K-Channels in Lymphocyte Function and Autoimmunity

描述（由申请人提供）：功能性阻断或消除抗原特异性免疫应答而不影响一般免疫功能必须被视为寻求治疗自身免疫性疾病的圣杯。实现这一崇高目标的一种方法是抑制和/或消除效应记忆（TEM）细胞，这些细胞与许多自身免疫性疾病的发病机制有关，而不影响其他免疫细胞。该提案的重点是TEM细胞中的电压门控Kv1.3钾通道作为自身免疫性疾病的治疗靶点。多发性硬化症（MS）、1型糖尿病（T1 DM）和类风湿性关节炎（RA）患者中的疾病相关自身反应性T细胞是Kv1.3通道表达升高的TEM细胞。我们已经开发了高度特异性的Kv1.3抑制剂，这些抑制剂选择性地抑制钙信号传导、细胞因子产生、增殖和TEM细胞的体内迁移，而不影响其他T细胞亚群。在概念验证研究中，Kv1.3阻滞剂预防和/或治疗MS、T1 DM、RA、接触性皮炎、迟发型超敏反应（DTH）和牙周炎相关骨吸收大鼠模型中的疾病。这些阻断剂在动物模型中具有优异的安全性。它们不会损害对病毒和细菌病原体的急性保护性免疫反应。特异性Kv1.3阻断剂提供了一种令人兴奋的新治疗方法，可以在不损害保护性免疫反应的情况下抑制自身反应性反应。本提案中概述的研究将为MS患者使用ShK-186（我们的主要候选药物）进行临床试验奠定基础。目的1将确定ShK-186在DA大鼠（人类MS的模型）中的慢性复发-缓解型实验性自身免疫性脑脊髓炎（CR-EAE）中的治疗作用的潜在机制。我们将测试ShK-186通过我们称之为“忽视死亡”的机制消除引起疾病的CNS自身抗原特异性TEM细胞的假设，而其他免疫细胞，特别是疾病抑制调节性T细胞，因为它们受到ShK-186抗性KCa3.1通道的保护，所以它们的增殖没有减弱。目的2将描述ShK-186治疗CR-EAE的治疗剂量、频率和持续时间，以更好地预测人体研究。在目标3中，我们将使用膜片钳记录和流式细胞术来确定Kv1.3高表达是否是MS的可靠生物标志物，以及它是否可用于跟踪Kv1.3阻断剂治疗。目的3的第二个目标是研究食蟹猴T细胞中的通道表达，因为该物种广泛用于人类免疫调节疗法的毒理学评价，并且是我们用于IND使能毒理学研究的非人灵长类动物选择。我们将定义食蟹猴T细胞亚群的K通道表型，并评估ShK-186抑制猴TEM细胞增殖的有效性。 公共卫生相关性：我们的目标是开发一种治疗方法，抑制或消除自身免疫性疾病中的致病免疫细胞，而不损害保护性免疫反应。我们的策略是选择性靶向效应记忆T细胞中的电压门控Kv1.3钾通道，这些通道是自身免疫性疾病的重要介质。

DESCRIPTION (provided by applicant): Functional blockade or elimination of antigen-specific immune responses without impacting general immune function must be considered a holy grail in the quest to treat autoimmune disease. One approach to achieving this lofty goal is to suppress and/or eliminate effector memory (TEM) cells that have been implicated in the pathogenesis of many autoimmune diseases without affecting other immune cells. This proposal focuses on the voltage-gated Kv1.3 potassium channel in TEM cells as a therapeutic target for autoimmune diseases. Disease-associated autoreactive T cells in patients with multiple sclerosis (MS), type-1 diabetes mellitus (T1DM) and rheumatoid arthritis (RA) are TEM cells with elevated expression of Kv1.3 channels. We have developed highly specific inhibitors of Kv1.3 and these selectively suppress calcium signaling, cytokine production, proliferation, and in vivo migration of TEM cells without affecting other T cell subsets. In proof-of-concept studies, Kv1.3 blockers prevent and/or treat disease in rat models of MS, T1DM, RA, contact dermatitis, delayed type hypersensitivity (DTH) and bone resorption associated with periodontitis. These blockers have excellent safety profiles in animal models. They do not compromise the acute protective immune response to viral and bacterial pathogens. Specific Kv1.3 blockers, provide an exciting new therapeutic approach to mute autoreactive responses without compromising the protective immune response. The studies outlined in this proposal will lay the groundwork for clinical trials in patients with MS with ShK-186, our lead candidate. Aim 1 will define the mechanisms underlying ShK-186's therapeutic effect in chronic relapsing-remitting experimental autoimmune encephalomyelitis (CR-EAE) in DA rats, a model for human MS. We will test the hypothesis that ShK-186 eliminates disease-causing CNS autoantigen-specific TEM cells by a mechanism we term "death by neglect" while other immune cells, particularly disease-suppressing regulatory T cells, proliferate unabated because they are protected by the ShK-186-resistant KCa3.1 channel. Aim 2 will characterize the therapeutic dose, frequency and duration of ShK-186 treatment in CR-EAE to better predict human studies. In Aim 3 we will use patch-clamp recording and flow cytometry to determine whether Kv1.3high expression is a reliable biomarker for MS, and whether it will be useful for tracking Kv1.3 blocker therapy. A second goal of aim 3 is to study channel expression in T cells from Cynomolgus monkeys because this species is widely used for toxicological evaluation of human immune modulating therapies and is our non-human primate choice for IND- enabling toxicological studies. We will define the K-channel phenotype of cynomolgus monkey T cell subsets, and we will evaluate ShK-186's effectiveness in suppressing monkey TEM cell-proliferation. PUBLIC HEALTH RELEVANCE: Our goal is to develop a therapy that suppresses or eliminates disease-causing immune cells in autoimmune diseases without compromising the protective immune response. Our strategy is to selectively target voltage-gated Kv1.3 potassium channels in effector memory T cells that are important mediators of autoimmune diseases.