Ex Vivo Nanoparticle Drug Delivery Targeted to Human Renal Allograft Endothelium

针对人肾同种异体移植物内皮的体外纳米颗粒药物输送

• 批准号: 10155842
• 负责人: JORDAN S POBER
• 金额: $ 2.94万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10155842
• 关键词: Acute; Address; Alleles; Allografting; Antibodies; Antibody titer measurement; Antibody-Producing Cells; Antigen Targeting; Antigens; Arteries; Binding; Biochemical; Blood; Blood Vessels; Blood capillaries; Body Temperature; Caliber; Cell Culture Techniques; Cells; Chronic; Clinical; Clinical Trials; Collaborations; Complement; Complement Activation; Complement Membrane Attack Complex; Data; Deposition; Development; Dose; Drug Delivery Systems; Early Endosome; Endothelial Cells; Endothelium; Erythrocytes; Ethics; Excision; Funding; Gamma globulin; Genes; Goals; Graft Survival; Grant; HLA Antigens; Harvest; Histocompatibility; Hour; Human; Immune; Immunodeficient Mouse; In Situ; In Vitro; Individual; Inflammation; Infusion procedures; Injury; Intervention; Intravenous; Intravenous Immunoglobulins; Ischemia; Kidney; Kidney Failure; Kidney Transplantation; Leukocytes; Mediating; Membrane Proteins; Methods; Microfluidics; Mus; Organ; Organ Harvestings; Outcome; PECAM1 gene; Patients; Perfusion; Perioperative; Pharmaceutical Preparations; Plasmapheresis; Polymers; Population; Postoperative Period; Prevention; Process; Protective Agents; Protein-Serine-Threonine Kinases; Proteins; Pump; Reagent; Recovery; Research; Resistance; Risk Factors; Signal Pathway; Signal Transduction; Small Interfering RNA; T-Lymphocyte; Testing; Therapeutic; Therapeutic Agents; Time; Transplant Recipients; Transplantation; Transplantation Surgery; United States National Institutes of Health; Universities; Validation; Vascular Endothelial Cell; anti-endothelial cell antibody; antibody conjugate; base; cell injury; cell type; cost; drug candidate; experimental study; fluorophore; gene product; graft failure; humanized mouse; immunoregulation; improved; improved outcome; in vivo; inflammatory milieu; inhibitor/antagonist; kidney allograft; kidney vascular structure; mouse model; nanomedicine; nanoparticle; nanoparticle delivery; nanoparticle drug; neutrophil; novel strategies; pre-clinical; prevent; recruit; response; small molecule; small molecule therapeutics; success; targeted delivery; targeted treatment

