Novel recombinant DNA and live virus vaccines to prevent or control HIV-1 infection

Abstract

Background Vaccination is the most cost effective and long-term solution to the global human immunodeficiency virus (HIV) pandemic. The HIV Gag and Tat proteins are attractive components of a HIV vaccine as immune responses targeting these proteins confer protective benefits against HIV infections in humans. This thesis has developed two innovative candidate HIV vaccines viz. a DNA vaccine encoding oligomerised and secreted Tat (pVAXsTat-IMX313), and a recombinant live human rhinovirus serotype A1 (HRV-A1)-based vaccine encoding Gag and Tat (rHRV-Gag/Tat). Methods To construct pVAX-sTat-IMX313, Tat was fused with the oligomerisation domain of IMX313 to form Tat heptamers and linked to the leader sequence of tissue plasminogen activator to ensure that the bulk of oligomerised protein is secreted. To develop the rHRVGag/Tat vaccine, initially, the full length tat gene and 5 discrete overlapping fragments corresponding to the full length gag gene were individually inserted into the junction between the HRV-A1 genes encoding structural and non-structural proteins (P1/P2 junction) to ensure that the exogenous HIV Gag or Tat proteins were separated from the recombinant polyprotein using the HRV encoded 2Aprotease enzyme. Thus, one recombinant HRV encoding Tat (rHRV-Tat) and 5 rHRVs each encoding a unique Gag fragment (rHRV-Gag1-5) were generated. The individual rHRVs were then mixed into a single cocktail vaccine (rHRVGag/Tat), purified and titrated for inoculation in mice. The immunogenicity of these vaccines was evaluated in female BALB/c mice that received up to five intradermal injections of pVAX-sTat-IMX313 (50 μg per dose) at 2 weekly intervals in one study. In another study, mice were vaccinated intranasally with 2 doses (5x106 TCID50/dose) of the rHRV-Gag/Tat followed by a single 50 μg booster dose of a cocktail DNA vaccine containing pVAX-sTat-IMX313 and pVAX-Gag-Perforin. Vaccine-induced immune responses were examined 2 weeks after the last dose by antibody ELISA, in-vitro Tat transactivation neutralization, IFN-γ ELISpot, KdGag197-205 tetramer staining and intracellular cytokine staining assays. Results Data showed that fusing Tat with IMX313 results in complete heptamerisation of Tat. Furthermore, the data suggested that pVAX-sTat-IMX313 vaccination elicited higher titers of serum neutralizing Tat-specific IgG, secretory IgA (sIgA) in the vagina and CMI responses, and showed superior control of ecotropic HIV (EcoHIV) infection, a surrogate murine HIV challenge model, compared with animals vaccinated with other DNA vaccines tested in this study. Human rhinovirus serotype A1 (HRV-A1) was successfully engineered into a replication-competent genetically stable recombinant vector to deliver a mucosally-targeted vaccine, rHRV-Gag/Tat, by inserting exogenous HIV gag and tat sequences into the HRV-A1 genome. Finally, intranasal administration of 2 doses of rHRV-Gag/Tat followed by a single DNA booster dose induced superior poly-functional Gag-specific CD8 T cell responses in the spleen (systemic) and mesenteric lymph nodes (mucosal), higher Tat-specific serum IgG and sIgA in the vagina, and effective control of EcoHIV infection compared to other vaccination regimens tested in this study. Conclusion First, the data support the inclusion of IMX313 as a molecular adjuvant for Tat-based HIV DNA vaccines. Second, the data demonstrated that intranasal vaccination with rHRV-Gag/Tat followed by a single DNA booster dose is effective in eliciting HIV-specific immunity panmucosally and systemically. Collectively, the data support further testing of the pVAX-sTat IMX313 and rHRV-Gag/Tat vaccines in macaques, preferably in a heterologous prime-boost vaccination strategy, and results from these studies might influence future HIV clinical trials.