KRAS inhibition and KRAS degradation: tools for the identification of on-target and off-target resistance to KRAS-G12C inhibitors.

KRAS is one of the most frequently mutated oncogenes in human cancer. In patients with LUAD, KRAS p.G12C is the most frequent driver mutation. It causes diminished regulation of the GTPase activity that results in the over-activation of the MAPK pathway and a deregulated cell proliferation and survival. A clinically approved, mutant selective KRASG12C therapy based on the KRASG12C inhibitor AMG-510 (Sotorasib) is now available following accelerated approval of an allele-specific covalent inhibitor granted by the FDA. In this study, we used a controlled genetic model, the Ras-less cell line and dTAG degradation system, to induce in vitro resistance to the G12C “off-inhibitor” AMG-510 and characterize the mechanisms of resistance using both inhibition and degradation of the target. This model gives us the opportunity to highlight different response in cells that maintain the expression of the wild-type allele of KRAS (WT/MUT) and in cells that undergo loss of heterozygosity (LOH). This works highlights the presence of off-mechanisms of resistance to AMG-510 inhibition that can rescue the signaling through the MAPK pathway under inhibition or degradation of KRASG12C. We also point out the role of wild-type KRAS as a biomarker for response to G12C “off-inhibitors”. Cells that still express this allele are more refractory to the inhibitors of KRASG12C, develop a faster and stronger resistance. Lastly, we studied the possibility that resistant cells to a first-line treatment retain sensitivity to a second-line treatment with another KRASG12C inhibitor, MRTX-849 (Adagrasib).