| A multiplexed cell assay in HepG2 cells for the identification of delta-5, delta-6, and delta-9 desaturase and elongase inhibitors. | |
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MedLine Citation:
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PMID: 20086206 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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A multiplexed cell assay has been optimized to measure the activities of fatty acyl-CoA elongase, delta-5 desaturase (Delta5D), delta-6 desaturase (Delta6D), and delta-9 desaturase (Delta9D) together using (14)C-labeled tracers in HepG2 cells, which express the human stearoyl-CoA desaturase-1 isoform (SCD1) exclusively. The Delta5 and Delta9 desaturase activities are indexed by the efficient conversion of [1-(14)C]-eicosatrienoic acid (C20:3, cis-8,11,14) to (14)C-arachidonic acid (C20:4, cis-5,8,11,14) and the conversion of [1-(14)C]-stearic acid to (14)C-oleic acid (C18:1, cis-9), respectively. CP-74006 potently blocks the Delta5D activity with an IC(50) value of 20 nM and simplifies the metabolism of [1-(14)C]-alpha-linolenate (C18:3, cis-9,12,15) by accumulating (14)C-eicosatetraenoic acid (C20:4, cis-8,11,14,17) as the major (14)C-eicosatrienoic acid (C20:3, cis-11,14,17) and (14)C-docosatetraenoic acid (C22:4, cis-10,13,16,19) as the minor metabolites through Delta6 desaturation and elongation. This simplified metabolite spectrum enables the delineation of the Delta6D activity by comparing the combined Delta6D/elongase activity index of the (14)C-(C20:4/C18:3) ratio with the corresponding elongation index of the (14)C-(C20:3/C18:3) ratio following compound treatment. SC-26196 and sterculic acid specifically inhibit the Delta6D and Delta9D activities with an IC(50) value of 0.1 microM and 0.9 microM, respectively. This medium-throughput cell assay provides an efficient tool in the identification of specific desaturase and elongase inhibitors. |
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Authors:
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Lei Zhang; Yeeman Ramtohul; Sebastien Gagné; Angela Styhler; Hao Wang; Jocelyne Guay; Zheng Huang |
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Publication Detail:
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Type: Journal Article Date: 2010-01-19 |
Journal Detail:
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Title: Journal of biomolecular screening Volume: 15 ISSN: 1552-454X ISO Abbreviation: J Biomol Screen Publication Date: 2010 Feb |
Date Detail:
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Created Date: 2010-02-05 Completed Date: 2010-04-27 Revised Date: 2011-05-23 |
Medline Journal Info:
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Nlm Unique ID: 9612112 Medline TA: J Biomol Screen Country: United States |
Other Details:
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Languages: eng Pagination: 169-76 Citation Subset: IM |
Affiliation:
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Merck Frosst Center for Therapeutic Research, Montreal, Canada. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Acetyltransferases
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antagonists & inhibitors*,
chemistry,
metabolism Acyl Coenzyme A / antagonists & inhibitors*, chemistry, metabolism Biological Assay* Carbon Radioisotopes / diagnostic use Fatty Acid Desaturases / antagonists & inhibitors*, chemistry, metabolism Hep G2 Cells Humans Inhibitory Concentration 50 Kinetics Linoleoyl-CoA Desaturase / antagonists & inhibitors*, chemistry, metabolism Models, Biological Models, Chemical |
| Chemical | |
Reg. No./Substance:
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0/Acyl Coenzyme A; 0/Carbon Radioisotopes; 362-66-3/stearoyl-coenzyme A; EC 1.14.19.-/Fatty Acid Desaturases; EC 1.14.19.3/Linoleoyl-CoA Desaturase; EC 1.14.99.-/delta-5 fatty acid desaturase; EC 2.3.1.-/Acetyltransferases; EC 2.3.1.-/fatty acid elongases |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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