Introduction: Polyphenols, naturally occurring compounds in plants, exhibit a wide range of pharmaceutical properties (1). Resveratrol, gallic acid, and kuromanin chloride are dietary polyphenols belonging to the stilbenes, phenolic acids, and flavonoids classes, respectively (2-4). Studies during the past decades have shown the promising effect of these compounds in the prevention and treatment of different diseases such as Alzheimer's, diabetes, cancer, etc. (1). Telomerase is a ribonucleoprotein complex responsible for the de novo synthesis of the telomere. The elevated activity of telomerase was detected in approximately 85% of cancer cells (5). Several anticancer drugs exert their effects by inhibiting telomerase activity (6). Previous studies have suggested that some polyphenolic compounds may also have anti-telomerase activity against cancers (7). No study has compared the effect of three polyphenols of resveratrol, gallic acid, and kuromanin chloride, on telomerase activity in prostate cancer cells. The present study was designed to investigate the anti-proliferative and anti-telomerase effects of these polyphenols on the PC3 human prostate cancer cell line.
Methods: The human prostate cancer (PC3) cell line was obtained from the National Cell Bank of Iran (NCBI, Pasteur Institute, Tehran). The cells were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum (FBS) and 1% penicillin-streptomycin. PC3 cancer cells were treated with an increasing concentration (2.5, 5,10, 20, 40, 80) of resveratrol and gallic acid in a 96-well plate for 72h. The effect of compounds on cell proliferation was determined using the tetrazolium-based colorimetric assay (MTT assay). The cancer cells were then exposed to different concentrations (10, 20, and 40 μM) of polyphenols for 72h. At the end of the incubation period, the protein was extracted using an NP-40 lysis buffer. Protein concentration was measured by the Bradford assay, and telomerase activity was assessed by telomeric repeat amplification protocol (TRAP assay). Also, a trypan blue exclusion assay was performed to determine polyphenols' effect on cell viability.
Results: Compared to the control group, resveratrol, and gallic acid significantly reduced PC3 cell proliferation. The IC50 value of resveratrol and gallic acid for PC3 cells was 16.69 ±1.502 μM and 78.36 ±14.05 μM, respectively. Kuromanin chloride had a negligible effect on the cell proliferation. The results of trypan blue exclusion assay indicated that all of these polyphenols reduce cell viability dose-dependently. Among the polyphenols, resveratrol had a substantial effect on reducing cell viability and proliferation. Compared to the control group, kuromanin chloride at all tested concentrations inhibited PC3 telomerase activity (p< 0.01). Gallic acid at 40 μM led to a significant reduction (40%) in telomerase activity (p<0.01). The telomerase activity in PC3 exhibited a reversed U-shaped response to different concentrations of resveratrol. Resveratrol at 20 μM significantly elevated telomerase activity by 34% (p<0.01), as compared to the non-treated control group. But, the change in the enzyme activity was not observed at other concentrations (10 and 40 μM).
Conclusion: The findings of the present study showed that all three polyphenols reduced cell proliferation and cell viability of PC3 cells. Resveratrol was the most effective polyphenol for inhibiting the proliferation of PC3 cancer cells. However, the change in telomerase activity was not correlated to the cell proliferation, which may be related to the other anticancer activity of these compounds. Previous studies have demonstrated that telomerase-dependent telomere shortening and consequently cell proliferation inhibition is dependent on several parameters such as initial telomere length and the required time for efficient telomere length shortening (8, 9). Our findings indicated that kuromanin chloride and gallic acid possess a considerable anti-telomerase effect. Further studies for evaluating the in vivo anti-telomerase activity of polyphenols against prostate cancer are needed.