Immunization with V987H-stabilized Spike glycoprotein protects K18-hACE2 mice and golden Syrian hamsters upon SARS-CoV-2 infection.

Safe and effective severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines are crucial to fight against the coronavirus disease 2019 pandemic. Most vaccines are based on a mutated version of the Spike glycoprotein [K986P/V987P (S-2P)] with improved stability, yield and immunogenicity. However, S-2P is still produced at low levels. Here, we describe the V987H mutation that increases by two-fold the production of the recombinant Spike and the exposure of the receptor binding domain (RBD). S-V987H immunogenicity is similar to S-2P in mice and golden Syrian hamsters (GSH), and superior to a monomeric RBD. S-V987H immunization confer full protection against severe disease in K18-hACE2 mice and GSH upon SARS-CoV-2 challenge (D614G or B.1.351 variants). Furthermore, S-V987H immunized K18-hACE2 mice show a faster tissue viral clearance than RBD- or S-2P-vaccinated animals challenged with D614G, B.1.351 or Omicron BQ1.1 variants. Thus, S-V987H protein might be considered for future SARS-CoV-2 vaccines development.

Immunisation efficacy of a stabilised SARS-CoV-2 spike glycoprotein in two geriatric animal models.

Age is associated with reduced efficacy of vaccines and linked to higher risk of severe COVID-19. Here we determined the impact of ageing on the efficacy of a SARS-CoV-2 vaccine based on a stabilised Spike glycoprotein (S-29) that had previously shown high efficacy in young animals. Thirteen to 18-month-old golden Syrian hamsters (GSH) and 22-23-month-old K18-hCAE2 mice were immunised twice with S-29 protein in AddaVaxTM adjuvant. GSH were intranasally inoculated with SARS-CoV-2 either two weeks or four months after the booster dose, while all K18-hACE2 mice were intranasally inoculated two weeks after the second immunisation. Body weight and clinical signs were recorded daily post-inoculation. Lesions and viral load were investigated in different target tissues. Immunisation induced seroconversion and production of neutralising antibodies; however, animals were only partially protected from weight loss. We observed a significant reduction in the amount of viral RNA and a faster viral protein clearance in the tissues of immunized animals. Infectious particles showed a faster decay in vaccinated animals while tissue lesion development was not altered. In GSH, the shortest interval between immunisation and inoculation reduced RNA levels in the lungs, while the longest interval was equally effective in reducing RNA in nasal turbinates; viral nucleoprotein amount decreased in both tissues. In mice, immunisation was able to improve the survival of infected animals. Despite the high protection shown in young animals, S-29 efficacy was reduced in the geriatric population. Our research highlights the importance of testing vaccine efficacy in older animals as part of preclinical vaccine evaluation.

Novel Spike-stabilized trimers with improved production protect K18-hACE2 mice and golden Syrian hamsters from the highly pathogenic SARS-CoV-2 Beta variant.

Most COVID-19 vaccines are based on the SARS-CoV-2 Spike glycoprotein (S) or their subunits. However, S shows some structural instability that limits its immunogenicity and production, hampering the development of recombinant S-based vaccines. The introduction of the K986P and V987P (S-2P) mutations increases the production and immunogenicity of the recombinant S trimer, suggesting that these two parameters are related. Nevertheless, S-2P still shows some molecular instability and it is produced with low yield. Here we described a novel set of mutations identified by molecular modeling and located in the S2 region of the S-2P that increase its production up to five-fold. Besides their immunogenicity, the efficacy of two representative S-2P-based mutants, S-29 and S-21, protecting from a heterologous SARS-CoV-2 Beta variant challenge was assayed in K18-hACE2 mice (an animal model of severe SARS-CoV-2 disease) and golden Syrian hamsters (GSH) (a moderate disease model). S-21 induced higher level of WH1 and Delta variants neutralizing antibodies than S-2P in K18-hACE2 mice three days after challenge. Viral load in nasal turbinate and oropharyngeal samples were reduced in S-21 and S-29 vaccinated mice. Despite that, only the S-29 protein protected 100% of K18-hACE2 mice from severe disease. When GSH were analyzed, all immunized animals were protected from disease development irrespectively of the immunogen they received. Therefore, the higher yield of S-29, as well as its improved immunogenicity and efficacy protecting from the highly pathogenic SARS-CoV-2 Beta variant, pinpoint the S-29 mutant as an alternative to the S-2P protein for future SARS-CoV-2 vaccine development.

Syphilis vaccine: challenges, controversies and opportunities.

