Name: Summary of probe development efforts to identify novel modulators of the Retinoic acid receptor-related Orphan Receptors (ROR). ..more
Targets
Depositor Specified Assays
Show more |
| AID | Name | Type | Probe | Comment |
|---|---|---|---|---|
| 561 | Primary Cell-based High Throughput Screening assay for inhibitors of the Retinoic Acid Receptor-related orphan receptor A (RORA) | screening | Primary screen (RORA inhibitors in singlicate) | |
| 610 | Dose-response cell-based assay for inhibitors of the Retinoic Acid Receptor-related orphan receptor A (RORA) | confirmatory | Dose response (RORA inhibitors in triplicate) | |
| 1901 | Summary of probe development efforts to identify inhibitors of the Retinoic Acid Receptor-related orphan receptor A (RORA). | summary | Summary (RORA inhibitors) | |
| 2117 | Late stage results from the probe development effort to identify novel modulators of the Retinoic acid receptor-related Orphan Receptors (ROR). | screening | Late stage results (ROR modulators). | |
| 2277 | Center Based Initiative to identify novel modulators of the Retinoic acid receptor-related Orphan Receptors (ROR): luminescence-based high throughput cell-based assay to identify modulators of human nuclear receptors. | screening | Primary screen (ROR modulators) | |
| 463078 | Late stage assay provider results from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): luminescent-based assay to identify RORA inhibitors | other | Late stage results (RORA inhibitors) | |
| 463143 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): luminescence-based cell-based assay to identify activators of the liver X receptor (LXR) | screening | Late stage counterscreen (LXR activators, 6 replicates) | |
| 463144 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): luminescence-based cell-based assay to identify inhibitors of glucose-6-phosphatase (G6PC) | other | Late stage counterscreen (G6PC inhibitors, 6 replicates) | |
| 463145 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): luminescence-based cell-based assay to identify activators of the farnesoid X receptor (FXR) | screening | Late stage counterscreen (FXR activators, 6 replicates) | |
| 463147 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): luminescence-based cell-based assay to identify RORG inhibitors | screening | Late stage counterscreen (RORG inhibitors, 6 replicates) | |
| 463148 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): luminescence-based cell-based assay to identify inhibitors of the human herpes virus VP16 transcriptional activator protein (VP16) | screening | Late stage counterscreen (VP16 inhibitors, 6 replicates) | |
| 463150 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): luminescence-based dose response assay to identify activators of the liver X receptor (LXR) | confirmatory | Late stage dose response counterscreen (LXR activators, 8 replicates) | |
| 463151 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): luminescent-based dose response assay to identify RORG inhibitors | confirmatory | Late stage dose response counterscreen (RORG inhibitors, 8 replicates) | |
| 463152 | Late stage assay provider results from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): luminescence-based dose response assay to identify RORA inhibitors | confirmatory | Late stage dose response (RORA inhibitors, 8 replicates) | |
| 463172 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): luminescence-based cell-based dose response assay to identify inhibitors of glucose-6-phosphatase (G6PC) | confirmatory | Late stage dose response counterscreen (G6PC inhibitors, 8 replicates) | |
| 504899 | Late stage assay provider results from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): Diet-Induced obesity (DIO) mouse model studies to assess the effect of probe candidate on hepatic glucose production | other | Late stage assay (DIO studies, hepatic glucose production) | |
| 504906 | Late stage assay provider results from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): radioligand binding assay for RORa using [3H]25-hydroxycholesterol to determine whether probe candidates bind directly to RORa | other | Late stage assay (radioligand binding RORa using [3H]25-hydroxycholesterol) | |
| 504907 | Late stage assay provider results from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptors (RORA): fluorescence-based real-time polymerase chain reaction assay to determine the effect of probe candidates on endogenous expression of glucose-6-phosphatase (G6PC) | other | Late stage assay (endogenous expression G6PC) | |
| 624276 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptor Gamma (RORC): radioligand binding for ROR gamma using Scintillation Proximity Assay (SPA) | confirmatory | 1 | Late stage counterscreen (radioligand binding for ROR gamma using SPA) |
| 624277 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptor Gamma (RORC): Luminescence-based cell-based assays using RORE-Luc, IL-17-Luc, and ABCA1-Luc Reporters | other | 1 | Late stage counterscreen (RORE-Luc, IL-17-Luc, and ABCA1-Luc reporter panels) |
