phospho-Akt levels. All mutants induced extensive cytoskeletal remodeling including formation of migratory and invasive structures. Finally, GDP docking simulations suggested increased nucleotide exchange as a mechanism of activation.