Engineering dendritic cells to target islet antigen to pro-tolerogenic subsets for prevention and treatment of Type 1 Diabetes

项目摘要 尽管通过将这些抗原靶向树突细胞（DC）来诱导抗原特异性耐受性， 尽管在多项临床前研究中有报道，但目前在临床中使用DC还没有有效的方法 以防止或逆转自身免疫。事实证明，发展中国家本身就具有恢复和 维持免疫耐受性，使它们成为治疗自身免疫性疾病的非常有吸引力的候选者。 然而，只有未活化的DC（其固有地具有致耐受性/抑制性表型）可以诱导免疫应答。 免疫耐受，而活化的DC可以触发免疫。值得注意的是，没有能够递送抗原的策略 只有未激活的DC。为了克服这一限制，本提案的长期目标是设计 树突状细胞仅通过致耐受性亚群呈递胰岛源性抗原治疗1型糖尿病 （T1D）。中心假设是通过确保胰岛衍生抗原仅由致耐受性细胞呈递， DC，这些DC将导致胰岛特异性自身反应性T细胞的消除和保护性T细胞的诱导。 本提案的总体目标将通过实现以下两个具体目标进行测试：（1）工程 使用将表达限制于促耐受原性DC的电路使DC表达胰岛衍生抗原，以及（2） 工程化DC以人工地实施促致耐受性不可逆表型，同时还驱动 胰岛衍生抗原。对于目标1，含有新启动子的构建体驱动表达至致耐受性DCs， 结合基于microRNA的电路，防止抗原表达对炎症的反应， 测试耐受性的诱导。对于Aim 2，可以强制DC进入高度亲核的转录因子， 致耐受性状态将用于编码胰岛衍生抗原的构建体中以在T1 D小鼠中工程化DC 模型这种转录调节因子在肿瘤内DCs中被鉴定，并且发现其强制激活， 将DC塑造成免疫抑制状态，即使在强烈的炎症反应中也不能恢复。 刺激。拟议的研究是创新的，因为它首次尝试设计DC，以确保 仅通过促耐受原性细胞呈递胰岛衍生抗原。这项建议的意义在于，如果 如果成功，它将确定一种安全地设计DC以诱导耐受的策略，并将提供一种 强大的科学框架，用于进一步研究，以设计这些免疫系统的关键协调机制， 系统最终，从这项工作中获得的知识有可能为创新奠定基础。 基于工程的疗法来阻止T1 D的自身免疫，这是实现治愈的重要步骤。

Project Summary Although induction of antigen-specific tolerance by means of targeting such antigens to dendritic cells (DCs) has been reported in multiple pre-clinical studies, there are currently no effective approaches using DCs in the clinic to prevent or reverse autoimmunity. DCs have been proven to intrinsically possess the ability to restore and maintain immune tolerance, making them very attractive candidates for treatment of autoimmune diseases. However, only non-activated DCs (which inherently possess a tolerogenic/suppressive phenotype) can induce tolerance, whereas activated DCs can trigger immunity. Notably, there is no strategy able to deliver antigen uniquely to non-activated DCs. To overcome this limitation, the long-term goal of this proposal is to engineer DCs to guarantee presentation of islet-derived antigen solely by tolerogenic subsets to treat Type 1 Diabetes (T1D). The central hypothesis is that by ensuring that islet-derived antigen is presented exclusively by tolerogenic DCs, these DCs will lead to elimination of islet-specific auto-reactive T cells and induction of protective Tregs. The overarching goal of this proposal will be tested by pursuing the following two specific aims: (1) Engineering DCs to express islet-derived antigen using circuits that limit expression to pro-tolerogenic DCs and (2) Engineering DCs to artificially enforce a pro-tolerogenic irreversible phenotype while also driving expression of islet-derived antigen. For Aim 1, constructs containing a new promoter driving expression to tolerogenic DCs in combination with microRNA-based circuits that prevent antigen expression in response to inflammation will be tested for induction of tolerance. For Aim 2, a transcription factor that can enforce DCs into a highly pro- tolerogenic state will be used in a construct encoding islet-derived antigen to engineer DCs in T1D mouse models. This transcriptional regulator was identified in intra-tumor DCs, and its enforced activation was found to shape DCs into an immune suppressive state that could not be reverted even in response to strong inflammatory stimuli. The proposed research is innovative because it attempts for the first time to engineer DCs to ensure presentation of islet-derived antigen only by pro-tolerogenic cells. The significance of this proposal is that, if successful, it will have identified a strategy to safely engineer DCs for induction of tolerance and will provide a strong scientific framework for further investigations to engineer these critical orchestrators of the immune system. Ultimately, the knowledge derived from this work has the potential to lay the ground for innovative engineering-based therapies to stop autoimmunity in T1D, which constitutes an essential step to achieve a cure.