Combined siRNA Therapy and In-vivo Imaging in Islet Transplantation

描述（由申请人提供）：糖尿病的特征在于产生胰岛素的β细胞的选择性破坏，这导致胰岛素分泌不足，并因此导致高血糖症。随着胰岛移植成为恢复1型糖尿病患者正常血糖的可接受的临床模式，由于手术后显著的移植物损失，迫切需要对胰岛移植物的命运进行非侵入性成像评估。同时，利用RNA干扰的基因治疗的潜力有望沉默有害基因，提高胰岛移植物对移植后损伤因素的抵抗力。在本申请中，我们首次提出将联合收割机治疗和体内成像结合起来，并合成和测试一种新的基因治疗成像探针，该探针除了其沉默特性之外，还可以用作能够检测和跟踪移植后体内胰岛移植物命运的成像造影剂。该建议基于先前开发的体内磁共振成像（MRI）方法，以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先前在合成用于siRNA转移至癌细胞并随后进行肿瘤体内成像的双重目的探针方面的经验（3）。在这里，我们建议合成和测试一种多功能siRNA成像探针，该探针由与模型或治疗靶点特异性siRNA分子共价连接的磁性纳米颗粒组成。作为siRNA靶点，我们建议使用负责潜在移植损伤的分子。具体来说，我们将靶向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吸收，无毒性和无特异性，我们期望siRNA治疗将保护待移植的移植物免受自身免疫攻击和同种异体排斥。因此，该提案的总体目标是两个方面：首先，我们提出基因操纵胰岛，目的是降低其移植后对损伤因素的易感性，其次，使用磁共振成像监测糖尿病动物移植后的体内存活率。新的“二合一”siRNA成像探针能够提供遗传和对比成像材料的胰岛将被用来实现这些目标。 公共卫生相关性随着胰岛移植成为治疗1型糖尿病患者的公认临床程序，胰岛移植的最大挑战是手术后显著的移植物丢失。这项建议将结合联合收割机治疗的基础上沉默某些基因负责胰岛死亡和在体内成像监测移植移植。我们建议合成和测试一种新的基因治疗成像探针，除了其沉默特性，也可以作为一种成像造影剂，能够检测和跟踪移植后体内胰岛移植物的命运。

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.