Comparison of Antibody Responses Induced by RV144, VAX003, and VAX004 Vaccination Regimens

Abstract

The RV144 prime-boost regimen demonstrated efficacy against HIV acquisition while VAX003 and VAX004 did not. Although these trials differed by risk groups, immunization regimens, and immunogens, antibody responses may have contributed to the differences observed in vaccine efficacy. We assessed HIV-specific IgG, both total and subclass, and IgA binding to HIV envelope (Env): gp120 proteins and Cyclic V2 (CycV2) and CycV3 peptides and gp70 V1 V2 scaffolds in these 3 HIV vaccine trials. After two protein immunizations, IgG responses to 92TH023 gp120 (contained in ALVAC-HIV vaccine) were significantly higher in RV144 but responses to other Env were higher in the VAX trials lacking ALVAC-HIV. IgG responses declined significantly between vaccinations. All trials induced antibodies to gp70 V1 V2 but VAX004 responses to 92TH023 gp70 V1 V2 were weak. All CycV2 responses were undetectable in VAX004 while 92TH023 gp70 V1 V2 was detected in both RV144 and VAX003 but MN CycV2 was detected only in VAX003. Multiple protein vaccinations in VAX trials did not improve magnitude or durability of V1 V2 and CycV2 antibodies. Herpes simplex virus glycoprotein D (gD) peptide at the N terminus of AIDSVAX^® B/E and B/B gp120 proteins induced antibodies in all trials, although significantly higher in VAX trials. gD peptide induced IgA, IgG1, IgG2, and IgG3 but not IgG4. Multiple protein vaccinations decreased IgG3 and increased IgG4 changing subclass contribution to total IgG. Although confounded by different modes of HIV transmission, higher Env-specific IgA and IgG4 binding antibodies induced in the VAX trials compared to RV144 raises the hypothesis that these differences may have contributed to different vaccine efficacy results.

Keywords: HIV vaccines; HSV; IgG; IgG subclasses IgA; RV144; V1 V2; V2; V3; VAX003; VAX004; antibodies; gp120.

FIG. 1.

RV144, VAX003, and VAX004 vaccination schedules and HIV-1 gp120 envelope variable loops 2 and 3 (V2 and V3). (A) RV144, VAX003 and VAX004 vaccination schedules and vaccine antigens; A = ALVAC-HIV (vCP1521), B/B = AIDSVAX^® B/B (gp120 MNgD and gp120 GNE8gD–600 μg), B/E = AIDSVAX B/E (gp120 MNgD and gp120 A244gD–600 μg). Visit samples used in study are in bold. In VAX trials immunizations are given at even numbers and odd numbers represent 2 weeks post immunization. (B) Alignments of HIV-1 gp120 envelope variable loops 2 and 3 (V2 and V3); V2 and V3 loops are shown in the top and bottom panel, respectively. Boxed amino acids show differences from reference strain HxB2. Numbers on top of each panel are positions of amino acids based on the HxB2 strain. (C) Percent identity and divergence of gp120 proteins and variable loops 2 and 3 (V2 and V3). gD, glycoprotein D.

FIG. 2.

IgG antibody binding to rgp120 envelope protein CRF01_AE A244gD, MNgD, and 92TH023 in RV144 and VAX trials. RV144 (red symbols, visits 1 and 8), VAX003 (blue symbols, visits A02, A05, and A09) and VAX004 (green symbols, visits A02, A05, and A09); symbols show GMTs of (A) A244gD, (B) MNgD, (C) 92TH023 with 95% confidence intervals and color coded numbers depict actual GMT and (D) RV144 (red symbols, visits 1, 8, and 9), VAX003 (blue symbols, visits A02-A13), and VAX004 (green symbols, visits A02-A13); solid symbols show 2 weeks post vaccination and hollow symbols show vaccination visits. Symbols show means of A244gD and bars are standard deviations at A405nm.VAX visit numbers are indicated below the x-axis with vaccination visits underlined. Responses above dotted line are considered positive. *p < .05; **p < .01; ***p < .001; ****p < .0001; ns, not significant. GMT, geometric mean titer; rgp120, recombinant gp120 proteins.

FIG. 3.

IgG antibodies to the gD peptide from RV144 and VAX samples. RV144 (red symbols), VAX003 (blue symbols), and VAX004 (green symbols) at 2 weeks post second AIDSVAX vaccination from visits 8 and A05, respectively. (A) Symbols are GMTs showing absorption of IgG antibodies binding to the gD peptide with 95% confidence intervals. Numbers indicate GMTs before (solid symbols) or after absorption (hollow symbols) of the gD binding antibodies. (B) IgG specificity to gD after multiple immunizations in VAX004 serum samples from second (visit A04), fourth (visit A08), and sixth (visit A12) AIDSVAX B/B vaccinations. Solid symbols show sera before absorption and hollow symbols show absorbed sera with the gD peptide. Recombinant gp120 CRF01_AE A244gD was the capturing antigen. The plots are means with standard deviations at absorbance 405 nm. x-axis shows visits. Numbers under x-axis show percentage of positive samples (A405 ≥ 0.25) and responses above dotted line are considered positive. **p < .01; ***p < .001; ****p < .0001.

FIG. 4.

