METHYLATION PROFILING AND RISK OF COLORECTAL CANCER

描述（由申请人提供）：结直肠癌（CRC）是美国最常见的胃肠道恶性肿瘤，非裔美国人（AA）的CRC发病率是白种人的1.5倍。分子医学领域的最新进展导致了微卫星不稳定性（MSI）、杂合性缺失（LOH）和甲基化特异性PCR技术的使用，这些技术可以检测结肠粘膜细胞的染色体和基因改变。标志物的使用有助于预测疾病进展和预后。表观遗传改变发生在导致结直肠癌的遗传改变序列的早期。随后的变化通常包括部分或整个染色体的丢失。肿瘤中的MSI在某些基因编码区内的微卫星中积累突变。这些数据表明，MSI、LOH和甲基化谱可能为恶性肿瘤的分子表型预测增加了重要的信息层。我们假设MSI-H，即DNA修复基因hMHL1、p16、APC、p53和DCC中缺失的基因沉默，可能改变肿瘤转化途径中的染色体行为，这些基因被认为是调节结肠粘膜细胞增殖的。MSI将使用5个微卫星位点进行测量，粘膜活检中p53、APC和DCC蛋白水平将通过免疫组化检测。通过测定250例病例的甲基化和突变/缺失谱，我们明确(1)阐明p16和hMLH1基因甲基化在AA合并结直肠癌患者正常组织和癌组织中肿瘤转化途径中的作用；(2)确定结肠黏膜中MSI的诱导作用，这可能反映在AA患者结肠腺瘤史中肿瘤转化和细胞增殖的生物标志物的表达中。无腺瘤病史者和有结直肠癌切除史者。这些实验可能有助于识别发生腺瘤和/或结直肠癌的高危人群，(3)确定结直肠癌患者正常组织和癌组织中APC、p53和DCC基因是否存在LOH或等位基因缺失，并确定发生其他腺瘤和/或结直肠癌的高危人群，(4)分析接受氟尿嘧啶治疗的AA型III期和高危II期结直肠癌患者的肿瘤组织。以及MSI和LOH标记物预测生存和/或治疗反应的能力。这些研究将有助于检测和分析AA中CRC通路的遗传变化。

DESCRIPTION (provided by applicant): Colorectal carcinoma (CRC) is the most common gastrointestinal malignancy in the U.S and the rate of CRC is 1.5 times higher in African Americans (AA) than Caucasians. Recent advances in the field of molecular medicine has led to the use of microsatellite instability (MSI), loss of heterozygosity (LOH), and methylation specific PCR techniques which permit detection of chromosomal and gene alterations of colonic mucosal cells. The use of markers has helped to predict disease progression and prognosis. Epigenetic changes are early in the sequence of genetic alterations leading to CRC. Subsequent changes often include the loss of portions or whole chromosomes. MSI in neoplasms accumulates mutations in microsatellites within the coding region of certain genes. These data suggest that MSI, LOH and methylation profiling may add an important layer of information in the molecular phenotyping of malignances for prognostic purposes. We postulate that MSI-H, gene silencing for DNA repair gene hMHL1, p16 and LOH of APC, p53 and deleted in colorectal cancer (DCC), which are known to modulate cellular proliferation in colonic mucosa, may alter chromosome behavior in the pathway of neoplastic transformation. MSI will be measured using five microsatellite loci, and the level of p53, APC and DCC protein will be determined by immunohistochemistry in mucosal biopsies. By determining the methylation and mutation/deletion profiles of 250 cases, we determine specifically (1) to elucidate the effect of p16 and hMLH1 gene methylation in the pathway of neoplastic transformation in normal and cancer tissue of AA patients with CRC, (2) to determine the induction of MSI in colonic mucosa that may be reflected in the expression of biomarkers of neoplastic transformation and cellular proliferation from AA patients history of colonic adenomas, those with no history of adenomas, and those with a history of resected CRC. These experiments may assist in identifying persons at risk of developing adenomas and/or CRC, (3) to determine whether LOH or allelic loss occurs in APC, p53, and DCC genes in normal and cancer tissue of CRC patients and identify persons at risk of developing additional adenomas and/or CRC, (4) to analyze tumor tissue in AA patients with stage III and high-risk stage II CRC who had been treated with fluorouracil, and the ability of MSI and LOH markers to predict survival and/or response to treatment. These studies will help in the detection and profiling of genetic changes in the pathway of CRC in AA.