Role of MAT and SAMe in Colon Cancer Pathogenesis and Treatment

描述（申请人提供）：甲硫氨酸腺苷转移酶（MAT）是催化S-腺苷甲硫氨酸（SAMe）形成的必需酶。在哺乳动物中，MAT的催化亚基由两个基因MAT 1A和MAT 2A编码，而第三个基因MAT 2？编码一个调节亚基调节MAT 2A编码的酶。MAT 1A在肝脏中表达，而MAT 2A广泛分布。在肝细胞中，增加MAT 2A和MAT 2？表达增加了生长。SAMe抑制肝细胞生长，在肝癌细胞中促细胞凋亡，但在正常肝细胞中抗细胞凋亡。MAT基因的调节以及MAT和SAMe是否调节结肠癌细胞的生长和死亡尚不清楚。我们发现增加MAT 2A和MAT 2？与邻近正常组织相比，切除的结肠癌标本中的mRNA水平。这是由于增加了转录。MAT 2A mRNA水平在Min小鼠的息肉中也增加。EGF，IGF-1和瘦素，三种有丝分裂原参与结肠癌的发病机制和侵袭性，诱导结肠癌细胞系RKO和HT 29中MAT 2A的表达和生长。通过RNAi降低MAT 2A表达降低了生长并阻断了这些生长因子的促有丝分裂作用。SAMe及其代谢产物甲硫腺苷（MTA）降低了MAT 2A的表达，并阻止了生长因子诱导MAT 2A和细胞增殖的能力。此外，SAMe和MTA通过诱导凋亡抑制结肠癌细胞的生长。鉴定的新靶点是cFLIP，其被SAMe和MTA下调。最后，过度表达增加，而减少表达的MAT 2？抑制生长并引起凋亡。目前的建议将扩展这些新的观察在五个具体目标：1。检查结肠癌细胞系中MAT 2A的转录调控-我们将确定MAT 2A在基线和响应生长因子SAMe和MTA时如何转录调控; 2.阐明MAT 2A和MAT 2？结肠癌中的上调-我们将使用切除的结肠癌和正常组织标本来阐明其上调的机制; 3.确定MAT基因如何影响结肠癌细胞的生长和死亡-我们将确定MAT 2A和MAT 2如何？调节生长和凋亡; 4.阐明SAMe和MTA诱导结肠癌细胞凋亡的机制-我们将定义分子机制并检查正常结肠上皮细胞是否受到影响;以及5.评估肿瘤形成过程中MAT表达的变化以及SAMe和MTA在肠道肿瘤的化学预防和治疗中的功效-我们将检查SAMe和MTA是否可以预防Min小鼠中的自发性息肉形成，并在用氧化偶氮甲烷处理的Min小鼠中停止进展并逆转已经建立的异常隐窝病灶。这些研究的成功完成将大大提高我们对MAT和SAMe在结肠癌发病机制中作用的理解，并可能为结肠息肉和癌症的化学预防和治疗提供一种新的治疗方法，这些主题与公共卫生高度相关。公共卫生相关性：结肠癌仍然是西方世界最常见的癌症之一。目前缺乏有效的无副作用的化学预防治疗。该项目的目标是研究结肠癌发展的新模式，并观察广泛使用的营养补充剂S-腺苷甲硫氨酸（SAMe）及其代谢产物甲硫腺苷（MTA）是否可以有效地化学预防和治疗结肠癌。

DESCRIPTION (provided by applicant): 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. PUBLIC HEALTH RELEVANCE: 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.