Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. Despite advances in diagnostic and therapeutic methods, survival of HNSCC patients remains unchanged over the last 30 years. Epigenetic alterations are commonly associated with several types of cancers including HNSCC. Studies have shown that DNA hypermethylation plays important roles in the risk of cancers by inactivating the tumor suppressor genes/proteins. Thus, epigenetic changes are considered as promising therapeutic targets for cancer treatment. Dietary phytochemicals offer promising options for the development of effective strategies for the prevention of cancer, and thus can be utilized as complementary and alternative medicine. In the present study, we investigated the effect of grape seed proanthocyanidins (GSPs) on DNA methylation and growth of HNSCC cells in vitro and in vivo models. Our analytical data revealed that the levels of global DNA methylation and DNA methyltransferase (DNMT) activity were higher in HNSCC cells compared with normal human bronchial epithelial cells (NHBEC). Treatment of HNSCC cells (FaDu and SCC-1) with GSPs for 6 days results in significant decrease in global DNA methylation (70-80%, P<0.001) level, DNMT activity (60-80%) as well as in levels of DNMT proteins (DNMT 3A and DNMT 3B) compared to non-GSPs-treated control cells. The observation on DNA methylation levels was verified by analyzing the levels of 5-methylcytosine (5-mC) using dot-blot analysis. Additionally, GSPs treatment also reactivated tumor suppressor proteins, such as Cip1/WAF1/p21 and p27 in HNSCC cells. Treatment of HNSCC cells with 5-Aza-dc, an inhibitor of DNA methylation, also resulted in suppression of DNA methylation as well as DNMT protein expression and DNMT activity in these cells. Tumor xenograft studies were performed to further determine the effect of GSPs on the growth of HNSCC cells using athymic nude mouse model. Our in vivo study demonstrated that administration of GSPs (0.5%, w/w) in supplementation with AIN76A control diet resulted in inhibition of xenograft growth of FaDu cells in nude mice (80%; P<0.01) compared to the tumor xenograft growth in mice fed with non-GSPs-supplemented control diet. The growth inhibitory effect of dietary GSPs on the HNSCC tumor xenografts was associated with the inhibition of DNA methylation, DNMT activity and reactivation of silenced tumor suppressor proteins in tumor tissues. Together, this preclinical study provides evidence that GSPs act as a DNA demethylating agent and able to reactivate an epigenetically silenced tumor suppressors to inhibit growth of HNSCC cells.

Citation Format: Santosh K. Katiyar, Ram Prasad. Proanthocyanidins from grape seeds inhibit growth of head and neck squamous cell carcinoma by decreasing dna methylation and reactivation of silenced tumor suppressors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1916. doi:10.1158/1538-7445.AM2015-1916