Targeting Janus kinases in the treatment of autoimmune disease

细胞因子组成一个分泌蛋白大家族，调节细胞生长和多种类型细胞的分化。这些因子在调节免疫和炎症反应以及调节淋巴样细胞的发育和分化方面尤为重要。毫不奇怪，细胞因子在许多自身免疫性疾病的发病机制中起着关键作用，如类风湿性关节炎、系统性红斑狼疮、炎症性肠病和牛皮癣。了解细胞因子作用的分子基础有助于深入了解免疫介导性疾病的发病机制，并提供新的治疗靶点。 我们发现了人类JAK3，一种通过结合共同伽马链的细胞因子GC(IL-2、IL-4、IL-7、IL-9、IL-15和IL-21)进行信号传递所必需的激酶。我们发现JAK3的突变会导致一种称为严重联合免疫缺陷(SCID)的原发性免疫缺陷疾病。我们已经获得了两项与将JAK3作为新型免疫调节药物基础的专利，并与辉瑞公司建立了合作研究和开发协议(CRADA)，以产生第一代JAK拮抗剂。一种名为tofacitinib的化合物由辉瑞公司开发，并被发现在临床前模型中有效。托法替尼现在被批准用于类风湿性关节炎、牛皮癣关节炎和炎症性肠病。托法替尼和其他JAK抑制剂(Jakinibs)也在研究中，用于治疗强直性脊柱炎、青少年关节炎和许多皮肤病，包括牛皮癣、斑秃和特应性皮炎。其他几种Jakinibs已经开发出来，也在进行临床试验，包括后期关键试验。 与辉瑞公司的CRADA进行了更新，旨在更好地了解tofacitinib和相关抑制剂的作用机制。在考虑Jakinibs可能有用的其他临床情况时，我们认为SLE可能是这类药物的合适候选者。许多影响天然免疫和获得性免疫的细胞因子被认为在SLE的免疫发病机制中起作用，包括干扰素IL-6、IL-21和其他干扰素。此外，系统性红斑狼疮患者免疫细胞调节失调也与过早的血管损伤有关。到目前为止，还没有药物被证明可以同时针对SLE的疾病活动和增加心血管风险。在临床前模型中，我们发现托法替尼的治疗导致肾炎、皮肤炎症和自身抗体产生的改善。此外，托法替尼治疗显著降低了相关细胞因子的血清水平。托法替尼还调节中性粒细胞功能障碍和内皮异常。因此，我们认为托法替尼可以调节狼疮小鼠的先天免疫反应和获得性免疫反应，改善血管功能。基于这些临床前发现，与卡普兰实验室和NIAMS狼疮临床试验合作，在NIH临床中心启动了轻中度SLE的临床试验。这项1b/2a期随机、双盲、安慰剂对照临床试验对30名SLE受试者使用tofacitinib(药物与安慰剂的比例为2：1)，根据STAT4风险等位基因的存在与否进行分层。这项研究显示，与安慰剂组相比，托法替尼治疗组在第56天时I型干扰素基因特征、循环低密度粒细胞水平和中性粒细胞胞外陷阱显著减少，同时不同免疫细胞的pSTAT磷酸化水平显著降低。托法替尼的使用导致服用托法替尼的患者的高密度脂蛋白和高密度脂蛋白颗粒数量显著增加，同时卵磷脂：胆固醇酰基转移酶(LCAT)浓度和胆固醇流出能力显著改善。与安慰剂组相比，治疗组的动脉僵硬和内皮功能障碍有所改善。大多数这些变化在研究结束时，即停用托法替尼4周后恢复到基线水平。其中一些变化与STAT4风险等位基因的存在或不存在有关。此外，托法替尼耐受性良好，SLE疾病活动没有恶化，没有严重的不良反应、机会性感染、血栓栓塞事件或肝功能异常。因此，在一项短期试验中，托法替尼的使用显著改善了与SLE动脉粥样硬化加速相关的心脏代谢和免疫学疾病参数。需要长期的研究来确定托法替尼对SLE的各种表现的疗效，包括心血管风险。 为了更好地了解第一代和第二代jakinibs的使用，特别是在炎症性肠病中的意义，我们还研究了这些药物对小鼠模型中适应性和先天淋巴样细胞动态平衡的影响。我们发现，在接受jakinibs治疗的小鼠中，与肝、脾和骨髓中存在的NK细胞池相比，肝脏ILC1的动态平衡池受到的影响较小。JAK抑制对两个亚群的转录组有重叠作用，主要影响调控细胞周期和细胞凋亡的基因。然而，JAK抑制的不同影响与ILC1表达的抗凋亡基因Bcl2的高水平有关。我们的发现为JAK抑制剂对NK细胞和ILC1的影响提供了机制上的解释，这可能具有重要的临床意义。

Cytokines comprise a large family of secreted proteins that regulate cell growth and differentiation of many types of cells. These factors are especially important in regulating immune and inflammatory responses, and regulating lymphoid development and differentiation. Not surprisingly, cytokines are critical in the pathogenesis of many autoimmune diseases such as rheumatoid arthritis, SLE, IBD and psoriasis. Understanding the molecular basis of cytokine action provides important insights into the pathogenesis of immune-mediated disease and offers new therapeutic targets. We discovered human Jak3, a kinase essential for signaling by cytokines that bind the common gamma chain, gc (IL-2, IL-4, IL-7, IL-9, IL-15 and IL-21). We found that mutation of Jak3 results in a primary immunodeficiency disorder termed severe combined immunodeficiency (SCID). We have received two patents related to targeting Jak3 as the basis for a new class of immunomodulatory drugs and established a Cooperative Research and Development Agreement (CRADA) with Pfizer to generate the first-generation Jak antagonists. One compound, tofacitinib, was developed by Pfizer and found to be effective in preclinical models. Tofacitinib is now approved for rheumatoid arthritis, psoriatic arthritis and inflammatory bowel disease. Tofacitinib and other Jak inhibitors (jakinibs) are also being studied in ankylosing spondylitis, juvenile arthritis, and many dermatological conditions including psoriasis, alopecia areata and atopic dermatitis. Several other Jakinibs have been developed and are also in clinical trials, including late phase pivotal trials. The CRADA with Pfizer was renewed and is directed at better understanding the mechanisms of action of tofacitinib and related inhibitors. In considering other clinical circumstances in which Jakinibs might