IgG antibody binding to gp70 V1 V2 scaffold in RV144 and VAX trials. RV144 (red symbols), VAX003 (blue symbols), and VAX004 (green symbols); (A) and (B) symbols show GMTs of gp70 V1 V2 92TH023 and gp70 V1 V2 Case A2, respectively, with 95% confidence intervals and color coded numbers depict actual GMTs of RV144 (visits 1 and 8), VAX003 (visits A02, A05, and A09), and VAX004 (visits A02, A05, and A09); (C) and (D) show means and bars are standard deviations of RV144 (visits 1, 8, and 9), VAX003 (visits A02-A13), and VAX004 (visits A02-A13) at A405nm of gp70 V1 V2 92TH023 and gp70 V1 V2 Case A2, respectively. Solid symbols show 2 weeks post vaccination and hollow symbols show vaccination visits. VAX visit numbers are indicated below the x-axis with vaccination visits underlined. Responses above dotted line are considered positive. *p < .05; ***p < .001; ****p < .0001; ns, not significant.

FIG. 5.

IgG antibody responses to Cyclic V2 in RV144 and VAX trials. RV144 (red symbols) visits 1 and 8, VAX003 (blue symbols), and VAX004 (green symbols) visits A02, A05, and A09. Color coded numbers are the actual GMTs and symbols are GMTs of IgG binding to (A) cyclic V2 92TH023 and (B) cyclic V2 MN with 95% confidence intervals. Responses above dotted line are considered positive. **p < .01; ***p < .001; ****p < .0001.

FIG. 6.

Comparison of IgG subtypes binding to rgp120 HIV-1 CRF01_AE A244gD protein and gD peptide. (A) IgG subtype (1–4) binding to rgp120 A244gD; RV144 (red symbol) samples at 2-weeks post second (week 26, visit 8), VAX003 (blue symbol) and VAX004 (green symbol) samples at 2-weeks post fourth (week 50, visit A09) vaccination. Color coded numbers in the plot area show GMTs. Numbers under x-axis show percentage of positive samples (A405 ≥ 0.25) and responses above dotted line are considered positive. Symbols are GMTs with 95% confidence intervals. ***p < .001; ****p < .0001. (B) IgG3 antibodies binding to rgp120 A244gD; VAX003 (blue symbols) samples at 2-weeks post second (week 6, visit A05), fourth (week 50, visit A09) and fifth (week 74, visit A11), and RV144 at 2-weeks post second (week 26, visit 8) AIDSVAX B/E vaccination. Numbers under x-axis show percentage of positive samples (A405 ≥ 0.25) and responses above dotted line are considered positive. Symbols are means and bars indicate standard deviations. **p < .01; ***p < .001; ****p < .0001. (C) Samples were preincubated with HSV-1 gD peptide before adding to ELISA plates with gD peptide as the capturing antigen and compared to gD unabsorbed plasma. Antibody responses with A405 ≥ 0.25 (2.5 times the background, wells without capturing antigen) were considered positive. Solid colors: IgG1, checkered columns: IgG2, and stripped columns: IgG3. RV144 samples in red at 2-weeks post second injection (visit 8), VAX003 and VAX004 samples, are in blue and green respectively, at 2-weeks post fourth (visit A09) AIDSVAX vaccinations. HSV-1, herpes simplex virus 1.

FIG. 7.

IgG1 and IgG3 antibody responses to HIV-1 gp70 V1 V2 scaffolds in RV144 and VAX samples. Antibody binding to gp70 V1 V2 92TH023 (A) and Case A2 (B) scaffolds in RV144 plasma at 2-weeks post second (visit 8), VAX003 and VAX004 sera at 2-weeks post fourth (visit A09) AIDSVAX B/E vaccination. Hollow symbols show IgG1, solid symbols show IgG3, and color coded numbers show GMT. Numbers below the x-axis show percentage of positive samples (A405 ≥ 0.25) and responses above the dotted line are considered positive. Symbols are GMT with 95% confidence intervals. **p < .01; ***p < .001; ****p < .0001.

FIG. 8.

IgG antibody responses in RV144 and VAX samples to CycV3 92TH023 and MN peptides. CycV3 92TH023 peptide binding is shown on the left panel and CycV3 MN is on the right. Samples were tested at base line (hollow symbols), 2 weeks post second protein boost (solid symbols) and 2 weeks post forth protein boost (cross symbols). RV144 is in red, VAX003 in blue, and VAX004 in green. Numbers under x-axis show percentage of positive samples (A405 ≥ 0.25) and responses above dotted line are considered positive. Symbols are GMTs with 95% confidence intervals and actual GMT values are shown on the panels. *p < .05; ***p < .001; ****p < .0001.

FIG. 9.

Comparison of IgA antibody binding to rgp120 HIV-1 CRF01_AE A244gD in RV144 and VAX samples. Samples were from RV144 (red symbols) at 2 weeks (V8) post second protein boost; VAX003 (blue symbols) at 2 weeks (A05) and post second, and 2 weeks (A09) post fourth protein boost; and VAX004 (green symbols) at 2 weeks (A05) post second, and 2 weeks (A09) post fourth protein boost. Responses above dotted line are considered positive. Symbols are means with 95% confidence intervals. **p < .01; ***p < .001; ****p < .0001.