Abstract
The chemokine receptor CXCR2 is involved in different inflammatory diseases, like chronic obstructive pulmonary disease, psoriasis, rheumatoid arthritis, and ulcerative colitis; therefore, it is considered an attractive drug target. Different classes of small CXCR2 antagonists have been developed. In this study, we selected seven CXCR2 antagonists from the diarylurea, imidazolylpyrimide, and thiazolopyrimidine class and studied their mechanisms of action at human CXCR2. All compounds are able to displace 125I-CXCL8 and inhibit CXCL8-induced β-arrestin2 recruitment. Detailed studies with representatives of each class showed that these compounds displace and antagonize CXCL8, most probably via a noncompetitive, allosteric mechanism. In addition, we radiolabeled the high-affinity CXCR2 antagonist SB265610 [1-(2-bromophenyl)-3-(4-cyano-1H-benzo[d] [1,2,3]-triazol-7-yl)urea] and subjected [3H]SB265610 to a detailed analysis. The binding of this radioligand was saturable and reversible. Using [3H]SB265610, we found that compounds of the different chemical classes bind to distinct binding sites. Hence, the use of a radiolabeled low-molecular weight CXCR2 antagonist serves as a tool to investigate the different binding sites of CXCR2 antagonists in more detail.
Footnotes
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This work was performed within the framework of the Dutch Top Institute Pharma Project T101-3 and The Netherlands Organization for Scientific Research.
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P.d.K. and J.v.H. contributed equally to this work.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.108.148387.
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ABBREVIATIONS: GPCR, G protein-coupled receptor; SB332235, 6-chloro-3-(3-(2,3-dichlorophenyl)ureido)-2-hydroxybenzenesulfonamide; SB225002, 1-(2-bromophenyl)-3-(4-cyano-1H-benzo[d][1,2,3]triazol-7-yl)urea; SB265610, 1-(2-bromophenyl)-3-(4-cyano-1H-benzo[d] [1,2,3]triazol-7-yl)urea; VUF10948, (R)-5-(benzylthio)-7-(1-hydroxybutan-2-ylamino)thiazolo[4,5-d]pyrimidin-2-ol; DMEM, Dulbecco's modified Eagle's medium; HEK, human embryonic kidney; compound 1, (S)-2-(2-(1H-imidazol-1-yl)-6-(octylthio)pyrimidin-4-ylamino)-N-(3-ethoxypropyl)-4-methylpentanamide; compound 2, (S)-N-(2-(2,5-dihydro-1H-pyrrol-1-yl)ethyl)-4-methyl-2-(6-methyl-2-(4-(4-(trifluoromethoxy)phenyl)-1H-imidazol-1-yl)pyrimidin-4-ylamino)pentanamide); compound 3, (S)-2-(6-butyl-2-(4-(4-chloro-3-(trifluoromethyl)phenyl)-1H-imidazol-1-yl)pyrimidin-4-ylamino)-4-methyl-N-(4,4,4-trifluorobutyl)pentanamide); GTPγS, guanosine-5′-O-(3-thio)triphosphate; Sch527123, 2-hydroxy-N,N-dimethyl-3-{2-[[(R)-1-(5-methyl-furan-2-yl)-propyl]amino]-3,4-dioxo-cyclobut-1-enylamino}-benzamide; AMD3100, 1,1′-[1,4-phenylenebis(methylene)]bis [1,4,8,11-tetraazacyclotetradecane] octohydrobromide dehydrate; TAK-799 (N,N-dimethyl-N-[4-[[[2-(4-methylphenyl)-6,7-dihydro-5H-benzocyclohepten-8-yl]carbon-yl]amino]benzyl]-tetrahydro-2H-pyran4-aminium chloride; BX471 N-(5-chloro-2-(2-(4-((4-fluorophenyl)methyl)-2-methyl-1-piperazinyl)-2-oxoethoxy)phenyl)urea hydrochloric acid.
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↵1 Current affiliation: National Institutes of Health Chemical Genomics Center, National Institutes of Health, Bethesda, Maryland.
- Received November 6, 2008.
- Accepted February 2, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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