Consensus HIV-1 FSU-A Integrase Gene Variants Electroporated into Mice Induce Polyfunctional Antigen-Specific CD4+ and CD8+ T Cells

• DOI: 10.1371/journal.pone.0062720
• Publication Date: 2013-05-08T21:42:37Z
• Authors: Olga Krotova; Elizaveta Starodu...; Stefan Petkov; Linda Kostic; Julia Agapkina; David Hallengärd; Alecia Viklund; Oleg Latyshev; Eva Gelius; Tomas Dillenbeck; Vadim Karpov; Marina Gottikh; Igor M. Belyakov; Vladimir Lukashov; Maria G. Isaguliants
• Keywords: CD4-Positive T-Lymphocytes; Science; HIV Integrase; CD8-Positive T-Lymphocytes; Quinolones; Lymphocyte Activation; Cell Line; Mice; 03 medical and health sciences; Drug Resistance, Viral; Escherichia coli; Animals; Humans; HIV Integrase Inhibitors; Luciferases; Mice, Inbred BALB C; 0303 health sciences; Q; R; Flow Cytometry; 3. Good health; Electroporation; HIV-1; Mutagenesis, Site-Directed; Medicine; Research Article

Our objective is to create gene immunogens targeted against drug-resistant HIV-1, focusing on HIV-1 enzymes as critical components in viral replication and drug resistance. Consensus-based gene vaccines are specifically fit for variable pathogens such as HIV-1 and have many advantages over viral genes and their expression-optimized variants. With this in mind, we designed the consensus integrase (IN) of the HIV-1 clade A strain predominant in the territory of the former Soviet Union and its inactivated derivative with and without mutations conferring resistance to elvitegravir. Humanized IN gene was synthesized; and inactivated derivatives (with 64D in the active site mutated to V) with and without elvitegravir-resistance mutations were generated by site-mutagenesis. Activity tests of IN variants expressed in E coli showed the consensus IN to be active, while both D64V-variants were devoid of specific activities. IN genes cloned in the DNA-immunization vector pVax1 (pVaxIN plasmids) were highly expressed in human and murine cell lines (>0.7 ng/cell). Injection of BALB/c mice with pVaxIN plasmids followed by electroporation generated potent IFN-γ and IL-2 responses registered in PBMC by day 15 and in splenocytes by day 23 after immunization. Multiparametric FACS demonstrated that CD8+ and CD4+ T cells of gene-immunized mice stimulated with IN-derived peptides secreted IFN-γ, IL-2, and TNF-α. The multi-cytokine responses of CD8+ and CD4+ T-cells correlated with the loss of in vivo activity of the luciferase reporter gene co-delivered with pVaxIN plasmids. This indicated the capacity of IN-specific CD4+ and CD8+ T-cells to clear IN/reporter co-expressing cells from the injection sites. Thus, the synthetic HIV-1 clade A integrase genes acted as potent immunogens generating polyfunctional Th1-type CD4+ and CD8+ T cells. Generation of such response is highly desirable for an effective HIV-1 vaccine as it offers a possibility to attack virus-infected cells via both MHC class I and II pathways.