No abstract

Non–small cell lung cancers (NSCLCs) that harbor mutations within the epidermal growth factor receptor (EGFR) gene are sensitive to the tyrosine kinase inhibitors (TKIs) gefitinib and erlotinib. Unfortunately, all patients treated with these drugs will acquire resistance, most commonly as a result of a secondary mutation within EGFR (T790M). Because both drugs were developed to target wild-type EGFR, we hypothesized that current dosing schedules were not optimized for mutant EGFR or to prevent resistance. To investigate this further, we developed isogenic TKI-sensitive and TKI-resistant pairs of cell lines that mimic the behavior of human tumors. We determined that the drug-sensitive and drug-resistant EGFR-mutant cells exhibited differential growth kinetics, with the drug-resistant cells showing slower growth. We incorporated these data into evolutionary mathematical cancer models with constraints derived from clinical data sets. This modeling predicted alternative therapeutic strategies that could prolong the clinical benefit of TKIs against EGFR-mutant NSCLCs by delaying the development of resistance.

Genome-wide analyses determined previously that the receptor tyrosine kinase (RTK) EPHA2 is commonly

Despite the success of treating EGFR mutant lung cancer patients with EGFR tyrosine kinase inhibitors (TKIs), all patients eventually acquire resistance to these therapies. Although various resistance mechanisms have been described, there are currently no FDA-approved therapies that target alternative mechanisms to treat lung tumors with acquired resistance to first-line EGFR TKI agents. Here we found that EPHA2 is overexpressed in EGFR TKI resistant tumor cells. Loss of EPHA2 reduced the viability of erlotinib resistant tumor cells harboring EGFRT790M mutations in vitro and inhibited tumor growth and progression in an inducible EGFRL858R+T790M mutant lung cancer model in vivo. Targeting EPHA2 in erlotinib resistant cells decreased S6K1-mediated phosphorylation of cell death agonist BAD, resulting in reduced tumor cell proliferation and increased apoptosis. Furthermore, pharmacologic inhibition of EPHA2 by the small molecule inhibitor, ALW-II-41-27, decreased both survival and proliferation of erlotinib resistant tumor cells and inhibited tumor growth in vivo. ALW-II-41-27 was also effective in decreasing viability of cells with acquired resistance to the third generation EGFR TKI, AZD9291. Collectively, these data define a role for EPHA2 in the maintenance of cell survival of TKI resistant, EGFR mutant lung cancer and indicate that EPHA2 may serve as a useful therapeutic target in TKI resistant tumors.

BackgroundCancer genome sequencing efforts recently identified EPHA3, which encodes the EPHA3 receptor tyrosine kinase, as one of the most frequently mutated genes in lung cancer. Although receptor tyrosine kinase mutations often drive oncogenic conversion and tumorigenesis, the oncogenic potential of the EPHA3 mutations in lung cancer remains unknown. MethodsWe used immunoprecipitation, western blotting, and kinase assays to determine the activity and signaling of mutant EPHA3 receptors. A mutation-associated gene signature was generated from one large dataset, mapped to another training dataset with survival information, and tested in a third independent dataset. EPHA3 expression levels were determined by quantitative reverse transcription-polymerase chain reaction in paired normal-tumor clinical specimens and by immunohistochemistry in human lung cancer tissue microarrays. We assessed tumor growth in vivo using A549 and H1299 human lung carcinoma cell xenografts in mice (n = 7-8 mice per group). Tumor cell proliferation was measured by bromodeoxyuridine incorporation and apoptosis by multiple assays. All P values are from two-sided tests. ResultsAt least two cancer-associated EPHA3 somatic mutations functioned as dominant inhibitors of the normal (wild type) EPHA3 protein. An EPHA3 mutation-associated gene signature that was associated with poor patient survival was identified. Moreover, EPHA3 gene copy numbers and/or expression levels were decreased in tumors from large cohorts of patients with lung cancer (eg, the gene was deleted in 157 of 371 [42%] primary lung adenocarcinomas). Reexpression of wild-type EPHA3 in human lung cancer lines increased apoptosis by suppression of AKT activation in vitro and inhibited the growth of tumor xenografts (eg, for H1299 cells, mean tumor volume with wild-type EPHA3 = 437.4 mm 3 vs control = 774.7 mm 3 , P < .001). Tumor-suppressive effects of wild-type EPHA3 could be overridden in trans by dominant negative EPHA3 somatic mutations discovered in patients with lung cancer. ConclusionCancer-associated EPHA3 mutations attenuate the tumor-suppressive effects of normal EPHA3 in lung cancer.