Computationally informed discovery of scavenging-sparing inhibitors of CC chemokine receptor 2

G蛋白偶联CC趋化因子受体2(CCR2)在循环单核细胞上表达，并驱动单核细胞 募集到感染和受损的组织，在那里单核细胞分化为浸润性巨噬细胞 并促进形成炎性促纤维化环境。因为这一过程是 许多炎症性疾病的发病机制(类风湿性关节炎、多发性硬化症、神经性疼痛和 除其他外，CCR2作为一种治疗靶点已经被追求了几十年。然而，没有抗CCR2 到目前为止，治疗学已经进入临床，大多数都因为缺乏疗效而失败。 矛盾的是，所有已知的CCR2拮抗剂都会导致血浆中CCR2激动剂水平的显著升高 趋化因子CCL2，这是他们最初设计用来抑制的。它的治疗后果是 尚不清楚：升高可能是所需的(通过钝化趋化因子梯度和迁移反应)或 一种不受欢迎的现象(通过“抑制一种抑制剂”)。非迁移性带来的额外复杂性 CCL2在调节白细胞分化等方面的功能。令人担忧的是，拮抗剂诱导的CCL2升高 当拮抗剂本身的血浆浓度下降时达到峰值。因此，这种副作用可能 降低抗CCR2临床候选药物的安全性和有效性。不幸的是，系统的调查 由于缺乏适当的药理学工具，这种现象目前在体内是不可能的。 申请者的长期目标是破译CCR2信号的复杂性，并在临床上取得成功 CCR2-目标代理。申请人最近发现了拮抗剂诱导的CCL2的基础 并证明它是通过抑制一个重要但被低估的调节功能而发生的 在CCR2中，它清除组织结构性产生的CCL2，并将其从血浆中清除。拾荒者是 被抑制是因为所有已知的拮抗剂都阻止趋化因子与CCR2结合。这些发现概述了 CCR2介导的迁移的拮抗剂不会导致CCL2升高的概念上的可能性，通过 避免趋化因子与CCR2结合，从而清除趋化因子。本提案的目标是发现 这类拮抗剂通过计算机引导的方法，通过追求两个特定目标：(1)通过在硅胶中 针对CCR2：CCL2复杂模型的化合物文库筛选，鉴定化学物质 与CCR2同时与CCL2结合，并在体外表征其药理作用。(2) 计算阐明CCR2与CCL2形成的络合物的动力学，并鉴定G-蛋白- 不相容的状态有可能在VLS中富含清除节约型抑制剂。 拟议工作的结果将是发现第一个具有新药理的化学探针 与CCR2的关系：CCR2介导的细胞迁移的抑制剂，使CCL2免于清除。这些分子 将协助申请人的小组和其他人在体内调查拮抗剂诱导的CCL2升高。这个 这些发现还将为未来针对CCR2治疗炎症和癌症的努力提供信息。

The G protein-coupled CC chemokine receptor 2 (CCR2) is expressed on circulating monocytes and drives their recruitment to infected and damaged tissues, where the monocytes differentiate into infiltrating macrophages and promote the formation of an inflammatory pro-fibrotic environment. Because this process is key to the pathogenesis of many inflammatory diseases (rheumatoid arthritis, multiple sclerosis, neuropathic pain, and fibrosis, among others), CCR2 has been pursued for decades as a therapeutic target. However, no anti-CCR2 therapeutics have so far made it to the clinic, with most failing for lack of efficacy. Paradoxically, all known CCR2 antagonists cause a profound elevation in plasma levels of CCR2 agonist chemokine CCL2, which they were designed to inhibit in the first place. The therapeutic consequences of this are not clear: the elevation may be a desired (via blunting of chemokine gradients and migratory responses) or an unwanted (via “inhibition of an inhibitor”) phenomenon. Additional complexity arises from non-migratory functions of CCL2 in e.g. regulating leukocyte differentiation. Concerningly, antagonist-induced CCL2 elevation reaches its peak when the plasma concentration of the antagonist itself wanes. Therefore, this side effect may compromise the safety and efficacy of anti-CCR2 clinical candidates. Unfortunately, systematic investigation of this phenomenon in vivo is currently impossible, for the lack of proper pharmacological tools. The applicants’ long-term goal is to decipher the intricacies of CCR2 signaling and to develop clinically successful CCR2-targeting agents. The applicants have recently discovered the basis for antagonist-induced CCL2 elevation, and demonstrated that it occurs via inhibition of an important but underappreciated regulatory function of CCR2 where it scavenges CCL2 constitutively produced by tissues, and clears it from plasma. Scavenging is inhibited because all known antagonists prevent chemokine binding to CCR2. These findings outline the conceptual possibility of an antagonist of CCR2-mediated migration that does not cause CCL2 elevation, by sparing chemokine binding to CCR2 and hence its scavenging. The goal of the present proposal is to discover such antagonists via a computationally guided approach, by pursuing two Specific Aims: (1) Via in silico compound library screening against an ensemble of CCR2:CCL2 complex models, identify chemicals that bind CCR2 concurrently with CCL2, and characterize their pharmacology in vitro. (2) Computationally elucidate the dynamics of CCR2 in complex with CCL2, and identify G-protein- incompatible states with the potential to enrich for scavenging-sparing inhibitors in VLS. The outcome of the proposed work will be the discovery of the first chemical probes with novel pharmacology in relation to CCR2: the inhibitors of CCR2-mediated cell migration that spare CCL2 scavenging. These molecules will assist investigations of antagonist-induced CCL2 elevation in vivo by the applicant’s group and others. The findings will also inform future efforts of therapeutic targeting of CCR2 for inflammation and cancer.