Abstract 5648: The characteristics of an acquired dual resistance to EGFR & MET inhibitors in non-small cell lung cancer, harboring active mutation of EGFR

• Published in: Cancer research (Chicago, Ill.) Ill.), 2013-04, Vol.73 (8_Supplement), p.5648-5648
• Main Authors: Yamaoka, Toshimitsu, Ichihashi, Yoko, Murata, Yasunori, Oki, Yasunari, Sugiyama, Tomohide, Ishida, Hiroo, Shirai, Takao, Nakashima, Masanao, Okuda, Kentaro, Hirose, Takashi, Ohnishi, Tsukasa, Ohmori, Tohru
• Format: Article
• Language: English
• ISSN: 0008-5472; 1538-7445
• DOI: 10.1158/1538-7445.AM2013-5648

BACKGROUND. The selective inhibitors of EGFR tyrosine kinase, gefitinib and erlotinib, prevent binding of ATP to the ATP-binding pocket in a competitive manner and then leading to the loss of catalytic activity. They provided dramatic clinical responses and survival benefits for patients with pulmonary adenocarcinoma, harboring somatic mutations in the EGFR kinase domain. Despite initial responses, acquired resistance develops inevitably after a median of ∼10 months. The secondary EGFR mutation, T790M and the amplification of the MET gene are the two main molecular mechanisms responsible for acquired resistance to EGFR-TKI. AIMS. We established the acquired resistant cells to gefitinib from pulmonary adenocarcinoma, PC-9 (15 bp deletion in exon 19) cells with continuous exposure of gefitinib. The MET amplification was determined in this acquired resistant cell line, so named as PC-9MET. In PC-9MET cells, the secondary mutation of T790M was not detected. These cells are sensitive to the combination of EGFR and MET inhibitors. In this study, for understanding the extreme bypass signaling pathways, we challenged to establish the resistant PC-9 cells to both EGFR and MET inhibitors, which named PC-9DR (Dual Resistant). METHODS. For screening new bypass signals, we employed the phospho-receptor tyrosine kinase array. Cell proliferation and signal transduction was determined by MTT assay and Western-blot analysis, respectively. RESULTS. Like as the establishment of PC-9MET by step-wised dose escalation of gefitinib, PC-9MET cells were also treated with the increasing concentration of PHA665752, which is a MET-TKI, under gefitinib 1μM exposure. After about 12 month, PHA665752 was reached to 1μM and finally PC-9DR cells were survived in the condition of gefitinib 1μM and PHA665752 1μM and showed cross-resistance to the combination of erlotinib/crizotinib and also afatinib/crizotinib. Using both direct-sequence and cycleave PCR technology, PC-9DR cells were not detected the T790M in EGFR. The expression levels of EGFR, ErbB3 and MET were not different between PC-9MET and PC-9DR. However, PC-9DR cells were still activated in EGFR, ErbB3 and MET, even exposed with both gefitinib and PHA665752 and the downstream of AKT and ERK were also activated. For identifying the novel bypass survival signals, the phospho-receptor tyrosine kinase array was performed. In PC-9MET and PC-9DR cells, IGF-IRβ, RET and FGFR3 were slightly activated than PC-9, however there were no change