Purpose Prostate cancer (PCa) is characterized by deregulated expression of several tumor suppressor or oncogenic miRNAs. The objective of this study was the identification and characterization of miR-let-7c as a potential tumor suppressor in PCa. Experimental Design Levels of expression of miR-let-7c were examined in human PCa cell lines and tissues using qRT-PCR and in situ hybridization. Let-7c was overexpressed or suppressed to assess the effects on the growth of human PCa cell lines. Lentiviral-mediated re-expression of let-7c was utilized to assess the effects on human PCa xenografts. Results We identified miR-let-7c as a potential tumor suppressor in PCa. Expression of let-7c is downregulated in castration-resistant prostate cancer (CRPC) cells. Overexpression of let-7c decreased while downregulation of let-7c increased cell proliferation, clonogenicity and anchorage-independent growth of PCa cells in vitro. Suppression of let-7c expression enhanced the ability of androgen-sensitive PCa cells to grow in androgen-deprived conditions in vitro. Reconstitution of Let-7c by lentiviral-mediated intratumoral delivery significantly reduced tumor burden in xenografts of human PCa cells. Furthermore, let-7c expression is downregulated in clinical PCa specimens compared to their matched benign tissues, while the expression of Lin28, a master regulator of let-7 miRNA processing, is upregulated in clinical PCa specimens. Conclusions These results demonstrate that microRNA let-7c is downregulated in PCa and functions as a tumor suppressor, and is a potential therapeutic target for PCa.