De Filippo Elisabetta, Manga Prashiela, Schiedel Anke C
Pharmaceutical Chemistry I, PharmaCenter Bonn, University of Bonn, Bonn, Germany.
Ronald O. Perelman Department of Dermatology and Department of Cell Biology, New York University School of Medicine, New York, New York, United States.
Invest Ophthalmol Vis Sci. 2017 Jun 1;58(7):3118-3126. doi: 10.1167/iovs.16-21128.
GPR143 regulates melanosome biogenesis and organelle size in pigment cells. The mechanisms underlying receptor function remain unclear. G protein-coupled receptors (GPCRs) are excellent pharmacologic targets; thus, we developed and applied a screening approach to identify potential GPR143 ligands and chemical modulators.
GPR143 interacts with β-arrestin; we therefore established a β-arrestin recruitment assay to screen for compounds that modulate activity. Because GPR143 is localized intracellularly, screening with the wild-type receptor would be restricted to agents absorbed by the cell. For the screen we used a mutant receptor, which shows similar basal activity as the wild type but traffics to the plasma membrane. We tested two compound libraries and investigated validated hits for their effects on melanocyte pigmentation.
GPR143, which showed high constitutive activity in the β-arrestin assay, was inhibited by several compounds. The three validated inhibitors (pimozide, niclosamide, and ethacridine lactate) were assessed for impact on melanocytes. Pigmentation and expression of tyrosinase, a key melanogenic enzyme, were reduced by all compounds. Because GPR143 appears to be constitutively active, these compounds may turn off its activity.
X-linked ocular albinism type I, characterized by developmental eye defects, results from GPR143 mutations. Identifying pharmacologic agents that modulate GPR143 activity will contribute significantly to our understanding of its function and provide novel tools with which to study GPCRs in melanocytes and retinal pigment epithelium. Pimozide, one of three GPR143 inhibitors identified in this study, maybe be a good lead structure for development of more potent compounds and provide a platform for design of novel therapeutic agents.
GPR143调节色素细胞中黑素小体的生物发生和细胞器大小。受体功能的潜在机制尚不清楚。G蛋白偶联受体（GPCRs）是理想的药物靶点；因此，我们开发并应用了一种筛选方法来鉴定潜在的GPR143配体和化学调节剂。
GPR143与β-抑制蛋白相互作用；因此，我们建立了一种β-抑制蛋白募集试验来筛选调节活性的化合物。由于GPR143定位于细胞内，用野生型受体进行筛选将仅限于细胞吸收的药物。在筛选中，我们使用了一种突变受体，它显示出与野生型相似的基础活性，但能转运到质膜。我们测试了两个化合物库，并研究了经过验证的命中化合物对黑素细胞色素沉着的影响。
在β-抑制蛋白试验中表现出高组成性活性的GPR143被几种化合物抑制。评估了三种经过验证的抑制剂（匹莫齐特、氯硝柳胺和乳酸依沙吖啶）对黑素细胞的影响。所有化合物均降低了色素沉着和关键黑素生成酶酪氨酸酶的表达。由于GPR143似乎具有组成性活性，这些化合物可能会关闭其活性。
以发育性眼缺陷为特征的I型X连锁眼白化病是由GPR143突变引起的。鉴定调节GPR143活性的药物将极大地有助于我们对其功能的理解，并提供研究黑素细胞和视网膜色素上皮中GPCRs的新工具。匹莫齐特是本研究中鉴定出的三种GPR143抑制剂之一，可能是开发更有效化合物的良好先导结构，并为设计新型治疗药物提供平台。
