Combination of AT-101/cisplatin overcomes chemoresistance by inducing apoptosis and modulating epigenetics in human ovarian cancer cells Burcak Karaca • Harika Atmaca • Emir Bozkurt • Asli Kisim • Selim Uzunoglu • Bu ¨ lent Karabulut • Canfeza Sezgin • Ulus Ali Sanli • Ruchan Uslu Received: 26 July 2012 / Accepted: 18 December 2012 / Published online: 27 December 2012 Ó Springer Science+Business Media Dordrecht 2012 Abstract We investigated the effects of AT-101/cisplatin combination treatment on the expression levels of apoptotic proteins and epigenetic events such as DNA methyltrans- ferase (DNMT) and histone deacetylase (HDAC) enzyme activities in OVCAR-3 and MDAH-2774 ovarian cancer cells. XTT cell viability assay was used to evaluate cyto- toxicity. For showing apoptosis, both DNA Fragmentation and caspase 3/7 activity measurements were performed. The expression levels of apoptotic proteins were assessed by human apoptosis antibody array. DNMT and HDAC activities were evaluated by ELISA assay and mRNA levels of DNMT1 and HDAC1 genes were quantified by qRT- PCR. Combination of AT-101/cisplatin resulted in strong synergistic cytotoxicity and apoptosis in human ovarian cancer cells. Combination treatment reduced some pivotal anti-apoptotic proteins such as Bcl-2, HIF-1A, cIAP-1, XIAP in OVCAR-3 cells, whereas p21, Bcl-2, cIAP-1, HSP27, Clusterin and XIAP in MDAH-2774 cells. Among the pro-apoptotic proteins, Bad, Bax, Fas, phospho-p53 (S46), Cleaved caspase-3, SMAC/Diablo, TNFR1 and Cytochrome c were induced in OVCAR-3 cells, whereas, Bax, TRAILR2, FADD, p27, phospho-p53 (S46), Cleaved caspase-3, Cytochrome c, SMAC/Diablo and TNFR1 were induced in MDAH-2774 cells. Combination treatment also inhibited both DNMT and HDAC activities and also mRNA levels in both ovarian cancer cells. AT-101 exhibits great potential in sensitization of human ovarian cancer cells to cisplatin treatment in vitro, suggesting that the combination of AT-101 with cisplatin may hold great promise for development as a novel chemotherapeutic approach to overcome platinum-resistance in human ovarian cancer. Keywords AT-101 Á Cisplatin Á Apoptosis Á Epigenetics Á Ovarian cancer cells Introduction The gold standard of care for advanced ovarian cancer is currently a combination of platinum and taxane. However, the disease almost invariably progresses to a platinum- resistant state, and unfortunately, second or third line chemotherapy regimens do not provide sufficient results [1–3]. Thus, for platinum-resistant ovarian cancer patients, new agents or their combinations with current chemother- apeutic agents with different cellular targets are urgently needed. Failure of drug-induced apoptosis have been suggested to underlie the phenomenon of drug resistance in cancer treatment, in addition to alterations in DNA repair, drug uptake, cell cycle related genes and also epigenetic events [4–6]. Thus, in recent years, therapies triggering apoptosis in drug resistant cancer cells are currently undergoing clinical trials to overcome drug resistance phenomenon [7]. Cisplatin causes the formation of DNA adducts, conse- quentially triggering apoptosis in cancer cells. However, toxicity to normal cells and development of drug resistance are the significant problems during cisplatin treatment [8]. The addition of new agents that can sensitize tumor cells to B. Karaca (&) Á B. Karabulut Á C. Sezgin Á U. A. Sanli Á R. Uslu Division of Medical Oncology, Tulay Aktas Oncology Hospital, School of Medicine, Ege University, 35100 Bornova, Izmir, Turkey e-mail: karacaburcak@hotmail.com; burcak.karaca@ege.edu.tr H. Atmaca Á E. Bozkurt Á A. Kisim Á S. Uzunoglu Section of Molecular Biology, Department of Biology, Faculty of Science and Letters, Celal Bayar University, 45140 Muradiye, Manisa, Turkey 123 Mol Biol Rep (2013) 40:3925–3933 DOI 10.1007/s11033-012-2469-z