Syphilis is a sexually or vertically (mother to fetus) transmitted disease caused by the infection of Treponema pallidum subspecie pallidum (TPA). The incidence of syphilis has increased over the past years despite the fact that this bacterium is an obligate human pathogen, the infection route is well known, and the disease can be successfully treated with penicillin. As complementary measures to preventive campaigns and early treatment of infected individuals, development of a syphilis vaccine may be crucial for controlling disease spread and/or severity, particularly in countries where the effectiveness of the aforementioned measures is limited. In the last century, several vaccine prototypes have been tested in preclinical studies, mainly in rabbits. While none of them provided protection against infection, some prototypes prevented bacteria from disseminating to distal organs, attenuated lesion development, and accelerated their healing. In spite of these promising results, there is still some controversy regarding the identification of vaccine candidates and the characteristics of a syphilis-protective immune response. In this review, we describe what is known about TPA immune response, and the main mechanisms used by this pathogen to evade it. Moreover, we emphasize the importance of integrating this knowledge, in conjunction with the characterization of outer membrane proteins (OMPs), to expedite the development of a syphilis vaccine that can protect against TPA infection.

Therapeutic neutralizing monoclonal antibody administration protects against lethal yellow fever virus infection.

Yellow fever virus (YFV) is a reemerging global health threat, driven by several factors, including increased spread of the mosquito vector and rapid urbanization. Although a prophylactic vaccine exists, vaccine hesitancy, supply deficits, and distribution difficulties leave specific populations at risk of severe YFV disease, as evidenced by recent outbreaks in South America. To establish a treatment for patients with severe YFV infection, we tested 37 YFV-specific monoclonal antibodies isolated from vaccinated humans and identified two capable of potently neutralizing multiple pathogenic primary YFV isolates. Using both hamster and nonhuman primate models of lethal YFV infection, we demonstrate that a single administration of either of these two potently neutralizing antibodies during acute infection fully controlled viremia and prevented severe disease and death in treated animals. Given the potential severity of YFV-induced disease, our results show that these antibodies could be effective in saving lives and fill a much-needed void in managing YFV cases during outbreaks.

Kinetics of immune responses elicited after three mRNA COVID-19 vaccine doses in predominantly antibody-deficient individuals.

Mass vaccination campaigns reduced COVID-19 incidence and severity. Here, we evaluated the immune responses developed in SARS-CoV-2-uninfected patients with predominantly antibody-deficiencies (PAD) after three mRNA-1273 vaccine doses. PAD patients were classified based on their immunodeficiency: unclassified primary antibody-deficiency (unPAD, n = 9), common variable immunodeficiency (CVID, n = 12), combined immunodeficiency (CID, n = 1), and thymoma with immunodeficiency (TID, n = 1). unPAD patients and healthy controls (HCs, n = 10) developed similar vaccine-induced humoral responses after two doses. However, CVID patients showed reduced binding and neutralizing titers compared to HCs. Of interest, these PAD groups showed lower levels of Spike-specific IFN-γ-producing cells. CVID individuals also presented diminished CD8+T cells. CID and TID patients developed cellular but not humoral responses. Although the third vaccine dose boosted humoral responses in most PAD patients, it had limited effect on expanding cellular immunity. Vaccine-induced immune responses in PAD individuals are heterogeneous, and should be immunomonitored to define a personalized therapeutic strategies.

Limited Humoral and Specific T-Cell Responses After SARS-CoV-2 Vaccination in PWH With Poor Immune Reconstitution.

BACKGROUND: We analyzed humoral and cellular immune responses induced by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) messenger RNA (mRNA) vaccines in people with human immunodeficiency virus (HIV; PWH) who had CD4+ T-cell counts <200/µL (HIV<200 group). METHODS: This prospective cohort study included 58 PWH in the HIV<200 group, 36 with CD4+ T-cell counts >500/µL (HIV>500 group), and 33 HIV-1-negative controls (control group). Antibodies against the SARS-CoV-2 spike protein (anti-S immunoglobulin [Ig] G) and the receptor-binding domain (anti-RBD IgG) were quantified before and 4 weeks after the first and the second doses of BNT162b2 or mRNA-1273 (at week 8). Viral neutralization activity and T-cell responses were also determined. RESULTS: At week 8, anti-S/anti-RBD IgG responses increased in all groups (P < .001). Median (interquartile range) anti-S and anti-RBD IgG levels at week 8 were 153.6 (26.4-654.9) and 171.9 (61.8-425.8) binding antibody units (BAU)/mL, respectively, in the HIV<200 group, compared with 245.6 (145-824) and 555.8 (166.4-1751) BAU/mL in the HIV>500 group and 274.7 (193.7-680.4) and 281.6 (181-831.8) BAU/mL in controls (P < .05). Neutralizing capacity and specific T-cell immune responses were absent or reduced in 33% of those in the HIV<200 group, compared with 3.7% in the HIV>500 group (P < .01). CONCLUSIONS: One-third of PWH with CD4+ T-cell counts <200/µL show low anti-S/anti-RBD IgG levels, reduced in vitro neutralization activity against SARS-CoV-2, and no vaccine-induced T cells after receiving coronavirus disease 2019 mRNA vaccines.

