Background: The role of vitamin D3 in calcium and mineral metabolism is known and it is an effective regulator of cell differentiation, cell growth and angiogenesis and recently it has been demonstrated to affect, tumor invasiveness and apoptosis. Though, many evidences from epidemiological studies show the potential involvement of vitamin D3 in the treatment and prevention of certain cancers, although it is not completely identified how vitamin D3 inhibits the growth of cancer cells. More recently numerous studies have demonstrated the known pro-apoptotic and anti-proliferative properties of vitamin D3 which are mediated by binding of a receptor called vitamin D receptor (VDR). Furthermore, it is also studied that vitamin D3 is a potent antioxidant and it reduces the reactive oxygen species (ROS) by preventing the growth of cancer cells. Though, it is currently unidentified whether vitamin D3 prompted cancer cell death which is intermediated by its effect on reactive oxygen species. Therefore, the efficacy of vitamin D3 for inhibiting the growth of HeLa and HepG2 cell lines was determined. Methods: In the present research, cancerous (HeLa and HepG2) and normal (BHK-21) cell lines were used to check the proliferation, angiogenesis, apoptosis and anti-oxidative activity. Cells were cultured and treated with 10-3M, 10-4M and 10-5M of vitamin D3. Results: After the treatment of vitamin D3, ELISA with VEGF and p53 was performed to check angiogenesis and apoptosis respectively. Gene expression level was analyzed by using apoptotic and proliferative markers. Reduced proliferation via MTT, reduced angiogenesis via VEGF and increased apoptosis via p53 were observed in cancer cells after the treatment as compared to normal treated cells. Anti-oxidant (SOD, CAT, APOX, GSH and LDH) activity was increased and oxidative stress was reduced in treated cells as compared to normal cells after the treatment with vitamin D3. Conclusion: In short term vitamin D3 decreases the cell viability by apoptosis induction. While in long term it has anti-angiogenic effect that is independent to cell viability. Our data showed that vitamin D3 inhibited the growth HeLa and HepG2 cells in a dose dependent manner. However, more studies are required in this respect.