Mutations of EGFR kinase are linked to cancers such as non-small cell lung cancer (NSCLC), which as of today has one of the lowest survival rates. Competitive non-covalent EGFR inhibitors, such as gefitinib and erlotinib, which bind to the kinase domain hinge region, have been approved by FDA as the first-line NSCLC therapeutics, resulting in longer median patients’ survival. The cancer-linked L858R mutation in EGFR is particularly responsive to gefitinib and erlotinib. However, the long-term effectiveness of EGFR inhibitors is often limited because of acquired drug resistance. It is thus essential to identify the molecular mechanisms underlying the resistance, in order to develop alternative therapeutic strategies. One of the common resistance mechanisms is acquired “gatekeeper” mutation (T790M). Although its resistance was initially attributed to steric effects, recent experimental studies have demonstrated that differences in the binding affinities of inhibitors to T790M and L858RT790M mutants are more a result of the increased sensitivity of these mutants to ATP than to a decrease in the affinity to inhibitors. In addition, recent studies indicate altered redox state of C797 thiol upon the mutations, which is likely to contribute to the resistance mechanism. In this study, we combined atomistic molecular dynamics simulations with the pKa calculations to determine how the T790M, L858R, and L858RT790M mutation impacts the intrinsic dynamics and redox states of C797, relatively to wtEGFR. To assess the role of solvation effects on the observed behaviour and to improve the sampling on the generated ensembles, two different water models were used: established TIP3P model, and improved 3-point CAIP3P model, recently developed in our group. Detailed structural and energetic analysis provided novel information on how intrinsic protein dynamics tunes the binding affinity of non-covalent ligands (ATP, inhibitors). This helps to explain the observed resistance of T790M and L858RT790M mutants to non-covalent inhibitors, compared to L858R and wtEGFR. Simulations in CAIP3P water allowed for more exhaustive sampling of EGFR conformational space and helped us to explain differences in effects of EGFR dimerisation on the ligand binding affinity between mutants and wtEGFR. In addition, we have observed that EGFR mutations exert diverse effects on the C797 oxidation and reactivity to covalent EGFR inhibitors such as osimertinib relatively to wtEGFR.

Investigating the effects of mutations and post-translational modifications on intrinsic EGFR dynamics using molecular simulations in improved solvation model

2019 Fall National Meeting

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