Inhibition of HIV-Induced Synaptodentritic Damage

描述（由申请人提供）：联合抗逆转录病毒治疗（cART）减缓了发病，但没有改变hiv相关神经系统疾病（HAND）的患病率。在老年患者群体中，尽管cART能够将中枢神经系统中的病毒载量降低到无法检测的水平，但HAND的患病率正在增加。半数艾滋病毒感染者可能会出现手足口病的症状。开发更多具有中枢神经系统外显性的抗病毒药物可能适得其反，因为已有研究表明，具有良好中枢神经系统外显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，可能是由于诱导代谢状态有利于促炎性血管疾病。HAND的特征包括：1)炎症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)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。许多炎症介质与这一级联反应有关，但已确定在HAND患者的脑脊液和死后脑组织中有一部分介质被上调：TNF-?？包括单核细胞趋化剂MCP-1和混合谱系激酶3 (MLK3)，后者是MAPK激酶调节炎症通路的重要控制点。此前，我们与罗切斯特大学医学中心的Harris Gelbard博士和Stephen Dewhurst博士合作，确定了URMC-099，这是抑制MLK3和阻断MCP-1上调的第一代化合物，TNF-？和其他炎症介质刺激了人类巨噬细胞。在小鼠脑内暴露于HIV-1 Tat的体内脑成像实验表明，口服URMC-099可阻止Tat诱导的白细胞浸润和小胶质细胞活化，并逆转Tat诱导的突触结构损伤。突触结构损伤是HAND在认知和记忆障碍中的关键病理过程。URMC-099是一种“选择性非选择性”激酶抑制剂，可抑制大量蛋白激酶，这一特性可能会阻碍未来的发展。在我们的1期SBIR研究中，我们试图确定第二代，更特异性的MLK3抑制剂，它将保护HIV-Tat小胶质细胞激活的体外模型中的神经元。我们已经成功地发现了一系列有效且更具选择性的MLK3抑制剂，例如CLFB-1134，在神经元保护实验中具有出色的体外活性，具有出色的血脑屏障穿透性，并且在cyp450， hERG和遗传毒性方面具有令人满意的体外安全性。在二期研究中，我们将优化这些化合物在HIV-Tat攻击小鼠脑成像模型中的MLK3效价、特异性、口服生物利用度和活性。在动物模型中显示有效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分析，以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.