Targeting the immunoproteasome as a novel therapeutic strategy for hemophagocytic lymphohistiocytosis

项目总结 原发性噬血细胞淋巴组织细胞增多症包括一系列不断扩大的遗传性疾病 以严重的过度炎症为特征的免疫系统。尽管使用了糖皮质激素地塞米松， 化疗药物依托泊苷、特异性细胞因子靶向药物和异基因造血干细胞 移植，高达40%的PHLH患者死于无法控制的疾病或其并发症 治疗。因此，迫切需要开发新的、更有效的治疗方法。到目前为止， 我们对PHLH发病机制的大部分了解都来自于对人类和小鼠的研究 缺乏穿孔素的表达，穿孔素是CD8 T细胞的细胞毒功能所必需的孔形成蛋白 自然杀伤(NK)细胞。在感染淋巴细胞脉络膜脑膜炎病毒(LCMV)后，穿孔素缺乏 小鼠患上一种致命的HLH样疾病，其特征是器质性肥大、贫血、血小板减少、 高细胞分裂素血症、高铁素血症和暴发性组织炎症。由于它们的细胞溶解缺陷，穿孔素- 缺乏CD8T和NK细胞不能消除LCMV感染的抗原提呈细胞(APC)。一直以来 提出了这些激活的APC的持久性导致了一个前馈循环来进一步驱动T细胞 增殖和促炎细胞因子的产生，最终导致HLH的症状和体征。 在APC中，免疫蛋白酶体处理内源和外源蛋白，包括病毒蛋白， 被装载到主要组织相容性复合体I类(MHCI)分子上的多肽用于呈递 免疫系统的其他细胞。考虑到激活的APC在PHLH中的持久性和核心作用 对于抗原递呈在驱动免疫细胞激活中的作用，我们假设抑制 免疫蛋白酶体将成为一种合理且潜在有益的治疗方法。事实上，使用 体外LCMV感染模型观察到免疫蛋白酶体抑制剂ONX-914和KZR-616 显著降低LCMV特异性CD8 T细胞的增殖和干扰素-γ(IFNG)的产生。同样， KZR-616对LCMV感染穿孔素缺陷小鼠的治疗作用 包括脾肿大、CD8 T细胞扩增、组织免疫病理学和血清IFNG水平。在这 R21探索性研究补助金，是应特殊利益通知提交的： 免疫/原发免疫fi免疫缺陷的调查(非-AI-21-032)，我们将进一步 探讨穿孔素对免疫蛋白酶体抑制的治疗作用及其机制 缺血小鼠模型。在目标1中，我们将检查免疫蛋白酶体抑制改善的程度。 临床疾病，测试特定的药物方案和组合。目标2将探索其机理基础 采用体内和体外互补检测。如果成功，这些研究将阐明 免疫蛋白酶体抑制可减轻PHLH的炎症反应，并为未来的临床试验奠定基础 旨在改善患有这些往往致命的免疫系统疾病的个人的总体结果。

PROJECT SUMMARY Primary hemophagocytic lymphohistiocytosis (pHLH) comprises an expanding array of inherited disorders of the immune system characterized by severe hyperinflammation. Despite use of the glucocorticoid dexamethasone, chemotherapeutic agent etoposide, specific cytokine-targeting agents, and allogeneic hematopoietic stem cell transplantation, up to 40% of pHLH patients die due to uncontrolled disease or the complications of its treatment. Accordingly, there is a pressing need to develop new and more effective therapies. To date, much of what we have learned about the pathogenesis of pHLH has come from the study of humans and mice lacking expression of perforin, a pore forming protein essential for the cytotoxic function of CD8 T cells and natural killer (NK) cells. Following infection with Lymphocytic Choriomeningitis virus (LCMV), perforin-deficient mice develop a fatal HLH-like illness characterized by organomegaly, anemia, thrombocytopenia, hypercytokinemia, hyperferritinemia, and fulminant tissue inflammation. Due to their cytolytic defects, perforin- deficient CD8 T and NK cells cannot eliminate LCMV-infected antigen presenting cells (APCs). It has been proposed that the persistence of these activated APCs results in a feed-forward loop to further drive T cell proliferation and proinflammatory cytokine production, ultimately culminating in the signs and symptoms of HLH. Within APCs, the immunoproteasome processes endogenous and exogenous proteins, including viral proteins, into peptides that are loaded onto Major Histocompatibility Complex class I (MHCI) molecules for presentation to other cells of the immune system. Given the persistence of activated APCs in pHLH and the central role for antigen presentation in driving immune cell activation, we hypothesize that inhibiting the immunoproteasome will serve as a rational and potentially beneficial therapeutic maneuver. Indeed, using an in vitro model of LCMV infection, we observe that the immunoproteasome inhibitors ONX-914 and KZR-616 significantly reduce LCMV-specific CD8 T cell proliferation and interferon-gamma (IFNg) production. Similarly, treatment of LCMV-infected perforin-deficient mice with KZR-616 significantly diminishes the manifestations of HLH, including splenomegaly, CD8 T cell expansion, tissue immunopathology, and serum IFNg levels. In this R21 Exploratory Research Grant, which is being submitted in response to Notice of Special Interest: Investigations on Inborn Errors of Immunity/Primary Immunodeficiencies (NOT-AI-21-032), we will further explore the therapeutic effects and mechanisms of action of immunoproteasome inhibition using the perforin- deficient mouse model. In Aim 1, we will examine the extent to which immunoproteasome inhibition ameliorates clinical disease, testing specific drug schedules and combinations. Aim 2 will explore the mechanistic basis using complementary in vivo and in vitro assays. If successful, these studies will elucidate whether immunoproteasome inhibition lessens inflammation in pHLH and set the stage for future clinical trials aimed at improving the overall outcome for individuals with these often-fatal immune system disorders.