Beyond KRAS(G12C): biochemical and computational characterization of sotorasib and adagrasib binding specificity and the critical role of H95 and Y96

Mutated KRAS proteins are frequently expressed in some of the most lethal human cancers, thus having been a target of intensive drug discovery efforts for decades. Lately, KRAS(G12C) switch-II pocket (SII-P)-targeting covalent small molecule inhibitors have finally reached the clinical practice. Sotorasib (AMG-510) was the first FDA-approved covalent inhibitor to treat KRAS(G12C)-positive non–small cell lung cancer (NSCLC), followed soon by adagrasib (MRTX849). Both drugs target the GDP-bound state of KRAS(G12C), exploiting the strong nucleophilicity of the acquired cysteine. Here, we evaluate the similarities and differences between sotorasib and adagrasib in their RAS SII-P binding by applying biochemical, cellular, and computational methods. Exact knowledge on SII-P engagement can enable targeting this site by reversible inhibitors for KRAS mutants beyond G12C. We show that adagrasib is strictly KRAS- but not KRAS(G12C)-specific, due to its strong and unreplaceable interaction with H95. Unlike adagrasib, sotorasib is less dependent on H95 for its binding, making it a RAS isoform-agnostic compound. Our results emphasize the accessibility of the SII-P beyond oncogenic G12C and aid in understanding the molecular mechanism behind the clinically observed drug resistance, associated especially with secondary mutations on KRAS H95 and Y96.