A Single-shot ChAd3 Vaccine Provides Protection from Intramuscular and Aerosol Sudan Virus Exposure.

Infection with Sudan virus (SUDV) is characterized by an aggressive disease course with case fatality rates between 40-100% and no approved vaccines or therapeutics. SUDV causes sporadic outbreaks in sub-Saharan Africa, including a recent outbreak in Uganda which has resulted in over 100 confirmed cases in one month. Prior vaccine and therapeutic efforts have historically prioritized Ebola virus (EBOV), leading to a significant gap in available treatments. Two vaccines, Erbevo ® and Zabdeno ® /Mvabea ® , are licensed for use against EBOV but are ineffective against SUDV. Recombinant adenovirus vector vaccines have been shown to be safe and effective against filoviruses, but efficacy depends on having low seroprevalence to the vector in the target human population. For this reason, and because of an excellent safety and immunogenicity profile, ChAd3 was selected as a superior vaccine vector. Here, a ChAd3 vaccine expressing the SUDV glycoprotein (GP) was evaluated for immunogenicity and efficacy in nonhuman primates. We demonstrate that a single dose of ChAd3-SUDV confers acute and durable protection against lethal SUDV challenge with a strong correlation between the SUDV GP-specific antibody titers and survival outcome. Additionally, we show that a bivalent ChAd3 vaccine encoding the GP from both EBOV and SUDV protects against both parenteral and aerosol lethal SUDV challenge. Our data indicate that the ChAd3-SUDV vaccine is a suitable candidate for a prophylactic vaccination strategy in regions at high risk of filovirus outbreaks. One Sentence Summary: A single-dose of ChAd3 vaccine protected macaques from lethal challenge with Sudan virus (SUDV) by parenteral and aerosol routes of exposure.

Durable immunity to SARS-CoV-2 in both lower and upper airways achieved with a gorilla adenovirus (GRAd) S-2P vaccine in non-human primates.

SARS-CoV-2 continues to pose a global threat, and current vaccines, while effective against severe illness, fall short in preventing transmission. To address this challenge, there's a need for vaccines that induce mucosal immunity and can rapidly control the virus. In this study, we demonstrate that a single immunization with a novel gorilla adenovirus-based vaccine (GRAd) carrying the pre-fusion stabilized Spike protein (S-2P) in non-human primates provided protective immunity for over one year against the BA.5 variant of SARS-CoV-2. A prime-boost regimen using GRAd followed by adjuvanted S-2P (GRAd+S-2P) accelerated viral clearance in both the lower and upper airways. GRAd delivered via aerosol (GRAd(AE)+S-2P) modestly improved protection compared to its matched intramuscular regimen, but showed dramatically superior boosting by mRNA and, importantly, total virus clearance in the upper airway by day 4 post infection. GrAd vaccination regimens elicited robust and durable systemic and mucosal antibody responses to multiple SARS-CoV-2 variants, but only GRAd(AE)+S-2P generated long-lasting T cell responses in the lung. This research underscores the flexibility of the GRAd vaccine platform to provide durable immunity against SARS-CoV-2 in both the lower and upper airways.

Mucosal Adenoviral-vectored Vaccine Boosting Durably Prevents XBB.1.16 Infection in Nonhuman Primates.

