title: Specificity for a CCR5 Inhibitor Is Conferred by a Single Amino Acid Residue: ROLE OF ILE198.
abstract: The chemokine receptors CCR5 and CCR2b share 89% amino acid homology. CCR5 is a co-receptor for HIV and CCR5 antagonists have been investigated as inhibitors of HIV infection. We describe the use of two CCR5 antagonists, Schering-C (SCH-C), which is specific for CCR5, and TAK-779, a dual inhibitor of CCR5 and CCR2b, to probe the CCR5 inhibitor binding site using CCR5/CCR2b chimeric receptors. Compound inhibition in the different chimeras was assessed by inhibition of chemokine-induced calcium flux. SCH-C inhibited RANTES (regulated on activation, normal T cell expressed and secreted) (CCL5)-mediated calcium flux on CCR5 with an IC50 of 22.8 nM but was inactive against monocyte chemoattractant protein-1 (CCL2)-mediated calcium flux on CCR2b. However, SCH-C inhibited CCL2-induced calcium flux against a CCR5/CCR2b chimera consisting of transmembrane domains IV-VI of CCR5 with an IC50 of 55 nM. A sequence comparison of CCR5 and CCR2b identified a divergent amino acid sequence located at the junction of transmembrane domain V and second extracellular loop. Transfer of the CCR5 sequence KNFQTLKIV into CCR2b conferred SCH-C inhibition (IC50 of 122 nM) into the predominantly CCR2b chimera. Furthermore, a single substitution, R206I, conferred partial but significant inhibition (IC50 of 1023 nM) by SCH-C. These results show that a limited amino acid sequence is responsible for SCH-C specificity to CCR5, and we propose a model showing the interaction with CCR5 Ile(198).
authors.affiliation: From Anormed Inc., Langley, British Columbia V2Y 1N5, Canada.
authors.forename: Gloria
authors.initials: G
authors.lastname: Lau
authors.affiliation: From Anormed Inc., Langley, British Columbia V2Y 1N5, Canada.
authors.forename: Jean
authors.initials: J
authors.lastname: Labrecque
authors.affiliation: Lead Generation to Candidate Realization, Sanofi, Waltham, Massachusetts 02451, and.
authors.forename: Markus
authors.initials: M
authors.lastname: Metz
authors.affiliation: Lead Generation to Candidate Realization, Sanofi, Waltham, Massachusetts 02451, and.
authors.forename: Roy
authors.initials: R
authors.lastname: Vaz
authors.affiliation: Sanofi-Genzyme Research and Development Center, Framingham, Massachusetts 01701 simonpfricker@gmail.com.
authors.forename: Simon P
authors.initials: SP
authors.lastname: Fricker
mesh_terms: D000577:Amides / Q000737:chemistry*; D000595:Amino Acid Sequence; D000818:Animals; D065100:CCR5 Receptor Antagonists / Q000737:chemistry*; D020013:Calcium Signaling; D057809:HEK293 Cells; D006801:Humans; D007532:Isoleucine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008251:Macaca; D008958:Models, Molecular; D017434:Protein Structure, Tertiary; D000644:Quaternary Ammonium Compounds / Q000737:chemistry*; D054390:Receptors, CCR2 / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019713:Receptors, CCR5 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D000577:Amides; D065100:CCR5 Receptor Antagonists; C584933:CCR5 protein, human; D000644:Quaternary Ammonium Compounds; D054390:Receptors, CCR2; D019713:Receptors, CCR5; D011993:Recombinant Fusion Proteins; D007532:Isoleucine; C119369:TAK 779
keywords: CC Chemokine Receptor Type 2b (CCR2b); CC Chemokine Receptor Type 5 (CCR5); Drug Design; Human Immunodeficiency Virus (HIV); Molecular Modeling; Site-directed Mutagenesis