Differential protein stability of EGFR mutants determines responsiveness to tyrosine kinase inhibitors
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Paramita Ray1, Yee Sun Tan1, Vishal Somnay1, Ranjit Mehta1, Merna Sitto1, Aarif Ahsan1,4, Shyam Nyati1, John P. Naughton1,5, Alexander Bridges2, Lili Zhao3, Alnawaz Rehemtulla1, Theodore S. Lawrence1, Dipankar Ray1, Mukesh K. Nyati1
1Department of Radiation Oncology, University of Michigan, Ann Arbor, MI 48109, USA
2School of Pharmacy, University of Michigan, Ann Arbor, MI 48109, USA
3Department of Biostatistics, University of Michigan, Ann Arbor, MI 48109, USA
4Current address: Oncology Research Unit East, Pfizer, Pearl River, NY 10965, USA
5Current address: Department of Otorhinolaryngology-Head and Neck Surgery, Albert Einstein College of Medicine, Montefiore Medical Center, Bronx, NY 10467, USA
Mukesh K. Nyati, email: email@example.com
Dipankar Ray, email: firstname.lastname@example.org
Theodore S. Lawrence, email: email@example.com
Keywords: EGFR, erlotinib, protein stability, ubiquitination, TKI sensitivity
Received: April 29, 2016 Accepted: August 25, 2016 Published: September 06, 2016
Non-small cell lung cancer (NSCLC) patients carrying specific EGFR kinase activating mutations (L858R, delE746-A750) respond well to tyrosine kinase inhibitors (TKIs). However, drug resistance develops within a year. In about 50% of such patients, acquired drug resistance is attributed to the enrichment of a constitutively active point mutation within the EGFR kinase domain (T790M). To date, differential drug-binding and altered ATP affinities by EGFR mutants have been shown to be responsible for differential TKI response. As it has been reported that EGFR stability plays a role in the survival of EGFR driven cancers, we hypothesized that differential TKI-induced receptor degradation between the sensitive L858R and delE746-A750 and the resistant T790M may also play a role in drug responsiveness. To explore this, we have utilized an EGFR-null CHO overexpression system as well as NSCLC cell lines expressing various EGFR mutants and determined the effects of erlotinib treatment. We found that erlotinib inhibits EGFR phosphorylation in both TKI sensitive and resistant cells, but the protein half-lives of L858R and delE746-A750 were significantly shorter than L858R/T790M. Third generation EGFR kinase inhibitor (AZD9291) inhibits the growth of L858R/T790M-EGFR driven cells and also induces EGFR degradation. Erlotinib treatment induced polyubiquitination and proteasomal degradation, primarily in a c-CBL-independent manner, in TKI sensitive L858R and delE746-A750 mutants when compared to the L858R/T790M mutant, which correlated with drug sensitivity. These data suggest an additional mechanism of TKI resistance, and we postulate that agents that degrade L858R/T790M-EGFR protein may overcome TKI resistance.
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