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. 2003 May;77(10):5589-97.
doi: 10.1128/jvi.77.10.5589-5597.2003.

Intranasal vaccination using interleukin-12 and cholera toxin subunit B as adjuvants to enhance mucosal and systemic immunity to human immunodeficiency virus type 1 glycoproteins

Diana I Albu  1 , Agnes Jones-Trower, Amy M Woron, Kathleen Stellrecht, Christopher C Broder, Dennis W Metzger
Affiliations

Intranasal vaccination using interleukin-12 and cholera toxin subunit B as adjuvants to enhance mucosal and systemic immunity to human immunodeficiency virus type 1 glycoproteins

Diana I Albu et al. J Virol. 2003 May.

Abstract

We have investigated the induction of protective mucosal immunity to human immunodeficiency virus type 1 (HIV-1) isolate 89.6 by intranasal (i.n.) immunization of mice with gp120 and gp140 together with interleukin-12 (IL-12) and cholera toxin subunit B (CTB) as adjuvants. It was found that both IL-12 and CTB were required to elicit mucosal antibody responses and that i.n. immunization resulted in increased total, immunoglobulin G1 (IgG1), and IgG2a anti-HIV-1 antibody levels in serum; increased total, IgG1, IgG2a, and IgA antibody expression in bronchoalveolar lavage fluids; and increased IgA antibody levels in vaginal washes. Levels of anti-HIV-1 antibodies in both sera and secretions were higher in groups immunized with gp140 than in those immunized with gp120. However, only gp120-specific mucosal antibodies demonstrated neutralizing activity against HIV-1 89.6. Taken together, the results show that IL-12 and CTB act synergistically to enhance both systemic and local mucosal antibody responses to HIV-1 glycoproteins and that even though gp140 induces higher antibody titers than gp120, only gp120-specific mucosal antibodies interfere with virus infectivity.

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Figures

FIG. 1.
FIG. 1.
SDS-polyacrylamide gel electrophoresis analysis of purified gp120 and gp140 glycoproteins. Proteins were electrophoresed on an SDS-5% polyacrylamide gel under reducing conditions, and the gel was stained with Coomassie blue. Lane 1, standard protein markers; lanes 2 and 3, gp120 (10 μg each); lanes 4 and 5, gp140 (5 μg each).
FIG. 2.
FIG. 2.
Levels of serum gp120-specific antibodies in i.n. vaccinated mice. Sera were obtained from BALB/c mice 35 days after i.n. immunization with 5 μg of vaccine with or without 1 μg of IL-12 and 10 μg of CTB. Titers for individual mice are indicated by the symbols, and the average titer for each group is displayed as a line. Titers are expressed as the reciprocal serum dilutions corresponding to 25% maximal binding on the titration curve. Significant differences in antigen-specific antibody production were detected between the group immunized with gp120 plus IL-12 and CTB and all of the other groups (∗∗, P < 0.005). Pooled data from two experiments are shown.
FIG. 3.
FIG. 3.
Levels of gp120-specific antibodies in BAL fluids. BAL fluids were obtained from BALB/c mice 35 days after i.n. immunization with 5 μg of vaccine with or without 1 μg of IL-12 and 10 μg of CTB. Individual OD values are indicated by the symbols, and the average OD value for each group is displayed as a line. OD readings are the maximal values obtained from undiluted BAL fluid samples. Significant differences (∗∗, P < 0.005) in IgA production were detected between the group immunized with gp120 plus IL-12 and CTB and that immunized with gp120 as well as between the group immunized with gp120 plus IL-12 and CTB and that immunized with gp120 plus IL-12. Significant differences (∗∗, P < 0.005) in IgG1 and IgG2a production were detected between the group immunized with gp120 plus IL-12 and CTB and all of the other groups. Pooled data from two experiments are shown.
FIG. 4.
FIG. 4.
Levels of gp120-specific antibodies in vaginal washes. Vaginal washes were obtained from BALB/c mice 35 days after i.n. immunization with 5 μg of vaccine with or without 1 μg of IL-12 and 10 μg of CTB. Individual OD values are indicated by the symbols, and the average OD value for each group is displayed as a line. The values represent ODs obtained with 1:10 dilutions of vaginal wash samples. Significant differences (∗, P < 0.05) in IgA antibody production were detected between the group immunized with gp120 plus IL-12 and CTB and that immunized with gp120 as well as between the group immunized with gp120 plus IL-12 and CTB and that immunized with gp120 plus CTB. Significant differences in IgG1 antibody production were detected between the group immunized with gp120 plus IL-12 and CTB and that immunized with gp120 group (∗∗, P < 0.005) and between the group immunized with gp120 plus IL-12 and CTB and that immunized with gp120 plus IL-12 (∗, P < 0.05). Pooled data from two experiments are shown.
FIG. 5.
FIG. 5.
Comparison of gp120 and gp140 as mucosal immunogens for systemic immunity. Sera were obtained from BALB/c mice 35 days after i.n. immunization with 1 or 5 μg of gp120 or gp140 plus 1 μg of IL-12 and 10 μg of CTB. Titers for individual mice are indicated by the symbols, and the average titer for each group is displayed as a line. Titer values are expressed as the reciprocal serum dilutions corresponding to 25% maximal binding on the titration curve. The results show enhanced gp140-specific total, IgG1, and IgG2a antibodies in the sera of immune mice compared with gp120-specific antibody at both doses of antigens. For IgG2a antibody the difference is significant (∗, P < 0.05) at the 5-μg dose.
FIG. 6.
FIG. 6.
Comparison of gp120 and gp140 as mucosal immunogens for induction of lung antibodies. BAL fluids were obtained from BALB/c mice 35 days after i.n. immunization with 1 or 5 μg of gp120 or gp140 plus 1 μg of IL-12 and 10 μg of CTB. Titers for individual mice are indicated by the symbols, and the average titer for each group is displayed as a line. Titers are expressed as the reciprocal BAL dilutions corresponding to 25% maximal binding on the titration curve. The results show enhanced gp140-specific IgA (∗∗, P < 0.005), IgG1 (∗, P < 0.05), and IgG2a (∗, P < 0.05) antibodies in BAL fluids of immune mice compared with gp120-specific antibody at both doses of antigen.
FIG. 7.
FIG. 7.
Comparison of gp120 and gp140 as mucosal immunogens for induction of antibodies in the genital compartment. Vaginal washes were obtained from BALB/c mice 35 days after i.n. immunization with 1 or 5 μg of gp120 or gp140 plus 1 μg of IL-12 and 10 μg of CTB. Titers for individual mice are indicated by the symbols, and the average titer for each group is displayed as a line. Titers are expressed as the reciprocal vaginal wash dilutions corresponding to 25% maximal binding on the titration curve. The results show that induction of gp140-specific mucosal IgA in vaginal washes is significantly greater than induction of gp120-specific IgA at 5 μg of antigen (∗∗, P < 0.005).
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