Mechanistic and Therapeutic Role of the TLR4 Signaling Pathway in Type 1 Diabetes

TLR4 信号通路在 1 型糖尿病中的机制和治疗作用

基本信息

批准号：
10718841
负责人：
ANDREW B HERR
金额：
$ 70.32万
依托单位：
UNIVERSITY OF CALIFORNIA AT DAVIS
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-01 至 2027-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10718841
关键词：
Acute Adoptive Transfer Agonist Antibodies Antibody Affinity Antibody Therapy Autoimmunity Avidity Binding Biological Assay Biophysics Cell Maturation Cell surface Cells Clinical Trials Cryoglobulins Data Diabetes Mellitus Diabetic Ketoacidosis Endosomes Generations Human Hyperglycemia IRF3 gene IgG1 IgG3 IgG4 Immobilization Immune Immunoglobulin Class Switching Immunologics Immunology Inbred NOD Mice Infiltration Inflammation Inflammatory Insulin Insulin-Dependent Diabetes Mellitus Length Life Macrophage Mediating Molecular Mus Myelogenous Myeloid Cells Myeloid-derived suppressor cells Myelopoiesis Nobel Prize Paper Pathway interactions Patients Peripheral Phenotype Process Production Recombinants Role Signal Induction Signal Pathway Signal Transduction Site Specificity Structure Structure of beta Cell of islet Surface T-Cell Proliferation T-Lymphocyte TLR4 gene Test Result Testing Therapeutic Therapeutic Effect Tissues Work autoimmune pathogenesis biophysical analysis biophysical techniques clinical effect cytokine diabetes mellitus therapy diabetic glycosylation humanized mouse immunoregulation in vivo innate immune mechanisms islet monocyte mouse model novel prevent receptor receptor binding receptor internalization therapeutic evaluation translation to humans

项目摘要

Type 1 Diabetes (T1D) is caused by autoimmune destruction of insulin-producing pancreatic beta cells. T1D therapies have largely failed in human clinical trials and thus an urgent need exists for targeting novel immune pathways. We have exciting data showing that a TLR4/MD2 agonistic antibody (TLR4-Ab) permanently reversed T1D in 71%, and had a significant clinical effect in 90%, of acutely diabetic non-obese diabetic (NOD) mice. In our recent Diabetes paper, we showed that TLR4-Ab can mobilize and activate myeloid-derived suppressor cells (MDSC) in vivo, that suppress T cells and ameliorate acute T1D upon adoptive transfer. We showed that TLR4- Ab induces TLR4/MD-2 sequestration in endosomes, unlike the canonical TLR4 agonist LPS (which cannot reverse T1D); however, the mechanism by which TLR4-Ab reverses T1D remains unclear. We have now produced anti-human TLR4 antibodies, allowing us to apply these findings to human T1D. Our combined expertise in autoimmunity/T1D (Ridgway), molecular biophysics/Immunology (Herr) and human T1D (Dolan) is well suited to perform these studies. We will pursue three aims: Specific Aim 1. Mechanism of reversal of T1D by TLR4-Ab-induced MDSCs. TLR4-Ab endosomal sequestration may induce prolonged signaling via the TLR4-mediated TRIF pathway, which protects from T1D (2015, Chervonsky et al.). We show that immobilization of TLR4-Ab on a plate, which prevents endosomal sequestration, eliminated TLR4 signaling. Therefore, we hypothesize that TLR4-Ab-induced TLR4/MD2 endosomal sequestration causes sustained TRIF endosomal signaling that induces APCs to undergo MDSC maturation, inducing immune regulation and reversing T1D. Specific Aim 2. Mechanistic role of Fc structure in TLR4-Ab reversal of T1D and cell suppression. While antibody F(ab) structure determines specificity, the Fc region contributes to antibody function through receptor binding and glycosylation. TLR4-Ab is an IgG3 isotype, which has the longest hinge region and increased glycosylation sites. IgG3 antibodies also form cryoglobulins, impacting avidity and internalization. Our preliminary data show that TLR4-Ab F(ab) and F(ab’)2 fragments elicit decreased NFκB signaling compared to full-length TLR4-Ab. This shows a critical role for Fc structure in TLR4-Ab function. Therefore, we hypothesize that the IgG3 Fc portion of the TLR4-Ab is critical to its tolerizing function and we will test this with immunological and molecular biophysical approaches. Specific Aim 3. Testing therapeutic effects of a novel panel of human anti-TLR4 antibodies on human T1D APCs Since the TLR4/MD2 pathway is strongly evolutionarily conserved, we have developed agonistic human recombinant TLR4-Ab (hTLR4-Ab). We show here that these hTLR4-Abs bind to, and activate TLR4/MD2. Our hypothesis is that hTLR4-Ab treatment will induce MDSCs from myeloid precursors and that these huMDSCs will suppress human T-cell proliferation and activation. These studies will characterize novel innate immune mechanisms by which TLR4-Ab treatment can reverse acute T1D, and our studies on the novel human TLR4 antibodies are the first step in translation to human T1D.

