A cyclic dodecapeptide-multiple-antigen peptide conjugate from the undecapeptidyl arch (from Arg(168) to Cys(178)) of extracellular loop 2 in CCR5 asa novel human immunodeficiency virus type 1 vaccine

A cyclic closed-chain dodecapeptide (cDDR5) mimicking the conformation-spec ific domain of CCR5 was prepared in which Gly-Asp, as a dipeptide forming a spacer arm, links the amino and carboxyl termini of the decapeptidyl linea r chain (Arg(168) to Thr(177)) derived from the undecapeptidyl arch (UPA; A rg(168) to Cys(178)) of extracellular loop 2 (ECL2) in CCR5. Novel monoclon al antibodies were raised against cDDR5 conjugated with a multiple-antigen peptide (cDDR5-MAP), and the purified antibody [KB8C12, immunoglobulin M(ka ppa)] reacted with cDDR5, but not with linear DDR5, in real-time biomolecul ar interaction analysis using surface plasmon resonance. The antibody also reacted with cells expressing CCR5, but not with cells expressing CXCR4, an d the immunoreaction was competed by cDDR5-MAP. The antibody significantly interfered with chemotaxis induced by macrophage inflammatory protein, 1 be ta, and at a concentration of 1.67 nM it almost completely inhibited infect ion by human immunodeficiency virus type 1 (HIV-1) R5, but not by HIV-1 X4, as observed by use of a new phenotypic assay for drug susceptibility of HI V-1 using the CCR5-expressing HeLa CD4(+) cell clone 1-10 (MAGIC-5). Furthe rmore, cDDR5-MAP suppressed infection by HIV-1 R5 at relatively high concen trations (50 to 400 muM) in a dose-dependent manner but did not suppress in fection by HIV-1 X4. Taken together, these results indicate that the antibo dy is conformation specific and recognizes the conformation-specific domain of the UPA of ECL2. Moreover, both the antibody and its immunogen, the cDD R5-MAP conjugate, may be useful in developing a new candidate vaccine for H IV therapy.