靶向Insulin自身反应性CD8+T细胞改造肽在人源化糖尿病小鼠中重塑自身耐受的效应及机制研究

Type 1 diabetes (T1D) is an organ-specific autoimmune disease, which is lack of effective antigen-specific treatments methods, and represents a rapidly growing threat to the children and teenagers' health. Insulin-autoreactive CD8+ T cells are the essential key effectors involved in the initiation and development of T1D. The therapeutic vaccine strategies, by administration with self-antigenic peptides or altered peptide ligands (APLs) may effectively prevent and control the development of T1D through immunotolerance induction of Insulin-autoreactive CD8+T cells. We designed an APL (named H6F) for an Insulin B chain-derived HLA-A*0201-restricted immunodominant epitope InsB5-14 by an H→F unconservative amino acid substitution at TCR contact site p6. Our previous study showed this APL can efficiently antagonize wide-type peptide-induced activation and proliferation of autoreactive CD8+ T cells in vitro. Furthermore, repeated intraperitoneal injection of this APL can effectively prevent the development of T1D in humanized T1D mice (HHD-NOD). This project will explore the exact cellular and molecular mechanisms of H6F administration in the immunotolerance remodeling and T1D protection in HHD-NOD mice. This study will provide a new antigen-specific immune strategy for the prevention and control of T1D, and also provide a candidate therapeutic peptide vaccine for the future clinical translational research.

I型糖尿病（type 1 diabetes， T1D）是一种器官特异性自身免疫病，自身反应性CD8+T细胞是关键致病因素。Insulin是启动T 1D的关键靶抗原，重塑Insulin自身反应性CD8+T细胞免疫耐受，可能有效阻断T1D发生发展。前期我们以HLA-A*0201限制性人鼠同源优势表位InsB5-14为关键靶标，借助分子模拟进行TCR作用位点氨基酸替换得到若干改造肽（altered peptide ligand, APL)，并通过体外功能实验筛选到一条显著抑制天然肽特异性CD8+T细胞反应的APL—H6F。进一步体内实验证明腹腔注射H6F可显著延缓人源化T1D小鼠（HHD-NOD）的发病。本课题拟从免疫失能、克隆清除及免疫抑制等方面深入探讨H6F在T1D防治中重塑自身免疫耐受的细胞及分子机制，为基于T1D特异性免疫防治提供新策略和实验依据，并为将来临床转化提供候选治疗性肽疫苗。

我们以人源化糖尿病小鼠为模型，基于计算机辅助的分子动力学模拟及细胞免疫学功能实验筛选到若干基于人鼠同源免疫显性胰岛素表位肽的单TCR作用位点替换的拮抗性改造肽（APL），体内试验证实 H6F具有显著的T1D保护作用， 反复多次腹腔注射APL，可以延缓并减轻胰岛炎，包括削弱胰腺淋巴细胞浸润、降低组织促炎因子分泌等。从细胞机制上，进一步发现反复系统性给予可溶性APL H6F处理，较天然肽，有效诱导一群肽特异性CD8+Treg，具有典型的Treg特性及强的免疫抑制功能，其受到肽特异性激活后能发挥显著的免疫抑制功能。 因此，H6F是一种具有潜在临床应用价值的修饰肽配体。