| 624278 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptor Gamma (RORC): Fluorescence-based cell-based QPCR assays to determine endogenous expression of ROR gamma target genes, proinflammatory IL-17 and IL-23R | other | 1 | Late stage counterscreen (endogenous expression of ROR gamma target genes, proinflammatory IL-17 and IL-23R) |
| 624279 | Late stage assay provider counterscreen from the probe development effort to identify selective inverse agonists of the Retinoic acid receptor-related Orphan Receptor Gamma (RORC): luminescence-based cell-based dose response panel assay against ROR alpha, ROR gamma, LXR, FXR, and VP16 | confirmatory | 1 | Late stage dose response counterscreen (ROR alpha, ROR gamma, LXR, FXR, and VP16 panels) |
Description:
Source (MLPCN Center Name): The Scripps Research Institute Molecular Screening Center (SRISMC)
Center Affiliation: The Scripps Research Institute, TSRI
Assay Provider: Patrick Griffin, TSRI
Network: Molecular Library Probe Production Center Network (MLPCN)
Grant Proposal Number: U54 MH084512
Grant Proposal PI: Patrick Griffin, TSRI
External Assay ID: ROR_MOD_PROBES_SUMMARY
Name: Summary of probe development efforts to identify novel modulators of the Retinoic acid receptor-related Orphan Receptors (ROR).
Description:
Nuclear receptors are a family of small molecule and hormone-regulated transcription factors that share conserved DNA-binding and ligand-binding domains. Small pharmacological compounds able to bind to the cleft of the ligand-binding domain could alter its conformation and subsequently modify transcription of target genes. Such ligand agonists and/or antagonists have already been successfully designed for 23 nuclear receptors among the 48 previously identified in the human genome (1-3). RORalpha (RORalpha; RORA; NR1F1) is one of three related orphan nuclear receptors, including RORbeta (RORbeta]; RORB; NR1F2) and RORgamma (RORgamma; RORC; NR1F3), known as "Retinoic Acid Receptor-related orphan receptors" (4).
RORA has unusual potential as a therapeutic target for the "metabolic syndrome" which results in pathologies such as insulin resistance, dyslipidemia, hypertension, and a pro-inflammatory state, that greatly elevates the risk of diabetes and atherosclerosis (5).The related RORC demonstrates significant expression in metabolic tissues such as liver, adipose, and skeletal muscle (6). These two receptors are implicated in several key aspects of this metabolic pathogenesis. For instance, the staggerer mouse, which carries a homozygous germline inactivation of RORA, shows low body weight, high food consumption (7-9), elevated angiogenesis in response to ischemia (10), susceptibility to atherosclerosis (9), and an abnormal serum lipid profile (11). RORG null mice exhibit normal plasma cholesterol levels, but when bred with the RORA staggerer mice, the resulting RORalpha/gamma knockout exhibits hypoglycemia not found in the single mutant animals. These studies reveal the functional redundancy of RORa and RORg in regulating blood glucose levels and highlight the need for RORalpha/gamma ligands that can bind to these receptors and modulate their transcriptional activity.
Summary of Probe Development Effort:
Following primary compound profiling of a cell-based GAL4-human nuclear receptor profiling assay, certain compounds were identified as possible candidates for probe development. Compounds derived from these initial candidates were purchased as powders or synthesized at the SRIMSC and were tested for their ability to inhibit RORa and RORg in luciferase-based reporter assays performed at a single concentration of 10 micromolar or in dose response assays starting at a nominal concentration of 20 micromolar. Compounds were subsequently counterscreened against the liver X receptor (LXR) and the farnesoid X receptor (FXR) to determine selectivity in singlicate and in dose response assays. Finally, compounds of interest were tested at a single concentration of 10 micromolar against VP16 to determine whether they were non-selective or cytotoxic. These late stage efforts have been described in AID 2117.
The above probe development efforts resulted in the identification of two probes. The benzenesulfonamide compound SID 85257301 previously identified as a selective agonist of LXR (12) was identified here as a novel RORa/g inverse agonist probe that decreases the transcriptional activity of both ROR receptors (IC50 values = 2.0 and 1.73 micromolar, respectively). The second probe, SID 85257298 synthesized at the SRIMSC, was found to decrease RORa transcriptional activity (IC50 value = 2.47 micromolar). SID 85257298 represents an improvement over the prior art due to its lack of activity for LXR. Probe compound SID 85257298 does not have activity against RORg (IC50 > 20 micromolar). These two probes are useful tools for examining ROR biology.
