Overcoming EGFR T790M and C797S resistance with mutant-selective allosteric inhibitors

Abstract

EGFR tyrosine kinase inhibitors (TKIs) gefitinib, erlotinib and afatinib are approved treatments for non-small cell lung cancers harboring activating mutations in the EGFR kinase ^{1, 2} , but resistance arises rapidly, most frequently due to the secondary T790M mutation within the ATP-site of the receptor. ^{3, 4} Recently developed mutant-selective irreversible inhibitors are highly active against the T790M mutant ^{5, 6} , but their efficacy can be compromised by acquired mutation of C797, the cysteine residue with which they form a key covalent bond ⁷ . All current EGFR TKIs target the ATP-site of the kinase, highlighting the need for therapeutic agents with alternate mechanisms of action. Here we describe rational discovery of EAI045, an allosteric inhibitor that targets selected drug-resistant EGFR mutants but spares the wild type receptor. A crystal structure shows that the compound binds an allosteric site created by the displacement of the regulatory C-helix in an inactive conformation of the kinase. The compound inhibits L858R/T790M-mutant EGFR with low-nanomolar potency in biochemical assays, but as a single agent is not effective in blocking EGFR-driven proliferation in cells due to differential potency on the two subunits of the dimeric receptor, which interact in an asymmetric manner in the active state ⁸ . We observe dramatic synergy of EAI045 with cetuximab, an antibody therapeutic that blocks EGFR dimerization ^{9, 10} , rendering the kinase uniformly susceptible to the allosteric agent. EAI045 in combination with cetuximab is effective in mouse models of lung cancer driven by L858R/T790M EGFR and by L858R/T790M/C797S EGFR, a mutant that is resistant to all currently available EGFR TKIs. More generally, our findings illustrate the utility of purposefully targeting allosteric sites to obtain mutant-selective inhibitors.