The lymphocyte population was gated for CD8+ B220- cells (c) and CD4+ B220- cells (d). IFN+ CD44+ B220- CD8+/CD4+ cells of the lymphocytes with the number of counted lymphocytes per spleen. To obtain the percentage of double positive (IFN+ TNF+) cells of IFN+ CD8+ and CD4+ T-cells, CD8+/CD4+ B220- cells were also SC 57461A gated for CD44+ cells (e, both rectangles) in homologous prime-boost regimen and subsequently for IFN- TNF+ (f, left rectangle) and IFN+ TNF+ (f, right rectangle) cells. 12967_2019_1924_MOESM1_ESM.pptx (5.9M) GUID:?D1D823F7-617B-4A94-A7FD-2624815BC4FE Data Availability StatementAll data generated and analyzed during this study are included in this published article and its additional file. Unique materials generated in the study are available for non-commercial purposes. Abstract Background In non-human primates (NHPs) and humans, partial Mouse monoclonal to CD106 protection from HIV/SIV infection or suppression of replication is achievable by Env-binding antibodies and Gag-specific CD8+ T-cells targeting protective epitopes. Unfortunately, such T-cell responses are frequently dominated by responses to non-protective, variable epitopes. In this study we attempt to combine three independent approaches, each developed to prevent immunodominance of non-protective epitopes. These approaches were (1) vaccines consisting exclusively of putatively protective p24 Gag highly conserved elements (CEs), (2) vaccines using?solely subdominant antigens which were acutely protective in a recent NHP SC 57461A trial, and (3) virus-encoded virus-like particle vaccines (virus-like vaccines/VLVs) using SC 57461A heterologous Env and Gag sequences to enable selection of broadly cross-reactive responses and to avoid immunodominance of non-conserved sequences in prime-boost regimens as previously observed. Methods We vaccinated outbred CD1 mice with HIV-1 clade B Gag/Env encoded in an adenoviral prime and SIVmac239 Gag/Env in an MVA boost. We combined this completely heterologous immunization regimen and the homologous SIVmac239 Gag/Env immunization regimen with an additional prime encoding SIV CEs and accessory antigens Rev, Vif and Vpr (Ad-Ii-SIVCErvv). T-cell responses were analyzed by intracellular cytokine staining of splenocytes and antibody responses by trimer-specific ELISA, avidity and isotype-specific ELISA. Results Env dominance could be avoided successfully in the completely heterologous prime-boost regimen, but Env immunodominance reappeared when Ad-Ii-SIVCErvv was added to the prime. This regimen did however still induce more cross-reactive Gag-specific CD8+ T-cells and Env-specific antibodies. Including Ad-Ii-SIVCErvv in the homologous prime-boost not only elicited accessory antigen-specific CD8+ memory T-cells, but also significantly increased the ratio of Gag- to Env-specific CD8+ T-cells. The CD4+ T-cell response shifted away from structural antigens previously associated with infection-enhancement. Conclusion The homologous Gag/Env prime-boost with Ad-Ii-SIVCErvv prime combined acutely protective CD8+ T-cell responses to subdominant antigens and Env-binding antibodies with chronically protective Gag-specific CD8+ T-cells in outbred mice. This vaccine regimen should be tested in an NHP efficacy trial. Electronic supplementary material The online version of this article (10.1186/s12967-019-1924-1) contains supplementary material, which is available to authorized users. Keywords: Adenoviral vectors, Human immunodeficiency virus, Virus-like particles, Virus-like vaccines, T-cells, Antibodies, Heterologous viral vectored prime-boost immunization, Subdominant antigen vaccine Background The primary goal of a human immunodeficiency virus type 1 (HIV-1) vaccine is to protect from HIV-1 acquisition [1]. The partial protection in the HIV vaccination trial RV144 correlated with IgG antibodies targeting the V1V2 regions of the viral envelope (Env) protein [2, 3]. If prevention of infection is not possible an alternative goal is to induce CD8+ T-cells that can control the infection [1]. In chronically infected individuals CD8+ T-cells targeting group-specific antigen (Gag) correlated directly with a lower viral load while Env-specific CD8+ T-cells correlated inversely [4, 5]. These associations provide an SC 57461A incentive to increase antibody responses towards Env as well as CD8+ T-cell responses towards Gag and to reduce Env-specific CD8+ T-cell responses. A great impediment for the development of an HIV-1 vaccine that induces protective immune responses or such that can control HIV-1 infection is the immunodominance of non-protective, variable epitopes which the virus can easily mutate without fitness cost. An immune response towards these epitopes suppresses responses towards more conserved regions with a higher protective potential [6]. Kulkarni et al. addressed this problem by designing immunogens based on 7?highly conserved elements (CEs) in HIV-1 p24 Gag [7]. A DNA vaccine encoding this string of peptides raised higher T-cell responses to the CEs compared to vaccination with full-length Gag DNA in mice and macaques [7, 8]. A prime vaccination using CE DNA, followed by a boost with full-length Gag DNA, increased the magnitude and breadth of Gag-specific T-cell responses, including responses to the CEs [8]. In Hu et al..