fig4

Figure 4. HCT116 cells acquire resistance to selumetinib by amplifying their driving oncogene KRAS^G13D. KRAS^G13D-mutant HCT116 cells are addicted to ERK1/2 signalling (red) for proliferation and survival (top, left); inhibiting this pathway with the MEKi selumetinib blocks cell proliferation and initiates cell death (top, right). Selumetinib inhibits MEK1/2 by constraining the kinase domain catalytic sites in an inactive conformation, thereby inhibiting phosphorylation and activation of ERK1/2 (top, right). HCT116 cells also harbour an activating H1047R mutation in PIK3CA, which encodes the catalytic p110α subunit of PI3K. Following 6-8 weeks culture in the presence of selumetinib, resistant derivatives of HCT116 (H6244-R) cells emerge that proliferate normally and harbour amplification of KRAS^G13D (bottom). The consequent increase in KRAS^G13D expression results in activation of a larger pool of p-MEK1/2 with sufficient residual activity in the presence of selumetinib to reinstate ERK1/2 phosphorylation and pathway activity to parental HCT116 levels (bottom). Consistent with upregulation of KRAS^G13D, selumetinib-resistant HCT116 cells also exhibit elevated PI3K-PKB signalling (blue). P: phosphate group; PIP₃: phosphatidylinositol-3,4,5-trisphosphate