Identifying urinary biomarkers for early type 2 diabetic nephropathy

描述（由申请人提供）：候选人：Maryam Afkarian在圣路易斯的华盛顿大学获得医学博士学位，并在纽约长老会医院-威尔康奈尔医疗中心接受内科住院医师培训。她目前是马萨诸塞州总医院和布里格姆妇女医院联合项目的肾脏病学研究员。从2010年7月开始，她将被任命为马萨诸塞州总医院肾脏科的讲师。导师：Ravi Thadhani博士是国际公认的临床和翻译研究者，在指导新研究者方面有着良好的记录。他现在是，并将继续是一个密切参与的导师，致力于阿夫卡里安博士的培训，她提出的研究和职业发展的成功。 糖尿病肾病（diabetic nephropathy，DN）是糖尿病肾病的一种常见类型。然而，以目前的治疗标准，我们只能延缓这种疾病的进展。这部分是由于缺乏有效的治疗方法，部分是由于缺乏可靠的检测方法来诊断早期疾病，在肾损伤严重或不可逆之前。迫切需要新的诊断和治疗工具，这两者都需要更好地了解疾病的自然史，即确定新的发病和进展标志物。尿液的比较蛋白质组学分析（研究尿液蛋白质组成及其在疾病状态下如何变化）已用于鉴定几种肾脏疾病的标志物。Thadhani博士（导师）的研究小组以前曾使用这种方法来鉴定一种肽特征，这种肽特征在临床诊断前长达10年就能检测到早期糖尿病肾损伤的迹象。该提案提出了一种多阶段的方法来鉴定早期DN中改变的蛋白质。初步数据描述了用于尿液的灵敏蛋白质组学技术（iTRAQ：相对和绝对定量的同量异位素标签）的校准和适应。在目标1A中，iTRAQ将用于比较来自患有和不患有DN的患者（分别为病例和对照）的尿液样本。这种无假设和无偏倚的比较分析将产生早期DN的候选生物标志物的初步列表。在目标1B中，该列表将补充假设驱动的候选人，即具有参与DN的显著证据的候选人。实例是蛋白质，其（a）是导致DN的途径的一部分，（B）在DN中具有改变的mRNA表达或（c）在DN中具有改变的蛋白质水平。该步骤用于将所有当前可用的信息并入该生物标志物发现工作中。在目标3中，使用靶特异性和定量测定，例如ELISA，在独立队列中重新评价来自目标1A和1B的推定生物标志物的表达。这一步骤将有助于剔除假阳性，并确定一个子集的标志物，其差异表达在DN中得到确认。在目的4中，将在疾病过程中表征这些确认的标志物的表达。该提案旨在产生一组已在原始研究人群中验证的早期DN候选生物标志物。未来的一个步骤是在更广泛和更多样化的人群中测试这些候选人。从长远来看，我们希望验证的生物标志物将作为一种新的早期DN的诊断工具，也有助于阐明疾病的病理生理。 公共卫生相关性：糖尿病是美国终末期肾病和透析的主要原因。目前的医学测试只能在已经发生严重的、有时是不可逆的损害之后才能诊断出这种疾病。我们计划找到尿液标记蛋白，可用于检测糖尿病对肾脏损害的早期迹象，当它更有可能停止或逆转时。

DESCRIPTION (provided by applicant): Candidate: Maryam Afkarian received an MD-PhD from Washington University in St. Louis and underwent residency training in Internal Medicine at New York Presbyterian Hospital-Weill Cornell Medical Center. She is currently a Nephrology Research Fellow at the joint program of Massachusetts General Hospital and Brigham and Women's Hospital. Starting July 2010, she will be appointed as an instructor at the Nephrology Division of Massachusetts General Hospital. Mentor: Dr. Ravi Thadhani is an internationally recognized clinical and translational investigator with a strong track record of mentoring new investigators. He is, and will remain, a closely-involved mentor, committed to the success of Dr. Afkarian's training, her proposed studies and career development. Research: Diabetic nephropathy (DN) is the leading cause of end-stage kidney disease. However, with the current standards of care, we can only delay progression of this disease. This is partly due to a lack of effective therapies and partly due to absence of reliable assays for diagnosis of the early disease, before renal damage is severe or irreversible. New diagnostic and therapeutic tools are urgently needed, both of which require an improved understanding of the disease natural history, i.e. identification of new markers of onset and progression. Comparative proteomic analysis of urine (study of the urine protein composition and how it is changed in the disease state) has been used to identify markers for several kidney diseases. Dr. Thadhani (mentor)'s group has previously used this approach to identify a peptide signature that detected signs of early diabetic kidney injury up to 10 years prior to clinical diagnosis. This proposal lays out a multi-stage approach to identify the proteins that are altered in early DN. The preliminary data describes calibration and adaptation of a sensitive proteomic technology (iTRAQ: isobaric Tags for Relative and Absolute Quantitation) for application to urine. In Aim 1A, iTRAQ will be used to compare urine samples from patients with and without DN (cases and controls, respectively). This hypothesis-free and unbiased comparative analysis will generate a preliminary list of candidate biomarkers for early DN. In Aim 1B, this list will be supplemented with hypothesis-driven candidates, i.e. those with significant evidence of involvement in DN. Examples are proteins which (a) are part of pathways incriminated in DN, (b) have altered mRNA expression in DN or (c) have altered protein level in DN. This step serves to incorporate all currently available information into this biomarker discovery effort. In Aim 3, a target-specific and quantitative assay, e.g. ELISA, is used to re-evaluate the expression of putative biomarkers from Aims 1A and 1B in an independent cohort. This step will help weed out the false positives and identify a subset of markers whose differential expression in DN is confirmed. In Aim 4, the expression of these confirmed markers will be characterized over the course of disease. This proposal is designed to generate a group of candidate biomarkers for early DN which have been validated within the original study population. A future step would be to test these candidates in a broader and more varied population. In long term, we hope that the validated biomarkers would serve as a novel diagnostic tool for early DN and also help shed light on the disease pathophysiology. PUBLIC HEALTH RELEVANCE: Diabetes is the leading cause of end-stage kidney disease and dialysis in the US. The current medical tests can diagnose the disease only after significant and sometimes irreversible damage has already occurred. We plan to find urine marker proteins that can be used to detect early signs of diabetic damage to the kidneys, when it is more likely to be stopped or reversed.