The results of our probe development efforts can be found at http://mlpcn.florida.scripps.edu/index.php/probes/probe-reports.html. A probe report for SIDs 85257301 and 85257298 can be found in the Molecular Libraries Bookshelf (PubMed Books) (http://www.ncbi.nlm.nih.gov/books) under ML125 and ML124, respectively. A paper has been published (13).
References:
(1) Evans RM. The nuclear receptor superfamily: a rosetta stone for physiology. Mol Endocrinol 19:1429-1438, 2005.
(2) Kliewer SA, Lehmann JM, and Willson TM. Orphan nuclear receptors: shifting endocrinology into reverse. Science 284: 757-760, 1999.
(3) Li Y, Lambert MH, and Xu HE. Activation of nuclear receptors: a perspective from structural genomics. Structure (Camb) 11: 741-746., 2003.
(4) Jetten AM, Kurebayashi S, and Ueda E. The ROR nuclear orphan receptor subfamily: critical regulators of multiple biological processes. Prog Nucleic Acid Res Mol Biol 69: 205-247, 2001.
(5) Grundy SM, Brewer HB, Jr., Cleeman JI, Smith SC, Jr., and Lenfant C. Definition of metabolic syndrome: report of the National Heart, Lung, and Blood Institute/American Heart Association conference on scientific issues related to definition. Arterioscler Thromb Vasc Biol 24: e13-18, 2004.
(6) Medvedev A, Yan ZH, Hirose T, Giguere V, Jetten AM. Cloning of a cDNA encoding the murine orphan receptor RZR/ROR gamma and characterization of its response element. Gene. 1996 Nov 28;181(1-2):199-206.
(7) Bertin R, Guastavino JM, and Portet R. Effects of cold acclimation on the energetic metabolism of the staggerer mutant mouse. Physiol Behav 47: 377-380, 1990.
(8) Guastavino JM, Bertin R, and Portet R. Effects of the rearing temperature on the temporal feeding pattern of the staggerer mutant mouse. Physiol Behav 49: 405-409, 1991.
(9) Mamontova A, Seguret-Mace S, Esposito B, Chaniale C, Bouly M, Delhaye-Bouchaud N, Luc G, Staels B, Duverger N, Mariani J, and Tedgui A. Severe atherosclerosis and hypoalphalipoproteinemia in the staggerer mouse, a mutant of the nuclear receptor RORalpha. Circulation 98: 2738-2743., 1998.
(10) Besnard S, Silvestre J-S, Duriez M, Bakouche J, Lemaigre-Dubreuil Y, Mariani J, Levy BI, and Tedgui A. Increased ischemia-induced angiogenesis in the staggerer mouse, a mutant of the nuclear receptor RORa. Circ Res 89: 1209-1215, 2001.
(11) Raspe E, Duez H, Gervois P, Fievet C, Fruchart J-C, Besnard S, Mariani J, Tedgui A, and Staels B. Transcriptional regulation of apolipoprotein C-III gene expression by the orphan nuclear receptor RORalpha. J Biol Chem 276: 2865-2871, 2001.
(12) Schultz JR, Tu H, Luk A, Repa JJ, Medina JC, Li L, Schwendner S, Wang S, Thoolen M, Mangelsdorf DJ, Lustig KD, Shan B. Role of LXRs in control of lipogenesis. Genes Dev. 2000 Nov 15;14(22):2831-8.
(13) Kumar N, Solt LA, Conkright JJ, Wang Y, Istrate MA, Busby SA, Garcia-Ordonez R, Burris TP, Griffin PR. The benzenesulfoamide T0901317 is a novel RORalpha/gamma Inverse Agonist. Mol Pharm. in press.
Keywords:
Late stage, probes, RAR-related orphan receptor A, ROR alpha, RORa, RORA, nuclear receptor, library, counterscreen, low throughput assay, RZRA, ROR1, ROR2, ROR3, NR1F1, inhibitor, activator, transcriptional assay, luciferase, luminescence, ROR gamma, RORg, RORC, RORG, NR1F3, Primary screen, Scripps Florida, The Scripps Research Institute Molecular Screening Center, SRIMSC, Molecular Libraries Probe Production Centers Network, MLPCN.
