Improving islet transplantation outcome with Akt1

描述（由申请人提供）：摘要胰岛移植正在成为1型糖尿病（T1 D）的潜在治疗方法。然而，有限的供应和移植围手术期的胰岛损失是这种治疗策略的主要局限性。本研究旨在通过体外将组成型活性Akt 1（CA-Akt 1）导入产生胰岛素的β 2细胞来改善胰岛移植的治疗效果。丝氨酸/苏氨酸蛋白激酶Akt/PKB是PI 3激酶通路的直接下游靶点，具有抗凋亡和诱导细胞增殖的双重功能。Akt对2-细胞存活和增殖的相关性已经在转基因和敲除小鼠模型的研究中以及使用药理学方法证明。然而，为了实现CA-Akt 1的治疗潜力，需要一种安全、有效和特异性的载体来将Akt 1离体递送到胰岛2细胞中。在这方面，基于腺病毒血清型5（Ad 5）的载体引起了极大的兴趣。我们前期的研究已经证明，Ad 5载体AdRGDpK 7对人胰岛细胞的基因转移效率显著提高。因此，我们建议采用Ad 5 RGDpK 7将Akt 1递送到离体胰岛细胞中。为了进一步降低CA-Akt 1的潜在副作用，我们将通过使用2-细胞特异性启动子-大鼠胰岛素启动子（RIP）来驱动Akt 1的表达来限制外源性Akt 1在2-细胞中的表达。此外，我们建议共表达一种双重功能模式，HSV-TK，与CA-Akt 1，使其既可以用作一种非侵入性成像模式，以遵循移植的胰岛和自杀基因恶性肿瘤发生。因此，我们的具体目标是：1）开发2-细胞特异性的、感染性增强的Ad 5载体，其允许有效和特异性的CA-Akt 1和HSV-TK基因离体递送到2-细胞中; 2）检查上述开发的Ad 5载体促进胰岛存活和增殖同时使转化最小化的能力，从而增强胰岛移植的功效;以及3）评估上述开发的Ad 5载体在胰岛移植背景下的安全性。公共卫生相关性：很明显，胰岛移植在治愈1型糖尿病方面具有巨大的潜力。本研究旨在通过使用高效、特异和安全的基因递送载体，用保护性基因修饰胰岛，以改善胰岛移植的治疗效果。这项研究的成功预计将在胰岛移植治疗1型糖尿病领域产生重大影响。

DESCRIPTION (provided by applicant): Summary Islet transplantation is becoming a potential cure for type 1 diabetes (T1D). However, the limited supply and significant islet loss in the peritransplant period are cast as the major limitations of this treatment strategy. This project is aimed to improve the therapeutic outcome of islet transplantation by introducing constitutively active Akt1 (CA-Akt1) into the insulin producing 2-cells ex vivo. The serine/threonine protein kinase Akt/PKB is the direct downstream target of PI3 Kinase pathway, and has been found to have dual functions of anti-apoptosis and induction of cell proliferation. Relevance of Akt on 2-cell survival and proliferation has been demonstrated in studies of transgenic and knockout mouse models, as well as using pharmacological methods. Nonetheless, in order to realize the therapeutic potential of CA-Akt1, a safe, efficient and specific vector is needed to deliver Akt1 into islet 2-cells ex vivo. In this regard, adenovirus serotype 5 (Ad5)-based vector is of great interest. Our previous studies have demonstrated the modified Ad5 vector, AdRGDpK7, exhibited significantly higher gene transfer efficiency for the human islet cells. We thus propose to employ Ad5RGDpK7 to deliver Akt1 into islet cells ex vivo. To further diminish the potential adverse effect of CA-Akt1, we will restrict exogenous Akt1 expression in 2-cells by employment of 2-cell specific promoter-rat insulin promoter (RIP) to drive Akt1 expression. In addition, we propose to co-express a dual functional modality, HSV-TK, with CA-Akt1 so that it can be used as both a non-invasive imaging modality to follow the transplanted islets and a suicide gene should malignancy occur. Our specific aims are thus: 1) To develop a 2-cell specific, infectivity-enhanced Ad5 vector that allows efficient and specific CA-Akt1 and HSV-TK gene delivery into 2-cells ex vivo; 2) To examine the capacity of the Ad5 vector developed above to promote islet survival and proliferation while minimizing transformation, thus enhancing the efficacy of islet transplantation; and 3) To evaluate the safety of the Ad5 vector developed above in the context of islet transplantation. PUBLIC HEALTH RELEVANCE: It is clear that islet transplantation holds great potential for the cure of Type 1 Diabetes. This study seeks to improve the therapeutic outcome of islet transplantation by modifying the islets with protective genes using a highly efficient, specific and safe gene delivery vector. Success of this study is expected to have significant impact in the field of islet transplantation treatment for type 1 diabetes.