Waning immunity and continued virus evolution have limited the durability of protection from symptomatic infection mediated by intramuscularly (IM)-delivered mRNA vaccines against COVID-19 although protection from severe disease remains high. Mucosal vaccination has been proposed as a strategy to increase protection at the site of SARS-CoV-2 infection by enhancing airway immunity, potentially reducing rates of infection and transmission. Here, we compared protection against XBB.1.16 virus challenge 5 months following IM or mucosal boosting in non-human primates (NHP) that had previously received a two-dose mRNA-1273 primary vaccine regimen. The mucosal boost was composed of a bivalent chimpanzee adenoviral-vectored vaccine encoding for both SARS-CoV-2 WA1 and BA.5 spike proteins (ChAd-SARS-CoV-2-S) and delivered either by an intranasal mist or an inhaled aerosol. An additional group of animals was boosted by the IM route with bivalent WA1/BA.5 spike-matched mRNA (mRNA-1273.222) as a benchmark control. NHP were challenged in the upper and lower airways 18 weeks after boosting with XBB.1.16, a heterologous Omicron lineage strain. Cohorts boosted with ChAd-SARS-CoV-2-S by an aerosolized or intranasal route had low to undetectable virus replication as assessed by levels of subgenomic SARS-CoV-2 RNA in the lungs and nose, respectively. In contrast, animals that received the mRNA-1273.222 boost by the IM route showed minimal protection against virus replication in the upper airway but substantial reduction of virus RNA levels in the lower airway. Immune analysis showed that the mucosal vaccines elicited more durable antibody and T cell responses than the IM vaccine. Protection elicited by the aerosolized vaccine was associated with mucosal IgG and IgA responses, whereas protection elicited by intranasal delivery was mediated primarily by mucosal IgA. Thus, durable immunity and effective protection against a highly transmissible heterologous variant in both the upper and lower airways can be achieved by mucosal delivery of a virus-vectored vaccine. Our study provides a template for the development of mucosal vaccines that limit infection and transmission against respiratory pathogens.

Immune Activation Profiles Elicited by Distinct, Repeated TLR Agonist Infusions in Rhesus Macaques.

TLR agonists are a promising class of immune system stimulants investigated for immunomodulatory applications in cancer immunotherapy and viral diseases. In this study, we sought to characterize the safety and immune activation achieved by different TLR agonists in rhesus macaques (Macaca mulatta), a useful preclinical model of complex immune interactions. Macaques received one of three TLR agonists, followed by plasma cytokine, immune cell subset representation, and blood cell activation measurements. The TLR4 agonist LPS administered i.v. induced very transient immune activation, including TNF-α expression and monocyte activation. The TLR7/8 agonist 2BXy elicited more persistent cytokine expression, including type I IFN, IL-1RA, and the proinflammatory IL-6, along with T cell and monocyte activation. Delivery of 2BXy i.v. and i.m. achieved comparable immune activation, which increased with escalating dose. Finally, i.v. bacillus Calmette-Guérin (BCG) vaccination (which activates multiple TLRs, especially TLR2/4) elicited the most pronounced and persistent innate and adaptive immune response, including strong induction of IFN-γ, IL-6, and IL-1RA. Strikingly, monocyte, T cell, and NK cell expression of the proliferation marker Ki67 increased dramatically following BCG vaccination. This aligned with a large increase in total and BCG-specific cells measured in the lung. Principal component analysis of the combined cytokine expression and cellular activation responses separated animals by treatment group, indicating distinct immune activation profiles induced by each agent. In sum, we report safe, effective doses and routes of administration for three TLR agonists that exhibit discrete immunomodulatory properties in primates and may be leveraged in future immunotherapeutic strategies.

Investigation of the impact of clonal hematopoiesis on severity and pathophysiology of COVID-19 in rhesus macaques.

Clinical manifestations of COVID-19 vary widely, ranging from asymptomatic to severe respiratory failure with profound inflammation. Although risk factors for severe illness have been identified, definitive determinants remain elusive. Clonal hematopoiesis (CH), the expansion of hematopoietic stem and progenitor cells bearing acquired somatic mutations, is associated with advanced age and hyperinflammation. Given the similar age range and hyperinflammatory phenotype between frequent CH and severe COVID-19, CH could impact the risk of severe COVID-19. Human cohort studies have attempted to prove this relationship, but conclusions are conflicting. Rhesus macaques (RMs) are being utilized to test vaccines and therapeutics for COVID-19. However, RMs, even other species, have not yet been reported to develop late inflammatory COVID-19 disease. Here, RMs with either spontaneous DNMT3A or engineered TET2 CH along with similarly transplanted and conditioned controls were infected with SARS-CoV-2 and monitored until 12 days post-inoculation (dpi). Although no significant differences in clinical symptoms and blood counts were noted, an aged animal with natural DNMT3A CH died on 10 dpi. CH macaques showed evidence of sustained local inflammatory responses compared to controls. Interestingly, viral loads in respiratory tracts were higher at every timepoint in the CH group. Lung sections from euthanasia showed evidence of mild inflammation in all animals, while viral antigen was more frequently detected in the lung tissues of CH macaques even at the time of autopsy. Despite the lack of striking inflammation and serious illness, our findings suggest potential pathophysiological differences in RMs with or without CH upon SARS-CoV-2 infection.

