Faloon Patrick W., Dockendorff Chris, Germain Andrew, Yu Miao, Nag Partha Pratim, Youngsaye Willmen, Bennion Melissa, Lam Garrett, Paterson Conor, Dandapani Sivaraman, Munoz Benito, Perez Jose, Palmer Michelle, Nieland Thomas J.F., Krieger Monty, Schreiber Stuart L.
Chemical Biology Platform, Broad Institute
Biology Department, Massachusetts Institute of Technology
Scavenger receptor, class B, type I (SR-BI) mediates selective uptake of cholesterol from high density lipoprotein (HDL) particles, a poorly understood process that is distinct from endocytic uptake of lipoproteins, such as low density lipoprotein (LDL). We set out to find small molecules that modulate SR-BI function and could be used to characterize the mechanisms involved in HDL uptake. Using a cell-based DiI-HDL uptake assay, we performed a high-throughput screen (HTS) of the National Institutes of Health Molecular Libraries Probe Centers Network (NIH MLPCN) compound library. Of 319,533 compounds, 3,046 compounds (0.96%) were classified as inhibitors of DiI-HDL uptake. A bis-amide (CID 650573) was identified in the primary HTS as an inhibitor. It had potent activity upon retesting in the primary assay, a low hit rate in other MLPCN assays, and possessed structural properties suitable for analog synthesis. Structure activity relationship (SAR) studies were performed to improve potency and to minimize deleterious properties. These efforts generated a probe (CID 53393835/ML279), a low nanomolar inhibitor with improved potency and decreased metabolic liability. ML279 was tested for efflux inhibition, modulation of HDL binding to SR-BI, and inhibition of endocytosis. ML279 functions by inhibiting both SR-BI-mediated uptake and efflux of free cholesterol to HDL particles. As a tool compound, ML279 is superior to existing small-molecule inhibitors of SR-BI (e.g., BLT-1 and ITX-5061) and will be useful in elucidating how SR-BI mediates lipid transport. In addition, it could clarify the role of SR-BI in a number of biological processes where it plays a crucial role.
清道夫受体B类I型（SR-BI）介导从高密度脂蛋白（HDL）颗粒中选择性摄取胆固醇，这是一个了解甚少的过程，与诸如低密度脂蛋白（LDL）等脂蛋白的内吞摄取不同。我们着手寻找能够调节SR-BI功能且可用于表征HDL摄取所涉及机制的小分子。利用基于细胞的DiI-HDL摄取试验，我们对美国国立卫生研究院分子文库探针中心网络（NIH MLPCN）化合物库进行了高通量筛选（HTS）。在319,533种化合物中，有3,046种化合物（0.96%）被归类为DiI-HDL摄取抑制剂。在初次高通量筛选中鉴定出一种双酰胺（化合物标识符650573）为抑制剂。它在初次试验中复测时具有强效活性，在其他MLPCN试验中的命中率较低，并且具有适合类似物合成的结构特性。进行了构效关系（SAR）研究以提高效力并使有害特性最小化。这些努力产生了一种探针（化合物标识符53393835/ML279），一种低纳摩尔抑制剂，其效力提高且代谢负担降低。对ML279进行了外排抑制、HDL与SR-BI结合的调节以及内吞作用抑制的测试。ML279通过抑制SR-BI介导的摄取以及游离胆固醇向HDL颗粒的外排来发挥作用。作为一种工具化合物，ML279优于现有的SR-BI小分子抑制剂（例如BLT-1和ITX-5061），将有助于阐明SR-BI如何介导脂质转运。此外，它可以阐明SR-BI在许多发挥关键作用的生物学过程中的作用。