Center Affiliation: The Scripps Research Institute, TSRI
Assay Provider: Patrick Griffin, TSRI
Network: Molecular Library Probe Production Center Network (MLPCN)
Grant Proposal Number: U54 MH084512
Grant Proposal PI: Patrick Griffin, TSRI
External Assay ID: ROR_MOD_PROBES_SUMMARY
Name: Summary of probe development efforts to identify novel modulators of the Retinoic acid receptor-related Orphan Receptors (ROR).
Description:
Nuclear receptors are a family of small molecule and hormone-regulated transcription factors that share conserved DNA-binding and ligand-binding domains. Small pharmacological compounds able to bind to the cleft of the ligand-binding domain could alter its conformation and subsequently modify transcription of target genes. Such ligand agonists and/or antagonists have already been successfully designed for 23 nuclear receptors among the 48 previously identified in the human genome (1-3). RORalpha (RORalpha; RORA; NR1F1) is one of three related orphan nuclear receptors, including RORbeta (RORbeta]; RORB; NR1F2) and RORgamma (RORgamma; RORC; NR1F3), known as "Retinoic Acid Receptor-related orphan receptors" (4).
RORA has unusual potential as a therapeutic target for the "metabolic syndrome" which results in pathologies such as insulin resistance, dyslipidemia, hypertension, and a pro-inflammatory state, that greatly elevates the risk of diabetes and atherosclerosis (5).The related RORC demonstrates significant expression in metabolic tissues such as liver, adipose, and skeletal muscle (6). These two receptors are implicated in several key aspects of this metabolic pathogenesis. For instance, the staggerer mouse, which carries a homozygous germline inactivation of RORA, shows low body weight, high food consumption (7-9), elevated angiogenesis in response to ischemia (10), susceptibility to atherosclerosis (9), and an abnormal serum lipid profile (11). RORG null mice exhibit normal plasma cholesterol levels, but when bred with the RORA staggerer mice, the resulting RORalpha/gamma knockout exhibits hypoglycemia not found in the single mutant animals. These studies reveal the functional redundancy of RORa and RORg in regulating blood glucose levels and highlight the need for RORalpha/gamma ligands that can bind to these receptors and modulate their transcriptional activity.
Summary of Probe Development Effort:
Following primary compound profiling of a cell-based GAL4-human nuclear receptor profiling assay, certain compounds were identified as possible candidates for probe development. Compounds derived from these initial candidates were purchased as powders or synthesized at the SRIMSC and were tested for their ability to inhibit RORa and RORg in luciferase-based reporter assays performed at a single concentration of 10 micromolar or in dose response assays starting at a nominal concentration of 20 micromolar. Compounds were subsequently counterscreened against the liver X receptor (LXR) and the farnesoid X receptor (FXR) to determine selectivity in singlicate and in dose response assays. Finally, compounds of interest were tested at a single concentration of 10 micromolar against VP16 to determine whether they were non-selective or cytotoxic. These late stage efforts have been described in AID 2117.
The above probe development efforts resulted in the identification of two probes. The benzenesulfonamide compound SID 85257301 previously identified as a selective agonist of LXR (12) was identified here as a novel RORa/g inverse agonist probe that decreases the transcriptional activity of both ROR receptors (IC50 values = 2.0 and 1.73 micromolar, respectively). The second probe, SID 85257298 synthesized at the SRIMSC, was found to decrease RORa transcriptional activity (IC50 value = 2.47 micromolar). SID 85257298 represents an improvement over the prior art due to its lack of activity for LXR. Probe compound SID 85257298 does not have activity against RORg (IC50 > 20 micromolar). These two probes are useful tools for examining ROR biology.
The results of our probe development efforts can be found at http://mlpcn.florida.scripps.edu/index.php/probes/probe-reports.html. A probe report for SIDs 85257301 and 85257298 can be found in the Molecular Libraries Bookshelf (PubMed Books) (http://www.ncbi.nlm.nih.gov/books) under ML125 and ML124, respectively. A paper has been published (13).
References:
(1) Evans RM. The nuclear receptor superfamily: a rosetta stone for physiology. Mol Endocrinol 19:1429-1438, 2005.
