Document Detail

Development and validation of a high-throughput screening assay for human long-chain fatty acid transport proteins 4 and 5.
MedLine Citation:
PMID:  20448275     Owner:  NLM     Status:  MEDLINE    
Dietary long-chain fatty acid (LCFA) uptake across cell membranes is mediated principally by fatty acid transport proteins (FATPs). Six subtypes of this transporter are differentially expressed throughout the human and rodent body. To facilitate drugs discovery against FATP subtypes, the authors used mammalian cell lines stably expressing the recombinant human FATP4 and 5 and developed a high-throughput screening (HTS) assay using a 96-well fluorometric imaging plate reader (FLIPR). LCFA uptake signal-to-background ratios were between 3- and 5-fold. Two 4-aryl-dihydropyrimidinones, j3 and j5, produced inhibition of FATP4 with a half-maximal inhibitory concentration (IC(50)) of 0.21 and 0.63 microM, respectively, and displayed approximately 100-fold selectivity over FATP5. The US Drug Collection library was screened against the FATP5. A hit rate of around 0.4% was observed with a Z' factor of 0.6 +/- 0.2. Two confirmed hits are bile acids, chenodiol and ursodiol with an IC(50) of 2.4 and 0.22 microM, respectively. To increase throughput, a single time point measurement in a 384-well format was developed using the Analyst HT, and the results are comparable with the 96-well format. In conclusion, the FATP4 and 5 cell-based fluorescence assays are suitable for a primary drug screen, whereas differentiated cell lines are useful for a secondary drug screen.
Wei Zhou; Peter Madrid; Amy Fluitt; Andreas Stahl; Xinmin Simon Xie
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Validation Studies     Date:  2010-05-06
Journal Detail:
Title:  Journal of biomolecular screening     Volume:  15     ISSN:  1552-454X     ISO Abbreviation:  J Biomol Screen     Publication Date:  2010 Jun 
Date Detail:
Created Date:  2010-05-31     Completed Date:  2010-08-17     Revised Date:  2011-09-26    
Medline Journal Info:
Nlm Unique ID:  9612112     Medline TA:  J Biomol Screen     Country:  United States    
Other Details:
Languages:  eng     Pagination:  488-97     Citation Subset:  IM    
Biosciences Division, SRI International, Menlo Park, CA 94063, USA.
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MeSH Terms
Cell Line
Fatty Acid Transport Proteins / antagonists & inhibitors,  metabolism*
Fluorescent Dyes / metabolism
High-Throughput Screening Assays / methods*
Molecular Structure
Pyrimidinones / chemistry,  metabolism
Reproducibility of Results
Grant Support
R01 MH078194/MH/NIMH NIH HHS; R01 MH078194-01A2/MH/NIMH NIH HHS; R21 NS057052-01S1/NS/NINDS NIH HHS; R21NS57052/NS/NINDS NIH HHS
Reg. No./Substance:
0/Fatty Acid Transport Proteins; 0/Fluorescent Dyes; 0/Pyrimidinones; 0/SLC27A4 protein, human; 0/SLC27A5 protein, human

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