Ellagic acid is a phenolic compound found in organic products in nature. It is abundant in many members of the fruit family, has already displayed anti-cancerous effects in many previous studies, and is a potent tool for chemoprevention. However, only a few modes of action have been proposed for ellagic acid's influence on cancer cells. Cell culture studies indicate that pure ellagic acid works through a p53-independent mechanism. Although ellagic acid and its analogs have been shown to modulate non-specific receptor tyrosine kinases (RTKs), not much is known about their function as inhibitors of a particular tyrosine kinase called epidermal growth factor receptor (EGFR), which plays a key role in cell signal transduction and tumor growth. To determine whether ellagic acid is a tyrosine kinase inhibitor in cancer cells, we focused on the effects of ellagic acid on EGFR expression in two human cancer cell types. These were cervical carcinoma (Caski) and lung cancer (A549) cells. Caski cells lack a functioning tumor suppressor gene (p53) because of the Human papillomavirus's (HPV 16) ability to disable it. This made it a perfect candidate for identifying a p53-independent mechanism for ellagic acid. Lung cancer cells were chosen because of their high expression of EGFR. First, to monitor the effects of ellagic acid on both cell types, we completed a cell growth analysis. We exposed the cells to varying amounts of ellagic acid over a period of 48 hours. During treatment, the amount of living cells and cells going through mitosis significantly decreased within 48 hours of exposure. Next, a flow cytometry analysis showed a clear halt in cell cycle activity with an arrest in the G1 phase of the treated cells. With this finding, we hypothesized that ellagic acid must work through a cell cycle regulatory protein. Cyclin D1 is a protein that is also involved in the regulation of cell signal transduction and cell communication. In observing a decrease in cyclin D1 activity, we hoped to pinpoint a definite mechanism through which ellagic acid works to inhibit cell cycle activity in cancer cells. In this part of the study, using an RT PCR, we discovered a prominent decrease in the mRNA expression of cyclin D1 in the treated cells. Because of the similar roles held by cyclin D1 and EGFR in cancer cell growth, we hypothesized that a possible link could exist between the two. By using a Western blot analysis, we saw a significant decrease in the expression of phosphorylated EGFR (pTyr100 and pTyr102) protein. In conclusion, we have demonstrated that ellagic acid inhibited cancer cell proliferation by down-regulating the tyrosine kinase EGFR. We believe that more progress can be made in the field of chemoprevention with further research into ellagic acid and tyrosine kinase mechanisms.
[Fifth AACR International Conference on Frontiers in Cancer Prevention Research, Nov 12-15, 2006]