Inhibition of HIV-Induced Synaptodentritic Damage

描述(申请人提供)：联合抗逆转录病毒疗法(CART)延缓了发病，但并未改变HIV相关神经疾病(HAND)的流行率。在老年患者群体中，HAND的患病率正在增加，尽管CART能够将CNS中的病毒载量降低到无法检测的水平。一半的艾滋病毒携带者可能会出现手部症状。开发更多的中枢神经系统渗透性抗病毒药物可能会适得其反，因为已有研究表明，具有良好中枢神经系统渗透性的抗逆转录病毒药物与神经认知功能受损有关。这可能是直接或间接神经毒性的结果，可能是由于有利于促炎性血管疾病的诱导代谢状态造成的。HAND的特征包括：1)炎性细胞因子和趋化因子的失调，2)免疫细胞(单核细胞)向中枢神经系统募集，3)神经胶质细胞的病毒感染导致其正常功能中断，4)广泛的突触树突损伤。许多炎症介质参与了这一级联反应，但有一小部分介质在手部患者的脑脊液和死后脑组织中表达上调：单核细胞趋化因子单核细胞趋化因子MCP-1，以及MAPK激酶调节炎症通路的一个重要控制点--混合血统激酶3(MLK3)。此前，我们与罗切斯特大学医学中心的哈里斯·格尔巴德博士和斯蒂芬·杜赫斯特博士合作，已经确定了URMC-099，这是一种第一代化合物，可以抑制MLK3并阻止MCP-1、TNF-？和HIV-1TAT中的其他炎症介质刺激人巨噬细胞。脑内暴露于HIV-1TAT的小鼠的活体脑成像实验表明，I.P.给予URMC-099可阻止TAT诱导的白细胞浸润和小胶质细胞激活，并逆转TAT诱导的突触结构损伤。突触结构的损伤是手部认知和记忆受损的关键病理过程。URMC-099是一种“选择性非选择性”的激酶抑制剂，可以抑制大量的蛋白激酶，这一特性可能会阻碍未来的发展。在我们的第一阶段SBIR研究中，我们试图确定第二代更具特异性的MLK3抑制剂，它将在HIV-TAT激活小胶质细胞的体外模型中保护神经元。我们已经成功地发现了一系列有效的、更具选择性的MLK3抑制剂，以CLFB-1134为例，在神经元保护试验中具有出色的体外活性，出色的血脑屏障穿透性，以及令人满意的CYP450、HERG和遗传毒性的体外安全性。在第二阶段的研究中，我们将在HIV-TAT挑战的小鼠脑成像模型中优化这些化合物的MLK3效力、特异性、口服生物利用度和活性。在动物模型中显示有效性的化合物将在体外安全性和遗传毒性筛选以及剂量范围毒性研究中进行概述，以供选择作为后期临床前试验的开发化合物。

DESCRIPTION (provided by applicant): Combination antiretroviral therapy (cART) has slowed the onset, but has not altered the prevalence of HIV-associated neurologic disease (HAND). In aging patient populations, the prevalence of HAND is increasing, despite the ability of cART to reduce viral load in the CNS to undetectable levels. Half of the population living with HIV may experience symptoms of HAND. Development of more CNS penetrant antiviral drugs may be counter-productive, since it has been shown that antiretrovirals with good CNS penetrance are associated with impaired neurocognitive function. This may be a result of direct or indirect neurotoxicity, possibly resulting from induced metabolic states that favor pro-inflammatory vascular disease. The hallmarks of HAND include: 1) a dysregulation of inflammatory cytokines and chemokines, 2) the recruitment of immune cells (monocytes) to the CNS, 3) the viral infection of glia leading to interruption of their normal function, and 4) extensive synaptodendritc damage. A host of inflammatory mediators are implicated in this cascade, but a limited subset of mediators has been identified as being up-regulated in the cerebrospinal fluid and postmortem brain tissues of HAND patients: TNF-??, the monocyte chemoattractant MCP-1, and mixed-lineage kinase 3 (MLK3), an important control point in MAPK kinase regulated inflammation pathways. Previously in collaboration with Dr. Harris Gelbard and Stephen Dewhurst at the University of Rochester Medical Center, we have identified URMC-099, a first generation compound that inhibits MLK3 and blocks up-regulation of MCP-1, TNF-? and other inflammatory mediators in HIV-1 Tat stimulated human macrophages. In vivo brain imaging experiments in mice exposed intracerebrally to HIV-1 Tat have shown that i.p. administration of URMC-099 prevents Tat-induced leukocyte infiltration and microglial activation, and reverses Tat-induced damage to synaptic architecture. Damage to synaptic architecture is the key pathological process of HAND in cognition and memory impairment. URMC-099 is a "selectively non-selective" kinase inhibitor and inhibits a large number of protein kinases, a property that may hinder future development. In our Phase 1 SBIR research we sought to identify second generation, much more specific MLK3 inhibitors, that would protect neurons in in vitro models of microglial activation by HIV-Tat. We have successfully discovered a series of potent and much more selective MLK3 inhibitors, exemplified by CLFB-1134, with excellent in vitro activity in neuron protection assays, excellent blood brain barrier penetration, and satisfactory in vitro safety profiles for CYP450s, hERG and genotoxicity. In Phase 2 research we will optimize these compounds for MLK3 potency, specificity, oral bioavailability and activity in mouse brain imaging models of HIV-Tat challenge. Compounds showing efficacy in animal models will be profiled in in vitro safety and genotoxicity screens and dose ranging toxicity studies for selection as development compounds for late stage preclinical testing.