Identification of full, partial and inverse CC chemokine receptor 3 agonists using [35S]GTPgammaS binding

Eur J Pharmacol. 2002 Dec 5;456(1-3):1-10. doi: 10.1016/s0014-2999(02)02621-3.


Study of the CC chemokine receptor 3 (CCR3) has been limited to using radiolabeled agonist chemokines. A small molecule CCR3 antagonist, 2-[(6-amino-2-benzothiazolyl)thio]-N-[1-[(3,4-dichlorylphenyl)methyl]-4-piperidinyl]acetamide, Banyu (I), was tritiated and used for pharmacological studies. Banyu (I) has a K(d) of 5.0+/-0.4 and 4.3+/-1.8 nM on human CCR3 transfectants and eosinophils, and noncompetitively inhibits [125I]eotaxin binding and eotaxin-induced [35S]guanosine-5'-O-(3-thiotriphosphate) ([35S]GTPgammaS) binding. The proportion of [125I]eotaxin: [3H]Banyu (I) binding sites in eosinophils or transfectants was 35% or 13%, although both binding sites were overexpressed in transfectants. CCR3 spontaneously couples to G-proteins in CCR3 transfectants, demonstrated by changes in basal and eotaxin-induced [35S]GTPgammaS binding under reduced NaCl and GDP concentrations. Consequently, Banyu (I) was identified as an inverse agonist. In contrast, CCL18 and I-TAC (interferon-inducible T cell alpha-chemoattractant) were neutral antagonists, inhibiting eotaxin-induced [35S]GTPgammaS binding, with minimal effect on basal coupling of CCR3 to G proteins. Eotaxin, eotaxin-2 and monocyte chemoattractant protein (MCP)-4 are full agonists inducing [35S]GTPgammaS binding; eotaxin-3, MCP-3, RANTES (regulated on activation normal T cell expressed and secreted), vMIP-I (Kaposi's sarcoma-associated herpesvirus macrophage inflammatory protein-) and vMIP-II are partial agonists, indicating that this is a sensitive method to quantitate agonist efficacy.

MeSH terms

  • Animals
  • Binding, Competitive / drug effects
  • CHO Cells
  • Cell Line
  • Cell Membrane / drug effects
  • Cell Membrane / metabolism
  • Cells, Cultured
  • Chemokine CCL11
  • Chemokine CCL5 / metabolism
  • Chemokine CCL5 / pharmacology
  • Chemokines / metabolism
  • Chemokines / pharmacology
  • Chemokines, CC / metabolism
  • Chemokines, CC / pharmacology
  • Cricetinae
  • Dose-Response Relationship, Drug
  • Eosinophils / cytology
  • Eosinophils / drug effects
  • Eosinophils / metabolism
  • Guanosine 5'-O-(3-Thiotriphosphate) / metabolism*
  • Humans
  • Monocyte Chemoattractant Proteins / metabolism
  • Monocyte Chemoattractant Proteins / pharmacology
  • Radioligand Assay
  • Rats
  • Receptors, CCR3
  • Receptors, Chemokine / agonists*
  • Receptors, Chemokine / genetics
  • Receptors, Chemokine / metabolism
  • Sulfur Radioisotopes
  • Transfection


  • CCL11 protein, human
  • CCL13 protein, human
  • CCR3 protein, human
  • Ccl11 protein, rat
  • Ccr3 protein, rat
  • Chemokine CCL11
  • Chemokine CCL5
  • Chemokines
  • Chemokines, CC
  • Monocyte Chemoattractant Proteins
  • Receptors, CCR3
  • Receptors, Chemokine
  • Sulfur Radioisotopes
  • Guanosine 5'-O-(3-Thiotriphosphate)