A significant proportion of patients with renal failure who would benefit from kidney transplantation are highly pre-sensitized, i.e. they have high titers of pre-formed circulating antibodies (Abs) reactive with 80% or more of non-self allelic forms of class I and/or class II HLA antigens called panel reactive Abs (PRA). Upon transplantation, host PRA will bind to graft HLA antigens that are highly expressed on graft endothelial cells (ECs) where they activate host human complement resulting in deposition of membrane attack complex (MAC) on the ECs. Human MAC does not lyse human ECs, but instead alters them to express gene products that promote inflammation. The inflammatory milieu favors activation of adaptive immune effectors at the expense of protective immunoregulation. Consequently, if transplanted, patients with high titer PRA have increased episodes of acute and chronic rejection resulting in more graft failure and graft loss. Current therapeutic approaches include plasmapheresis to reduce the titer of circulating PRA, targeted elimination of Ab-producing cells and/or administration of high doses of intravenous gamma globulin to reduce inflammation, but PRA titers return and these interventions have had only limited impact on outcomes. We propose a novel strategy to complement these approaches, namely to reduce the response of graft ECs to PRA/MAC by reducing expression of HLA antigen targets and/or by inhibiting PRA/MAC signaling in ECs. The latter approach is based upon our elucidation of the relevant signaling pathways. To accomplish this, we will develop safe, polymeric nanoparticles (NPs) that are targeted towards graft ECs by means of conjugated anti-EC Abs and use these NPs to deliver siRNAs or small molecule therapeutics (“drugs”) during a period of ex vivo normothermic perfusion (EVNP), an approach that is being applied to improve energy stores in kidneys and other organs from deceased donors prior to transplantation. The NPs, which will be bound to and internalized by the graft ECs, will then serve as a depot for sustained release of the therapeutic agent for a period sufficient to allow graft accommodation and/or host immunoregulation to develop. In Specific Aim 1, we will use human EC cultures and human artery segments interposed into the aortae of immunodeficient mice to identify the optimal siRNAs or drugs that can protect ECs from PRA. Our initial target will be prevention of Akt activation, a key step in PRA/MAC signaling. In Specific Aim 2, we will identify optimal Abs for targeting renal human ECs and use these to identify conditions for efficient pan-EC delivery in human kidneys unsuitable for clinical transplantation that are subjected to EVNP by our collaborators at the University of Cambridge. (U01 support will be used only for experiments and analyses conducted at Yale; the costs of experimental EVNP will be provided by our Cambridge colleagues who are supported by a grant from the UK National Institute for Health Research and experimental EVNP will be conducted at the University of Cambridge under their Ethics Approval.) If successful, this approach can be extended to other uses and could justify a human clinical trial.

在肾脏移植中受益的肾衰竭患者中，很大一部分是高度的 预敏敏感，即​​它们具有高滴度的预成型循环抗体（ABS），有80％或以上的反应性 I类和/或II类HLA抗原的非自身的Allic形式称为反应性ABS（PRA）。之上 移植，宿主PRA将与高度表达移植物内皮细胞的移植物HLA抗原结合 （EC）在激活宿主的人体完成中，导致膜攻击复合物的沉积（MAC） 在EC上。人类Mac不会绑扎人类EC​​，而是改变它们以表达基因产品 促进炎症。炎症环境有利于自适应免疫作用的激活 受保护的免疫调节。因此，如果移植，高滴度PRA的患者增加了 急性和慢性排斥反应的发作，导致更多的移植失败和移植物丢失。当前的疗法 方法包括血浆置换以减少循环PRA的滴度，目标消除AB产生 细胞和/或高剂量的静脉γ球蛋白减少炎症，但PRA滴答 返回，这些干预措施对结果的影响只有有限的影响。我们提出了一种新颖的策略 补充这些方法，即减少移植EC对PRA/MAC的反应 HLA抗原靶标和/或通过抑制EC中的PRA/MAC信号的表达。以后的方法是 基于我们阐明相关信号通路。为此，我们将开发安全， 通过共轭抗EC ABS和 使用这些NP在离体期间提供siRNA或小分子疗法（“药物”） 正常热灌注（EVNP），这种方法正在应用于改善肾脏的能量存储和 移植前来自已故捐赠者的其他器官。 NP，将被绑定到并内在化 然后，通过移植ECS，将充当持续释放治疗剂的沉积物 允许嫁接适应和/或宿主免疫调节。在特定目标1中，我们将使用人类 EC培养物和人动脉段插入了免疫缺陷小鼠的主动脉中，以鉴定 可以保护EC免受PRA的最佳siRNA或药物。我们的最初目标将是预防AKT激活，A PRA/MAC信号的关键步骤。在特定目标2中，我们将确定针对肾脏人类EC的最佳ABS 并使用这些来识别不适合临床的人类肾脏中有效pan-ec递送的条件 我们在剑桥大学的合作者进行EVNP的移植。 （U01支持 仅用于在耶鲁大学进行的实验和分析；实验EVNP的成本将是 由我们的剑桥同事提供 研究和实验EVNP将根据其道德规范在剑桥大学进行 批准。）如果成功，则可以将这种方法扩展到其他用途，并可以证明人类的临床试验合理。