Reversing Autoimmune Diabetes with Gene Therapy Induced Islet Neogenesis

描述（由申请人提供）： 1型糖尿病（T1 D）是由胰腺胰岛素分泌细胞的自身免疫性破坏引起的。虽然外源性胰岛素替代可纠正高血糖，但生理葡萄糖反应性的恢复一直难以实现，直到胰岛替代疗法的出现，该疗法受到显著供体短缺、需要长期免疫抑制和胰岛长期存活率差的限制。我们已经成功地使用Ngn 3/β细胞素的体内基因转移作为在肝脏中产生胰岛素产生的胰岛样簇的方式，所述胰岛样簇在STZ糖尿病小鼠模型中逆转糖尿病。这一建议是基于这样的假设，即基因治疗诱导的胰岛，通过转基因介导的细胞因子信号转导的抑制来抵抗自身免疫性破坏，可以恢复T1 D中的eukaryotic。任何胰岛新生仍然容易受到细胞因子的破坏？从自身反应性T细胞中释放出来因此，阻断这些胰岛增生诱导细胞因子的细胞内信号传导可以防止这些新胰岛的破坏。SOCS-1和SOCS-3已经显示负调节胰岛中这些细胞因子的信号传导。因此，我们推断，Ngn 3诱导的胰岛新生可以通过NOD小鼠（1型糖尿病的自身免疫小鼠模型）中SOCS-1和SOCS-3的过表达来保护免受苦参碱诱导的细胞凋亡。因此，该提议的广泛目标是在自身免疫性糖尿病小鼠模型中诱导对细胞因子介导的细胞凋亡有抗性的内源性胰岛新生，作为恢复正常血糖和葡萄糖耐受的手段。这项研究的数据将为体内基因治疗诱导的工程胰岛新生作为自身免疫性1型糖尿病的可行治疗方法提供概念验证。本研究的具体目的是：1.通过将SOCS-1/SOCS-3通过辅助依赖性腺病毒（HDAd）载体递送到肝脏，通过保护神经生成素3（Neurogenin 3，Ngn 3）诱导的异位胰岛新生，恢复糖尿病NOD小鼠的正常血糖和糖耐量，并在体内评估葡萄糖刺激的胰岛素分泌。2.通过直接评估血浆和组织细胞因子水平沿着自身反应性和调节性T细胞功能，研究这些小鼠治疗后的免疫系统，阐明Ngn 3-BTC-SOCS治疗应答的潜在机制。还将通过将其脾细胞过继转移到NOD-Scid小鼠中来确定这些治疗小鼠中自身免疫性是否持续的评估。

DESCRIPTION (provided by applicant): Type 1 diabetes (T1D) results from an autoimmune destruction of pancreatic insulin-producing (-cells. Though exogenous insulin replacement corrects hyperglycemia, a restoration of physiological glucose responsiveness has been elusive until the advent of islet replacement therapy which is limited by a significant donor shortage, a requirement for long-term immunosuppression and poor long-term islet survival. We have successfully used in vivo gene transfer of Ngn3/Betacellulin as a modality to generate insulin-producing islet-like clusters in the liver that reverse diabetes in the STZ-diabetic mouse model. This proposal is based on the hypothesis that gene therapy-induced islets, engineered to resist autoimmune destruction by transgene-mediated suppression of cytokine signaling, can restore euglycemia in T1D. Any islet neogenesis is still susceptible to destruction by cytokines ?released from auto-reactive T-cells. Blocking the intracellular signaling of these apoptosis-inducing cytokines could therefore prevent the destruction of these new islets. SOCS-1 & SOCS-3 have been shown to negatively regulate the signaling of these cytokines in islets. We, therefore, reasoned that Ngn3-induced islet neogenesis could be protected from cytokine-induced apoptosis by over-expression of SOCS-1 and SOCS-3 in the NOD mouse, an autoimmune mouse model of type 1 diabetes. The broad goal of this proposal, therefore, is to induce endogenous islet neogenesis that is resistant to cytokine mediated apoptosis, as a means to restore euglycemia and glucose tolerance, in an autoimmune diabetes mouse model. The data from this research will provide a proof-of-concept for in vivo gene therapy-induced engineered islet neogenesis as a viable curative therapy for autoimmune type 1 diabetes. The specific aims of this proposal are: 1.To restore euglycemia and glucose tolerance in diabetic NOD mice by protecting the ectopic islet neogenesis, induced using Neurogenin3 (Ngn3), with SOCS-1/SOCS-3 delivered to the liver via a helper dependent adenoviral (HDAd) vector and assess glucose-stimulated insulin secretion in vivo. 2. To elucidate the mechanism underlying the response to Ngn3-BTC-SOCS treatment by studying the immune system after therapy in these mice by directly assessing plasma and tissue cytokine levels along with autoreactive and regulatory T-cell functions. An assessment whether autoimmunity persists in these treated mice will also be established by adoptive transfer of their splenocytes into NOD-Scid mice.