Investigation of the Impact of Clonal Hematopoiesis on Severity and Pathophysiology of COVID-19 in Rhesus Macaques.

Clinical manifestations of COVID-19 vary widely, ranging from asymptomatic to severe respiratory failure with profound inflammation. Although risk factors for severe illness have been identified, definitive determinants remain elusive. Clonal hematopoiesis (CH), the expansion of hematopoietic stem and progenitor cells bearing acquired somatic mutations, is associated with advanced age and hyperinflammation. Given the similar age range and hyperinflammatory phenotype between frequent CH and severe COVID-19, CH could impact the risk of severe COVID-19. Human cohort studies have attempted to prove this relationship, but conclusions are conflicting. Rhesus macaques (RMs) are being utilized to test vaccines and therapeutics for COVID-19. However, RMs, even other species, have not yet been reported to develop late inflammatory COVID-19 disease. Here, RMs with either spontaneous DNMT3A or engineered TET2 CH along with similarly transplanted and conditioned controls were infected with SARS-CoV-2 and monitored until 12 days post-inoculation (dpi). Although no significant differences in clinical symptoms and blood counts were noted, an aged animal with natural DNMT3A CH died on 10 dpi. CH macaques showed evidence of sustained local inflammatory responses compared to controls. Interestingly, viral loads in respiratory tracts were higher at every timepoint in the CH group. Lung sections from euthanasia showed evidence of mild inflammation in all animals, while viral antigen was more frequently detected in the lung tissues of CH macaques even at the time of autopsy. Despite the lack of striking inflammation and serious illness, our findings suggest potential pathophysiological differences in RMs with or without CH upon SARS-CoV-2 infection. Highlights: No evidence of association between CH and COVID-19 clinical severity in macaques.The presence of CH is associated with prolonged local inflammatory responses in COVID-19.SARS-CoV-2 persists longer in respiratory tracts of macaques with CH following infection.

A single-shot ChAd3-MARV vaccine confers rapid and durable protection against Marburg virus in nonhuman primates.

Marburg virus (MARV) causes a severe hemorrhagic fever disease in primates with mortality rates in humans of up to 90%. MARV has been identified as a category A bioterrorism agent by the Centers for Disease Control and Prevention (CDC) and priority pathogen A by the National Institute of Allergy and Infectious Diseases (NIAID), needing urgent research and development of countermeasures because of the high public health risk it poses. The recent cases of MARV in West Africa underscore the substantial outbreak potential of this virus. The potential for cross-border spread, as had occurred during the 2014-2016 Ebola virus outbreak, illustrates the critical need for MARV vaccines. To support regulatory approval of the chimpanzee adenovirus 3 (ChAd3)-MARV vaccine that has completed phase 1 trials, we showed that the nonreplicating ChAd3 vector, which has a demonstrated safety profile in humans, protected against a uniformly lethal challenge with MARV/Ang. Protective immunity was achieved within 7 days of vaccination and was maintained through 1 year after vaccination. Antigen-specific antibodies were an immune correlate of protection in the acute challenge model, and their concentration was predictive of protection. These results demonstrate that a single-shot ChAd3-MARV vaccine generated a protective immune response that was both rapid and durable with an immune correlate of protection that will support advanced clinical development.

Early and Long-Term HIV-1 Immunogenicity Induced in Macaques by the Combined Administration of DNA, NYVAC and Env Protein-Based Vaccine Candidates: The AUP512 Study.

