Mechanistic insights into KRASG12D inhibitor binding revealed by molecular dynamics simulations of multiple crystal structures

Abstract

The discovery of selective and potent KRAS^G12D inhibitors remains a critical priority in oncology drug development. Here, we performed comparative all-atom molecular dynamics (MD) simulations on four KRAS^G12D–inhibitor complexes (PDB IDs: 7RPZ, 7RT2, 7EWB, 7EW9) spanning a wide affinity range (IC₅₀ = 2 nM–14 µM). By integrating high-resolution crystallographic data with ex-tensive all-atom MD simulations, we aimed to elucidate the structural and dynamic determinants that differentiate high- and low-affinity inhibitor binding. Alchemical free-energy calculations yielded ΔG values of −11.3 to −6.2 kcal mol⁻¹, which showed a strong correlation with experimental pIC₅₀ (R² = 0.92). Per-residue energy decomposition revealed five dominant polar interaction hotspots (D12 ≈ −34, G60 ≈ −13, E62 ≈ −20, D69 ≈ −17, and D92 ≈ −6 kcal mol⁻¹) driving stable bind- ing, whereas weak inhibitors exhibited markedly reduced contributions at these residues. Structural dynamics analysis further showed that strong binders maintained compact binding pockets (RMSD: 1.8–2.2 Å) and reduced ligand flexibility, whereas weak binders sampled expanded and less stable conformations (RMSD: 2.7–3.4 Å). These findings delineate the structural and energetic determinants underlying KRAS^G12D inhibitor potency and provide quantitative guidelines for the rational design of next-generation KRAS^G12D inhibitors.