Klotho is an anti-aging transmembrane protein, which can be shed and function as a hormone. Accumulating data indicate klotho as a tumor suppressor in a wide array of malignancies, and we identified klotho as an inhibitor of the insulin-like growth factor (IGF-1) pathway in cancer cells. As this pathway is significant in the development of epithelial ovarian cancer (EOC) we studied klotho expression and activity in this tumor. Klotho mRNA levels were reduced in 16 of 19 EOC cell lines and immunohistochemistry analysis revealed high expression in normal ovaries, and reduced expression in 100 of 241 high grade papillary-serous adenocarcinoma of the ovaries, fallopian tubes and peritoneum. Reduced expression was associated with wild-type BRCA status. Klotho reduced EOC cell viability, enhanced cisplatin sensitivity, and reduced expression of mesenchymal markers. Finally, klotho inhibited IGF-1 pathway activation and inhibited transcriptional activity of the estrogen receptor. In conclusion, klotho is silenced in a substantial subset of the tumors and restoring its expression slows growth of EOC cells and inhibits major signaling pathways. As klotho is a hormone, treatment with klotho may serve as a novel treatment for EOC.