Epidermal Growth Factor Receptors Harboring Kinase Domain Mutations Associate with the Heat Shock Protein 90 Chaperone and Are Destabilized following Exposure to Geldanamycins

Abstract

Somatic mutations in the kinase domain of the epidermal growth factor receptor (EGFR), including L858R and exon 19 deletions, underlie responsiveness to gefitinib and erlotinib in non-small cell lung cancer (NSCLC). Acquired resistance to these tyrosine kinase inhibitors is in some cases mediated by a second mutation, T790M. Ansamycin antibiotics, such as geldanamycin, potently inhibit heat shock protein 90 (Hsp90), promoting ubiquitin-mediated degradation of oncogenic kinases that require the chaperone for proper conformational folding. Here, we show that L858R and deletion mutant EGFR proteins found in NSCLC interact with the chaperone and are sensitive to degradation following Hsp90 inhibition. In NIH/3T3 cells expressing either wild-type or mutant EGFR, diminution of expression of both L858R and EGFR delL747-S752, P753S occurred following exposure to 50 nmol/L geldanamycin over 24 hours, whereas partial diminution of wild-type EGFR required a minimum of 200 nmol/L drug. In time course experiments, mutant EGFR expression was depleted after only 4 hours of exposure to 1 micromol/L geldanamycin, whereas diminution of wild-type EGFR was less substantial and seen only following 12 hours. Similarly, EGFR proteins in NSCLC cell lines harboring EGFR mutations, including NCI-H1650, NCI-H3255, and NCI-H1975, were also more sensitive to geldanamycin-induced degradation compared with the protein in wild-type cells. Exposure of EGFR-mutant cell lines to geldanamycin induced marked depletion of phospho-Akt and cyclin D1 as well as apoptosis. These data suggest mutational activation of EGFR is associated with dependence on Hsp90 for stability and that Hsp90 inhibition may represent a novel strategy for the treatment of EGFR-mutant NSCLC.