Improved Innate and Adaptive Immunostimulation by Genetically Modified HIV-1 Protein Expressing NYVAC Vectors

Abstract

Attenuated poxviruses are safe and capable of expressing foreign antigens. Poxviruses are applied in veterinary vaccination and explored as candidate vaccines for humans. However, poxviruses express multiple genes encoding proteins that interfere with components of the innate and adaptive immune response. This manuscript describes two strategies aimed to improve the immunogenicity of the highly attenuated, host-range restricted poxvirus NYVAC: deletion of the viral gene encoding type-I interferon-binding protein and development of attenuated replication-competent NYVAC. We evaluated these newly generated NYVAC mutants, encoding HIV-1 env, gag, pol and nef, for their ability to stimulate HIV-specific CD8 T-cell responses in vitro from blood mononuclear cells of HIV-infected subjects. The new vectors were evaluated and compared to the parental NYVAC vector in dendritic cells (DCs), RNA expression arrays, HIV gag expression and cross-presentation assays in vitro. Deletion of type-I interferon-binding protein enhanced expression of interferon and interferon-induced genes in DCs, and increased maturation of infected DCs. Restoration of replication competence induced activation of pathways involving antigen processing and presentation. Also, replication-competent NYVAC showed increased Gag expression in infected cells, permitting enhanced cross-presentation to HIV-specific CD8 T cells and proliferation of HIV-specific memory CD8 T-cells in vitro. The recombinant NYVAC combining both modifications induced interferon-induced genes and genes involved in antigen processing and presentation, as well as increased Gag expression. This combined replication-competent NYVAC is a promising candidate for the next generation of HIV 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

“…(81), but an additional fifth deletion of the uracil-DNA glycosylase gene (MVA101R) decreased the responses (81). Moreover, NYVAC vectors with a single deletion of a VACV gene encoding a TLR inhibitor (A46R [83]) and with single or double deletions in VACV genes encoding type I and II IFN binding proteins (B19R and B8R, respectively [84][85][86]) were also able to enhance the immune responses to HIV-1 antigens in mice.…”

Removal of Vaccinia Virus Genes That Block Interferon Type I and II Pathways Improves Adaptive and Memory Responses of the HIV/AIDS Vaccine Candidate NYVAC-C in Mice

“…One approach to improve the vector immunogenicity is to remove from the viral DNA genes that antagonize host-specific immune responses. This strategy has being successfully used for MVA and NYVAC vectors by targeting several immunomodulatory genes (8,10,15,16,24,42,47). Since virus recognition and induction of IFNs are critical components of the innate immune system, here we evaluated in a mouse model how the single or double deletion of the viral genes B8R and B19R affected the immunogenicity of the HIV/AIDS vaccine candidate NYVAC-C.…”

“…We recently reported the generation and in vitro characterization of an improved NYVAC-C recombinant expressing the HIV-1 Env and Gag-Pol-Nef antigens from clade C, by deletion of the viral gene B19R (NYVAC-C-⌬B19R) (24,42). In contrast to the parental NYVAC-C, the deletion mutant NYVAC-C-⌬B19R induced in human DCs enhanced IFN-␣ production, maturation, and expression of IFN-induced pathways and IFN-regulated transcription factors as well as multiple inflammatory cytokines.…”

CD4 T follicular helper cell dynamics during SIV infection

“…RNA samples were prepared using the Illumina beads station assay, as previously described (59,60), and hybridized to the Illumina HumanHT-12 version 4 Expression BeadChip, according to the Illumina's instruction. The data were normalized with the quantile normalization method of Bioconductor package limma (61).…”