Abstract A224: CS-7017, a selective PPARγ agonist, attenuates the TGF-β/Smad-mediated enhanced cell motility of EGFR-tyrosine kinase inhibitor-resistant non-small cell lung cancer cells.
Authors: Serizawa Masakuni , Watanabe Masaru , Suzuki Junko , Naruoka Akane , Murakami Haruyasu , Yamamoto Nobuyuki , Koh Yasuhiro 
Published: June 2012

Abstract
Background: Peroxisome proliferator-activated receptor gamma (PPARγ) agonists have been shown to possess antitumor property in preclinical models of human cancers including non-small cell lung cancer (NSCLC). CS-7017, a novel selective PPARγ agonist of thiazolidinedione class, has been reported to induce morphological change and cellular differentiation as well as its antiproliferative effects. A phase II clinical trial of CS-7017 in combination with epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) erlotinib is currently ongoing in advanced NSCLC patients. Though EGFR-TKIs are effective for NSCLCs with EGFR-activating mutations, the majority of responders eventually develop acquired resistance. We have previously reported that EGFR-TKI-resistant NSCLC cells acquire enhanced cell motility via activation of TGF-β/Smad pathway. In this study, we aim to investigate the effects of CS-7017 in EGFR-TKI-resistant NSCLC cells with a focus on cell motility. Materials and Methods: Erlotinib-resistant PC-9/ER and gefitinib-resistant PC-9/ZD cells, established from PC-9 NSCLC cells with EGFR-activating mutation, were used for this study and constitutive PI3K/Akt activation and secondary T790M mutation have been previously shown to account for the acquired resistance, respectively. Growth-inhibitory effect of CS-7017 was evaluated by MTT assay. Wound closure assay and transwell assay were performed to evaluate the cell motility. Secretion level of TGF-β ligands was quantified by ELISA. Transcriptional activity of Smad and PPARγ was measured by luciferase assay. Effects of CS-7017 on the molecules relevant to EGFR, TGF-β and PPARγ pathways were observed by immunoblot analysis. Results: Both PC-9/ER and PC-9/ZD resistant cells showed enhanced cell motility compared with PC-9 parental cells. Significantly higher amount of secreted TGF-β2 was detected in both resistant cells than in PC-9 cells and induction of phosphorylated Smad2 was observed in the resistant cell lines. Activation of TGF-β/Smad pathway by the increased TGF-β2 secretion was suggested to confer the enhanced cell motility of the resistant cells. CS-7017 showed significant inhibitory effect on the cell motility of the resistant cells but showed no growth-inhibitory effect. Moreover, combined treatment with CS-7017 and erlotinib resulted in better inhibition of the cell motility of PC-9/ER cells. CS-7017 treatment was found to suppress the TGF-β2 secretion significantly and attenuated Smad2 phosphorylation was observed in the resistant cells. Smad-mediated transcriptional activity was also suppressed by CS-7017 treatment. CS-7017 treatment induced PPARγ-mediated transcriptional activity in PC-9 cells but not in PC-9/ER cells, suggesting that the inhibitory effect of CS-7017 on the cell motility of PC-9/ER cells may not be due to direct activation of PPARγ signaling by CS-7017. Conclusions: These results imply that CS-7017 may serve as a potential therapeutic agent to prevent metastasis in NSCLC patients who develop acquired resistance to EGFR-TKIs regardless of the types of resistance mechanisms. The mechanism of TGF-β2 downregulation by CS-7017 needs to be further elucidated.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr A224.