Alpha cells conversion to beta cells in non-human primates

非人类灵长类动物中的α细胞转化为β细胞

• 批准号: 9789262
• 负责人: GEORGE K. GITTES
• 金额: $ 39.17万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-30 至 2020-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9789262
• 关键词: Address; Alloxan; Alpha Cell; Anatomy; Animals; Autoimmune Process; Autoimmunity; Automobile Driving; Beta Cell; Biology; Blood Glucose; CMV promoter; Cell physiology; Cells; Clinical; Clinical Trials; Data; Dependovirus; Development; Diabetes Mellitus; Diabetic mouse; Disease; Duct (organ) structure; Ductal; Electrons; Endocrine; Endoscopic Retrograde Cholangiopancreatography; Feasibility Studies; Gene Delivery; Gene Expression; Gene Expression Profile; Gene therapy trial; Generations; Genetic; Glare; Glucagon; Histology; Human; Hyperplasia; Immunosuppression; Immunosuppressive Agents; In Situ; In Vitro; Inbred NOD Mice; Infusion procedures; Injections; Insulin-Dependent Diabetes Mellitus; Intervention; Islet Cell; Islets of Langerhans Transplantation; Kidney; Location; Mechanics; Methods; Microscopic; Modeling; Monitor; Mus; Oral cavity; Pancreas; Pancreatic duct; Patients; Phenotype; Physiology; Positioning Attribute; Preparation; Primates; Procedures; Quality Control; Recovery; Recurrence; Replacement Therapy; Reporting; Residual state; Signal Transduction; Source; System; Texture; Therapeutic; Tissues; Viral; Viral Genes; Virus; alpha Toxin; beta cell replacement; blood glucose regulation; capsule; clinical application; diabetic; euglycemia; experimental study; expression vector; gene therapy; gene therapy clinical trial; glucose metabolism; islet; mouse model; neutralizing antibody; nonhuman primate; prevent; promoter; single-cell RNA sequencing; transcriptome sequencing; transdifferentiation; vector; viral gene delivery

PROJECT SUMMARY AND RELEVANCE An ideal solution to the treatment or cure of type 1 diabetes mellitus would be the formation of new functioning β-cells from the patient’s own tissues that are not attacked by the autoimmunity, thereby avoiding the need for any immunosuppression. Abundant recent data have suggested that α-cells are a viable potential source for endogenous transdifferentiation into β-cells. Here, we describe a pancreatic intraductal viral delivery system in the mouse, wherein a single infusion of an adeno-associated virus (AAV), carrying a pdx1/mafA expression vector, is given to a toxin-induced (alloxan) diabetic mouse. This AAV gene therapy induced robust and durable α-cell transdifferentiation into β-cell-like cells through neogenesis, with recovery of over 60% of the β-cell mass within 4 weeks, and with persistent, durable euglycemia. Serendipitously, when this β-cell-like cell neogenesis was similarly induced in the autoimmune NOD mouse model, the mice became euglycemic for 4 months or more, without any additional therapy or immunosuppression. To our knowledge, no clinically applicable β-cell replacement therapy in NOD mice has been successful without immunosuppression. We suspect that the neogenic β-cell-like cells may not be attacked by the autoimmunity because they are “imperfect” β-cells by RNA-seq analysis. Since pancreatic duct injection is routinely performed in humans as a relatively simple, non-surgical procedure, and since numerous viral gene therapy trials are currently ongoing for several diseases, we feel that our approach may be rapidly translatable to humans with type 1 diabetes mellitus. In this proposal, we will perform important proof-of-principle studies in non-human primates as last steps in preparation for human gene therapy clinical trials. The primate pancreas has a very different texture and consistency than the mouse pancreas (and is very similar to the human pancreas). Thus, the mechanics of the viral delivery will likely require substantial alterations. In addition, a glucagon promoter is preferable to the CMV promoter for expression of pdx1 and mafA, so we will strive to develop and optimize a glucagon promoter vector that is effective in primates. Further studies will investigate this pancreatic ductal infusion approach in the context of AAV neutralizing antibodies. We will also perform detailed analyses of the new β-cell-like cells, including physiology, gene expression phenotype, and anatomy. In summary, we feel that the proposed studies, if successful, should position us well in preparation for clinical trials in humans with type 1 diabetes mellitus.

项目摘要和相关性 治疗或治愈 1 型糖尿病的理想解决方案是形成新的功能性 β 细胞 来自患者自身未被自身免疫攻击的组织，从而避免了任何需要 免疫抑制。最近的大量数据表明，α 细胞是内源性细胞的可行的潜在来源。 转分化为β细胞。在这里，我们描述了小鼠的胰腺导管内病毒递送系统，其中 单次输注携带 pdx1/mafA 表达载体的腺相关病毒 (AAV)，可诱导毒素诱导 （四氧嘧啶）糖尿病小鼠。这种 AAV 基因疗法诱导稳健且持久的 α 细胞转分化为 β 细胞样细胞 细胞通过新生，在 4 周内恢复了 60% 以上的 β 细胞质量，并且具有持久、持久的作用 血糖正常。偶然地，当这种 β 细胞样细胞新生在自身免疫性 NOD 小鼠中被类似地诱导时 模型中，小鼠的血糖持续 4 个月或更长时间保持正常，无需任何额外治疗或免疫抑制。致我们的 据了解，临床上适用的 NOD 小鼠 β 细胞替代疗法在没有 免疫抑制。我们怀疑新生的 β 细胞样细胞可能不会受到自身免疫的攻击，因为它们 通过RNA-seq分析，它们是“不完美”的β细胞。由于胰管注射是在人类中常规进行的 相对简单的非手术程序，并且由于目前正在进行许多病毒基因治疗试验 疾病，我们认为我们的方法可能会迅速应用于患有 1 型糖尿病的人类。在这个提案中， 我们将在非人类灵长类动物中进行重要的原理验证研究，作为人类基因准备的最后步骤 治疗临床试验。灵长类动物胰腺的质地和稠度与小鼠胰腺非常不同（并且是 与人类胰腺非常相似）。因此，病毒传递的机制可能需要进行重大改变。在 此外，对于 pdx1 和 mafA 的表达，胰高血糖素启动子优于 CMV 启动子，因此我们将努力 开发和优化对灵长类动物有效的胰高血糖素启动子载体。进一步的研究将调查这一点 AAV 中和抗体背景下的胰腺导管输注方法。我们也会进行详细的分析 新的 β 细胞样细胞的详细信息，包括生理学、基因表达表型和解剖学。综上所述，我们认为 拟议的研究如果成功，将使我们能够为人类 1 型糖尿病临床试验做好准备 梅利图斯。