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Antigenicity and immunogenicity of HIV-1 consensus subtype B envelope glycoproteins - PubMed

  • ️Mon Jan 01 2007

Comparative Study

. 2007 Mar 30;360(1):218-34.

doi: 10.1016/j.virol.2006.10.017. Epub 2006 Nov 13.

Affiliations

Comparative Study

Antigenicity and immunogenicity of HIV-1 consensus subtype B envelope glycoproteins

Denise L Kothe et al. Virology. 2007.

Abstract

"Centralized" (ancestral and consensus) HIV-1 envelope immunogens induce broadly cross-reactive T cell responses in laboratory animals; however, their potential to elicit cross-reactive neutralizing antibodies has not been fully explored. Here, we report the construction of a panel of consensus subtype B (ConB) envelopes and compare their biologic, antigenic, and immunogenic properties to those of two wild-type Env controls from individuals with early and acute HIV-1 infection. Glycoprotein expressed from full-length (gp160), uncleaved (gp160-UNC), truncated (gp145), and N-linked glycosylation site deleted (gp160-201N/S) versions of the ConB env gene were packaged into virions and, except for the fusion defective gp160-UNC, mediated infection via the CCR5 co-receptor. Pseudovirions containing ConB Envs were sensitive to neutralization by patient plasma and monoclonal antibodies, indicating the preservation of neutralizing epitopes found in contemporary subtype B viruses. When used as DNA vaccines in guinea pigs, ConB and wild-type env immunogens induced appreciable binding, but overall only low level neutralizing antibodies. However, all four ConB immunogens were significantly more potent than one wild-type vaccine at eliciting neutralizing antibodies against a panel of tier 1 and tier 2 viruses, and ConB gp145 and gp160 were significantly more potent than both wild-type vaccines at inducing neutralizing antibodies against tier 1 viruses. Thus, consensus subtype B env immunogens appear to be at least as good as, and in some instances better than, wild-type B env immunogens at inducing a neutralizing antibody response, and are amenable to further improvement by specific gene modifications.

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Figures

Fig. 1
Fig. 1

Generation of consensus subtype B (ConB) envelopes. (A) Alignment of the deduced protein sequence of full length (unmodified) ConB Env with those of recently transmitted, contemporary subtype B Env controls (CAAN5342.A2, WITO4160.27). Sequences are compared to ConB gp160, with dots indicating sequence identity, and dashes indicating gaps introduced for optimal alignment. Potential N-linked glycosylation sites are highlighted in red. The locations of signal peptide (SP) and gp120/gp41 cleavage sites are indicated, as are the positions of the variable loops (V1–V5) and the transmembrane (TM) domain. (B) Location of specific mutations within the ConB Env sequence. Amino acid substitutions are highlighted (see text for design and construction details of ConB gp160-201N/S, ConB gp160-UNC and ConB gp145).

Fig. 2
Fig. 2

Processing and virion incorporation of ConB Envs. Consensus (ConB gp160, ConB gp160-UNC, ConB gp145, and ConB gp160-201N/S) and wildtype (CAAN5342.A2 and WITO41260.27) env genes were co-transfected with the HIV-1/SG3Δenv backbone vector. Purified particles were examined by Western blot analysis using antibodies specific for HIV-1 Env (polyclonal anti-gp120; upper panel) and Gag (monoclonal anti-p24; lower panel) proteins (the SG3Δenv backbone vector was included for control). The position of the uncleaved gp160 precursor, the mature gp120 protein, and the p24 Gag protein are indicated.

Fig. 3
Fig. 3

Infectivity of virions containing ConB Envs. Virus stocks were generated by co-transfection of ConB (ConB gp160, ConB gp160-UNC, ConB gp145, ConB gp160-201N/S) and contemporary (CAAN5342.A2, WITO41260.27) env genes with HIV-1/SG3Δenv. Infectivity was determined in JC53-BL cells by determining the number of blue cells (infectious units, IU) per nanogram of p24. Bars indicated standard errors (values are averaged from 3 independent experiments). Infectivity values of virions containing YU-2 and NL4.3 Envs are shown for control.

Fig. 4
Fig. 4

Co-receptor preference of ConB Envs. JC53-BL cells were pretreated with AMD3100 (inhibitor of CXCR4), TAK779 (inhibitor of CCR5), both or neither (media only) prior to addition of virions containing the Env proteins indicated. Virus infectivity is plotted on the vertical axis as a percentage of the untreated control. The CCR5-tropic YU-2 and the CXCR4-tropic NL4.3 Envs were included as controls.

Fig. 5
Fig. 5

Neutralization of ConB Env containing virions by “high reactive” (A) and “low reactive” (B) plasma pools. Neutralization of viral infectivity in JC53-BL cells (y-axis) was scored as the plasma dilution (x-axis) required to reduce virus infectivity by 50% (IC50) (grey line).

Fig. 6
Fig. 6

Neutralization of ConB Env containing virions by patient plasma. Plasma samples from seven subtype B infected individuals were tested for their ability to neutralize ConB or contemporary Env containing virions. Neutralization was scored as the plasma dilution required to reduce virus infectivity by 50% (IC50). Vertical boxes represent the 25th and 75th percentiles of the IC50 values, the line in the box the median, and the lines emerging from the box the highest and lowest serum dilutions observed for the group, respectively.

Fig. 7
Fig. 7

Neutralization of ConB Env containing virions by soluble CD4. Neutralization of viral infectivity in JC53-BL cells (y-axis) was scored as the concentration of sCD4 (x-axis) required to reduce virus infectivity by 50% (IC50) (grey line).

Fig. 8
Fig. 8

Neutralization of ConB Env containing virions by monoclonal antibodies. Neutralization of viral infectivity in JC53-BL cells (y-axis) by monoclonal antibodies IgGb12 (A), 2G12 (B), 2F5 (C), 4E10 (D), 17b (E) and E51 (F) was scored as the antibody concentration (μg/ml; x-axis) required to reduce virus infectivity by 50% (IC50) (grey line).

Fig. 9
Fig. 9

Humoral immune responses to DNA vaccination. Guinea pigs were vaccinated three times at four-week intervals with plasmid DNA containing ConB (ConB gp160, ConB gp160-UNC, ConB gp145, ConB gp160-201N/S) or contemporary env genes (CAAN5342.A2-opt, WITO4160.27-opt). Two-weeks following the third vaccination, sera were assayed for the presence of binding antibodies to their respective (cognate) gp120 proteins. Sera were serially diluted and the last dilution giving absorbance values greater than twice the optical density (OD) value of the negative control was identified as the endpoint titer. Vertical boxes indicate the mean endpoint titer ± SEM for each of the groups indicated.

Fig. 10
Fig. 10

Relative potency of ConB and wildtype env vaccines as determined by comparison of neutralization responses against the entire panel of viruses (A), tier 1 viruses (B), and tier 2 viruses (C), as indicated in Table 1. Neutralization data were analyzed by performing multiple pairwise comparisons (Tukey’s Honestly Significant Difference procedure). An estimated difference between neutralization coefficients (solid circles) as well as a 95% confidence interval for that difference (brackets) are shown for all pairwise comparisons of immunogens (as indicated on the left). A negative difference (circle to the left of the vertical dotted line) indicates the first named immunogen of the pair was better at eliciting a neutralizing antibody response than the second named immunogen. Pairwise comparison differences with 95% confidence intervals that do not cross the dotted vertical line indicate statistically significant differences in potency.

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