Molecular insights into antibody-mediated protection against the prototypic simian immunodeficiency virus.

SIVmac239 infection of macaques is a favored model of human HIV infection. However, the SIVmac239 envelope (Env) trimer structure, glycan occupancy, and the targets and ability of neutralizing antibodies (nAbs) to protect against SIVmac239 remain unknown. Here, we report the isolation of SIVmac239 nAbs that recognize a glycan hole and the V1/V4 loop. A high-resolution structure of a SIVmac239 Env trimer-nAb complex shows many similarities to HIV and SIVcpz Envs, but with distinct V4 features and an extended V1 loop. Moreover, SIVmac239 Env has a higher glycan shield density than HIV Env that may contribute to poor or delayed nAb responses in SIVmac239-infected macaques. Passive transfer of a nAb protects macaques from repeated intravenous SIVmac239 challenge at serum titers comparable to those described for protection of humans against HIV infection. Our results provide structural insights for vaccine design and shed light on antibody-mediated protection in the SIV model.

T-Follicular-Like CD8+ T Cell Responses in Chronic HIV Infection Are Associated With Virus Control and Antibody Isotype Switching to IgG.

T cell responses are considered critical for the in vivo control of HIV, but the contribution of different T cell subsets to this control remains unclear. Using a boosted flow cytometric approach that is able to differentiate CD4+ and CD8+ T cell Th1/Tc1, Th2/Tc2, Th17/Tc17, Treg and Tfh/Tfc-like HIV-specific T cell populations, we identified CD8+ Tfc responses that were related to HIV plasma viral loads and associated with rate of antibody isotype class switching to IgG. This favorable balance towards IgG responses positively correlated with increased virus neutralization, higher avidity of neutralizing antibodies and more potent antibody-dependent cell cytotoxicity (ADCC) in PBMCs from HIV controllers compared to non-controllers. Our results identified the CD8+ Tfc-like T-cell response as a component of effective virus control which could possibly be exploited therapeutically.

Immunoescape of HIV-1 in Env-EL9 CD8 + T cell response restricted by HLA-B*14:02 in a Non progressor who lost twenty-seven years of HIV-1 control.

BACKGROUND: Long-Term Non-Progressors (LTNPs) are untreated Human Immunodeficiency virus type 1 (HIV-1) infected individuals able to control disease progression for prolonged periods. However, the LTNPs status is temporary, as viral load increases followed by decreases in CD4 + T-cell counts. Control of HIV-1 infection in LTNPs viremic controllers, have been associated with effective immunodominant HIV-1 Gag-CD8 + T-cell responses restricted by protective HLA-B alleles. Individuals carrying HLA-B*14:02 control HIV-1 infection is related to an immunodominant Env-CD8 + T-cell response. Limited data are available on the contribution of HLA-B*14:02 CD8 + T -cells in LTNPs. RESULTS: In this study, we performed a virological and immunological detailed analysis of an HLA-B*14:02 LNTP individual that lost viral control (LVC) 27 years after HIV-1 diagnosis. We analysed viral evolution and immune escape in HLA-B*14:02 restricted CD8 + T -cell epitopes and identified viral evolution at the Env-EL9 epitope selecting the L592R mutation. By IFN-γ ELISpot and immune phenotype, we characterized HLA- B*14:02 HIV-1 CD8 + T cell responses targeting, Gag-DA9 and Env-EL9 epitopes before and after LVC. We observed an immunodominant response against the Env-EL9 epitope and a decreased of the CD8 T + cell response over time with LVC. Loss of Env-EL9 responses was concomitant with selecting K588R + L592R mutations at Env-EL9. Finally, we evaluated the impact of Env-EL9 escape mutations on HIV-1 infectivity and Env protein structure. The K588R + L592R escape variant was directly related to HIV-1 increase replicative capacity and stability of Env at the LVC. CONCLUSIONS: These findings support the contribution of immunodominant Env-EL9 CD8 + T-cell responses and the imposition of immune escape variants with higher replicative capacity associated with LVC in this LNTP. These data highlight the importance of Env-EL9 specific-CD8 + T-cell responses restricted by the HLA-B*14:02 and brings new insights into understanding long-term HIV-1 control mediated by Env mediated CD8 + T-cell responses.

