Methionine adenosyltransferase (MAT) is an essential enzyme that catalyzes the formation of S- adenosylmethionine (SAMe). In mammals, the catalytic subunit of MAT is encoded by two genes, MAT1A and MAT2A, while a third gene MAT2?, encodes for a regulatory subunit ? that regulates MAT2A-encoded enzyme. MAT1A is expressed in liver while MAT2A is widely distributed. In hepatocytes, increased MAT2A and MAT2? expression increased growth. SAMe inhibits hepatocyte growth, is pro-apoptotic in liver cancer cells but anti- apoptotic in normal hepatocytes. Regulation of MAT genes and whether MAT and SAMe regulate growth and death in colon cancer cells are unknown. We found increased MAT2A and MAT2? mRNA levels in resected colon cancer specimens as compared to adjacent normal tissue. This occurred due to increased transcription. MAT2A mRNA levels are also increased in the polyps of Min mice. EGF, IGF-1 and leptin, three mitogens implicated in the pathogenesis and invasiveness of colon cancer, induced MAT2A expression and growth in colon cancer cell lines RKO and HT29. Lowering MAT2A expression by RNAi reduced growth and blocked the mitogenic effects of these growth factors. SAMe and its metabolite methylthioadenosine (MTA) lowered MAT2A expression and prevented the ability of growth factors to induce MAT2A and cell proliferation. Furthermore, SAMe and MTA inhibited growth of colon cancer cells by inducing apoptosis. A novel target identified is cFLIP, which is down-regulated by both SAMe and MTA. Finally, overexpression increased, while reduced expression of MAT2? inhibited growth and caused apoptosis. The current proposal will extend these novel observations in five specific aims: 1. Examine transcriptional regulation of MAT2A in colon cancer cell lines ? we will determine how MAT2A is transcriptionally regulated at baseline and in response to growth factors, SAMe and MTA;2. elucidate the molecular mechanism(s) of MAT2A and MAT2? up-regulation in colon cancer ? we will use resected colon cancer and normal tissue specimens to elucidate the mechanisms of their up-regulation;3. determine how MAT genes influence growth and death of colon cancer cells ? we will determine how MAT2A and MAT2? regulate growth and apoptosis;4. elucidate the mechanisms of SAMe and MTA-induced apoptosis in colon cancer cells ? we will define the molecular mechanisms and examine whether normal colon epithelial cells are affected;and 5. Assess changes in MAT expression during tumor formation and efficacy of SAMe and MTA in chemoprevention and treatment of intestinal tumors ? we will examine whether SAMe and MTA can prevent spontaneous polyp formation in Min mice, and halt progression and reverse already established aberrant crypt foci in Min mice treated with azoxymethane. Successful completion of these studies should greatly enhance our understanding of the role of MAT and SAMe in colon cancer pathogenesis and may provide a novel therapeutic approach for both chemoprevention and treatment of colon polyps and cancer, topics that are highly relevant to public health. PROJECT NARRATIVE Colon cancer remains one of the most common cancers in the western world. At the present time effective chemopreventive therapy that is devoid of side effects is lacking. The goals of this project are to investigate a new paradigm in colon cancer development and see if S-adenosylmethionine (SAMe), a nutritional supplement widely available, and its metabolite methylthioadenosine (MTA) can be effective in the chemoprevention and treatment of colon cancer.
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