Construction and immunogenicity of replication-competent adenovirus 5 host range mutant recombinants expressing HIV-1 gp160 of SF162 and TV1 strains

Abstract

An HIV Env immunogen capable of eliciting broad immunity is critical for a successful vaccine. We constructed and characterized adenovirus 5 host range mutant (Ad5hr) recombinants encoding HIV(SF162) gp160 (subtype B) and HIV(TV1) gp160 (subtype C). Immunization of mice with one or both induced cellular immunity to subtype B and C peptides by ELISpot, and antibody responses with high binding titers to HIV Env of subtypes A, B, C, and E. Notably, Ad5hr-HIV(TV1) gp160 induced better cellular immunity than Ad5hr-HIV(SF162) gp160, either alone or following co-administration. Thus, the TV1 Env recombinant alone may be sufficient for eliciting immune responses against both subtype B and C envelopes. Further studies of Ad5hr-HIV(TV1) gp160 in rhesus macaques will evaluate the suitability of this insert for a future phase I clinical trial using a replication-competent Ad4 vector.

Figure 1. Construction of Ad5hr HIV-1 gp160 recombinants

(A) Western blot of QBI 293 cell lysate after being transfected with plasmid expressing HIV_TV1 gp160 (lane 2) or HIV_SF162 gp160 (lane 3). The Env band was not observed in mock transfected cells (lane 1) or untransfected cells (lane 4). (B) Schematic diagram of the Ad5hr-shuttle plasmid and the left hand Ad5hr fragment used to create replication-competent Ad5hr-recombinant virus by homologous recombination. (C) Expression of HIV-1 gp160 in QBI 293 cells after being transduced with 1 MOI of Ad5hr-HIV_TV1 (lane 1) or Ad5hr-HIV_SF162 (lane 2).

Figure 2. HIV-1 gp160 specific ELISpot responses following immunization with Ad5hr recombinants

Spleen cells of immunized mice were harvested at the indicated time points and tested for IFN-γ secretion after stimulation with the indicated HIV-1 gp160 peptides. Significant differences in comparison to the baseline control are indicated as follows: a, p <0.01; b, p <0.02; c, p <0.03; d, p < 0.04; e, p <0.05.

Figure 3. Mean vaccine-induced IFN-γ-secreting cells by immunization group

SFC are plotted as a function of time post-immunization, in response to the indicated Env peptide stimuli. Mean values and standard errors of the mean are shown.

Figure 4. Anti-HIV Env binding antibody in immunized mouse sera

Sera collected from immunized mice in each group at indicated time points were pooled and tested for specific binding to HIV Env proteins of several different HIV-1 subtypes. Results are the average of 2 independent measurements of 450nm absorbance at serum dilutions of 1:50.

Figure 5. Antibody binding titers against HIV Env proteins of different subtypes

Results of sera pooled according to immunization group and time post-immunization. Means of each HIV-Env subtype are shown.