Combined siRNA Therapy and In-vivo Imaging in Islet Transplantation

描述（由申请人提供）：糖尿病的特点是选择性地破坏产生胰岛素的β细胞，导致胰岛素分泌不足，从而导致高血糖。随着胰岛移植成为1型糖尿病患者恢复正常血糖的一种可接受的临床方式，由于手术后胰岛移植的严重损失，因此迫切需要对胰岛移植的预后进行无创影像学评估。与此同时，利用RNA干扰的基因治疗的潜力有望沉默有害基因并提高移植后胰岛移植物对损伤因素的抵抗力。在本应用中，我们首次提出将治疗与体内成像相结合，合成并测试一种新型基因治疗成像探针，该探针除了具有沉默特性外，还可以作为成像造影剂，能够检测和跟踪移植后胰岛移植物的体内命运。该建议基于先前开发的体内磁共振成像（MRI）方法来评估胰岛移植物的命运（1;2;4）。该提议的治疗部分是基于我们之前合成用于siRNA转移到癌细胞并随后进行肿瘤体内成像的双用途探针的经验(3)。在这里，我们提出合成和测试一种多功能siRNA成像探针，该探针由磁性纳米颗粒共价连接到特定于模型或治疗靶点的siRNA分子。作为siRNA靶标，我们建议使用负责潜在移植物损伤的分子。具体来说，我们将针对2-2微球蛋白（以防止可能的免疫攻击），凋亡蛋白caspase 3， caspase 8和Fas。由于纳米颗粒被胰岛细胞高效、无毒、无特异性地吸收，我们预计siRNA疗法将保护即将移植的移植物免受自身免疫攻击和异体排斥反应。因此，本研究的总体目标有两个方面：首先，我们提出对胰岛进行遗传操作，以降低其移植后对损伤因素的易感性；其次，利用磁共振成像技术监测糖尿病动物移植后胰岛的体内存活。新型的“二合一”sirna成像探针能够将遗传和对比成像材料传递到胰岛，将用于实现这些目标。

DESCRIPTION (provided by applicant): Diabetes mellitus is characterized by the selective destruction of insulin-producing beta- cells, which leads to a deficiency in insulin secretion and as a result, to hyperglycemia. As islet transplantation becomes an acceptable clinical modality for restoring normoglycemia in Type 1 diabetic patients, there is a critical need for non-invasive imaging assessment of the fate of islet grafts because of significant graft loss after the procedure. At the same time, the potential of gene therapy utilizing RNA interference holds promise for silencing harmful genes and improving islet graft resistance to damaging factors after transplantation. In this application we propose for the first time to combine therapy and in vivo imaging and synthesize and test a novel gene therapy imaging probes that in addition to its silencing properties can also serve as an imaging contrast agent capable of detecting and following the fate of islet grafts in vivo after transplantation. This proposal is based on a previously developed in vivo magnetic resonance imaging (MRI) approach to assess the fate of islet grafts (1; 2; 4). The therapeutic part of this proposal is based on our prior experience in synthesizing dual- purpose probes for siRNA transfer to cancer cells with subsequent in vivo imaging of tumors (3). Here we propose to synthesize and test a multifunctional siRNA-imaging probe consisting of magnetic nanoparticles covalently linked to an siRNA molecule(s) specific for model or therapeutic targets. As siRNA targets we propose to use molecules responsible for potential graft damage. Specifically, we will target 2-2microglobulin (to prevent possible immune attack), the apoptotic proteins caspase 3, caspase 8 and Fas. Since nanoparticles are taken up by islet cells with high efficiency, no toxicity and no specificity, we expect that siRNA therapy will protect a to-be transplanted graft from autoimmune attack and allogeneic rejection. Therefore, the overall goal of this proposal is two fold: first, we propose to genetically manipulate pancreatic islets with the purpose to reduce their susceptibility to damaging factors after transplantation, and second, to monitor in vivo their survival after transplantation in diabetic animals using magnetic resonance imaging. The novel "two-in-one" siRNA-imaging probe capable of delivering genetic and contrast imaging material to the islets will be used to achieve these goals. PUBLIC HEALTH RELEVANCE The biggest challenge in pancreatic islet transplantation as it becoming an accepted clinical procedure to treat patients with Type 1 Diabetes is significant graft loss after the procedure. This proposal will combine therapy based on silencing certain genes responsible for islet death and in vivo imaging to monitor transplanted grafts. We propose to synthesize and test a novel gene therapy imaging probes that in addition to its silencing properties can also serve as an imaging contrast agent capable of detecting and following the fate of islet grafts in vivo after transplantation.