Suite of Biochemical\nand Cell-Based Assays for the\nCharacterization of Kirsten Rat Sarcoma (KRAS) Inhibitors and Degraders

KRAS is an important\noncogenic driver which is mutated in numerous\ncancers. Recent advances in the selective targeting of KRAS mutants\nvia small molecule inhibitors and targeted protein degraders have\ngenerated an increase in research activity in this area in recent\nyears. As such, there is a need for new assay platforms to profile\nnext generation inhibitors which improve on the potency and selectivity\nof existing drug candidates, while evading the emergence of resistance.\nHere, we describe the development of a new panel of biochemical and\ncell-based assays to evaluate the binding and function of known chemical\nentities targeting mutant KRAS. Our assay panels generated selectivity\nprofiles and quantitative binding interaction dissociation constants\nfor small molecules and degraders against wild type, G12C, G12D, and\nG12V KRAS, which were congruent with published data. These assays\ncan be leveraged for additional mutants of interest beyond those described\nin this study, using both overexpressed cell-free systems and cell-based\nsystems with endogenous protein levels.