Ibrutinib targets mutant-EGFR kinase with a distinct binding conformation

Area of Science:

• Pharmacology and Molecular Biology
• Oncology Drug Development

Background:

• Ibrutinib, an approved Bruton's tyrosine kinase (BTK) inhibitor for B-cell malignancies, also shows activity against mutant epidermal growth factor receptor (EGFR) kinases.
• Its potential in Non-Small Cell Lung Cancer (NSCLC) driven by EGFR mutations is under clinical investigation.

Purpose of the Study:

• To investigate the binding mechanism and efficiency of Ibrutinib against wild-type (wt) and mutant EGFR.
• To compare Ibrutinib's covalent binding to EGFR with established irreversible EGFR inhibitors.
• To elucidate the structural basis for Ibrutinib's interaction with EGFR and inform future drug design.

Main Methods:

• Utilized engineered isogenic BaF3 cell lines expressing wild-type and mutant EGFR.
• Performed washing-out experiments to assess covalent binding efficiency.
• Conducted biochemical binding affinity assays (Kd measurements) for EGFR L858R/T790M.
• Determined the X-ray crystal structure of EGFR (T790M) in complex with Ibrutinib.

Main Results:

• Confirmed Ibrutinib's irreversible binding to EGFR wt/mutant via Cys797.
• Demonstrated significantly less efficient covalent binding and lower biochemical affinity (Kd: 0.18 μM) compared to WZ4002 (Kd: 0.074 μM).
• Revealed a unique DFG-in/c-Helix-out inactive binding conformation of EGFR (T790M) with Ibrutinib.

Conclusions:

• Ibrutinib's unique binding conformation partially explains its less efficient covalent inhibition of EGFR.
• These findings suggest that sustained effective concentrations of Ibrutinib may be necessary for maximal efficacy in EGFR-driven NSCLC.
• Provides structural insights for developing more potent irreversible EGFR inhibitors.