Expression Profiling In Acute and Chronic Cardiac Allograft Rejection

急性和慢性心脏同种异体移植排斥反应中的表达谱

• 批准号: 8565288
• 负责人: Michael Solomon
• 金额: --
• 依托单位: CLINICAL CENTER
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8565288
• 关键词: Acute; Allograft Tolerance; Allografting; Bioinformatics; Biological Markers; Biopsy; Blood Vessels; Cardiac; Cardiac Catheterization Procedures; Chronic; Clinical; Diagnosis; Diagnostic tests; Dyspnea; Echocardiography; Electrocardiogram; Enrollment; Fatigue; Future; Gene Expression Profile; Genes; Gold; Heart Transplantation; Heart failure; Histopathology; Image; Immunohistochemistry; Immunologics; Individual; Institutional Review Boards; Investigation; Laboratories; Laboratory Procedures; Low Grade Fever; Methodology; Methods; Microarray Analysis; Molecular Profiling; Morbidity - disease rate; Myocardial Infarction; Nuclear; Oligonucleotide Microarrays; Pathogenesis; Pathway interactions; Patients; Peripheral Blood Mononuclear Cell; Phase; Phosphorus; Procedures; Process; Proteomics; Protocols documentation; RNA; Regulatory Pathway; Research Infrastructure; Reverse Transcriptase Polymerase Chain Reaction; Risk; Sampling; Source; Spectrum Analysis; Survivors; Symptoms; T-Lymphocyte; Techniques; Testing; Therapeutic immunosuppression; Translating; Transplant Recipients; Transplantation; Ultrasonography; Universities; Vascular Diseases; Ventricular; Western Blotting; allograft rejection; base; density; diagnosis standard; drug development; experience; functional genomics; heart allograft; mortality; programs; success; sudden cardiac death; tool

Acute cardiac allograft cellular rejection remains a significant source of morbidity and mortality within the first year after heart transplantation. Afterwards, cardiac allograft vasculopathy (CAV), as a result of chronic vascular rejection, is the major cause of morbidity and mortality. Within the first year post-transplantation, almost two-thirds of recipients will experience at least one rejection episode. At five years post-transplantation, nearly 50% of survivors will have CAV. In the first year after transplantation, nearly 63% of patients experience at least one episode of cardiac rejection and approximately one third of these patients will have multiple episodes. The clinical symptoms of acute cardiac rejection are relatively nonspecific (fatigue, dyspnea, low grade fever). Most CAV patients remain asymptomatic until they develop serious problems such as myocardial infarction, heart failure, ventricular dysrhythmias or sudden cardiac death. No widely accepted noninvasive method exists for the accurate diagnosis of acute or chronic cardiac rejection. Noninvasive methods such as electrocardiography, echocardiography, and nuclear studies all have been studied, but have been unsuccessful, thus far, for either condition. Several methodologies have been studied including electrocardiography, echocardiography, nuclear imaging, and phosphorus spectroscopy without success. The current gold standard for the diagnosis of acute cellular rejection remains right ventricular endomyocardial biopsy. This is an invasive method of diagnosis subject to morbidity and random sampling and interpretation error. Likewise, the gold standard for diagnosing CAV is cardiac catheterization with intravascular ultrasound, an invasive procedure also subject to morbidity. We are applying functional genomics to study acute cardiac allograft cellular rejection and CAV. Our group has developed and tested standard laboratory procedures for sample processing and if necessary amplification. We have established laboratory and bioinformatics infrastructure to support oligonucleotide microarray investigations. Two major local transplant programs (Johns Hopkins University and INOVA-Fairfax) have been or are currently collaborating. We have an IRB approved protocol and are currently actively enrolling patients (184 individuals enrolled to date). We hypothesize that large scale expression profiling of circulating peripheral blood mononuclear cells (PBMC, predominantly T lymphocytes) will identify genes that can serve as reliable biomarkers of acute and chronic cardiac cellular rejection. In the initial bench phase of the project, PBMCs are obtained from heart transplant recipients during periods of immunological tolerance of the allograft (no acute rejection) and immunologic intolerance of the allograft (acute rejection) and from heart transplant recipients with and without CAV to determine whether unique gene expression patterns are associated with each state. Other analytic tools that may be employed include proteomics, RT-PCR, Western blot, insitu-hybridization, immunohistochemistry, and histopathology. In the latter phase of the project we hope to translate these profiles into an acceptable bedside test for acute and chronic cardiac allograft cellular rejection. In addition to developing a biomarker approach to the diagnosis of rejection in cardiac transplant patients, expression profiling has the potential to identify immunoregulatory pathways that can serve as new targets for immunosuppressive therapy (rational drug development). In 2010, we processed 64 samples for high density oligonucleotide microarray analysis. In the latter part of 2010, a second batch containing 168 samples was processed to TRNA, the required step before going to microarrays. In 2011, an additional 139 samples were processed to TRNA and 89 samples were further processed to high density oligonucleotide microarray analysis. Since October 2011 through August 2012 we have processed an additional 44 samples to TRNA and processed 397 TRNA samples to an RNA stable state. The protocol continues to actively enroll subjects.

