Vitamin K4Induces Tumor Cytotoxicity In Human Prostate Carcinoma PC-3Cells Via The Mitochondria-related Apoptotic Pathway

Prostate cancer is the second most frequently diagnosed cancer worldwide, behind onlylung cancer, and is the second most common diagnosed cancer of American male. Prostatecancer occurs mainly in older men. Nearly two thirds are diagnosed in men aged65or older,and it is rare before age40. There are about approximately240,000new cases and28,000people die of this malignancy as estimated by a survey report.Despite recent progress in diagnostic and multi-modal therapies of initial prostate cancer,there is no effective therapy in treating androgen-independent or hormone-refractory recurrentcases, the mortality rate associated with recurrent prostate cancer is still very high. So, there isurgent need to identify new therapeutic agents to improve the overall survival of patients withprostate cancer.Vitamin K4(VK4) is a water-soluble menadione which is synthetic derivatives ofVK2, it was primarily used as a clinical coagulation. Recent studies have shown that VKshave intimate connection with vascular health and bone metabolism.VKs play important rolesin normal blood coagulation system, in which they act as cofactors for the synthesis of gammacarboxyglutamate (Gla) protein family such as prothrombin and factors VII, IX, and X. Sincethere’re three VK-dependent proteins (Osteocalcin, Matrix Gla-protein and Protein S) whichmay be invoved in bone metabolism, therefore, impairment of the function of these proteinsmay lead to adverse effects of bone. Some reports suggest that VKs may also play a role inimmunology, and were found to exhibit antitumor activity against a number of cell lines,including hepatic, leukemia, colon, lung, oral, breast and bladder cancers. The goal of thisstudy is to investigate the inhibitory activity of VK4on androgen-independent prostate cancercell line PC-3proliferation and the mechanisms involved.Inhibitory effect of VK4on human prostate PC-3cells was measured using MTTmethod.Cell apoptosis, cell cycle distribution, intracellular ROS, mitochondrial membranepotential (MMP) were evaluated by flow cytometry method. Hoechst33258staining was usedto evaluate nuclear DNA fragmentation. Trypan blue was used to observe the cytotoxicity ofVK4on mouse spleen cells. Western blot analysis was used to investigate effect of VK4onmajor mitochondrial apoptosis regulatory proteins. MTT results showed that VK4dose-dependently decreased the viability of PC-3cells. IC50value of the inhibition wasaround20.94μM. Hoechst33258results showed that VK4caused significant nuclear condensation in PC-3cells. Annexin V-FITC/PI double staining showed that VK4inducedapoptosis in PC-3cells. PI staining indicated that VK4arrested the cell cycle at S phase in adose-dependent way. Further mechanistic studies revealed that VK4-mediated induction ofapoptosis in PC-3cells is associated with disruption of mitochondrial membrane potential,down-regulation of Bcl-2, and up-regulation of Bax, release of cytochrome c frommitochondria, and activation of caspase-3and PARP.In conclusion, our findings add new pharmaceutical activity to VK4, and VK4maybecome a potential leading drug in the therapy of prostate carcinoma.