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Effects of oncogenic mutations on the conformational free-energy landscape of EGFR kinase.

ABSTRACT: Activating mutations in the epidermal growth factor receptor (EGFR) tyrosine kinase are frequently found in many cancers. It has been suggested that changes in the equilibrium between its active and inactive conformations are linked to its oncogenic potential. Here, we quantify the effects of some of the most common single (L858R and T790M) and double (T790M-L858R) oncogenic mutations on the conformational free-energy landscape of the EGFR kinase domain by using massive molecular dynamics simulations together with parallel tempering, metadynamics, and one of the best force-fields available. Whereas the wild-type EGFR catalytic domain monomer is mostly found in an inactive conformation, our results show a clear shift toward the active conformation for all of the mutants. The L858R mutation stabilizes the active conformation at the expense of the inactive conformation and rigidifies the ?C-helix. The T790M gatekeeper mutant favors activation by stabilizing a hydrophobic cluster. Finally, T790M with L858R shows a significant positive epistasis effect. This combination not only stabilizes the active conformation, but in nontrivial ways changes the free-energy landscape lowering the transition barriers.

SUBMITTER: Sutto L

PROVIDER: S-EPMC3696764 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature

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Effects of oncogenic mutations on the conformational free-energy landscape of EGFR kinase.

Sutto Ludovico L Gervasio Francesco Luigi FL

Proceedings of the National Academy of Sciences of the United States of America 20130610 26

Activating mutations in the epidermal growth factor receptor (EGFR) tyrosine kinase are frequently found in many cancers. It has been suggested that changes in the equilibrium between its active and inactive conformations are linked to its oncogenic potential. Here, we quantify the effects of some of the most common single (L858R and T790M) and double (T790M-L858R) oncogenic mutations on the conformational free-energy landscape of the EGFR kinase domain by using massive molecular dynamics simula ...[more]

PMID: 23754386

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Project description:BackgroundEpidermal growth factor receptor (EGFR) mutations are present in the majority of patients with non-small cell lung cancer (NSCLC) responsive to the EGFR tyrosine kinase inhibitors (TKIs) gefitinib or erlotinib. These EGFR-dependent tumors eventually become TKI resistant, and the common secondary T790M mutation accounts for half the tumors with acquired resistance to gefitinib. However, the key proapoptotic proteins involved in TKI-induced cell death and other secondary mutations involved in resistance remain unclear. The objective of this study was to identify the mechanism of EGFR TKI-induced apoptosis and secondary resistant mutations that affect this process.Methods and findingsTo study TKI-induced cell death and mechanisms of resistance, we used lung cancer cell lines (with or without EGFR mutations), Ba/F3 cells stably transfected with EGFR mutation constructs, and tumor samples from a gefitinib-resistant patient. Here we show that up-regulation of the BH3-only polypeptide BIM (also known as BCL2-like 11) correlated with gefitinib-induced apoptosis in gefitinib-sensitive EGFR-mutant lung cancer cells. The T790M mutation blocked gefitinib-induced up-regulation of BIM and apoptosis. This blockade was overcome by the irreversible TKI CL-387,785. Knockdown of BIM by small interfering RNA was able to attenuate apoptosis induced by EGFR TKIs. Furthermore, from a gefitinib-resistant patient carrying the activating L858R mutation, we identified a novel secondary resistant mutation, L747S in cis to the activating mutation, which attenuated the up-regulation of BIM and reduced apoptosis.ConclusionsOur results provide evidence that BIM is involved in TKI-induced apoptosis in sensitive EGFR-mutant cells and that both attenuation of the up-regulation of BIM and resistance to gefitinib-induced apoptosis are seen in models that contain the common EGFR T790M and the novel L747S secondary resistance mutations. These findings also suggest that induction of BIM may have a role in the treatment of TKI-resistant tumors.

Dataset Information

Effects of oncogenic mutations on the conformational free-energy landscape of EGFR kinase.

Publications

Effects of oncogenic mutations on the conformational free-energy landscape of EGFR kinase.

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