Abstract
Abstract Objectives: The multitargeted antifolate pemetrexed is approved as a first-line or second-line treatment of nonsquamous nonsmall cell lung cancer. The expression of the thymidylate synthase (TS) gene may mediate resistance to pemetrexed. In this study, we investigated whether the pemetrexed in combination with SAHA enhances the cytotoxicity of squamous carcinoma cells of lung. Methods: Squamous carcinoma cells of lung (NCI-H157, HCC-95 and HCC-1588) were exposed to 10-3M to 10-7M of pemetrexed with various HDAC inhibitors and their cytotoxicities were analyzed using the colorimetric assay. Cell cycle and apoptosis analyses were evaluated by flow cytometry. The mRNA and cellular protein levels were detected by RT-PCR and western blot assay. Results: After NCI-H157 was exposed to SAHA, TS mRNA and protein expression levels were strongly suppressed by time and dose-dependent manner, respectively. Additionally, although pemetrexed activated phosphorylation of AMPK-α, secondary target (AICAR) of pemetrexed, co-treatment of pemetrexed and SAHA decreased phosphorylation of AMPK-α and mTOR pathway. NCI-H157, HCC-95, and HCC-1588 treated with sequential SAHA (24h) followed by SAHA plus pemetrexed (72h) represented more synergistic cytotoxicities than those treated with simultaneous 96h exposure combination (p<0.05). This sequential treatment was significantly associated with TS inhibition as well as apoptosis of squamous carcinoma cells. While pemetrexed increased expressions of p21, PARP, and cleaved caspase 3 and reduced expression of bcl-2, the combination of pemetrexed and SAHA additionally increased the pro-apoptotic molecules. Sequential treatment of SAHA followed by pemetrexed resulted in 10-fold and 1.6-fold higher sub-G1 portions than control and pemetrexed alone groups, respectively in NCI-H157 cell. In particular, the sequential administration produced more apoptotic index as compared to pemetrexed alone in NCI-H157 cell (27% vs 8%). Conclusion: Our results demonstrate SAHA reduces TS expression and sequential treatment with pemetrexed has a synergistic cytotoxicity in squamous cell carcinoma of lung. Sequential treatment of SAHA and pemetrexed might be a new strategy to overcome pemetrexed resistance in patients with squamous cell carcinoma of lung. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3500. doi:10.1158/1538-7445.AM2011-3500
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