Novel estrogen receptor ligands and their structure-activity relationship evaluated by scintillation proximity assay for high-throughput screening

Abstract

The estrogen receptor (ER) is an important drug target with allosteric characteristics that binds orthotopic hormones and other ligands. A recently developed scintillation proximity (SPA)-based assay for high-throughput screening (HTS) of compound libraries was used to identify novel estrogen receptor ligands that might have ER subtype selective binding activity. Radioligand binding was determined in a multi-detector scintillation counter designed for microtitration plates. Equilibrium binding experiments and kinetic competition tests were performed with [ 3 H]-estradiol and human ERa and ERb receptors. A library of 6,000 structurally diverse compounds was screened. From this, several novel ligands were identified that showed pronounced subtype-selective differences in ligand binding for ERa and ERb. The observed equilibrium dissociation constant (K d ) for the binding of [ 3 H]estradiol to ERa and ERb receptors were approximately 0.25 and 0.64 nM, respectively. When 17b-estradiol, raloxifene and daidzein were tested for binding affinity to ERa in a competition assay, the IC 50 values were 0.34, 1.31, and 75.6 nM, respectively. When tested for binding affinity to ERb, the IC 50 values were 1.05, 11.4, and 10.6 nM, respectively. The results obtained show that the methodology is valid in comparison to previously published data regarding estradiol and other standard compounds (raloxifene and daidzein) binding characteristics of estrogen receptors. The assay is also well suited to applied research as a tool in HTS of compound libraries in the search of ER ligands. Several novel active compounds were identified and selected as potent ER subtype ligands. Drug Dev Res 64:203-212, 2005.

A multiplexed screening method for agonists and antagonists of the estrogen receptor protein

“…Two commercial drugs, tamoxifen and raloxifene, were used as positive controls. The IC 50 values of tamoxifen and raloxifene obtained using this protein chip method were consistent with those reported in the literature [37][38][39][40][41][42]. The estrogen protein chips were also used to screen two libraries containing 29 synthetic compounds and 384 natural products, respectively.…”

Total Synthesis and Establishment of the Stereochemistry of Spiniferin-1, a Rare Planar Chiral Marine Natural Product with a 1,6-Methano[10]annulene Skeleton

“…18 The ligand competitive inhibition binding assays of (±)-dihydrospiniferin-1, as well as its analogues, on estrogen receptors ER α and ER β have also been reported. 19…”

Deoxyfluorination of Peracetylated Pyranose Hemiacetal with Fluoroalkanesulfonyl Fluoride

“…羟基睾酮的高烯丙位重排反应 [11] 及应用于海洋天然产 物 spiniferin 1 [12] 和新型雌激素拮抗剂 [13] 的全合成中; 和 赤式邻位二醇的反应制备顺式环氧化合物并应用于桥 环黄皮酰胺的全合成 [14] ; 和烯烃作用生成环氧化合物 的反应 [15] ; 作为缩合试剂诱导羧酸与醇和胺的酯化反 应和酰胺化反应生成羧酸酯和酰胺等 [16] . 在我们所考…”