Abstract Objective. Development of cisplatin resistance by cancer cells is a major hurdle in successful treatment of human ovarian cancer. However, the mechanism of cisplatin resistance in ovarian cancer cells remains unclear. In this study, we have examined the possible role of X-linked inhibitor of apoptosis protein (Xiap) in the development of cisplatin resistance in ovarian cancer cells and investigate if suppressed cytosolic high temperature required protein A (HtrA2/Omi) level is an important factor in cisplatin resistance in ovarian cancer. Methods. Cisplatin-sensitive (A2780 and COC1) and -resistant (A2780/DDP and COC1/DDP) ovarian cancer cells were cultured for different durations (0–24 h) and with different cisplatin concentrations (0–20 μM). Xiap content and cytosolic HtrA2/Omi content were analyzed by Western blot. Antisense oligonucleotides were used to downregulate Xiap content in ovarian cancer cells. Results. Cisplatin decreased Xiap content and increased cytosolic HtrA2/Omi content and caspase-3 activity in cisplatin-sensitive ovarian cancer cell lines (A2780 and COC1), but not in the resistant variants (A2780/DDP and COC1/DDP). Downregulation of Xiap by antisense Xiap oligonucleotides increased caspase-3 activity and sensitized cisplatin-resistant cells to cisplatin treatment. Cytosolic HtrA2/Omi level increased while Xiap was downregulated in cisplatin-resistant ovarian cancer cells. Conclusion. Development of cisplatin resistance may be due to Xiap neutralizing caspase-3 activation and lower cytosolic HtrA2/Omi level in response to cisplatin in human ovarian cancer cells. Cytosolic HtrA2/Omi level is partly regulated by Xiap in ovarian cancer cells.