Biomarker Discovery in Chronic Graft-vs-Host Disease

描述（由申请人提供）： 慢性移植物抗宿主病（cGVHD）发生在异基因造血干细胞移植（HSCT）后的50-70%的患者中，并且是发病率和死亡率的主要原因。cGVHD的临床表现是多样的和可变的，并且诊断、分期和对治疗干预的反应的评估一直难以标准化。没有迹象或症状或诊断工具可以预测cGVHD的发展或严重程度，或需要免疫抑制治疗。该提案的目标是通过鉴定与cGVHD的发作和严重程度相关的新型生物标志物来解决这些未满足的需求。研究的初始阶段将包括一个平行的发现策略，该策略采用外周血单核细胞（PBMC）中转录变化的全基因组分析以及高分辨率，高灵敏度的定量蛋白质组学分析，以确定血浆中与疾病活动相关的信息蛋白。这些转录变化和蛋白质将被优先考虑，以开发用于第二阶段验证研究的候选物列表，以确认并进一步定义一组标记物，这些标记物一起为cGVHD的诊断和严重程度和/或治疗反应的预测提供高灵敏度和特异性。本研究的目的1是使用双重方法鉴定外周血中用于诊断cGVHD的新生物标志物：1a）分析单核白细胞中的全局基因表达和可变剪接;和1b）定量深入分析血浆中的循环蛋白。将使用Affytron Human Exon 1.0 ST阵列对PBMC进行转录分析，并将通过质谱法对血浆进行蛋白质组学分析。目标2将集中于验证在cGVHD的独立、多中心队列研究中鉴定的候选生物标志物。候选基因和替代剪接变体将通过基于PCR的试验进行验证，候选蛋白质将通过基于抗体的试验进行验证。本申请是为了响应标题为“临床试验中的辅助研究（R 01）”的RFA-HL-08-001而提交的。“我们正在提出一个生物标志物发现和验证项目，重点是在allo HSCT后发生的cGVHD。本研究的受试者（病例和对照）将入组一项由NIH资助的单独多中心研究，该研究题为“改善慢性GVHD的结局评估”（R 01-CA 118953; Stephanie Lee博士，PI）。参与中心包括丹娜-法伯癌症研究所（DFCI）、西雅图癌症护理联盟（SCCA）/弗雷德哈钦森癌症研究中心（FHCRC）、斯坦福大学和明尼苏达大学。各研究中心的总入组人数将达到336例事件和336例流行cGVHD病例。将对患者进行为期三年的随访，并系统评价cGVHD状态和对cGVHD治疗的反应。慢性移植物抗宿主病（cGVHD）发生在异基因造血干细胞移植（HSCT）后的50-70%的患者中，并且是发病率和死亡率的主要原因。随着HSCT的使用和HSCT幸存者数量的增加，cGVHD的问题不仅对患者及其家属而且对负责其护理的专业医生和卫生保健机构都变得更加关注。本文提出的研究旨在改善cGVHD的诊断方法，监测疾病活动和cGVHD治疗的有效性。这些研究也可能深入了解cGVHD的潜在疾病，从而提出新的治疗策略。(End摘要）

DESCRIPTION (provided by applicant): Chronic graft-versus-host disease (cGVHD) occurs in 50-70% of patients following allogeneic hematopoietic stem cell transplantation (HSCT) and is a major cause of morbidity and mortality. The clinical manifestations of cGVHD are diverse and variable, and the diagnosis, staging and assessment of response to therapeutic interventions have been difficult to standardize. There are no signs or symptoms or diagnostic tools that predict the development or severity of cGVHD, or the need for immune suppression therapy. The goal of this proposal is to address these unmet needs by identifying novel biomarkers that are associated with the onset and severity of cGVHD. The initial phase of study will consist of a parallel discovery strategy that employs a genome-wide analysis of transcriptional changes in peripheral blood mononuclear cells (PBMC) together with a high-resolution, high-sensitivity quantitative proteomic analysis to identify informative proteins in plasma that correlate with disease activity. These transcriptional changes and proteins will be prioritized to develop a list of candidates for second phase validation studies to confirm and further define a panel of markers that together provide high sensitivity and specificity for the diagnosis of cGVHD and the prediction of severity and/or treatment response. Aim 1 of this study is to identify novel biomarkers in peripheral blood for the diagnosis of cGVHD using dual approaches: 1a) analysis of global gene expression and alternate splicing in mononuclear leucocytes; and 1b) quantitative in-depth profiling of circulating proteins in plasma. The transcriptional analysis will be performed on PBMC using the Affymetrix Human Exon 1.0 ST array, and the proteomic analysis will be performed on plasma by mass spectrometry. Aim 2 will focus on the validation of candidate biomarkers identified in an independent, multicenter cohort study of cGVHD. Candidate genes and alternate splice variants will be validated by PCR-based assays, and candidate proteins will be validated by antibody-based assays. This application is submitted in response to RFA-HL-08-001, titled "Ancillary Studies in Clinical trials (R01)." We are proposing a biomarker discovery and validation project that is focused on cGVHD occurring after allo HSCT. The subjects for this study, cases and controls, will be enrolled in a separate NIH funded multi-center study titled "Improving outcomes assessment in chronic GVHD" (R01-CA118953; Dr. Stephanie Lee, PI). Participating Centers include the Dana-Farber Cancer Institute (DFCI), the Seattle Cancer Care Alliance (SCCA)/Fred Hutchinson Cancer Research Center (FHCRC), Stanford University and the University of Minnesota. Total enrollment across the sites will reach 336 incident and 336 prevalent cGVHD cases. Patients will be followed for three years, and systematically evaluated for cGVHD status and response to cGVHD therapy. Chronic graft-versus-host disease (cGVHD) occurs in 50-70% of patients following allogeneic hematopoietic stem cell transplantation (HSCT) and is a major cause of morbidity and mortality. As the utilization of HSCT and number of HSCT survivors increases, the issue of cGVHD becomes a greater concern not only to patients and their families but also to the specialized physicians and health care facilities responsible for their care. The studies proposed here are aimed at improving methods for the diagnosis of cGVHD, monitoring disease activity and the effectiveness of cGVHD therapy. These studies may also yield insights into the underlying disorders responsible for cGVHD and thereby suggest new treatment strategies. (End of Abstract)