Receptor transactivation assay

Zhu ZR. 2007. Use of cryopreserved transiently transfected cells in high-throughput pregnane X receptor transactivation assay. J. Biomol. Screen. 12, 248-254. 

Organon disclosed A510 steroids as ERa-selective agonists [50]. Compound 10 showed 290-fold ERa selectivity in a receptor-binding assay and over 50-fold ERa selectivity in a CHO cell-based transactivation assay. 

Benzothiophenes 2-Phenyl-benzothiophenes were described as ERP-selective agonists by Organon [71]. It was found that dibutyryl analog 42 exhibits 15-fold ERP selectivity in a CHO cell-based transactivation assay. Similar observations were made by Novartis [72]. Receptor binding studies showed 12-fold ERP selectivity for 43. Introduction of substituents at C3 or C4 reduced ERP selectivity. 

At least two other chemotypes, tetrahydroquinolines (Fig. 25.1.5, 25 [87-88]) and amidoketones (Fig. 25.1.5, 26 [89, 90] also reviewed in Ref. [6]) have been shown to directly (ligand binding assays) or indirectly (cell based reporter gene transactivation assays) interact with the ecdysone receptor. These new ecdysone receptor binding chemistries could lead to new products for control of insect pests not controlled by current BAH insecticides. 

The constitutive androstane receptor (CAR) and pregnane X receptor (PXR) are important nuclear receptors involved in the regulation of cellular responses from exposure to many xenobiotics and various physiological processes. In recent years, CAR and PXR transactivation assays have freqnently been used during the discovery stage as a screening tool for CYP induction is PXR reporter assays (Sinz et al., 2008). 

CYP, cytochrome P450 PAMPA, parallel artificial membrane permeability assay Caco-2, human colonic adenocarcinoma cell MDCK, Madin Darby canine kidney cell rCYP, recombinant cytochrome P450 HLM, human liver microsomes PXR-TA, pregnane X receptor transactivation Fa2N-4, Fa2N-4 cryopreserved immortalized hepatocytes P-gp, P-glycoprotein. 

Receptor transactivation activity ED 50, nM (activation at 1 pM, %) Cotransfection assay conditions ... 

Organization for Economic Cooperation and Development (2012) Performance-based test guideline for stably transfected transactivation in vitro assays to detect estrogen receptor agonists 455... 

Zhu Z, Kim S, Chen T, Lin JH, Bell A, Bryson J, Dubaquie Y, Yan N, Yanchunas J, Xie D, Stoffel R, Sinz M, Dickinson K. Correlation of high-throughput pregnane X receptor (pxr) transactivation and binding assays. J Biomol Screen 2004 9 533-540. 

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