URINARY BIOMARKERS OF RENAL CELL CARCINOMAS

描述(申请人提供)：肾癌通常是无症状的，通常是致命的。仅在美国，2009年就有超过57,000例新的肾癌被诊断出来，近13,000例死亡归因于这种疾病。早期诊断，引发手术切除，通常是治愈的。体征和症状出现较晚，当诊断为肾癌时，通常已经转移到淋巴结或邻近器官，并进展到治愈阶段，生存机会很小。偶然的早期发现可能发生在腹部成像(CT、MRI、超声)过程中，原因无关。尽管如此，目前还没有人群筛查和诊断肾癌的方法，特别是在治愈阶段。未得到满足的需求是一种简单、高吞吐量、成本效益高的肾癌筛查测试，具有每年防止超过13,000人死亡的公共卫生意义，特别是在高危人群中。这项拟议的研究将检验一种假设，即肾癌细胞表达并分泌或活跃地分泌蛋白质到尿液中，这些蛋白质是标志性分子标志物(生物标志物)，这些生物标志物是早期疾病指标，当肿瘤处于可治疗阶段时可以检测到。[在一份报刊中，我们发现肾癌患者尿液中水通道蛋白1(AQP1)和脂联素(ADFP)显著增加，从而提供了该疾病的第一个敏感和特异的生物标志物。]因此，进一步假设，尿液外体，即由肾单位所有部分释放到尿液中的囊泡结构，将是[发现其他新的生物标志物蛋白的工具，并将作为肾细胞肿瘤发生和转移的非侵入性活检]。总体目标是在肾癌患者的尿外切体中识别出这种疾病所特有的蛋白质，这些蛋白质可以发展成为一种诊断试验，以筛查[全尿]以在早期治愈阶段发现肾癌，并跟踪患者术后是否复发。其具体目标是：1)在肾癌患者的尿液外体中识别引起疾病的候选生物标记物蛋白；2)开发灵敏和特异的ELISA以测量Aim1中确定的新生物标记物蛋白的水平；以及验证阶段3)表征候选生物标记物蛋白以确定其诊断的敏感性和特异性。唯一上调或下调的蛋白质将通过无偏的自上而下和有监督的自下而上的质谱学(LC/MS/MS)和蛋白印迹蛋白质组分析尿液外体蛋白来鉴定。一旦通过质谱学鉴定了患者训练组中的蛋白质，将开发出精确但经济的基于ELISA法的免疫分析方法，给出当天的结果，用于验证患者组中[全尿和血清]的蛋白质定量。总体而言，我们的研究将把肾癌的诊断提高到足够早的阶段，以便在最大限度地提高未来肾功能的同时，有很大的治愈机会[术后跟踪患者的复发情况，并确定未来选择性干预的目标]。 公共卫生相关性：肾癌通常是无症状的，通常是致命的。仅在美国，2009年就有超过57000例新诊断的肾癌和近13000例死亡可归因于这种疾病。体征和症状出现较晚，当肾癌确诊时，通常已转移到淋巴结或邻近器官，并进展到治愈阶段，生存机会很小。未得到满足的需求是一种简单、高吞吐量、成本效益高的肾癌筛查和复发测试，具有每年防止13,000多人死亡的公共卫生意义。我们的研究将在足够早的阶段改善肾癌的诊断，以便有很大的机会治愈，保留剩余的肾功能[并跟踪术后患者的复发和治疗，同时为肾癌的发生和转移提供重要的洞察力]。

DESCRIPTION (provided by applicant): Renal cancer is typically silent and frequently fatal. In the United States in 2009 alone over 57,000 new kidney cancers were diagnosed and almost 13, 000 deaths were attributable to this disease. Early diagnosis, triggering surgical excision, is usually curative. Physical signs and symptoms occur late, and by the time renal cancer is consequently diagnosed, it has usually metastasized to lymph nodes or adjacent organs and advanced beyond a curative stage with minimal chance for survival. Incidental early discovery may occur fortuitously during abdominal imaging (CT, MRI, ultrasound) for unrelated causes. Nonetheless, there is presently no method for population screening and diagnosis of renal cancer, particularly at a curative stage. The unmet need is a simple, high through-put, cost-effective test for kidney cancer screening, with the public health significance of preventing over 13,000 deaths annually especially in at risk populations. The proposed research will test the hypothesis that renal cancer cells express and shed or actively secrete proteins into the urine that are signature molecular markers (biomarkers), and that these biomarkers are an early disease indicator, detectable when the tumor is at a treatable stage. [In an in press publication, we have shown that aquaporin 1 (AQP1) and adipophilin (ADFP) are significantly increased in urinary exosomes of patients with kidney cancer thus providing the first sensitive and specific biomarkers of this disease.] Therefore, it is further hypothesized that urine exosomes, vesicular structures released into the urine by all parts of the nephron, will be a [discovery tool for additional novel biomarker proteins and will serve as a non-invasive biopsy of kidney cell oncogenesis and metastasis]. The overall objective is to identify proteins in the urine exosomes of kidney cancer patients which are unique to this disease, and which can be developed into a diagnostic test to screen [whole urine] to detect kidney cancer at an early curable stage and to follow patients post-operatively for recurrence. The specific aims are a discovery stage to: 1) Identify candidate biomarker proteins in the urine exosomes of patients with kidney cancer which are pathogneumonic of the disease; a development stage to 2) Develop sensitive and specific ELISAs to measure levels of novel biomarker proteins identified in Aim1; and a validation stage to 3) Characterize the candidate biomarker proteins to determine their diagnostic sensitivity and specificity. Unique up- or down-regulated proteins will be identified by unbiased top-down and supervised bottom-up mass spectral (LC/MS/MS) and Western blot proteomic analysis of the urine exosomal proteins. Once proteins are identified in training sets of patients by mass spectrometry, precise but economical ELISA-based immunologic assays that give same-day results will be developed, for protein quantification [in whole urine and serum] in validation sets of patients. Overall, our study will improve the diagnosis of kidney cancer to a stage early enough to allow a significant chance for cure while maximizing future kidney function, [follow patients post-operatively for recurrence and identify future targets for selective intervention]. PUBLIC HEALTH RELEVANCE: Renal cancer is typically silent and frequently fatal. In the United States alone in 2009 there were over 57,000 new kidney cancers diagnosed and almost 13,000 deaths attributable to this disease. Physical signs and symptoms occur late, and by the time renal cancer diagnosed, it has usually metastasized to lymph nodes or adjacent organs and advanced beyond a curative stage with minimal chance for survival. The unmet need is a simple, high through-put, cost-effective test for kidney cancer screening and recurrence with the public health significance of preventing over 13,000 deaths annually. Our study will improve the diagnosis of kidney cancer at a stage early enough to allow a significant chance for cure, preserve remaining kidney function [and follow post-operative patients for recurrence and treatment while providing important insight into kidney cancer oncogenesis and metastasis].