Abstract 746: Activation of EGFR bypass signaling through TGFα overexpression induces acquired resistance to alectinib in ALK-translocated lung cancer cells
Authors: Tani Tetsuo , Yasuda Hiroyuki , Hamamoto Junko , Kuroda Aoi , Arai Daisuke , Ishioka Kota , Ohgino Keiko , Kawada Ichiro , Naoki Katsuhiko , Yuichiro Hayashi , Betsuyaku Tomoko , Soejima Kenzo 
Published: August 2015

Purpose: Alectinib is a highly selective ALK inhibitor and showed a striking efficacy in non-small cell lung cancers (NSCLC) harboring EML4-ALK gene rearrangement. Recently, it was reported that alectinib had an objective response rate of 93.5% in ALK positive patients in a clinical study. However, cancer cells usually acquire resistance to molecular-targeted drugs. The mechanisms of acquired resistance to alectinib are not yet well clarified. The purpose of this study is to clarify the mechanism of acquired resistance to alectinib in ALK translocated lung cancer cells.

Experimental design: We have established alectinib resistant cells (H3122-AR) from H3122, which harbors EML4-ALK rearrangement, by long term exposure to alectinib. The mechanism of acquired resistance to alectinib in H3122-AR cells was evaluated in vitro by cDNA sequencing of ALK tyrosine kinase (TK) domain, or by phospho-RTK array to estimate activation of bypass pathway. The efficacy of combination therapy targeting ALK and EGFR was evaluated in vitro and in vivo by using alectinib, afatinib, or siRNA for EGFR. For in vivo experiment, we used mouse xenograft model. Human tumor samples were evaluated by immunohistochemistry.

Results: No additional mutation in ALK TK domain was found by cDNA sequencing. Phospho-RTK array revealed that the phosphorylation level of EGFR was increased in H3122-AR cells compared with H3122 parental cells, which was subsequently confirmed by western blotting. We found no additional mutation in EGFR TK domain by cDNA sequencing. Among 5 ligands for EGFR, the expression of TGFα was significantly increased in H3122-AR cells compared with H3122 parental cells. We found the phosphorylation level of EGFR was decreased in TGFα gene knockdown, therefore TGFα and EGFR contributed to the acquired resistance to alectinib. The combination therapy targeting ALK and EGFR by using alectinib, afatinib in vitro and in vivo, or siRNA for EGFR in vitro showed efficacy. We found the increased phosphorylation of EGFR after long term ALK-TKI treatment was also detected in human tumor samples.

Conclusion: Activation of EGFR signaling pathway through TGFα overexpression induces acquired resistance to alectinib in ALK-translocated lung cancer cells.

Citation Format: Tetsuo Tani, Hiroyuki Yasuda, Junko Hamamoto, Aoi Kuroda, Daisuke Arai, Kota Ishioka, Keiko Ohgino, Ichiro Kawada, Katsuhiko Naoki, Hayashi Yuichiro, Tomoko Betsuyaku, Kenzo Soejima. Activation of EGFR bypass signaling through TGFα overexpression induces acquired resistance to alectinib in ALK-translocated lung cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 746. doi:10.1158/1538-7445.AM2015-746