Role of MAT and SAMe in Colon Cancer Pathogenesis and Treatment

说明(申请人提供)：蛋氨酸腺苷转移酶(MAT)是催化S-腺苷蛋氨酸(SAME)形成的重要酶。在哺乳动物中，MAT的催化亚基由两个基因MAT1A和MAT2A编码，而第三个基因MAT2？编码调节亚基？它调节由MAT2A编码的酶。Mat1a在肝脏中表达，而Mat2a在肝脏中广泛分布。在肝细胞中，MAT2A和MAT2？表达促进了生长。Same抑制肝细胞生长，在肝癌细胞中是促凋亡的，在正常肝细胞中是抗凋亡的。MAT基因的调控以及MAT和SAME是否调控结肠癌细胞的生长和死亡尚不清楚。我们发现MATA和MAT2升高了？结肠癌切除标本与癌旁正常组织的mRNA水平比较。这是由于转录增加所致。Min小鼠息肉中MAT2AmRNA水平也升高。EGF、IGF-1和Leptin是与结肠癌的发生和侵袭有关的三种有丝分裂原，它们诱导了结肠癌细胞株RKO和HT29中MAT2A的表达和生长。通过RNAi降低MAT2A的表达减少了生长，并阻断了这些生长因子的有丝分裂作用。Same及其代谢物甲硫腺苷(MTA)降低MAT2A的表达，阻止生长因子诱导MAT2A和细胞增殖的能力。此外，SAME和MTA通过诱导细胞凋亡来抑制结肠癌细胞的生长。发现了一个新的靶点是cFLIP，它被Same和MTA下调。最后，MAT2？的过度表达增加，而表达降低。抑制生长并导致细胞凋亡。目前的建议将在五个具体目标上扩展这些新的观察结果：1.研究MAT2A在结肠癌细胞系中的转录调控--我们将确定MAT2A在基线以及对生长因子、SAME和MTA的反应中如何转录调控；2.阐明MAT2A和MAT2的分子机制(S)？在结肠癌中上调-我们将使用切除的结肠癌和正常组织标本来阐明它们上调的机制；3.确定MAT基因如何影响结肠癌细胞的生长和死亡-我们将如何确定MAT2A和MAT2？4.阐明Same和MTA诱导结肠癌细胞凋亡的机制--我们将确定分子机制并检测正常结肠上皮细胞是否受到影响；以及5.评估MAT在肿瘤形成过程中的表达变化以及Same和MTA在化学预防和治疗肠道肿瘤中的有效性-我们将研究Same和MTA是否可以防止Min小鼠自发性息肉的形成，并阻止和逆转偶氮甲烷处理的Min小鼠已经建立的异常隐窝病灶。这些研究的成功完成将大大加深我们对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，这是与公共卫生高度相关的主题。与公共卫生相关：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。该项目的目标是探索结肠癌发展的新模式，并观察广泛使用的营养补充剂S-腺苷蛋氨酸(SAME)及其代谢物甲硫腺苷(MTA)是否能有效地用于结肠癌的化学预防和治疗。

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.