To control HIV infection there is a need for vaccines to induce broad, potent and long-term B and T cell immune responses. With the objective to accelerate and maintain the induction of substantial levels of HIV-1 Env-specific antibodies and, at the same time, to enhance balanced CD4 and CD8 T cell responses, we evaluated the effect of concurrent administration of MF59-adjuvanted Env protein together with DNA or NYVAC vectors at priming to establish if early administration of Env leads to early induction of antibody responses. The primary goal was to assess the immunogenicity endpoint at week 26. Secondary endpoints were (i) to determine the quality of responses with regard to RV144 correlates of protection and (ii) to explore a potential impact of two late boosts. In this study, five different prime/boost vaccination regimens were tested in rhesus macaques. Animals received priming immunizations with either NYVAC or DNA alone or in combination with Env protein, followed by NYVAC + protein or DNA + protein boosts. All regimens induced broad, polyfunctional and well-balanced CD4 and CD8 T cell responses, with DNA-primed regimens eliciting higher response rates and magnitudes than NYVAC-primed regimens. Very high plasma binding IgG titers including V1/V2 specific antibodies, modest antibody-dependent cellular cytotoxicity (ADCC) and moderate neutralization activity were observed. Of note, early administration of the MF59-adjuvanted Env protein in parallel with DNA priming leads to more rapid elicitation of humoral responses, without negatively affecting the cellular responses, while responses were rapidly boosted after repeated immunizations, indicating the induction of a robust memory response. In conclusion, our findings support the use of the Env protein component during priming in the context of an heterologous immunization regimen with a DNA and/or NYVAC vector as an optimized immunization protocol against HIV infection.

Potent anti-viral activity of a trispecific HIV neutralizing antibody in SHIV-infected monkeys.

Broadly neutralizing antibodies (bNAbs) represent an alternative to drug therapy for the treatment of HIV-1 infection. Immunotherapy with single bNAbs often leads to emergence of escape variants, suggesting a potential benefit of combination bNAb therapy. Here, a trispecific bNAb reduces viremia 100- to 1000-fold in viremic SHIV-infected macaques. After treatment discontinuation, viremia rebounds transiently and returns to low levels, through CD8-mediated immune control. These viruses remain sensitive to the trispecific antibody, despite loss of sensitivity to one of the parental bNAbs. Similarly, the trispecific bNAb suppresses the emergence of resistance in viruses derived from HIV-1-infected subjects, in contrast to parental bNAbs. Trispecific HIV-1 neutralizing antibodies, therefore, mediate potent antiviral activity in vivo and may minimize the potential for immune escape.

Vaccine nanodiscs plus polyICLC elicit robust CD8+ T cell responses in mice and non-human primates.

Conventional cancer vaccines based on soluble vaccines and traditional adjuvants have produced suboptimal therapeutic efficacy in clinical trials. Thus, there is an urgent need for vaccine technologies that can generate potent T cell responses with strong anti-tumor efficacy. We have previously reported the development of synthetic high-density protein (sHDL) nanodiscs for efficient lymph node (LN)-targeted co-delivery of antigen peptides and CpG oligonucleotides (a Toll-like receptor-9 agonist). Here, we performed a comparative study in mice and non-human primates (NHPs) to identify an ideal vaccine platform for induction of CD8+ T cell responses. In particular, we compared the efficacy of CpG class B, CpG class C, and polyICLC (a synthetic double-stranded RNA analog, a TLR-3 agonist), each formulated with antigen-carrying sHDL nanodiscs. Here, we report that sHDL-Ag admixed with polyICLC elicited robust Ag-specific CD8+ T cell responses in mice, and when used in combination with α-PD-1 immune checkpoint inhibitor, sHDL-Ag + polyICLC eliminated large established (~100 mm3) MC-38 tumors in mice. Moreover, sHDL-Gag + polyICLC induced robust Simian immunodeficiency virus Gag-specific, polyfunctional CD8+ T cell responses in rhesus macaques and could further amplify the efficacy of recombinant adenovirus-based vaccine. Notably, while both sHDL-Ag-CpG-B and sHDL-Ag-CpG-C generated strong Ag-specific CD8+ T cell responses in mice, their results were mixed in NHPs. Overall, sHDL combined with polyICLC offers a strong platform to induce CD8+ T cells for vaccine applications.

Fusion peptide priming reduces immune responses to HIV-1 envelope trimer base.

Soluble "SOSIP"-stabilized envelope (Env) trimers are promising HIV-vaccine immunogens. However, they induce high-titer responses against the glycan-free trimer base, which is occluded on native virions. To delineate the effect on base responses of priming with immunogens targeting the fusion peptide (FP) site of vulnerability, here, we quantify the prevalence of trimer-base antibody responses in 49 non-human primates immunized with various SOSIP-stabilized Env trimers and FP-carrier conjugates. Trimer-base responses account for ∼90% of the overall trimer response in animals immunized with trimer only, ∼70% in animals immunized with a cocktail of SOSIP trimer and FP conjugate, and ∼30% in animals primed with FP conjugates before trimer immunization. Notably, neutralization breadth in FP-conjugate-primed animals correlates inversely with trimer-base responses. Our data provide methods to quantify the prevalence of trimer-base responses and reveal that FP-conjugate priming, either alone or as part of a cocktail, can reduce the trimer-base response and improve the neutralization outcome.

