Defining Inhibitory Mechanisms of Novel CCRS-targeted Microbicide Candidates

项目1-项目1的目标是确定三种新的趋化因子类似物的作用机制 具有很强的阻止HIV-1通过CCR5进入的活性。这些完全重组的分子显示了三个 不同的活性特征：(I组)CCR5阻断，没有信号活性或受体内化；(组 II)，CCR5快速内化，具有信号活性；和(III)，适度内化和 封锁而不发出信号。这些新分子在以下方面具有显著的潜在优势 安全和生产成本……超过币值[b^fw，irrtpr.pvej Tients.，p，n 这些杀微生物剂需要对其作用机制有更详细的了解。我们会 确定第二组分子的细胞内隔离途径，并与第三组分子的路线进行比较 分子。我们将检验一些假说来解释在缺乏药物的情况下持续的抗病毒活性。 细胞内受体隔离(组I)或中度内化(组III)，包括 CCR5二聚体的形成、膜上受体定位的改变、变构效应和受体 不依赖于G蛋白连接信号的内化。我们还将研究CCR5、CCL5、 CCL3L1基因多态性对CCR5蛋白质合成和周转率的影响。这些研究是 旨在研究正常人类捐赠者的初级靶细胞易感性的变异 每一种新的抑制剂。我们还将使用一组突变的CCR5分子来研究结构 活跃度的关联性。我们将利用这些关于机制和靶细胞可变性的信息来开发新的 具有更好的活性分布的分子，我们将与其他项目进行协调实验 在这个项目中，将我们在基于细胞的模型中的发现与现有和新的CCR5抑制剂的影响联系起来 在组织外植体和全动物模型中。这种方法将产生更好和更安全的CCR5抑制剂 可按适合于阻止艾滋病毒/艾滋病在受影响最严重地区传播的规模生产。

Project 1 - The goal of Project 1 is to define the mechanism of action of three new chemokine analogues with potent activity in blocking HIV-1 entry via CCR5. These fully recombinant molecules display three distinct activity profiles: (group I) CCR5 blockade without signaling activity or receptor internalization; (group II), CCR5 rapid internalization with signaling activity; and (group III), moderate CCR5 internalization and blockade without signaling activity. These new molecules have significant potential advantages in terms of safety and cost of production ..over currenynh[b^ fwither, irrtpr.pvejTients.,p,n these bahdidate'microbicides requires more detailed knowledge of their mechanisms of action. We will define the route to intracellular sequestration for group II molecules, and compare with that of the group III molecules. We will examine a number of hypotheses to explain prolonged antiviral activity in the absence of intracellular receptor sequestration (group I) or with moderate internalization (group III), including changes in CCR5 dimer formation, altered receptor localization in the membrane, allosteric effects, and receptor internalization independent of G-protein-linked signaling. We will also examine the impact of CCR5, CCL5, and CCL3L1 genetic polymorphisms on CCR5 protein synthesis and turnover rate. These studies are designed to investigate variability in the susceptibility of primary target cells from normal human donors to each of the new inhibitors. We will also use a panel of mutant CCR5 molecules to examine structural correlates of activity. We will use this information on mechanism and target cell variability to develop new molecules with even better activity profiles, and we will perform coordinated experiments with other projects in this Program to relate our findings in cell-based models to the effects of current and new CCR5 inhibitors in tissue explant and whole animal models. This approach will generate better and safer CCR5 inhibitors that can be produced on a scale suitable for stopping the spread of HIV/AIDS in the most-impacted areas.