Purpose: iASPP is a specific regulator of p53-mediated apoptosis. Herein, we provided the first report on the expression profile of iASPP in ovarian epithelial tumor and its effect on paclitaxel chemosensitivity. Experimental Design: Expression and amplification status of iASPP was examined in 203 clinical samples and 17 cell lines using immunohistochemistry, quantitative real-time PCR, and immunoblotting, and correlated with clinicopathologic parameters. Changes in proliferation, mitotic catastrophe, apoptosis, and underlyingmechanism in ovarian cancer cells of different p53 status following paclitaxel exposure were also analyzed. Results: The protein and mRNA expression of iASPP was found to be significantly increased in ovarian cancer samples and cell lines. High iASPP expression was significantly associated with clear cell carcinoma subtype (P1⁄40.003), carboplatin andpaclitaxel chemoresistance (P1⁄40.04), shorter overall (P1⁄40.003), and disease-free (P 1⁄4 0.001) survival. Multivariate analysis confirmed iASPP expression as an independent prognostic factor. Increased iASPP mRNA expression was significantly correlated with gene amplification (P 1⁄4 0.023). iASPP overexpression in ovarian cancer cells conferred resistance to paclitaxel by reducing mitotic catastrophe in a p53-independent manner via activation of separase, whereas knockdown of iASPP enhanced paclitaxel-mediated mitotic catastrophe through inactivating separase. Both securin and cyclin B1/CDK1 complex were involved in regulating separase by iASPP. Conversely, overexpressed iASPP inhibited apoptosis in a p53-dependent mode. Conclusions:Our data show an association of iASPP overexpression with gene amplification in ovarian cancer and suggest a role of iASPP in poor patient outcome and chemoresistance, through blocking mitotic catastrophe. iASPP shouldbe explored further as a potential prognosticmarker and target for chemotherapy. Clin Cancer Res; 17(21); 6924–33. 2011 AACR.