Grape seed extract induces cell cycle arrest via up-regulation of p21^Waf1 and down-regulation of cyclins-cyclin dependent kinases, and causes apoptotic death of human colorectal carcinoma LoVo, HT29 and SW480 cells

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Colorectal cancer (CRC) is the second most frequently diagnosed malignancy in the United States as well as the second most common cause of cancer-related deaths. Primary treatment of patients diagnosed with CRC has been surgical resection, which is often limited by relapse of disease, leading to increased morbidity. Accumulating evidences have suggested the beneficial effects of fruit and vegetable consumption in lowering the risk of various cancers including CRC. Consistent with this, various phytochemicals present in fruits and vegetables, and those consumed as dietary supplements have been shown to exhibit anticancer activity against various cancer sites including CRC in preclinical models and human cancer cell lines in vitro. Current study was conducted to investigate whether grape seed extract, a rich source of proanthocyanidins, exerts anticancer effects in human CRC cell lines, and if that is the case, what is its molecular mechanism of efficacy. Based on completed studies, here we report for the first time that grape seed extract, irrespective of manufacturer sources, inhibits the growth of a battery of human CRC cell lines, namely LoVo, HT29 and SW480. Treatment of these cells with GSE (0-100 microgram/ml) for 12-48h resulted in a significant dose- and time- dependent inhibition of cell growth with concomitant increase in cell death, albeit at different level. With regard to its cell growth inhibitory effect, a G1 cell cycle arrest was observed in LoVo and HT29 cells following grape seed extract treatment, which possibly was associated with an observed up-regulation of p21^Waf1 and down-regulation of cyclin D1, E, A and cyclin dependent kinase (CDK-2, -4 and -6) levels. In case of HT29 cells, up-regulation of p27 ^Kip1 was also observed following grape seed extract treatment of the cells, though the levels of this CDKI remained unaltered in case of LoVo cells under similar treatment conditions. An induction of p21^Waf1 by grape seed extract seems to be independent of p53 status in these CRC cell lines where LoVo cells are p53(+/+), while SW480 and HT29 cells are p53(-/0). In SW480 cells, grape seed extract induced S and/or G2-M phase cell cycle arrest together with a strong induction of p21^Waf1 and down regulation of cyclin D1, B1, A and CDK-1, -2, -4, -6 levels. An induction of apoptosis was also observed in all three CRC cell lines following grape seed extract treatment as measured by annexin-V/PI staining and western blot analysis for cleaved caspase-3 and PARP. Taken together, based on these findings, we conclude that grape seed extract could be an effective agent against colorectal cancer. More studies, specifically in pre-clinical CRC models, are needed in future to assess and establish GSE efficacy against CRC.