Prostate cancer is a common type of cancer. Contemporary treatment modalities require that new, effective methods or chemicals should be developed. Tolfenamic acid (TA) is a nonsteroidal anti-inflammatory medicine stimulating apoptosis in cancer cells like pancreas, oesophagus and lungs. This study aims to determine the role of TA in cell viability and apoptosis in glioma cells (T-98G) and androgen-sensitive human prostate cancer cell line (LNCaP). In our study, 1, 5, 10, 25, 50, and 100 μM doses of TA were applied to T-98G and LNCaP cell lines for periods of 24 and 48 h. Subsequent cell vitality was tested with MTT Atest. The effective doses were analysed by flow cytometry and real-time PCR techniques. MTT spectrophotometric tests showed no significant effects on the survival rate of T-98G cells in 24 and 48 h. Therefore, T-98G cells were not analysed by flow cytometry and RT-PCR. When LNCaP cells were exposed to 1, 5, 10, 25, 50 and 100 μM doses of TA for 24 h, the rates of living cells were determined as 91%, 83%, 82%, 76%, 61% and 49%, respectively. When the same doses were applied for 48 h, the living cell rates were 90%, 83%, 82%, 78%, 52%, and 47%, respectively. Following applications of 25, 50 and 100 μM TA to LNCaP cells for 24 h, the apoptotic values (0.60% for the control group) were found 2%, 4%, and 17%, whereas the values for 48 h were 10%, 10% and 64% (0.48% for the control group), respectively. TA induced apoptosis of LNCaP cells depending on the dose and duration of exposure. According to PCR results, caspase-9 activity increased incrementally in groups given 50 μM of TA. The increase in the apoptosis of LNCaP cells may be due to the rising amount of TA concentration. In our study, in 25, 50, and 100 μM TA groups, cyclooxygenase-2 (COX-2) decreased incrementally compared to the control group. Considering overstimulation of COX-2 paths in emergence of cancer or in metastasis, this study shows TA has an antiproliferative and stimulating effects on apoptosis of LNCaP cells based on the dose and duration of exposure.