Epigentic Alterations in Pre-malignant Tissue of the Bladder

描述（由申请人提供）：膀胱癌是美国第五大常见癌症。虽然这些肿瘤可以经尿道切除，但约50-80%会复发。由于复发是浅表性膀胱癌的一个共同特征，人们通常认为这可能是由于膀胱的癌前改变，尽管它还没有很好的分子定义。异常DNA甲基化已被提出作为一个有吸引力的候选人潜在的改变控制癌前病变。LINE-1逆转录转座因子的低甲基化在包括膀胱癌在内的许多癌症中很常见。我们最近在膀胱癌细胞中发现了一种特定的LINE-1元件的低甲基化，这种低甲基化也存在于相应的正常细胞中，而与肿瘤部位的距离无关。这种特殊的LINE-1元件（MET- line）位于原癌基因MET的内含子区域，我们已经在MET- line启动子低甲基化的细胞中检测到癌基因的截断形式的表达。这些结果可能提示膀胱癌前病变，其中有特定line1甲基化减少和癌基因表达的可能性增加。在本研究中，我们计划首先通过焦磷酸测序法和实时RT-PCR检测DNA甲基化和基因表达，进一步研究特异性LINE-1的低甲基化与截断MET异位表达之间的相关性，作为癌前变化的介质之一；其次，检测手术切除前后患者尿液沉积物中MET-LINE的甲基化变化，通过高灵敏度的检测方法（MethyLight）研究MET-LINE低甲基化作为复发标志物的潜在作用；最后，表征截断MET在肿瘤发生过程中的作用。完成这些特定目标将使我们能够在分子水平上获得膀胱癌患者膀胱上皮细胞癌前变化的证据，而没有恶性证据，并解决高度复发的膀胱癌的潜在机制。最后，这个转化项目将汇集基础科学家和临床医生，将临床发现转化为实验室问题，并利用这些基础发现改善患有这种疾病的患者的生活。公共卫生相关性：在美国，膀胱癌是男性中第四大最常见的癌症诊断，在女性中排名第八，50-80%的病例复发，2007年有67160例新病例和13750例死亡，根据国家癌症研究所。这项研究的完成将使我们能够阐明膀胱癌频繁复发的潜在原因，并开发出一种高度特异性和敏感性的生物标志物，不仅可以用于膀胱癌的诊断，还可以用于监测膀胱癌的复发。

DESCRIPTION (provided by applicant): Bladder cancer is the fifth most common cancer in the United States. Although these tumors can be removed by transurethral resection, about 50-80% will recur. Since recurrence is a common feature of superficial bladder cancer, it is often thought that this may be due to pre malignant changes in the bladder, although it has not been well defined molecularly. Aberrant DNA methylation has been proposed as an attractive candidate for the underlying alterations governing pre malignant changes. Hypomethylation of LINE-1 retrotransposable elements is quite common in many cancers including bladder cancer. We recently found hypomethylation of a specific LINE-1 element in bladder cancer cells that was also present in the corresponding normal appearing cells regardless of the distance from the site of the tumor. This particular LINE-1 element (MET-LINE) is located in the intronic region of the proto- oncogene MET and we have detected the expression of a truncated form of the oncogene in cells with hypomethylated promoter of MET-LINE. These results may indicate pre malignant changes in the bladder where there is a reduction of methylation of a specific LINE 1 and an increased likelihood of expression of an oncogene. In this proposal, we plan first to further investigate the correlation between hypomethylation of the specific LINE-1 and ectopic expression of truncated MET as one of mediators of pre milignant changes by the Pyrosequencing assay and real time RT-PCR to measure DNA methylation and gene expression; second to detect methylation changes of the MET-LINE in urine sediments of patients before and after surgical excision to study the potential role of hypomethylation of the MET-LINE as a marker of recurrence by a highly sensitive assay (MethyLight); and finally, to characterize the role of truncated MET during tumorigenesis. Accomplishment of these specific aims will allow us to obtain evidence of premalignant changes at the molecular level in bladder epithelial cells from bladder cancer patients without evidence of malignancy and address the potential mechanism of highly recurrent bladder cancers. Finally this translational project will bring together basic scientists and clinicians to translate clinical findings into laboratory questions and use these fundamental discoveries to better the lives of patients with this disease. PUBLIC HEALTH RELEVANCE: In the United States, bladder cancer is the fourth most common cancer diagnosis in men and the eighth most common in women with 50-80% of cases recurring, in 2007 there were 67160 new cases and 13750 deaths according to the Natinal Cancer Institute. Accomplishment of this proposal will allow us to elucidate the potential cause of the frequent recurrence of bladder cancer and develop a highly specific and sensitive biomarker not only for diagnosis of bladder cancer but also for monitoring the recurrence of bladder cancer.