Distinguishing inflammatory Th17 subsets through using an autoimmune Th17-selective inhibitor

项目摘要 Th 17细胞是产生IL-17的辅助性T细胞CD 4 + T细胞的一个亚群，在急性淋巴细胞白血病的致病性中起关键作用。 各种自身免疫性疾病，这是影响超过2300万美国人无法治愈。知识 Th 17细胞的调节机制可能为治疗这些疾病提供新的机会。基于 由于IL-23的发育需要，Th 17细胞目前被分为IL-23非依赖性稳态 和IL-23依赖性炎性Th 17细胞，其中后者执行生理（清除）和免疫调节功能。 细胞外病原体感染）和致病（引发自身免疫）功能。大量的努力， 已被用于定义区分稳态和炎性Th 17细胞的基因调控网络。 然而，如何解剖炎性Th 17细胞的亚群在很大程度上是未知的。这种知识是 因为非选择性抑制炎性Th 17细胞，如新批准的IL-17， 23单克隆抗体为基础的治疗自身免疫性疾病，不可避免地增加易感性真菌 和细菌感染。在此，自身免疫和抗感染Th 17亚群显示出对以下的不同应答： 小分子氯法齐明：Th 17亚群的自身免疫作用被抑制，而抗感染作用被抑制。 Th 17亚群功能未受影响。本申请的总体目标是阐明 区分自身免疫和抗感染Th 17亚群的表达和代谢程序。中央 假设自身免疫和抗感染Th 17亚群是可区分的，SHMT 1是检查点 自身免疫性Th 17细胞中的细胞内丝氨酸水平。该项目的基本原理是， 自身免疫和抗感染Th 17亚群之间的差异可能提供一个强有力的科学框架 由此，具有显著增加的选择性和减少的免疫性疾病治疗的新策略， 可能产生不良影响。中心假设将通过追求两个具体目标来检验：目标1） 鉴定体内抗感染和自身免疫性Th 17亚群的基因表达特征;目的2） 确定SHMT 1在自身免疫性Th 17细胞中调节细胞内丝氨酸的作用。根据目标1，单一- 细胞RNA-seq分析将与使用基于Cas9的敲除测定的验证一起使用。对于目标2， 将在存在下评价自身免疫Th 17细胞的致病性及其细胞内丝氨酸水平 SHMT 1调节：药理学抑制、基因敲除和酶失活的点突变 活动本申请中提出的研究是创新的，因为它关注的是监管网络 区分抗感染和自身免疫性Th 17亚群，这是一种迄今为止尚未研究的机制。的 拟议的研究意义重大，因为它有望为发展自身免疫性疾病提供新的机会。 用于自身免疫性疾病的Th 17选择性治疗剂。这对那些 已经感染或暴露于某些病原体，如结核病，因为现有的Th 17抑制剂不能 由于降低患者对感染的控制的风险，

Project Summary Th17 cells are a subset of IL-17-producing T helper CD4+ T cells and play a pivotal role in the pathogenicity of a variety of autoimmune diseases, which are affecting over 23 million Americans with no cure. Knowledge about regulatory mechanisms of Th17 cells might offer new opportunities for treating these diseases. Based on the requirement of IL-23 for the development, Th17 cells are currently divided into IL-23-independent homeostatic and IL-23-dependent inflammatory Th17 cells, of which the latter execute both physiological (clearing extracellular pathogen infections) and pathogenic (eliciting autoimmunity) functions. Substantial efforts have been made to define gene regulatory networks that discriminate homeostatic and inflammatory Th17 cells. However, how to dissect the subsets of inflammatory Th17 cells is largely unknown. This knowledge is critically needed because non-selective suppression of inflammatory Th17 cells, as by the newly approved IL- 23 monoclonal antibody-based therapy for autoimmune diseases, inevitably increases susceptibility to fungal and bacterial infections. Herein, autoimmune and anti-infection Th17 subsets display differential responses to the small molecule Clofazimine: the autoimmune effect of Th17 subsets was repressed while the anti-infection function of Th17 subsets was not disturbed. Our overall objectives in this application are to elucidate the gene expression and metabolic programs discriminating autoimmune and anti-infection Th17 subsets. The central hypothesis is that autoimmune and anti-infection Th17 subsets are distinguishable and SHMT1 is a checkpoint of intracellular serine levels in autoimmune Th17 cells. The rationale for this project is that determination of the differences between autoimmune and anti-infection Th17 subsets is likely to offer a strong scientific framework whereby new strategies to autoimmune diseases therapy with substantially increased selectivity and reduced adverse effects can be developed. The central hypothesis will be tested by pursuing two specific aims: Aim 1) To identify gene expression signatures of anti-infection and autoimmune Th17 subsets in vivo; and Aim 2) To determine the role of SHMT1 in regulating intracellular serine in autoimmune Th17 cells. Under Aim 1, single- cell RNA-seq analysis will be utilized with verification using Cas9-based knockout assay. For Aim 2, the pathogenicity of autoimmune Th17 cells and their intracellular serine levels will be evaluated in the presence of SHMT1 modulation: pharmacologic inhibition, genetic knockout, and point mutation with inactive enzymatic activities. The research proposed in this application is innovative, because it focuses on the regulatory network discriminating anti-infection and autoimmune Th17 subsets, a heretofore-unexamined mechanism. The proposed research is significant because it is expected to provide novel opportunities to develop autoimmune Th17-selective therapeutics for autoimmune diseases. This would be extraordinarily important for patients that have been infected or exposed to certain pathogens, such as tuberculosis, since existing Th17 inhibitors cannot be used due to the risks of decreasing the patients’ control of the infection.