Abstract
The steroidogenic factor 1 (SF-1, also known as NR5A1) is a transcription factor belonging to the nuclear receptor superfamily. Whereas most of the members of this family have been extensively characterized, the therapeutic potential and pharmacology of SF-1 still remains elusive. Described here is the identification and characterization of selective inhibitory chemical probes of SF-1 by a rational ultra-high-throughput screening (uHTS) strategy. A set of 64,908 compounds from the National Institute of Health's Molecular Libraries Small Molecule Repository was screened in a transactivation cell-based assay employing a chimeric SF-1 construct. Two analogous isoquinolinones, ethyl 2-[2-[2-(2,3-dihydro-1,4-benzodioxin-7-ylamino)-2-oxoethyl]-1-oxoisoquinolin-5-yl]oxypropanoate (SID7969543) and ethyl 2-[2-[2-(1,3-benzodioxol-5-ylmethylamino)-2-oxoethyl]-1-oxoisoquinolin-5-yl]oxypropanoate and (SID7970631), were identified as potent submicromolar inhibitors, yielding IC50 values of 760 and 260 nM. The compounds retained their potency in a more physiologic functional assay employing the full-length SF-1 protein and its native response element, yielding IC50 values of 30 and 16 nM, respectively. The selectivity of these isoquinolinones was confirmed via transactivation-based functional assays for RAR-related orphan receptor A (RORA), Herpes simplex virus transcriptional activator protein Vmw65 (VP16), and liver receptor homolog 1 (LRH-1). Their cytotoxicity, solubility, permeability and metabolic stability were also measured. These isoquinolinones represent valuable chemical probes to investigate the therapeutic potential of SF-1.
Footnotes
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The SF-1 assay was initially developed with the National Institutes of Health grant CA099875. National Institutes of Health grant MH077624-01 supported the transfer of the assay to the MLSCN. The efforts of M.C., P.C., P.G., and F.M. were supported by the National Institutes of Health Molecular Library Screening Center Network grant U54-MH074404.
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ABBREVIATIONS: NR, nuclear receptor; SF-1, steroidogenic factor 1; SID7969543, ethyl 2-[2-[2-(2,3-dihydro-1,4-benzodioxin-7-ylamino)-2-oxoethyl]-1-oxoisoquinolin-5-yl]oxypropanoate; SID7970631, ethyl 2-[2-[2-(1,3-benzodioxol-5-ylmethylamino)-2-oxoethyl]-1-oxoisoquinolin-5-yl]oxypropanoate; SID4243980, 3-[(4-methoxyphenoxy)methyl]benzoic acid; SID4255902, 3-[(4-ethoxyphenoxy)methyl]benzoic acid; SID4261716, 3-{[2-chloro-4-(hydroxymethyl)phenoxy]methyl}benzoic acid; SID4263122, 3-[(2-Chlorophenoxy)methyl]benzoic acid; AC-45594, 4 (heptyloxy)phenol; aa, amino acid(s); LBD, ligand-binding domain; DBD DNA-binding domain; RORA, RAR-related orphan receptor A; CMV, cytomegalovirus; VP16, Herpes simplex virus transcriptional activator protein Vmw65; CHO, Chinese hamster ovary; PBS, phosphate-buffered saline; RH, relative humidity; uHTS, ultra-high-throughput screening; -NR, not cotransfected nuclear receptor; +NR, with cotransfected nuclear receptor; DMSO, dimethyl sulfoxide; HTS, high-throughput screening; HEK, human embryonic kidney; SFRE, steroidogenic factor 1 response element; MLSCN, Molecular Libraries Screening Center Network; LRH-1, liver receptor homolog 1.
- Received February 1, 2008.
- Accepted March 10, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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