title: Therapeutic strategies to overcome crizotinib resistance in non-small cell lung cancers harboring the fusion oncogene EML4-ALK.
abstract: The echinoderm microtubule-associated protein-like 4 (EML4)-anaplastic lymphoma kinase (ALK) fusion oncogene represents a molecular target in a small subset of non-small cell lung cancers (NSCLCs). This fusion leads to constitutive ALK activation with potent transforming activity. In a pivotal phase 1 clinical trial, the ALK tyrosine kinase inhibitor (TKI) crizotinib (PF-02341066) demonstrated impressive antitumor activity in the majority of patients with NSCLC harboring ALK fusions. However, despite these remarkable initial responses, cancers eventually develop resistance to crizotinib, usually within 1 y, thereby limiting the potential clinical benefit. To determine how cancers acquire resistance to ALK inhibitors, we established a model of acquired resistance to crizotinib by exposing a highly sensitive EML4-ALK-positive NSCLC cell line to increasing doses of crizotinib until resistance emerged. We found that cells resistant to intermediate doses of crizotinib developed amplification of the EML4-ALK gene. Cells resistant to higher doses (1 μM) also developed a gatekeeper mutation, L1196M, within the kinase domain, rendering EML4-ALK insensitive to crizotinib. This gatekeeper mutation was readily detected using a unique and highly sensitive allele-specific PCR assay. Although crizotinib was ineffectual against EML4-ALK harboring the gatekeeper mutation, we observed that two structurally different ALK inhibitors, NVP-TAE684 and AP26113, were highly active against the resistant cancer cells in vitro and in vivo. Furthermore, these resistant cells remained highly sensitive to the Hsp90 inhibitor 17-AAG. Thus, we have developed a model of acquired resistance to ALK inhibitors and have shown that second-generation ALK TKIs or Hsp90 inhibitors are effective in treating crizotinib-resistant tumors harboring secondary gatekeeper mutations.
authors.affiliation: Massachusetts General Hospital Cancer Center, Boston, MA 02129, USA.
authors.forename: Ryohei
authors.initials: R
authors.lastname: Katayama
authors.forename: Tahsin M
authors.initials: TM
authors.lastname: Khan
authors.forename: Cyril
authors.initials: C
authors.lastname: Benes
authors.forename: Eugene
authors.initials: E
authors.lastname: Lifshits
authors.forename: Hiromichi
authors.initials: H
authors.lastname: Ebi
authors.forename: Victor M
authors.initials: VM
authors.lastname: Rivera
authors.forename: William C
authors.initials: WC
authors.lastname: Shakespeare
authors.forename: A John
authors.initials: AJ
authors.lastname: Iafrate
authors.forename: Jeffrey A
authors.initials: JA
authors.lastname: Engelman
authors.forename: Alice T
authors.initials: AT
authors.lastname: Shaw
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy* / Q000235:genetics; D045744:Cell Line, Tumor; D000077547:Crizotinib; D017931:DNA Primers / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D017404:In Situ Hybridization, Fluorescence; D051379:Mice; D008819:Mice, Nude; D009154:Mutation / Q000235:genetics; D015514:Oncogene Proteins, Fusion / Q000037:antagonists & inhibitors / Q000235:genetics*; D009943:Organophosphorus Compounds / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011720:Pyrazoles / Q000494:pharmacology* / Q000627:therapeutic use; D011725:Pyridines / Q000494:pharmacology* / Q000627:therapeutic use; D011743:Pyrimidines / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D016019:Survival Analysis; D014162:Transfection
chemical_list: D000970:Antineoplastic Agents; D017931:DNA Primers; C522504:EML4-ALK fusion protein, human; C516714:NVP-TAE684; D015514:Oncogene Proteins, Fusion; D009943:Organophosphorus Compounds; D011720:Pyrazoles; D011725:Pyridines; D011743:Pyrimidines; D034741:RNA, Small Interfering; D000077547:Crizotinib; C000598580:brigatinib