Genotype-specific features reduce the susceptibility of South American yellow fever virus strains to vaccine-induced antibodies.

The resurgence of yellow fever in South America has prompted vaccination against the etiologic agent, yellow fever virus (YFV). Current vaccines are based on a live-attenuated YF-17D virus derived from a virulent African isolate. The capacity of these vaccines to induce neutralizing antibodies against the vaccine strain is used as a surrogate for protection. However, the sensitivity of genetically distinct South American strains to vaccine-induced antibodies is unknown. We show that antiviral potency of the polyclonal antibody response in vaccinees is attenuated against an emergent Brazilian strain. This reduction was attributable to amino acid changes at two sites in central domain II of the glycoprotein E, including multiple changes at the domain I-domain II hinge, which are unique to and shared among most South American YFV strains. Our findings call for a reevaluation of current approaches to YFV immunological surveillance in South America and suggest approaches for updating vaccines.

Non-neutralizing Antibodies May Contribute to Suppression of SIVmac239 Viremia in Indian Rhesus Macaques.

The antiviral properties of broadly neutralizing antibodies against HIV are well-documented but no vaccine is currently able to elicit protective titers of these responses in primates. While current vaccine modalities can readily induce non-neutralizing antibodies against simian immunodeficiency virus (SIV) and HIV, the ability of these responses to restrict lentivirus transmission and replication remains controversial. Here, we investigated the antiviral properties of non-neutralizing antibodies in a group of Indian rhesus macaques (RMs) that were vaccinated with vif, rev, tat, nef, and env, as part of a previous study conducted by our group. These animals manifested rapid and durable control of viral replication to below detection limits shortly after SIVmac239 infection. Although these animals had no serological neutralizing activity against SIVmac239 prior to infection, their pre-challenge titers of Env-binding antibodies correlated with control of viral replication. To assess the contribution of anti-Env humoral immune responses to virologic control in two of these animals, we transiently depleted their circulating antibodies via extracorporeal plasma immunoadsorption and inhibition of IgG recycling through antibody-mediated blockade of the neonatal Fc receptor. These procedures reduced Ig serum concentrations by up to 80% and temporarily induced SIVmac239 replication in these animals. Next, we transferred purified total Ig from the rapid controllers into six vaccinated RMs one day before intrarectal challenge with SIVmac239. Although recipients of the hyperimmune anti-SIV Ig fraction were not protected from infection, their peak and chronic phase viral loads were significantly lower than those in concurrent unvaccinated control animals. Together, our results suggest that non-neutralizing Abs may play a role in the suppression of SIVmac239 viremia.

Rectal Acquisition of Simian Immunodeficiency Virus (SIV) SIVmac239 Infection despite Vaccine-Induced Immune Responses against the Entire SIV Proteome.

Given the complex biology of human immunodeficiency virus (HIV) and its remarkable capacity to evade host immune responses, HIV vaccine efficacy may benefit from the induction of both humoral and cellular immune responses of maximal breadth, potency, and longevity. Guided by this rationale, we set out to develop an immunization protocol aimed at maximizing the induction of anti-Envelope (anti-Env) antibodies and CD8+ T cells targeting non-Env epitopes in rhesus macaques (RMs). Our approach was to deliver the entire simian immunodeficiency virus (SIV) proteome by serial vaccinations. To that end, 12 RMs were vaccinated over 81 weeks with DNA, modified vaccinia Ankara (MVA), vesicular stomatitis virus (VSV), adenovirus type 5 (Ad5), rhesus monkey rhadinovirus (RRV), and DNA again. Both the RRV and the final DNA boosters delivered a near-full-length SIVmac239 genome capable of assembling noninfectious SIV particles and inducing T-cell responses against all nine SIV proteins. Compared to previous SIV vaccine trials, the present DNA-MVA-VSV-Ad5-RRV-DNA regimen resulted in comparable levels of Env-binding antibodies and SIV-specific CD8+ T-cells. Interestingly, one vaccinee developed low titers of neutralizing antibodies (NAbs) against SIVmac239, a tier 3 virus. Following repeated intrarectal marginal-dose challenges with SIVmac239, vaccinees were not protected from SIV acquisition but manifested partial control of viremia. Strikingly, the animal with the low-titer vaccine-induced anti-SIVmac239 NAb response acquired infection after the first SIVmac239 exposure. Collectively, these results highlight the difficulties in eliciting protective immunity against immunodeficiency virus infection.IMPORTANCE Our results are relevant to HIV vaccine development efforts because they suggest that increasing the number of booster immunizations or delivering additional viral antigens may not necessarily improve vaccine efficacy against immunodeficiency virus infection.

