Abstract: Objective To investigate the effects and mechanism of lactate dehydrogenase A (LDHA)gene knockout combined with mitomycin C(MMC)on apoptosis of prostate cancer cell line PC3. Methods Western blotting was used to validate the LDHA gene knockout PC3 cells(LDHA PC3). Cell invasion assay was used to examine the invasiveness of LDHA PC3 cells and PC3 cells. Flow cytometry was used to investigate the effects of MMC treatment and LDHA gene knockout on PC3 apoptosis. Western blotting was employed to determine the altered expression of proteins related to apoptosis pathway. Results Western blotting validated complete LDHA gene knockout in 3 clones of KOLDHA PC3 cell line. LDHA gene knockout was shown to inhibit the invasiveness of PC3 cells,and to promote MMC?induced PC3 cell apoptosis via the caspase?3/poly ADP?ribose polymerase (PARP) pathway. Conclusion LDHA gene knockout and MMC may work synergistically to induce apoptosis of PC3 cells.