急性心脏同种异体移植细胞排斥仍然是心脏移植后第一年内发病率和死亡率的重要来源。此后，慢性血管排斥导致的同种异体心脏移植血管病（CAV）是发病和死亡的主要原因。 在移植后的第一年内，几乎三分之二的受者会经历至少一次排斥反应。 移植后五年，近 50% 的幸存者将患有 CAV。在移植后的第一年，近 63% 的患者至少经历一次心脏排斥反应，其中大约三分之一的患者会多次发作。急性心脏排斥反应的临床症状相对不具有特异性（疲劳、呼吸困难、低烧）。大多数 CAV 患者在出现心肌梗塞、心力衰竭、心室心律失常或心源性猝死等严重问题之前一直没有症状。 目前还没有广泛接受的非侵入性方法来准确诊断急性或慢性心脏排斥反应。心电图、超声心动图和核研究等非侵入性方法都已被研究过，但迄今为止对于这两种情况均未成功。 已经研究了几种方法，包括心电图、超声心动图、核成像和磷光谱，但没有成功。目前诊断急性细胞排斥反应的金标准仍然是右心室心内膜心肌活检。 这是一种侵入性诊断方法，容易出现发病率、随机抽样和解释错误。 同样，诊断 CAV 的黄金标准是采用血管内超声进行心导管插入术，这是一种侵入性手术，也容易引起发病。 我们正在应用功能基因组学来研究急性心脏同种异体移植细胞排斥和 CAV。我们的团队开发并测试了用于样品处理和必要时扩增的标准实验室程序。 我们建立了实验室和生物信息学基础设施来支持寡核苷酸微阵列研究。 两个主要的当地移植项目（约翰·霍普金斯大学和 INOVA-Fairfax）已经或正在合作。 我们拥有 IRB 批准的方案，目前正在积极招募患者（迄今为止已招募 184 名患者）。 我们假设，循环外周血单核细胞（PBMC，主要是 T 淋巴细胞）的大规模表达谱将鉴定出可作为急性和慢性心脏细胞排斥的可靠生物标志物的基因。在该项目的初始试验阶段，从同种异体移植物的免疫耐受（无急性排斥）和同种异体移植物的免疫不耐受（急性排斥）期间的心脏移植受者以及有或没有CAV的心脏移植受者中获得PBMC，以确定独特的基因表达模式是否与每个状态相关。 可以使用的其他分析工具包括蛋白质组学、RT-PCR、蛋白质印迹、原位杂交、免疫组织化学和组织病理学。 在该项目的后期阶段，我们希望将这些概况转化为可接受的急性和慢性心脏同种异体移植细胞排斥的床边测试。除了开发诊断心脏移植患者排斥反应的生物标志物方法外，表达谱还有可能识别免疫调节途径，从而作为免疫抑制治疗（合理药物开发）的新靶点。 2010 年，我们处理了 64 个样品进行高密度寡核苷酸微阵列分析。 2010 年下半年，第二批包含 168 个样本的样本被处理为 TRNA，这是进入微阵列之前所需的步骤。 2011 年，另外 139 个样品被处理为 TRNA，89 个样品被进一步处理为高密度寡核苷酸微阵列分析。 自 2011 年 10 月至 2012 年 8 月，我们已将另外 44 个样本处理为 TRNA，并将 397 个 TRNA 样本处理为 RNA 稳定状态。 该协议继续积极招募受试者。