Measurement of leukocyte trafficking kinetics in macaques by serial intravascular staining.

Leukocyte trafficking enables detection of pathogens, immune responses, and immune memory. Dysregulation of leukocyte trafficking is often found in disease, highlighting its important role in homeostasis and the immune response. Whereas some of the molecular mechanisms mediating leukocyte trafficking are understood, little is known about the regulation of trafficking, including trafficking kinetics and its impact on immune homeostasis. We developed a method of serial intravascular staining (SIVS) to measure trafficking kinetics in nonhuman primates using infusions of fluorescently labeled antibodies to label circulating leukocytes. Because antibody infusions labeled only leukocytes in the blood, cells were "barcoded" according to their location at the time of each infusion, providing positional histories that could be used to infer trafficking kinetics. We used SIVS and multiparameter flow cytometry to quantitate cellular trafficking into lymphoid tissues of healthy animals at homeostasis and to identify perivascular cells that could be unique to nonlymphoid organs. To investigate how these parameters could be influenced during disease, SIVS was used to quantify lymphocyte trafficking in macaques infected with the bacterial pathogen Mycobacterium tuberculosis and to enumerate intravascular leukocytes in lung granulomas. We showed that whereas most cells in lung granulomas were localized there for more than 24 hours, granulomas were dynamic with a slow continual cellular influx, the rate of which predicted clearance of M. tuberculosis from the granulomas. SIVS, in combination with intracellular staining and multiparametric flow cytometry, is a powerful method to quantify the kinetics of leukocyte trafficking in nonhuman primates in vivo.

Immune Monitoring Reveals Fusion Peptide Priming to Imprint Cross-Clade HIV-Neutralizing Responses with a Characteristic Early B Cell Signature.

The HIV fusion peptide (FP) is a promising vaccine target. FP-directed monoclonal antibodies from vaccinated macaques have been identified that neutralize up to ∼60% of HIV strains; these vaccinations, however, have involved ∼1 year with an extended neutralization-eclipse phase without measurable serum neutralization. Here, in 32 macaques, we test seven vaccination regimens, each comprising multiple immunizations of FP-carrier conjugates and HIV envelope (Env) trimers. Comparisons of vaccine regimens reveal FP-carrier conjugates to imprint cross-clade neutralizing responses and a cocktail of FP conjugate and Env trimer to elicit the earliest broad responses. We identify a signature, appearing as early as week 6 and involving the frequency of B cells recognizing both FP and Env trimer, predictive of vaccine-elicited breadth ∼1 year later. Immune monitoring of B cells in response to vaccination can thus enable vaccine insights even in the absence of serum neutralization, here identifying FP imprinting, cocktail approach, and early signature as means to improve FP-directed vaccine responses.

Blocking α4ß7 integrin binding to SIV does not improve virologic control.

A study in nonhuman primates reported that infusions of an antibody against α4ß7 integrin, in combination with antiretroviral therapy, showed consistent, durable control of simian immunodeficiency virus (SIV) in rhesus macaques. The antibody used has pleiotropic effects, so we set out to gain insight into the underlying mechanism by comparing this treatment to treatment with non-neutralizing monoclonal antibodies against the SIV envelope glycoprotein that only block α4ß7 binding to SIV Env but have no other host-directed effects. Similar to the initial study, we used an attenuated strain of SIV containing a stop codon in nef. The study used 30 macaques that all began antiretroviral therapy and then were divided into five groups to receive different antibody treatments. Unlike the published report, we found no sustained virologic control by these treatments in vivo.

Antibody Lineages with Vaccine-Induced Antigen-Binding Hotspots Develop Broad HIV Neutralization.

