Endogenous oncogenic KRAS expression increases cell proliferation and motility in near-diploid hTERT RPE-1 cells

About 18% of all human cancers carry a mutation in the KRAS gene making it among the most sought-after anti-cancer targets. However, mutant KRas protein has proved remarkably undruggable. The recent approval of the first generation of RAS inhibitors therefore marks a seminal milestone in the history of cancer research. Inevitably though, it also raises the predictable challenges of limited drug efficacies and acquired resistance. Hence, new approaches that improve our understanding of the tumorigenic mechanisms of oncogenic RAS within more physiological settings continue to be essential. Here, we have employed the near-diploid human hTERT RPE-1 cells to generate isogenic cell lines in which one of the endogenous KRAS alleles carries an oncogenic KRAS mutation at glycine 12. Cells with a KRAS ^G12V/+ , KRAS ^G12C/+ , or KRAS ^G12D/+ genotype, together with wild-type KRAS ^G12G(WT)/+ cells, reveal that oncogenic KRAS.G12X mutations increase cell proliferation rate, while further analyses showed that KRAS ^G12V/+ cells had increased cell motility and reduced focal adhesions. EGF-induced ERK phosphorylation was marginally increased in KRAS ^G12V/+ cells, while EGF-induced AKT phosphorylation was comparable between KRAS ^G12V/+ and KRAS ^G12G(WT)/+ cells. Interestingly, the KRAS ^G12V/+ cells were more sensitive to hydroxyurea and a MEK inhibitor, U0126, but more resistant to a PI3K inhibitor, PIK-90, than the KRAS ^G12G(WT)/+ cells. A combination of low doses of hydroxyurea and U0126 showed an additive inhibition on growth rate that was greater in KRAS ^G12V/+ than wild-type cells. Collectively, these cell lines will be a valuable resource for studying oncogenic RAS signalling and developing effective anti-KRAS reagents with minimum cytotoxicity on wild-type cells.