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

基本信息

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
项目类别：
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

项目摘要

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（由适用提供）：我们建议将生物工程和人类免疫学的方法和见解应用于手术疗法，即组织的移植。我们的目标是通过开发siRNA的离体靶向纳米颗粒转化，从而在人类移植物内皮细胞（EC）内产生安全，高效，选择性和持续的敲低免疫刺激性蛋白质，从而减少人性化的临床前模型中的合金排斥。排斥反应仍然是移植物丧失和宿主免疫抑制的当前方案的重要原因。我们的新方法将通过修改移植物的同种抗原性来减少排斥。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局限性。首先，成年人，但不是实验性啮齿动物，具有循环效应的记忆T细胞，能够直接识别非层次的组织兼容性复合物（MHC）分子，并在激活后引起图形排斥。人们认为，同种异体记忆细胞的高频率可以解释啮齿动物成功的许多疗法的失败。其次，与啮齿动物EC不同，人类内皮细胞（EC）表达并直接呈现非级别的II MHC分子以循环效应的记忆T细胞，启动拒绝并绕过嫁接的树突状细胞的需求，“乘客“乘客“乘客”白血细胞”）与人类的模型中的培养型托管型托管。揭示了EC表达的共刺激剂和EC衍生的细胞因子以及EC表达的MHC分子在T细胞激活中的关键作用。与移植ECS的初步接触。换句话说，在周期期间，EC的免疫刺激性或调节分子的表达变化可能会对移植结果产生持久影响。在当前的临床实践中，移植脉管系统用器官保存冲洗解决方案使整个移植物中的EC与完美接触。我们将使用可生物降解的聚合物纳米颗粒进行体内递送的条件，以有效地翻译人体血管内部的EC，并通过当前转化方法实现EC表型的持续变化（特定的目标1）；我们将使用这种方法来击倒特定的免疫调节分子，例如CIITA，LFA-3，RAPTOR和/或IL-1A，在培养的人ECS中，以及与常规EC转换相比，对同种异体记忆T细胞的激活对同种异体记忆T细胞激活的影响（特定目标2）；并且我们将使用纳米颗粒介导的转化来击倒在EC 2中识别为AIM 2的分子，然后在人类动脉衬里的EC 2中，在植入与人类免疫系统对动脉供体的同种异体系统中重构的小鼠之前，在体内离体，评估了对急性和亚肌脱落的作用（特定的目标3）。这些临床前的研究将为我们的新方法提供概念证明，以改善同倍解植物的结果。