IgG binding antibody responses to gp120 A244gD and gp70V1V2 scaffolds in cervico-vaginal mucus (CVM).

<p>Reciprocal titers of IgG binding antibody responses to (A) gp120 A244gD (CRF01_AE), (B) gp70V1V2 92TH023 (CRF01_AE) and (C) gp70V1V2 Case A2 (subtype B) in CVM are shown along with numeric depiction of geometric mean titers above panels. Each group is color coded; red, ALVAC-HIV/AIDSVAX^®B/E (ALVAC/AIDSVAX); green, AIDSVAX^®B/E (AIDSVAX); blue, ALVAC-HIV (ALVAC). Error bars depict 95% confidence intervals. The cut-off level of responses (0.5-fold the reciprocal titers of initial dilution of specimens) is shown by the dotted line. RV305 vaccine administration time points are indicated by black arrows (weeks 0 and 24). The non-parametric Mann-Whitney U Test was used to assess within-group comparison of IgG responses between time points indicated by horizontal black bars. Comparisons reaching statistical significance at the level of p<0.05 are shown. *p<0.05 to 0.001, ^&p<0.001.</p

Correlation of IgG responses in anogenital secretions and plasma.

<p>Spearman’s rank correlations of IgG responses for gp120 A244gD (CRF01_AE), gp70V1V2 92TH023 (CRF01_AE) and gp70V1 V2 Case A2 (subtype B) at weeks 2, 26, 48 and 72 in matched CVM (A-C), SP (D-E) and RS (F), and plasma of RV305 vaccine recipients are shown. Each group is color coded; red, ALVAC-HIV/AIDSVAX^®B/E; green, AIDSVAX^®B/E; blue, ALVAC-HIV. Numeric values above each plot depict r- and p-values. Significant p-value <0.05, GMT = Geometric Mean Titer.</p

IgG binding antibody responses to gp120 A244gD and gp70V1V2 scaffolds in seminal plasma (SP).

<p>Reciprocal titers of IgG binding antibody responses to (A) gp120 A244gD (CRF01_AE), (B) gp70V1V2 92TH023 (CRF01_AE) and (C) gp70V1V2 Case A2 (subtype B) in SP are shown along with numeric depiction of geometric mean titers above panels. Each group is color coded; red, ALVAC-HIV/AIDSVAX^®B/E (ALVAC/AIDSVAX); green, AIDSVAX^®B/E (AIDSVAX); blue, ALVAC-HIV (ALVAC). Error bars depict 95% confidence intervals. The cut-off level of responses (0.5-fold the reciprocal titers of the initial dilution of specimens) is shown by the dotted line. RV305 vaccine administration time points are indicated by black arrows (weeks 0 and 24). The non-parametric Mann-Whitney U Test was used to assess within-group comparison of IgG responses between time points indicated by black bars. Comparisons reaching statistical significance at the level of p<0.05 are shown. *p<0.05 to 0.001, ^&p<0.001.</p

Concentration of total IgG and IgA in anogenital secretions that did not contain blood.

<p>Concentration of total IgG and IgA in anogenital secretions that did not contain blood.</p

Percent of positive IgG responders to all HIV antigen tested in anogenital secretions.

<p>Percent of positive IgG responders to gp120 A244gD (CRF01_AE), gp70V1V2 92TH023 (CRF01_AE) and gp70V1V2 Case A2 (subtype B) in CVM (A-C), SP (D-F), and RS (G-I) are shown. Each time point is color coded; red, week 0; orange, week 2; green, week 26; blue, week 48; magenta, week 72. CVM and RS were not collected from placebo recipient of groups AIDSVAX^®B/E and ALVAC-HIV, respectively, at any time point. VAC = vaccine recipients; PLB = placebo recipients; ALVAC/AIDSVAX = ALVAC-HIV/AIDSVAX^®B/E group; AIDSVAX = AIDSVAX^®B/E group; ALVAC = ALVAC-HIV group.</p

Specimen collection algorithm.

<p>Blood specimens were collected from each study participant to obtain plasma. Cervico-vaginal mucus (CVM) was collected from consenting female participants. Seminal plasma (SP) and rectal secretions (RS) were collected from consenting male participants. Blood contamination was tested on both CVM and RS using Hemoccult® SENSA® test kit (Beckmann Coulter, Brea, CA). All blood contaminated specimens were excluded from the analysis. Blood contamination was not tested on SP.</p

Boosting of RV144 vaccinees increased V_H chain gene mutation frequency and caused a repertoire shift, increasing the frequency of antibodies with Heavy Chain Complementary Determinant Region 3 (HCDR3) ≥ 22 amino acids.

<p>PBMCs from four vaccinees post-RV144 (A&B) and post-RV305 (C&D) were antigen-specific single-cell sorted with fluorophore labeled conjugates. The V_H/V_L chain genes were PCR-amplified and screened for Env-reactivity by ELISA. The V_H chain gene mutation frequency and HCDR3 lengths of 145 Env-reactive antibodies from RV135/144 and 242 Env-reactive antibodies from RV305 were analyzed with Cloanalyst[<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006182#ppat.1006182.ref056" target="_blank">56</a>]. (A&C) Error bars represent the standard error of the mean.</p

Negative stained electron microscopy of DH576 in complex with CH505 SOSIP.664.

<p>(A) 2D class averages of one, two, or three DH576 Fabs bound to the trimer and (B) Side and top views of the 3D EM reconstruction with the indicated Ab or sCD4 superimposed on the gp120 subunit for comparison with DH576 binding. Arrows indicate the angle at which the indicated antibody approaches the trimer.</p

The long Heavy Chain Determinant Region 3 (HCDR3) antibodies that neutralize virus bind the CD4 binding site (CD4 bs).

<p>Purified recombinant monoclonal antibodies (mAbs) were assayed by ELISA for (A) blocking the binding of soluble CD4 to AE.A244gp120, and (B) sensitivity to the CD4 binding site mutations Δ371I/P363N, D368R, N276A, and T278A in the AE.A244gp120 protein. (C) Epitope mapping of mAbs on yeast displayed YU2gp120 with point mutations within the inner domain, outer domain and CD4 binding site. (D) Assaying the long HCDR3 CD4 bs antibodies for HIV-1 neutralization in the TZM-bl neutralization assay.</p