be useful, we considered that SLE might be an appropriate candidate for this class of drugs. A number of cytokines that impact both innate and adaptive immunity have been suggested to contribute to the immunopathogenesis of SLE, including interferons IL-6, IL-21, and other interferons. In addition, immune cell dysregulation in SLE is also associated with premature vascular damage. To date, no drug has proven to target both disease activity and enhanced cardiovascular risk in SLE. In a preclinical model, we found that treatment with tofacitinib led to improvement in nephritis, skin inflammation, and autoantibody production. In addition, tofacitinib treatment significantly reduced serum levels of relevant cytokines. Tofacitinib also modulated neutrophil dysfunction and endothelial abnormalities. Thus, we concluded that tofacitinib can modulate the innate and adaptive immune responses in murine lupus and improve vascular function. Based on these preclinical findings, in collaboration with the Kaplan lab and the NIAMS Lupus Clinical Trials clinical trial in mild-moderate SLE was launched at the NIH Clinical Center. This phase 1b/2a randomized, double-blind, placebo-controlled clinical trial of using tofacitinib in 30 SLE subjects (2:1 drug to placebo ratio) was stratified by the presence or absence of STAT4 risk allele. This study showed the type I Interferon gene signature, circulating levels of low-density granulocytes and neutrophil extracellular traps significantly decreased in the tofacitinib treated group compared to the placebo group by day 56, accompanied by significant decreases in pSTAT phosphorylation of different immune cells. The use of tofacitinib resulted in a significant increase in HDL-C and HDL particle number in tofacitinib-treated patients accompanied by significant improvements in lecithin: cholesterol acyltransferase (LCAT) concentration and cholesterol efflux capacity. Arterial stiffness and endothelial dysfunction improved in the treatment group as compared to the placebo group. Most of these changes reverted towards baseline values at the end of study, 4 weeks after withdrawal of tofacitinib. Some of these changes were associated to presence or absence of STAT4 risk allele. In addition, tofacitinib was well tolerated with no worsening of SLE disease activity, no severe AEs, opportunistic infections, thromboembolic events or liver function abnormalities. Thus, in a short-term trial, use of tofacitinib resulted in significant improvements in cardiometabolic and immunologic disease parameters associated with accelerated atherosclerosis in SLE. Long-term studies are needed to determine the efficacy of tofacitinib in the various manifestations of SLE including cardiovascular risk. To better understand the implications of the use of first and second generation jakinibs, especially in inflammatory bowel disease, the effects these agents on the homeostasis of adaptive and innate lymphoid cells in murine models was also investigated. We found that in mice treated with jakinibs, the homeostatic pool of liver ILC1 was less affected compared to the pool of NK cells present in the liver, spleen and bone marrow. JAK inhibition had overlapping effects on the transcriptome of both subsets, mainly affecting genes regulating cell cycle and apoptosis. However, the differential impact of JAK inhibition was linked to the high levels of the antiapoptotic gene Bcl2 expressed by ILC1. Our findings provide mechanistic explanations for the effects of JAK inhibitors on NK cells and ILC1 which could be of major clinically relevance.