The vaccine-mediated elicitation of antibodies (Abs) capable of neutralizing diverse HIV-1 strains has been a long-standing goal. To understand how broadly neutralizing antibodies (bNAbs) can be elicited, we identified, characterized, and tracked five neutralizing Ab lineages targeting the HIV-1-fusion peptide (FP) in vaccinated macaques over time. Genetic and structural analyses revealed two of these lineages to belong to a reproducible class capable of neutralizing up to 59% of 208 diverse viral strains. B cell analysis indicated each of the five lineages to have been initiated and expanded by FP-carrier priming, with envelope (Env)-trimer boosts inducing cross-reactive neutralization. These Abs had binding-energy hotspots focused on FP, whereas several FP-directed Abs induced by immunization with Env trimer-only were less FP-focused and less broadly neutralizing. Priming with a conserved subregion, such as FP, can thus induce Abs with binding-energy hotspots coincident with the target subregion and capable of broad neutralization.

Structure-based design of a quadrivalent fusion glycoprotein vaccine for human parainfluenza virus types 1-4.

Parainfluenza virus types 1-4 (PIV1-4) are highly infectious human pathogens, of which PIV3 is most commonly responsible for severe respiratory illness in newborns, elderly, and immunocompromised individuals. To obtain a vaccine effective against all four PIV types, we engineered mutations in each of the four PIV fusion (F) glycoproteins to stabilize their metastable prefusion states, as such stabilization had previously enabled the elicitation of high-titer neutralizing antibodies against the related respiratory syncytial virus. A cryoelectron microscopy structure of an engineered PIV3 F prefusion-stabilized trimer, bound to the prefusion-specific antibody PIA174, revealed atomic-level details for how introduced mutations improved stability as well as how a single PIA174 antibody recognized the trimeric apex of prefusion PIV3 F. Nine combinations of six newly identified disulfides and two cavity-filling mutations stabilized the prefusion PIV3 F immunogens and induced 200- to 500-fold higher neutralizing titers in mice than were elicited by PIV3 F in the postfusion conformation. For PIV1, PIV2, and PIV4, we also obtained stabilized prefusion Fs, for which prefusion versus postfusion titers were 2- to 20-fold higher. Elicited murine responses were PIV type-specific, with little cross-neutralization of other PIVs. In nonhuman primates (NHPs), quadrivalent immunization with prefusion-stabilized Fs from PIV1-4 consistently induced potent neutralizing responses against all four PIVs. For PIV3, the average elicited NHP titer from the quadrivalent immunization was more than fivefold higher than any titer observed in a cohort of over 100 human adults, highlighting the ability of a prefusion-stabilized immunogen to elicit especially potent neutralization.

Epitope-based vaccine design yields fusion peptide-directed antibodies that neutralize diverse strains of HIV-1.

A central goal of HIV-1 vaccine research is the elicitation of antibodies capable of neutralizing diverse primary isolates of HIV-1. Here we show that focusing the immune response to exposed N-terminal residues of the fusion peptide, a critical component of the viral entry machinery and the epitope of antibodies elicited by HIV-1 infection, through immunization with fusion peptide-coupled carriers and prefusion stabilized envelope trimers, induces cross-clade neutralizing responses. In mice, these immunogens elicited monoclonal antibodies capable of neutralizing up to 31% of a cross-clade panel of 208 HIV-1 strains. Crystal and cryoelectron microscopy structures of these antibodies revealed fusion peptide conformational diversity as a molecular explanation for the cross-clade neutralization. Immunization of guinea pigs and rhesus macaques induced similarly broad fusion peptide-directed neutralizing responses, suggesting translatability. The N terminus of the HIV-1 fusion peptide is thus a promising target of vaccine efforts aimed at eliciting broadly neutralizing antibodies.

Early antibody therapy can induce long-lasting immunity to SHIV.

