title: Structural basis for the altered drug sensitivities of non-small cell lung cancer-associated mutants of human epidermal growth factor receptor.
abstract: The epidermal growth factor receptor (EGFR) has an essential role in multiple signaling pathways, including cell proliferation and migration, through extracellular ligand binding and subsequent activation of its intracellular tyrosine kinase (TK) domain. The non-small cell lung cancer (NSCLC)-associated EGFR mutants, L858R and G719S, are constitutively active and oncogenic. They display sensitivity to TK inhibitors, including gefitinib and erlotinib. In contrast, the secondary mutation of the gatekeeper residue, T790M, reportedly confers inhibitor resistance on the oncogenic EGFR mutants. In this study, our biochemical analyses revealed that the introduction of the T790M mutation confers gefitinib resistance on the G719S mutant. The G719S/T790M double mutant has enhanced activity and retains high gefitinib-binding affinity. The T790M mutation increases the ATP affinity of the G719S mutant, explaining the acquired drug resistance of the double mutant. Structural analyses of the G719S/T790M double mutant, as well as the wild type and the G719S and L858R mutants, revealed that the T790M mutation stabilizes the hydrophobic spine of the active EGFR-TK conformation. The Met790 side chain of the G719S/T790M double mutant, in the apo form and gefitinib- and AMPPNP-bound forms, adopts different conformations that explain the accommodation of these ligands. In the L858R mutant structure, the active-site cleft is expanded by the repositioning of Phe723 within the P-loop. Notably, the introduction of the F723A mutation greatly enhanced the gefitinib sensitivity of the wild-type EGFR in vivo, supporting our hypothesis that the expansion of the active-site cleft results in enhanced gefitinib sensitivity. Taken together, our results provide a structural basis for the altered drug sensitivities caused by distinct NSCLC-associated EGFR mutations.
authors.affiliation: RIKEN Systems and Structural Biology Center, Yokohama Institute, Yokohama, Japan.
authors.forename: S
authors.initials: S
authors.lastname: Yoshikawa
authors.forename: M
authors.initials: M
authors.lastname: Kukimoto-Niino
authors.forename: L
authors.initials: L
authors.lastname: Parker
authors.forename: N
authors.initials: N
authors.lastname: Handa
authors.forename: T
authors.initials: T
authors.lastname: Terada
authors.forename: T
authors.initials: T
authors.lastname: Fujimoto
authors.forename: Y
authors.initials: Y
authors.lastname: Terazawa
authors.forename: M
authors.initials: M
authors.lastname: Wakiyama
authors.forename: M
authors.initials: M
authors.lastname: Sato
authors.forename: S
authors.initials: S
authors.lastname: Sano
authors.forename: T
authors.initials: T
authors.lastname: Kobayashi
authors.forename: T
authors.initials: T
authors.lastname: Tanaka
authors.forename: L
authors.initials: L
authors.lastname: Chen
authors.forename: Z-J
authors.initials: ZJ
authors.lastname: Liu
authors.forename: B-C
authors.initials: BC
authors.lastname: Wang
authors.forename: M
authors.initials: M
authors.lastname: Shirouzu
authors.forename: S
authors.initials: S
authors.lastname: Kawa
authors.forename: K
authors.initials: K
authors.lastname: Semba
authors.forename: T
authors.initials: T
authors.lastname: Yamamoto
authors.forename: S
authors.initials: S
authors.lastname: Yokoyama
mesh_terms: D000970:Antineoplastic Agents / Q000627:therapeutic use*; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy* / Q000235:genetics; D004354:Drug Screening Assays, Antitumor; D066246:ErbB Receptors / Q000737:chemistry / Q000235:genetics*; D000077156:Gefitinib; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000235:genetics; D009154:Mutation; D011487:Protein Conformation; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D011799:Quinazolines / Q000627:therapeutic use*
chemical_list: D000970:Antineoplastic Agents; D011799:Quinazolines; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D000077156:Gefitinib