PDL1-based rAAV-mediated gene therapy for mouse T1D

描述(申请人提供)：PD-1/PDL-1信号已被证明是激活T细胞功能的关键调节因子。PD-L1与PD-1的结合可下调效应T细胞的增殖和细胞因子的产生，诱导T细胞周期停滞和凋亡，或促进产生IL-10的调节性T细胞应答。然而，PDL1相关信号在T细胞对3细胞抗原的应答和T1D过程中的作用知之甚少。最近的研究表明，NOD小鼠APC上PDL1的表达缺失，胰岛特异的PDL1转基因表达显著减少了NOD小鼠的胰腺炎和预防糖尿病，阻断PD-1/PDL1信号通路可迅速加速NOD小鼠糖尿病的发生。我们的初步研究表明，激动型PDL1-LG在体外抑制了自发性Th1对p细胞抗原的反应，阻止了致糖尿病的T细胞周期，并在体内延缓了过继转移的T1D的发生。此外，重组腺相关病毒(RAAV)-PDLI-LG(而不是rAAV-LG)转导的同源胰岛移植延长了糖尿病NOD小鼠的胰岛移植物存活时间，rAAV-PDL1-LG治疗6周龄NOD小鼠诱导了PDL1-LG的稳定表达，抑制了疾病的进展，并与抑制自发T细胞自身免疫有关。因此，我们推测，用rAAV-PDL1-LG诱导PDL1-LG稳定表达可能灭活效应T细胞和/或诱导调节性T细胞反应，在NOD小鼠T1D前期抑制疾病进展，逆转T1D。由于AAV是一种非致病性病毒，免疫原性较弱，因此我们选择以AAV作为载体将PDL1-Lg或对照Ig基因转移到NOD小鼠体内，具有临床应用价值。AAV可以有效地感染多种类型的细胞，导致靶基因的长期表达。在这项研究中，我们将研究在NOD小鼠自身免疫过程的后期使用rAAV-PDL1-LG对NOD小鼠疾病进展和逆转新发糖尿病的影响，并确定PDL1相关信号在NOD小鼠T细胞自身免疫过程中作用的机制(S)。我们的研究将解决与PD-1/PDL1信号对自身免疫T细胞反应的调节功能有关的基本问题。我们的发现可能为抑制/逆转人类T1D的新的免疫疗法提供基础。外行人摘要：我们将集中研究一种T细胞自身免疫工程抑制剂对抑制/逆转小鼠模型1型糖尿病的治疗效果。我们的研究结果可能为设计针对T1D患者的特异性免疫疗法提供依据。

DESCRIPTION (provided by applicant): The PD-1/PDL-1 signaling has been shown to be a crucial regulator for activated T cell function. Engagement of the PD-1 by PD-L1 down-regulates effector T cell proliferation and cytokine production, and induces T cell cycle arrest and apoptosis or promotes IL-10-producing regulatory T cell responses. However, little is known the role of PDL1 -related signaling in T cell responses to (3-cell antigens and the T1D process. Recent studies have shown that NOD mice are deficient in expression of PDL1 on APCs, islet-specific transgenic expression of PDL1 significantly reduced insulitis and prevented diabetes in NOD mice, and treatment with blockade for PD-1/PDL1 signaling rapidly precipitated diabetes onset in young NOD mice. Our preliminary studies showed that agonistic PDL1-lg inhibited spontaneous Th1 response to p-cell antigens, arrested diabetogenic T cell cycling in vitro, and treatment with PDL1-lg retarded the onset of adoptively transferred T1D in vivo. In addition, transplantation with syngenic islets transduced by recombinant adeno-associated virus (rAAV)-PDLI-lg, but not rAAV-lg, prolonged islet-graft survival in diabetic NOD mice and treatment of 6 weeks old NOD mice with rAAV-PDL1-lg induced stable expression of PDL1-lg, inhibited disease progression, associated with inhibition of spontaneous T cell autoimmunity. Hence, we hypothesize that treatment with rAAV-PDL1-lg to induce PDL1-lg stable expression may inactivate effector T cells and/or induce regulatory T cell responses, inhibiting disease progression at advanced pre-T1D and reverse T1D in NOD mice. We choose to use AAV as the vector to deliver PDL1-lg or control Ig genes to NOD mice because AAV is a non-pathogenic virus and has little immunogenicity, making it attractive for clinical applications. AAV can effectively infect broad types of cells, leading to a long-term expression of target genes. In this proposal, we will examine the effect of treatment with rAAV- PDL1-lg at the late stage of autoimmune process on disease progression and on reversal of newly diabetes in NOD mice and determine the mechanism(s) underlying the action of PDL1-related signaling on the process of T cell autoimmunity in NOD mice. Our studies will address fundamental questions concerning the regulatory function of PD-1/PDL1 signaling on autoimmune T cell responses. Our findings may provide the basis for novel immunotherapies for the inhibition/reversal of T1D in human. Layperson's abstract: We will center on examining the therapeutic effect of an engineered inhibitor for T cell autoimmunity on inhibition/reversal of type 1 diabetes in mouse model. Our findings may provide a basis for the design of specific immunotherapies for T1D patients.