Bioengineered siRNA/Nanoparticles to Prevent Human Transplant Rejection

生物工程 siRNA/纳米颗粒可防止人体移植排斥

• 批准号: 8693080
• 负责人: JORDAN S POBER
• 金额: $ 29.1万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-01 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8693080
• 关键词: Accounting; Acute; Address; Adult; Alloantigen; Allogenic; Allografting; Antigens; Arteries; Artificial nanoparticles; Biomedical Engineering; Blood Circulation; Blood Vessels; Bypass; CD58 gene; CD8B1 gene; Cell Line; Cells; Clinical Trials; Coculture Techniques; Cues; Cultured Cells; Development; Dose-Limiting; Drug usage; Endothelial Cells; Failure; Flushing; Frequencies; Gene Mutation; Goals; Graft Rejection; Graft Survival; Health; Histocompatibility Antigens Class II; Human; Immune; Immune response; Immune system; Immunodeficient Mouse; Immunologics; Immunology; Immunosuppression; Immunosuppressive Agents; In Situ; In Vitro; Injury; Interleukin-1; Interleukin-2; Interleukin-6; Investigation; Leukocytes; Major Histocompatibility Complex; Mediating; Memory; Methods; Modeling; Mus; Neoadjuvant Therapy; Operative Surgical Procedures; Organ Preservation Solutions; Organ Transplantation; Outcome; Pathologic; Pathway interactions; Patients; Pattern; Perfusion; Perioperative; Phenotype; Plastics; Population; Pre-Clinical Model; Preservation Technique; Proteins; RNA; Raptors; Regimen; Regulatory T-Lymphocyte; Rest; Risk; Rodent; Rodent Model; Role; Shapes; Signal Transduction; Signaling Protein; Simulate; Small Interfering RNA; T cell differentiation; T cell response; T memory cell; T-Cell Activation; T-Lymphocyte; Temperature; Testing; Time; Toxic effect; Transfection; Transplant Recipients; Transplantation; Work; allograft rejection; base; biodegradable polymer; clinical practice; clinically relevant; cytokine; ex vivo perfusion; experience; immunogenic; implantation; improved; injured; insight; knock-down; mouse model; nanoparticle; novel strategies; preclinical study; pressure; prevent; protein expression; reconstitution; research study; response; uptake

DESCRIPTION (provided by applicant): We propose to apply methods and insights of bioengineering and human immunology to a surgical therapy, namely organ transplantation. Our goal is to produce safe, efficient, selective and sustained knock down of immunostimulatory proteins within human graft endothelial cells (EC) by developing ex vivo targeted nanoparticle transfection of siRNA so as to reduce allograft rejection in humanized pre-clinical models. Rejection remains an important cause of graft loss and current regimens of host immunosuppression produce significant complications. Our novel approach will reduce rejection instead by modifying the alloantigenicity of the graft. By focusing on human-based models, we address two fundamental limitations of most rodent transplant models. First, adult humans, but not experimental rodents, have circulating effector memory T cells capable of directly recognizing non-self-major histocompatibility complex (MHC) molecules and, upon activation, causing graft rejection. The high frequency of alloreactive memory cells is thought to account for the failure in humans of many therapies successful in rodents. Second, human endothelial cells (ECs), unlike rodent ECs, express and directly present non-self-class II MHC molecules to circulating effector memory T cells, initiating rejection and bypassing the need for graft dendriti cells ("passenger leukocytes") to activate na¿ve host T cells seen in rodent models. Our experiments with cultured human ECs and with humanized mouse models of allograft rejection have revealed crucial roles for EC-expressed co-stimulators and EC- derived cytokines as well as EC-expressed MHC molecules in T cell activation. Furthermore, human effector memory T cells are still somewhat plastic and can be irreversibly directed along different pathways by their initial contact with graft ECs. In other words, changes in the expression of immune stimulatory or regulatory molecules by ECs in the perioperative period can have lasting effects on graft outcomes. In current clinical practice, the graft vasculature is flushed with an organ preservation solution so that ECs throughout the graft come in contact with the perfusate. We will optimize conditions for ex vivo delivery of siRNAs using biodegradable polymer nanoparticles engineered to efficiently transfect ECs lining human blood vessels and to produce a more sustained change in the EC phenotype achieved by current transfection approaches (specific aim 1); we will use this approach to knock down specific immunomodulatory molecules, examples being CIITA, LFA-3, raptor and/or IL-1a, in cultured human ECs and assess effects on the activation of allogeneic memory T cells in vitro, compared to conventional EC transfections (specific aim 2); and we will use nanoparticle- mediated transfection to knock down molecules identified as important in aim 2 in the ECs lining human artery segments ex vivo prior to implantation into mice reconstituted with a human immune system allogeneic to the artery donor, assessing the effect on acute and subacute graft rejection (specific aim 3). These pre-clinical studies will provide proof of concept for our novel approach to improve the outcome of allotransplantation.

描述（由申请人提供）：我们建议将生物工程和人体免疫学的方法和见解应用于外科治疗，即器官移植。我们的目标是通过体外靶向siRNA纳米颗粒转染，在人移植内皮细胞（EC）中产生安全、高效、选择性和持续的免疫刺激蛋白，从而减少人源化临床前模型中的异体移植排斥反应。排斥反应仍然是移植物损失的重要原因，目前的宿主免疫抑制方案产生明显的并发症。我们的新方法将减少排斥，而不是通过修改移植物的同种抗原。通过关注基于人类的模型，我们解决了大多数啮齿动物移植模型的两个基本限制。首先，成年人（而非实验啮齿类动物）具有循环效应记忆T细胞，能够直接识别非自我主要组织相容性复合体（MHC）分子，并在激活后引起移植物排斥反应。同种异体反应记忆细胞的高频率被认为是许多在啮齿动物身上成功的治疗方法在人类身上失败的原因。其次，与啮齿类动物的内皮细胞不同，人类内皮细胞（ECs）表达并直接向循环效应记忆T细胞呈递非自我II类MHC分子，启动排斥反应，绕过移植物树突细胞（“客运白细胞”）激活啮齿动物模型中所见的na - ve宿主T细胞的需要。我们对培养的人EC和同种异体移植排斥的人源化小鼠模型的实验揭示了EC表达的共刺激物和EC衍生的细胞因子以及EC表达的MHC分子在T细胞活化中的关键作用。此外，人类效应记忆T细胞仍然具有一定的可塑性，并且可以通过它们的免疫系统沿着不同的途径不可逆地引导