Abstract
The interaction between CC chemokine receptor 2 (CCR2) with monocyte chemoattractant proteins, such as MCP-1, regulates the activation and recruitment of inflammatory leukocytes. In this study, we characterized (S)-3-[3,4-difluoro-phenyl)-propyl]-5-isoxazol-5-yl-2-thioxo-2,3-dihydro-1H-imidazole-4-carboxyl acid methyl ester (JNJ-27141491) as a noncompetitive and orally active functional antagonist of human (h)CCR2. JNJ-27141491 strongly suppressed hCCR2-mediated in vitro functions, such as MCP-1-induced guanosine 5′-O-(3-[35S]thio)triphosphate binding; MCP-1, -3, and -4-induced Ca2+ mobilization; and leukocyte chemotaxis toward MCP-1 (IC50 = 7–97 nM), whereas it had little or no effect on the function of other chemokine receptors tested. The inhibition of CCR2 function was both insurmountable and reversible, consistent with a noncompetitive mode of action. JNJ-27141491 blocked the binding of 125I-MCP-1 to human monocytes (IC50 = 0.4 μM), but it failed to affect MCP-1 binding to mouse, rat, and dog cells (IC50 > 10 μM). Therefore, transgenic mice, in which the mouse (m)CCR2 gene was replaced by the human counterpart, were generated for in vivo testing. In these mice, oral administration of JNJ-27141491 dose-dependently [5–40 mg/kg q.d. (once daily) or b.i.d.] inhibited monocyte and neutrophil recruitment to the alveolar space 48 h after intratracheal mMCP-1/lipopolysaccharide instillation. Furthermore, treatment with JNJ-27141491 (20 mg/kg q.d.) significantly delayed the onset and temporarily reduced neurological signs in an experimental autoimmune encephalomyelitis model of multiple sclerosis. Taken together, these results identify JNJ-27141491 as a noncompetitive, functional antagonist of hCCR2, capable of exerting oral anti-inflammatory activity in transgenic hCCR2-expressing mice.
Footnotes
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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.108.140723.
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ABBREVIATIONS: CCR, CC chemokine receptor; MCP, monocyte chemoattractant protein; MS, multiple sclerosis; EAE, experimental autoimmune encephalomyelitis; GTPγS, guanosine 5′-O-(3-thio)triphosphate; h, human; CHO, Chinese hamster ovary; JNJ-27141491, (S)-3-[3,4-difluoro-phenyl)-propyl]-5-isoxazol-5-yl-2-thioxo-2,3-dihydro-1H-imidazole-4-carboxyl acid methyl ester; m, mouse; PBMC, peripheral blood mononuclear cells; DMSO, dimethyl sulfoxide; rAM, resident alveolar macrophages; PBS, phosphate-buffered saline; LPS, lipopolysaccharide; BAL, bronchoalveolar lavage; FSC, forward scatter; FL, fluorescence channel; MOG, myelin oligodendrocyte glycoprotein; MIP, macrophage inflammatory protein; IL, interleukin; q.d., quaque die, once daily; CNS, central nervous system.
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↵ The online version of this article (available at http://jpet.aspetjournals.org) contains supplemental material.
- Received May 2, 2008.
- Accepted July 2, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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