Immunogenic Profiling in Mice of a HIV/AIDS Vaccine Candidate (MVA-B) Expressing Four HIV-1 Antigens and Potentiation by Specific Gene Deletions

Abstract

BackgroundThe immune parameters of HIV/AIDS vaccine candidates that might be relevant in protection against HIV-1 infection are still undefined. The highly attenuated poxvirus strain MVA is one of the most promising vectors to be use as HIV-1 vaccine. We have previously described a recombinant MVA expressing HIV-1 Env, Gag, Pol and Nef antigens from clade B (referred as MVA-B), that induced HIV-1-specific immune responses in different animal models and gene signatures in human dendritic cells (DCs) with immunoregulatory function.Methodology/Principal FindingsIn an effort to characterize in more detail the immunogenic profile of MVA-B and to improve its immunogenicity we have generated a new vector lacking two genes (A41L and B16R), known to counteract host immune responses by blocking the action of CC-chemokines and of interleukin 1β, respectively (referred as MVA-B ΔA41L/ΔB16R). A DNA prime/MVA boost immunization protocol was used to compare the adaptive and memory HIV-1 specific immune responses induced in mice by the parental MVA-B and by the double deletion mutant MVA-B ΔA41L/ΔB16R. Flow cytometry analysis revealed that both vectors triggered HIV-1-specific CD4+ and CD8+ T cells, with the CD8+ T-cell compartment responsible for >91.9% of the total HIV-1 responses in both immunization groups. However, MVA-B ΔA41L/ΔB16R enhanced the magnitude and polyfunctionality of the HIV-1-specific CD4+ and CD8+ T-cell immune responses. HIV-1-specific CD4+ T-cell responses were polyfunctional and preferentially Env-specific in both immunization groups. Significantly, while MVA-B induced preferentially Env-specific CD8+ T-cell responses, MVA-B ΔA41L/ΔB16R induced more GPN-specific CD8+ T-cell responses, with an enhanced polyfunctional pattern. Both vectors were capable of producing similar levels of antibodies against Env.Conclusions/SignificanceThese findings revealed that MVA-B and MVA-B ΔA41L/ΔB16R induced in mice robust, polyfunctional and durable T-cell responses to HIV-1 antigens, but the double deletion mutant showed enhanced magnitude and quality of HIV-1 adaptive and memory responses. Our observations are relevant in the immune evaluation of MVA-B and on improvements of MVA vectors as HIV-1 vaccines.

Deletion of the Vaccinia Virus N2L Gene Encoding an Inhibitor of IRF3 Improves the Immunogenicity of Modified Vaccinia Virus Ankara Expressing HIV-1 Antigens

“…For simplicity, throughout this work, we use the open reading frame nomenclature of the Copenhagen strain to refer to the MVA genes). pGem-RG-N2L wm was obtained by sequential cloning of N2L flanking sequences into plasmid pGem-RG wm (4,540 bp), whose generation was previously described (63), and contains the genes for DsRed2 and red-shifted green fluorescent protein (rsGFP) under the control of the synthetic early/late (sE/L) promoter. The MVA-B genome was used as the template to amplify the right flank of the N2L gene (474 bp) with oligonucleotides RFN2L-AatII-F (5=-GTCTAGGACGTCTAATTTT CATATGG-3=) (AatII site underlined) and RFN2L-XbaI-R (5=-AAGTAT TCTAGAAATTAATGATGC-3=) (XbaI site underlined).…”

“…The MVA-B ⌬N2L deletion mutant generated in this work contains a deletion of the VACV gene N2L and was constructed by transfecting plasmid transfer vector pGem-RG-N2L wm into DF-1 cells infected with the parental virus MVA-B and then screening for transient Red2-GFP coexpression with the genes for DsRed2 and rs-GFP as the transiently selectable markers as previously described (42,63). MVA-B ⌬N2L was finally obtained after six consecutive rounds of plaque purification in DF-1 cells.…”

“…Primers RFN2L-AatII-F and LFN2L-BamHI-R (described above), spanning N2L flanking regions, were used for PCR analysis of the N2L locus. The amplification protocol was previously described (63). PCR products were run in 1% agarose gel and visualized by SYBR Safe staining (Invitrogen).…”

“…This was done by deleting the VACV N2L gene from the vector backbone of MVA-B, an HIV/AIDS vaccine candidate based on an attenuated recombinant MVA vector expressing HIV-1 Env and Gag-Pol-Nef (GPN) antigens from clade B (60). MVA-B has been extensively studied in vitro and in different animal models (42,(60)(61)(62)(63)(64)(65)(66), where it triggers strong and durable immune responses to HIV-1 antigens. Furthermore, MVA-B was safe and highly immunogenic when tested in a phase I clinical trial with healthy human volunteers, inducing humoral responses to Env and HIV-1-specific CD4 ϩ and CD8 ϩ T cell responses that were high, broad, polyfunctional, and long-lasting (12,14).…”

Deletion of Specific Immune-Modulatory Genes from Modified Vaccinia Virus Ankara-Based HIV Vaccines Engenders Improved Immunogenicity in Rhesus Macaques

“…The biological activities of these factors can be inferred by the fact that deletions of their orthologs from replication-competent strains of vaccinia virus result in altered pathogenic phenotypes in murine models (2,62,74,77,80). Additionally, the observation that MVA recombinants lacking MVA184R and/or MVA153L may elicit modestly augmented CD8 T cell responses in mice indicates that the deletion of immune-modulatory genes from MVA-based vaccines is a relevant approach toward improving vaccine immunogenicity (18,21,32,78).…”