Highly potent and broadly neutralizing anti-HIV-1 antibodies (bNAbs) have been used to prevent and treat lentivirus infections in humanized mice, macaques, and humans. In immunotherapy experiments, administration of bNAbs to chronically infected animals transiently suppresses virus replication, which invariably returns to pre-treatment levels and results in progression to clinical disease. Here we show that early administration of bNAbs in a macaque simian/human immunodeficiency virus (SHIV) model is associated with very low levels of persistent viraemia, which leads to the establishment of T-cell immunity and resultant long-term infection control. Animals challenged with SHIVAD8-EO by mucosal or intravenous routes received a single 2-week course of two potent passively transferred bNAbs (3BNC117 and 10-1074 (refs 13, 14)). Viraemia remained undetectable for 56-177 days, depending on bNAb half-life in vivo. Moreover, in the 13 treated monkeys, plasma virus loads subsequently declined to undetectable levels in 6 controller macaques. Four additional animals maintained their counts of T cells carrying the CD4 antigen (CD4+) and very low levels of viraemia persisted for over 2 years. The frequency of cells carrying replication-competent virus was less than 1 per 106 circulating CD4+ T cells in the six controller macaques. Infusion of a T-cell-depleting anti-CD8ß monoclonal antibody to the controller animals led to a specific decline in levels of CD8+ T cells and the rapid reappearance of plasma viraemia. In contrast, macaques treated for 15 weeks with combination anti-retroviral therapy, beginning on day 3 after infection, experienced sustained rebound plasma viraemia when treatment was interrupted. Our results show that passive immunotherapy during acute SHIV infection differs from combination anti-retroviral therapy in that it facilitates the emergence of potent CD8+ T-cell immunity able to durably suppress virus replication.

HIV/AIDS Vaccine Candidates Based on Replication-Competent Recombinant Poxvirus NYVAC-C-KC Expressing Trimeric gp140 and Gag-Derived Virus-Like Particles or Lacking the Viral Molecule B19 That Inhibits Type I Interferon Activate Relevant HIV-1-Specific B and T Cell Immune Functions in Nonhuman Primates.

The nonreplicating attenuated poxvirus vector NYVAC expressing clade C(CN54) HIV-1 Env(gp120) and Gag-Pol-Nef antigens (NYVAC-C) showed limited immunogenicity in phase I clinical trials. To enhance the capacity of the NYVAC vector to trigger broad humoral responses and a more balanced activation of CD4+ and CD8+ T cells, here we compared the HIV-1-specific immunogenicity elicited in nonhuman primates immunized with two replicating NYVAC vectors that have been modified by the insertion of the K1L and C7L vaccinia virus host range genes and express the clade C(ZM96) trimeric HIV-1 gp140 protein or a Gag(ZM96)-Pol-Nef(CN54) polyprotein as Gag-derived virus-like particles (termed NYVAC-C-KC). Additionally, one NYVAC-C-KC vector was generated by deleting the viral gene B19R, an inhibitor of the type I interferon response (NYVAC-C-KC-ΔB19R). An immunization protocol mimicking that of the RV144 phase III clinical trial was used. Two groups of macaques received two doses of the corresponding NYVAC-C-KC vectors (weeks 0 and 4) and booster doses with NYVAC-C-KC vectors plus the clade C HIV-1 gp120 protein (weeks 12 and 24). The two replicating NYVAC-C-KC vectors induced enhanced and similar HIV-1-specific CD4+ and CD8+ T cell responses, similar levels of binding IgG antibodies, low levels of IgA antibodies, and high levels of antibody-dependent cellular cytotoxicity responses and HIV-1-neutralizing antibodies. Small differences within the NYVAC-C-KC-ΔB19R group were seen in the magnitude of CD4+ and CD8+ T cells, the induction of some cytokines, and the neutralization of some HIV-1 isolates. Thus, replication-competent NYVAC-C-KC vectors acquired relevant immunological properties as vaccine candidates against HIV/AIDS, and the viral B19 molecule exerts some control of immune functions.IMPORTANCE It is of special importance to find a safe and effective HIV/AIDS vaccine that can induce strong and broad T cell and humoral immune responses correlating with HIV-1 protection. Here we developed novel replicating poxvirus NYVAC-based HIV/AIDS vaccine candidates expressing clade C HIV-1 antigens, with one of them lacking the vaccinia virus B19 protein, an inhibitor of the type I interferon response. Immunization of nonhuman primates with these novel NYVAC-C-KC vectors and the protein component gp120 elicited high levels of T cell and humoral immune responses, with the vector containing a deletion in B19R inducing a trend toward a higher magnitude of CD4+ and CD8+ T cell responses and neutralization of some HIV-1 strains. These poxvirus vectors could be considered HIV/AIDS vaccine candidates based on their activation of potential immune correlates of protection.