1 型糖尿病 (T1D) 是由产生胰岛素的胰腺 β 细胞的自身免疫性破坏引起的。疗法在人体临床试验中基本失败，因此迫切需要针对新型免疫疗法我们有令人兴奋的数据表明 TLR4/MD2 激动性抗体 (TLR4-Ab) 永久逆转。 71% 的急性糖尿病非肥胖糖尿病 (NOD) 小鼠患有 T1D，并且对 90% 的小鼠具有显着的临床效果。我们最近的糖尿病论文表明，TLR4-Ab 可以动员并激活骨髓源性抑制细胞（MDSC）在体内抑制 T 细胞并改善过继转移后的急性 T1D。 Ab 诱导 TLR4/MD-2 在内体中隔离，这与经典的 TLR4 激动剂 LPS（不能逆转 T1D）；然而，TLR4-Ab 逆转 T1D 的机制目前仍不清楚。产生了抗人 TLR4 抗体，使我们能够将这些发现应用于人类 T1D。自身免疫/T1D (Ridgway)、分子生物物理学/免疫学 (Herr) 和人类 T1D (Dolan) 方面的专业知识我们将追求三个目标：具体目标 1. T1D 逆转机制。 TLR4-Ab 诱导的 MDSC 可能会通过 TLR4-Ab 内体隔离诱导延长的信号传导。 TLR4 介导的 TRIF 通路可预防 T1D（2015，Chervonsky 等人）。平板上的 TLR4-Ab 可以防止内体隔离，从而消除了 TLR4 信号传导。 TLR4-Ab 诱导的 TLR4/MD2 内体隔离导致持续的 TRIF 内体诱导 APC 经历 MDSC 成熟、诱导免疫调节和逆转 T1D 的信号传导。具体目标 2. Fc 结构在 TLR4-Ab 逆转 T1D 和细胞抑制中的机制作用。抗体F(ab)结构决定特异性，Fc区通过受体贡献抗体功能 TLR4-Ab 是一种 IgG3 同种型，具有最长的铰链区并增加。糖基化位点。IgG3 抗体也会形成冷球蛋白，影响亲合力和内化。数据显示，与全长相比，TLR4-Ab F(ab) 和 F(ab')2 片段引起 NFκB 信号传导减少 TLR4-Ab。这显示了 Fc 结构在 TLR4-Ab 功能中的关键作用。 TLR4-Ab 的 IgG3 Fc 部分对其耐受功能至关重要，我们将通过免疫学和具体目标 3. 测试一组新的人类治疗效果。人类 T1D APC 上的抗 TLR4 抗体由于 TLR4/MD2 通路在进化上高度保守，我们开发了激动性人类重组 TLR4-Ab (hTLR4-Ab)，我们在此展示了这些 hTLR4-Ab。我们的假设是 hTLR4-Ab 治疗会诱导骨髓中的 MDSC。这些研究将抑制人类 T 细胞的增殖和活化。描述了 TLR4-Ab 治疗可逆转急性 T1D 的新型先天免疫机制，我们的研究对新型人类 TLR4 抗体的研究是转化为人类 T1D 的第一步。