Analyzing Genes for Autoimmune Thyroiditis and Diabetes: Translation to Therapy

项目总结 自身免疫性甲状腺疾病(AITD)经常与1型糖尿病(T1D)一起发展，这是一种组合 被认为是自身免疫性多腺体综合征3型(APS3v)的变种。我们的目标是确定 AITD+T1D(APS3v)的联合易感基因，分析它们致病的机制，以及 将这些知识转化为新的治疗方法。在上一个赠款周期，我们在以下方面取得了实质性进展 这些目标：(1)我们使用全基因组方法定位了新的APS3v易感基因/位点；(2)我们 发现了一个对APS3v的发展至关重要的独特的HLA-DR口袋签名；(3)我们鉴定了4个 致病的甲状腺和胰岛多肽与APS3v特异性的HLA-DR结合并激活T细胞；(4)我们鉴定了 遗传-表观遗传相互作用引发AITD&T1D；(5)我们发现了一种化合物，Cepharanthine，它 阻断甲状腺球蛋白多肽递呈，预防小鼠自身免疫性甲状腺炎。以这些发现为基础 我们建议：(1)我们定位的序列3 APS3v基因座，以识别易患APS3v的罕见变异；以及(2) 通过阻止这个口袋，将我们发现的APS3v-DR口袋转化为新的疗法。我们的假设 除了常见的变异外，罕见的非人类白细胞抗原变异对AITD+T1D的联合病因也是至关重要的， 而识别它们将揭示新的机制和治疗靶点；此外，我们假设 病原肽与APS3v-DR口袋的结合及其对T细胞的提呈可触发APS3v，并且 阻断与人类白细胞抗原DR结合的致病多肽可用于治疗或预防APS3v。我们的具体目标是： 具体目标1：使用靶向NEXT识别与APS3v连锁的3个基因座(2Q、8P、XP)的罕见变异 在我们独特的AITD+T1D聚类的家庭数据集中进行世代测序(NGS)。已确定的变种 这些被预测会改变基因功能的基因将在两个APS3v患者和家庭的大型队列中复制。 特定目标2：筛选阻断致病多肽的小分子抑制物(SMI‘s)大文库 绑定到APS3v-DR口袋以识别命中。HIT SMI将在体外、体外和体内得到确认。铅 SMI将通过药物化学进行优化，以提高效力、选择性和药代动力学特性。 特定目标3：设计与APS3v人类白细胞抗原-DR结合的致病多肽的D-氨基酸肽阻断剂 口袋里。我们将在硅胶中设计D-氨基酸多肽(D-多肽)，预计将阻止结合和呈现 致病的甲状腺/胰岛多肽。预测的D-肽阻滞剂将在体外、体外和体内得到证实。 总而言之，我们的多学科翻译项目建立在上一次赠款期间获得的知识的基础上。 我们的目标是继续我们成功的基因发现，使用定位的APS3v基因座的靶向NGS来识别 新的治疗靶点，并通过阻断APS3v的 我们鉴定出的APS3v-DR口袋。我们治疗方法的优势在于，它既是选择性的，也是唯一的T- 识别致病肽的细胞是有针对性的和个性化的，因为只有携带APS3v的患者- DR将接受治疗。我们的研究有望为APS3v以及AITD和T1D带来新的治疗方法。

PROJECT SUMMARY Autoimmune thyroid diseases (AITD) frequently develop together with type 1 diabetes (T1D), a combination considered a variant of the Autoimmune Polyglandular Syndrome type 3 (APS3v). Our goals are to identify the joint susceptibility genes for AITD+T1D (APS3v), to dissect the mechanisms by which they cause disease, and to translate this knowledge into new therapies. In the last grant cycle we made substantial progress toward these goals: (1) We mapped new susceptibility genes/loci for APS3v using whole genome approaches; (2) We discovered a unique HLA-DR-pocket signature critical for the development of APS3v; (3) We identified 4 pathogenic thyroid & islet peptides that bind to the APS3v-specific HLA-DR & activate T-cells; (4) We identified genetic-epigenetic interactions that trigger AITD & T1D; (5) We discovered a compound, Cepharanthine, that blocks thyroglobulin peptide presentation & prevents autoimmune thyroiditis in mice. Building on these findings we propose to: (1) Sequence 3 APS3v loci we mapped to identify rare variants predisposing to APS3v; and (2) Translate our discovery of the APS3v-DR pocket into new therapies by blocking this pocket. Our hypothesis is that in addition to common variants, rare non-HLA variants are also critical to the joint etiology of AITD+T1D, and that identifying them will reveal new mechanisms & therapeutic targets; furthermore, we hypothesize that binding of pathogenic peptides to the APS3v-DR pocket & their presentation to T-cells triggers APS3v, and that blocking pathogenic peptide binding to HLA-DR can be used to treat or prevent APS3v. Our specific aims are: Specific Aim 1: Identifying rare variants in 3 loci (2q, 8p, Xp) that are linked with APS3v using targeted next generation sequencing (NGS) in our unique dataset of families in which AITD+T1D cluster. Identified variants that are predicted to alter gene function will be replicated in 2 large cohorts of APS3v patients and families. Specific Aim 2: Screening of a large library for small molecule inhibitors (SMI's) that block pathogenic peptide binding to the APS3v-DR pocket to identify hits. Hit SMI's will be confirmed in vitro, ex vivo and in vivo. Lead SMI's will be optimized by medicinal chemistry to improve potency, selectivity, & pharmacokinetic properties. Specific Aim 3: Designing D-amino acid peptide blockers of pathogenic peptide binding to the APS3v HLA-DR pocket. We will design in silico D-amino acid peptides (D-peptides) predicted to block binding and presentation of pathogenic thyroid/islet peptides. Predicted D-peptide blockers will be confirmed in vitro, ex vivo, and in vivo. In summary, our multidisciplinary translational project builds on the knowledge gained in the last grant period. Our goals are to continue our successful gene discovery using targeted NGS of mapped APS3v loci to identify new therapeutic targets, and to pursue pre-clinical development of novel therapies for APS3v by blocking the APS3v-DR pocket we identified. The strength of our therapeutic approach is that it is both selective as only T- cells recognizing pathogenic peptides are targeted and personalized since only patients carrying the APS3v- DR will be treated. Our studies will hopefully lead to new therapies for APS3v, as well as for AITD and T1D.