Abstract 3160: Sustained MAPK activation as a mechanism of resistance to osimertinib plus selumetinib in models of EGFR-mutant cancer
Authors: Westover David , Meador Catherine B. , Ichihara Eiki , Byrd Hayden F. , Eberlein Cath , Yan Yingjun , Cross Darren A. , Lovly Christine M. 
Published: July 2017

Abstract
INTRODUCTION: Osimertinib is a third-generation, mutant-selective epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) recently approved for the treatment of T790M-positive EGFR-mutant lung cancer. The mutant-selective nature of osimertinib improves its tolerability by limiting its inhibition of wild-type EGFR compared to first- and second-generation EGFR inhibitors, making it a good candidate for rationally-designed combination therapies. One such combination, osimertinib plus the MEK inhibitor selumetinib (AZD6244; ARRY-142886), is currently being studied as part of the phase 1b TATTON trial (NCT02143466). Previous work by our group demonstrated that selumetinib may delay or reverse resistance to osimertinib in some cases; however, we anticipate that resistance to this combination will ultimately develop in patients. Here, we investigate acquired resistance to osimertinib and selumetinib combination therapy in vitro and in vivo.
DESIGN: In cells sensitive to osimertinib plus selumetinib (hereafter referred to as combination-sensitive cells), ERK phosphorylation was monitored via Western blot over the course of a 7-day treatment with 200 nM osimertinib plus 1 µM selumetinib. To detect RAS-GTP, an active RAS precipitation was performed in combination-sensitive and combination-resistant cell lines following treatment with osimertinib. In combination-resistant cells, pharmacologic inhibitors of various MAPK pathway components were used to assess whether they could restore potency. Western blot analysis was used to confirm on-target effects. Lastly, an analysis of differences in gene expression between four isogenic sets of combination-sensitive and combination-resistant cell lines is currently ongoing.
RESULTS: In cell lines that are sensitive to growth inhibition by the combination of osimertinib plus selumetinib, we observed complete re-activation of the MAPK pathway after 5 days of continuous exposure to both inhibitors. Additionally, RAS activity was elevated in combination-resistant cell lines, and these cells remained sensitive to the ERK inhibitor SCH772984. Likewise, the addition of a pan-RAF inhibitor restored the growth inhibitory effects of osimertinib plus selumetinib in combination-resistant cells, suggesting that incomplete inhibition of MAPK is responsible for resistance in these cells. Furthermore, an alternative MEK inhibitor, trametinib, was efficacious in combination with osimertinib in cell lines that were resistant to osimertinib plus selumetinib.
CONCLUSION: These data identify a potential mechanism of resistance to a combination therapy that is currently being tested in the clinic. Specifically, these data demonstrate that re-activation of the MAPK pathway is a mechanism of resistance to osimertinib plus selumetinib and that this pathway is still targetable in combination-resistant cells.
Citation Format: David Westover, Catherine B. Meador, Eiki Ichihara, Hayden F. Byrd, Cath Eberlein, Yingjun Yan, Darren A. Cross, Christine M. Lovly. Sustained MAPK activation as a mechanism of resistance to osimertinib plus selumetinib in models of EGFR-mutant cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3160. doi:10.1158/1538-7445.AM2017-3160