title: Pharmacological profile of a bifunctional ligand of the formyl peptide receptor1 fused to the myc epitope.
abstract: In human peripheral blood neutrophils or in myeloid PLB-985 cells differentiated towards a neutrophil-like phenotype, the peptide N-formyl-L-norleucyl-L-leucyl-L-phenylalanyl-L-norleucyl-L-tyrosyl-L-leucyl-fluorescein isothiocyanate (f-Nle-Leu-Phe-Nle-Tyr-Lys-FITC) binds to and activates formyl peptide receptor1 (FPR1) and is submitted to receptor-mediated endocytosis (microscopy, cytofluorometry). This peptide may be considered a C-terminally extended version of f-Met-Leu-Phe which carries a fluorescent cargo into cells. By analogy to other peptide hormones for which we have evaluated epitope-tagged agonists as carriers of antibody cargoes, we have designed and evaluated f-Nle-Leu-Phe-Nle-Tyr-Lys-myc, C-terminally extended with the 10-residue myc tag. This peptide is as potent as f-Met-Leu-Phe to compete for f-Nle-Leu-Phe-Nle-Tyr-Lys-FITC uptake by PLB-985 cells, but did not mediate (10-1000nM) the internalization of the fluorescent anti-myc monoclonal antibody 4A6 added to the extracellular fluid at ~7nM (microscopy). The nonfluorescent version of the antibody (28nM) acts as a pre-receptor antagonist of f-Nle-Leu-Phe-Nle-Tyr-Lys-myc, but not of f-Met-Leu-Phe (superoxide release assay in differentiated PLB-985 cells). A further prolonged analog, f-Nle-Leu-Phe-Nle-Tyr-Lys-(Asn-Gly)5-myc, designed to decrease the possible steric hindrance between FPR1 and the bound anti-myc antibody, has little affinity for the receptor, precluding a direct assessment of this issue. Thus, the relatively low-affinity anti-myc antibody used at a high concentration functionally behaves as a selective pre-receptor antagonist of the agonist f-Nle-Leu-Phe-Nle-Tyr-Lys-myc.
authors: Centre de Recherche en Rhumatologie et Immunologie, Centre Hospitalier Universitaire de Québec, Québec, QC, Canada, G1V 4G2.
authors: Xavier
authors: X
authors: Charest-Morin
authors: Centre de Recherche en Rhumatologie et Immunologie, Centre Hospitalier Universitaire de Québec, Québec, QC, Canada, G1V 4G2.
authors: Caroline
authors: C
authors: Roy
authors: Centre de Recherche en Rhumatologie et Immunologie, Centre Hospitalier Universitaire de Québec, Québec, QC, Canada, G1V 4G2.
authors: Maria J G
authors: MJ
authors: Fernandes
authors: Centre de Recherche en Rhumatologie et Immunologie, Centre Hospitalier Universitaire de Québec, Québec, QC, Canada, G1V 4G2. Electronic address: francois.marceau@crchul.ulaval.ca.
authors: François
authors: F
authors: Marceau
mesh_terms: D000911:Antibodies, Monoclonal / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D004705:Endocytosis / Q000235:genetics; D000939:Epitopes / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D008024:Ligands; D022423:Myeloid Cells / Q000276:immunology*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000031:analogs & derivatives / Q000378:metabolism*; D009504:Neutrophils / Q000276:immunology*; D011485:Protein Binding; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D044042:Receptors, Formyl Peptide / Q000819:agonists*; D011993:Recombinant Fusion Proteins / Q000235:genetics; D013237:Stereoisomerism
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; C469986:FPR1 protein, human; D008024:Ligands; C489427:MYC protein, human; D016271:Proto-Oncogene Proteins c-myc; D044042:Receptors, Formyl Peptide; D011993:Recombinant Fusion Proteins; D009240:N-Formylmethionine Leucyl-Phenylalanine
keywords: Formylated peptide; Formylated peptide receptor(1); Neutrophil; Receptor-mediated endocytosis