None of the HIV T-cell vaccine candidates that have reached advanced clinical testing have been able to induce protective T cell immunity. A major reason for these failures may have been suboptimal T cell immunogen designs. To overcome this problem, we used a novel immunogen design approach that is based on functional T cell response data from more than 1,000 HIV-1 clade B and C infected individuals and which aims to direct the T cell response to the most vulnerable sites of HIV-1. Our approach identified 16 regions in Gag, Pol, Vif and Nef that were relatively conserved and predominantly targeted by individuals with reduced viral loads. These regions formed the basis of the HIVACAT T-cell Immunogen (HTI) sequence which is 529 amino acids in length, includes more than 50 optimally defined CD4+ and CD8+ T-cell epitopes restricted by a wide range of HLA class I and II molecules and covers viral sites where mutations led to a dramatic reduction in viral replicative fitness. In both, C57BL/6 mice and Indian rhesus macaques immunized with an HTI-expressing DNA plasmid (DNA.HTI) induced broad and balanced T-cell responses to several segments within Gag, Pol, and Vif. DNA.HTI induced robust CD4+ and CD8+ T cell responses that were increased by a booster vaccination using modified virus Ankara (MVA.HTI), expanding the DNA.HTI induced response to up to 3.2% IFN-γ T-cells in macaques. HTI-specific T cells showed a central and effector memory phenotype with a significant fraction of the IFN-γ+ CD8+ T cells being Granzyme B+ and able to degranulate (CD107a+). These data demonstrate the immunogenicity of a novel HIV-1 T cell vaccine concept that induced broadly balanced responses to vulnerable sites of HIV-1 while avoiding the induction of responses to potential decoy targets that may divert effective T-cell responses towards variable and less protective viral determinants.

An effective AIDS vaccine continues to be of paramount importance for the control of the pandemic, and it has been proven to be an elusive target. Vaccine efficacy trials and macaque challenge studies indicate that protection may be the result of combinations of many parameters. We show that a combination of DNA and protein vaccinations applied at the same time provides rapid and robust cellular and humoral immune responses and evidence for a reduced risk of infection. Vaccine-induced neutralizing antibodies and Env V2-specific antibodies at mucosal sites contribute to the delay of SIV _smE660 acquisition, and genetic makeup (TRIM-5α) affects the effectiveness of the vaccine. These data are important for the design of better vaccines and may also affect other vaccine platforms.

HIV sequence diversity and the propensity of eliciting immunodominant responses targeting variable regions of the HIV proteome are hurdles in the development of an effective AIDS vaccine. An HIV-derived conserved element (CE) p24^gag plasmid DNA (pDNA) vaccine is able to redirect immunodominant responses to otherwise subdominant and often more vulnerable viral targets. By homology to the HIV immunogen, seven CE were identified in SIV p27^Gag. Analysis of 31 rhesus macaques vaccinated with full-length SIV gag pDNA showed inefficient induction (58% response rate) of cellular responses targeting these CE. In contrast, all 14 macaques immunized with SIV p27CE pDNA developed robust T cell responses recognizing CE. Vaccination with p27CE pDNA was also critical for the efficient induction and increased the frequency of Ag-specific T cells with cytotoxic potential (granzyme B⁺ CD107a⁺) targeting subdominant CE epitopes, compared with the responses elicited by the p57^gag pDNA vaccine. Following p27CE pDNA priming, two booster regimens, gag pDNA or codelivery of p27CE+gag pDNA, significantly increased the levels of CE-specific T cells. However, the CE+gag pDNA booster vaccination elicited significantly broader CE epitope recognition, and thus, a more profound alteration of the immunodominance hierarchy. Vaccination with HIV molecules showed that CE+gag pDNA booster regimen further expanded the breadth of HIV CE responses. Hence, SIV/HIV vaccine regimens comprising CE pDNA prime and CE+gag pDNA booster vaccination significantly increased cytotoxic T cell responses to subdominant highly conserved Gag epitopes and maximized response breadth.

We evaluated the antibody responses in 259 potential convalescent plasma donors for Covid-19 patients. Different assays were used: a commercial ELISA detecting antibodies against the recombinant spike protein (S1); a multiplex assay detecting total and specific antibody isotypes against three SARS-CoV-2 antigens (S1, basic nucleocapsid (N) protein and receptor-binding domain (RBD)); and an in-house ELISA detecting antibodies to complete spike, RBD and N in 60 of these donors. Neutralizing antibodies (NAb) were also evaluated in these 60 donors. Analyzed samples were collected at a median time of 62 (14–104) days from the day of first symptoms or positive PCR (for asymptomatic patients). Anti-SARS-CoV-2 antibodies were detected in 88% and 87.8% of donors using the ELISA and the multiplex assay, respectively. The multivariate analysis showed that age ≥50 years (p < 0.001) and need for hospitalization (p < 0.001) correlated with higher antibody titers, while asymptomatic status (p < 0.001) and testing >60 days after symptom onset (p = 0.001) correlated with lower titers. Interestingly, pseudotype virus-neutralizing antibodies (PsNAbs) significantly correlated with spike and with RBD antibodies by ELISA. Sera with high PsNAb also showed a strong ability to neutralize active SARS-CoV-2 virus, with hospitalized patients showing higher titers. Therefore, convalescent plasma donors can be selected based on the presence of high RBD antibody titers.

HIV sequence diversity and the propensity of eliciting immunodominant responses targeting inessential variable regions are hurdles in the development of an effective AIDS vaccine. We developed a DNA vaccine comprising conserved elements (CE) of SIV p27^Gag and HIV-1 Env and found that priming vaccination with CE DNA is critical to efficiently overcome the dominance imposed by Gag and Env variable regions. Here, we show that DNA vaccinated macaques receiving the CE prime/CE+full-length DNA co-delivery booster vaccine regimens developed broad, potent and durable cytotoxic T cell responses targeting conserved protein segments of SIV Gag and HIV Env. Gag CE-specific T cells showed robust anamnestic responses upon infection with SIV_mac239 which led to the identification of CE-specific cytotoxic lymphocytes able to recognize epitopes covering distinct CE on the surface of SIV infected cells in vivo. Though not controlling infection overall, we found an inverse correlation between Gag CE-specific CD8⁺ T cell responses and peak viremia. The T cell responses induced by the HIV Env CE immunogen were recalled in some animals upon SIV infection, leading to the identification of two cross-reactive epitopes between HIV and SIV Env based in sequence homology. These data demonstrate that a vaccine combining Gag and Env CE DNA subverted the normal immunodominance patterns, eliciting immune responses that included subdominant, highly conserved epitopes. These vaccine regimens augment cytotoxic T cell responses to highly conserved epitopes in the viral proteome and maximize response breadth. The vaccine-induced CE-specific T cells were expanded upon SIV infection, indicating that the predicted CE epitopes incorporated in the DNA vaccine are processed and exposed by infected cells in their natural context within the viral proteome.