CCR5-specific mucosal IgA in saliva and genital fluids of HIV-exposed seronegative subjects.

CCR5 is a chemokine receptor expressed on blood T lymphocytes and monocyte-macrophages; in the genital tract,1 it works as the main HIV coreceptor.2,3 CCR5 mediates HIV entry following sexual transmission. Many studies have addressed the role of the CCR5 molecule as a putative target to prevent HIV infection. Serum antibodies to CCR5, found in subpopulations of HIVexposed seronegative subjects (ESNs), have been considered to play a role in natural HIV resistance.4,5 We studied samples from 118 HIV-exposed subjects, sexual partners of HIV-seropositive patients (mean age, 34.7; range, 19-51 years). Pools of saliva and genital fluids from 10 healthy blood donors (HDs) were used as negative controls. Immunoglobulinenriched fractions were purified from blood, genital secretions (vaginal and seminal fluids), and saliva samples and tested on a CCR5-transfected cell line. Positive binding was found in serum immunoglobulins (IgG and IgA) from 9 of 118 ESN subjects but in no HDs. All 9 sera contained anti–CCR5 IgG; notably, 8 ESN sera (except ESN no. 34) also contained anti–CCR5 IgA antibodies. One individual (ESN no. 108) only presented serum IgA and serum IgG (Table 1). CCR5 binding of all 8 IgA from positive ESNs were significantly higher than those observed with HD IgA (P .0001). Interestingly, all 6 subjects (5 males and 1 female) presenting anti–CCR5 IgA in genital fluids did also possess anti–CCR5 IgA in saliva. Mucosal anti–CCR5 antibodies were further tested for binding specificity on a panel of 5 synthetic peptides spanning the extracellular loops of CCR5 and a sixth, unrelated, control peptide. Both anti–CCR5 IgG and IgA recognized a 13-mer peptide, corresponding to the second extracellular loop of CCR5 (Table 1). This 90-103 peptide contains a conformational epitope, uniquely recognized by anti–CCR5 antibodies from ESN subjects under native conditions (data not shown). Anti–CCR5 immunoglobulins from each ESN sample and a pool of 10 CCR5-negative immunoglobulins from HDs also were tested in virus entry-neutralization assays, as previously reported.6 Briefly, plasmid (pCAGGS) was used to express membrane-bound envelope (env) of the primary R5 isolate JR-FL. Vesicular stomatitis virus G (VSV-G) was used as a negative control virus. Pseudoviruses (kind gift of J. Binley and D. Burton) were produced by transfection of 293T cells with pNL4-3.Luc.R-E– and Envexpressing pCAGGS-based plasmids. Single-round infections were performed using U87.CD4.CCR5, and luciferase activity was