(2) Kliewer SA, Lehmann JM, and Willson TM. Orphan nuclear receptors: shifting endocrinology into reverse. Science 284: 757-760, 1999.
(3) Li Y, Lambert MH, and Xu HE. Activation of nuclear receptors: a perspective from structural genomics. Structure (Camb) 11: 741-746., 2003.
(4) Jetten AM, Kurebayashi S, and Ueda E. The ROR nuclear orphan receptor subfamily: critical regulators of multiple biological processes. Prog Nucleic Acid Res Mol Biol 69: 205-247, 2001.
(5) Grundy SM, Brewer HB, Jr., Cleeman JI, Smith SC, Jr., and Lenfant C. Definition of metabolic syndrome: report of the National Heart, Lung, and Blood Institute/American Heart Association conference on scientific issues related to definition. Arterioscler Thromb Vasc Biol 24: e13-18, 2004.
(6) Medvedev A, Yan ZH, Hirose T, Giguere V, Jetten AM. Cloning of a cDNA encoding the murine orphan receptor RZR/ROR gamma and characterization of its response element. Gene. 1996 Nov 28;181(1-2):199-206.
(7) Bertin R, Guastavino JM, and Portet R. Effects of cold acclimation on the energetic metabolism of the staggerer mutant mouse. Physiol Behav 47: 377-380, 1990.
(8) Guastavino JM, Bertin R, and Portet R. Effects of the rearing temperature on the temporal feeding pattern of the staggerer mutant mouse. Physiol Behav 49: 405-409, 1991.
(9) Mamontova A, Seguret-Mace S, Esposito B, Chaniale C, Bouly M, Delhaye-Bouchaud N, Luc G, Staels B, Duverger N, Mariani J, and Tedgui A. Severe atherosclerosis and hypoalphalipoproteinemia in the staggerer mouse, a mutant of the nuclear receptor RORalpha. Circulation 98: 2738-2743., 1998.
(10) Besnard S, Silvestre J-S, Duriez M, Bakouche J, Lemaigre-Dubreuil Y, Mariani J, Levy BI, and Tedgui A. Increased ischemia-induced angiogenesis in the staggerer mouse, a mutant of the nuclear receptor RORa. Circ Res 89: 1209-1215, 2001.
(11) Raspe E, Duez H, Gervois P, Fievet C, Fruchart J-C, Besnard S, Mariani J, Tedgui A, and Staels B. Transcriptional regulation of apolipoprotein C-III gene expression by the orphan nuclear receptor RORalpha. J Biol Chem 276: 2865-2871, 2001.
(12) Schultz JR, Tu H, Luk A, Repa JJ, Medina JC, Li L, Schwendner S, Wang S, Thoolen M, Mangelsdorf DJ, Lustig KD, Shan B. Role of LXRs in control of lipogenesis. Genes Dev. 2000 Nov 15;14(22):2831-8.
(13) Kumar N, Solt LA, Conkright JJ, Wang Y, Istrate MA, Busby SA, Garcia-Ordonez R, Burris TP, Griffin PR. The benzenesulfoamide T0901317 is a novel RORalpha/gamma Inverse Agonist. Mol Pharm. in press.
Keywords:
Late stage, probes, RAR-related orphan receptor A, ROR alpha, RORa, RORA, nuclear receptor, library, counterscreen, low throughput assay, RZRA, ROR1, ROR2, ROR3, NR1F1, inhibitor, activator, transcriptional assay, luciferase, luminescence, ROR gamma, RORg, RORC, RORG, NR1F3, Primary screen, Scripps Florida, The Scripps Research Institute Molecular Screening Center, SRIMSC, Molecular Libraries Probe Production Centers Network, MLPCN.
Protocol
Please see Related Bioassays and reference (13) for all protocols performed in this Center-based probe development effort.
Comment
Probes were identified.
Result Definitions
| TID | Name | Description | Histogram | Type | Unit | |
|---|---|---|---|---|---|---|
| Outcome | The BioAssay activity outcome | Outcome | ||||
| Score | The BioAssay activity ranking score |  | Integer | |||
| 1 | ML # | Unique alphanumeric identifier assigned to a chemical probe molecule within the Molecular Libraries Probe Production Centers Network (MLPCN). | String |
Additional Information
Grant Number: U54 MH084512
Data Table (Concise)
Classification
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