by Juan Zeng, YiXuan Lan, Fei XiaRas proteins are prominent oncogenes, with KRas mutations found in approximately 80% of cancer cells harboring Ras mutations. The mechanism by which Ras mutations cause cancer remains unclear. Human Son of Sevenless (SOS) promotes the GDP-to-GTP exchange in the inactive GDP-bound Ras (RasGDP) by interacting with RasGDP conformation, thereby leading to the development of human cancer. Elucidating the Ras-SOS interaction mechanism can guide the drug design for Ras and SOS proteins. Based on our previously sampled special structure KRasGDP·Mg2+S1.2, this study constructs a functional ternary complex (KRasGDP·Mg2+)·SOS1·(KRasGTP·Mg2+). Furthermore, the KRas-SOS1 interactions regulated by the KRas G12D mutation and the SOS1 inhibitor BI-3406 that reportedly exhibits synergistic effects with G12D-mutant Ras inhibitors, are investigated through molecular dynamics (MD) simulations. The findings reveal that the G12D mutation and BI-3406 both affect the KRas-SOS1 interaction via the Switch-II (SW2) region of KRas. The negatively charged Asp12 has a repulsive effect on KRas, particularly on SW2, altering the interfacial electrostatic landscapes and diminishing the binding affinities by approximately 25 kcal/mol for both KRasGDP·Mg2+ and KRasGTP·Mg2+. BI-3406 forms a hydrogen-bond bridge between SW2 and SOS1 in wild type (WT) KRas, interrupting the interactions among the N-terminal residues of SW2 and SOS1. Moreover, BI-3406 was found here to attenuate the binding affinity of both WT and G12D-mutant KRasGDP·Mg2+ to SOS1, Interestingly, BI-3406 hardly affects the binding affinity of WT KRasGTP·Mg2+, while enhances the binding affinity of G12D-mutant KRasGTP·Mg2+. The change of binding affinity makes the catalytic pocket of SOS1 prefer to KRasGTP·Mg2+ and inhibits the growth of G12D-mutant KRas-driven tumors. These mechanistic insights provide valuable information for designing SOS1-co-targeting inhibitors to potentiate antitumor efficacy against G12D-mutated KRas.