Patients whose lung adenocarcinomas harbor specific mutations within the exons encoding the tyrosine kinase domain of the epidermal growth factor receptor (EGFR) frequently experience clinical and radiographic responses to the selective EGFR tyrosine kinase inhibitors (TKIs), gefitinib (Iressa) or erlotinib (Tarceva). However, after about one year, these patients develop progression of disease. We and others have shown that in addition to primary drug-sensitive EGFR mutations, tumor cells from about half of patients with such """"""""acquired resistance"""""""" contain a second mutation in the EGFR kinase domain. From crystal structure analyses, the resulting amino acid change (T790M) is predicted to block binding of drug to the ATP pocket via steric clash resulting from introduction of the bulky methionine residue. The T790M mutation is analogous to common secondary mutations in other kinases (e.g. BCR-ABL, T315I) found in patients with acquired resistance to another kinase inhibitor, imatinib (Gleevec). The overall goals of this revised proposal are to use human tumor specimens, genetically engineered mice, and various molecular and biochemical techniques to enhance knowledge about the subset of EGFR- mutant harboring lung adenocarcinomas that develop acquired resistance to gefitinib and erlotinib. An improved understanding of acquired resistance to these agents will hopefully allow us to both treat progressive disease and suppress the development of acquired resistance. Thus, we aim to: 1) Determine the mutational status of the EGFR kinase domain in tumor cells from patients with (up to 80 individuals) with acquired resistance to gefitinib or erlotinib, and establish how newly """"""""acquired"""""""" mutations affect biochemical properties of EGFR, such as kinase activity and sensitivity to EGFR TKIs, and 2) Characterize transgenic animals carrying tetracycline-inducible transgenes that encode the common T790M resistance mutation by itself and in the context of a drug-sensitive EGFR mutation (the exon 21 amino acid substitution L858R or the exon 19 deletion L747-S752), comparing them to mice that express a drug-sensitive EGFR mutation alone.
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