An Automated Fluorescence-Based Method to Isolate Bone Marrow-Derived Plasma Cells from Rhesus Macaques Using SIVmac239 SOSIP.664.

Simian immunodeficiency virus (SIV) infection of Indian rhesus macaques (RMs) is one of the best-characterized animal models for human immunodeficiency virus (HIV) infection. Monoclonal antibodies (mAbs) have shown promise for prevention and treatment of HIV infection. However, it has been difficult to isolate mAbs that potently neutralize the highly pathogenic SIVmac239 strain. This has been largely due to the low frequency of circulating B cells encoding neutralizing Abs. Here we describe a novel technique to isolate mAbs directly from bone marrow-derived, Ab-secreting plasma cells. We employed an automated micromanipulator to isolate single SIVmac239 SOSIP.664-specific plasma cells from the bone marrow of a SIVmac239-infected RM with serum neutralization titers against SIVmac239. After picking plasma cells, we obtained 44 paired Ab sequences. Ten of these mAbs were SIV specific. Although none of these mAbs neutralized SIVmac239, three mAbs completely neutralized the related SIVmac316 strain. The majority of these mAbs bound to primary rhesus CD4+ T cells infected with SIVmac239 and induced Ab-dependent cellular cytotoxicity. This method is a first step in successful isolation of antigen-specific bone marrow-derived plasma cells from RMs.

Rhesus Cytomegalovirus-Specific CD8+ Cytotoxic T Lymphocytes Do Not Become Functionally Exhausted in Chronic SIVmac239 Infection.

CD8+ cytotoxic T lymphocytes (CTLs) exert potent antiviral activity after HIV/SIV infection. However, efforts to harness the antiviral efficacy of CTLs for HIV/SIV prophylaxis and therapy have been severely hindered by two major problems: viral escape and exhaustion. By contrast, CTLs directed against human cytomegalovirus (HCMV), a ubiquitous chronic herpesvirus, seldom select for escape mutations and remain functional and refractory to exhaustion during chronic HCMV and HIV infection. Recently, attempts have been made to retarget HCMV-specific CTLs for cancer immunotherapy. We speculate that such a strategy may also be beneficial in the context of HIV/SIV infection, facilitating CTL-mediated control of HIV/SIV replication. As a preliminary assessment of the validity of this approach, we investigated the phenotypes and functionality of rhesus CMV (RhCMV)-specific CTLs in SIVmac239-infected Indian rhesus macaques (RMs), a crucial HIV animal model system. We recently identified two immunodominant, Mamu-A∗02-restricted CTL epitopes derived from RhCMV proteins and sought to evaluate the phenotypic and functional characteristics of these CTL populations in chronic SIVmac239 infection. We analyzed and directly compared RhCMV- and SIVmac239-specific CTLs during SIVmac239 infection in a cohort of Mamu-A∗01+ and Mamu-A∗02+ RMs. CTL populations specific for at least one of the RhCMV-derived CTL epitopes were detected in ten of eleven Mamu-A∗02+ animals tested, and both populations were detected in five of these animals. Neither RhCMV-specific CTL population exhibited significant changes in frequency, memory phenotype, granzyme B expression, exhaustion marker (PD-1 and CTLA-4) expression, or polyfunctionality between pre- and chronic SIVmac239 infection timepoints. In chronic SIVmac239 infection, RhCMV-specific CTLs exhibited higher levels of granzyme B expression and polyfunctionality, and lower levels of exhaustion marker expression, than SIVmac239-specific CTLs. Additionally, compared to SIVmac239-specific CTLs, greater proportions of RhCMV-specific CTLs were of the terminally differentiated effector memory phenotype (CD28- CCR7-) during chronic SIVmac239 infection. These results suggest that, in contrast to SIVmac239-specific CTLs, RhCMV-specific CTLs maintain their phenotypes and cytolytic effector functions during chronic SIVmac239 infection, and that retargeting RhCMV-specific CTLs might be a promising SIV immunotherapeutic strategy.

Longitudinal dynamics of the human B cell response to the yellow fever 17D vaccine.

A comprehensive understanding of the development and evolution of human B cell responses induced by pathogen exposure will facilitate the design of next-generation vaccines. Here, we utilized a high-throughput single B cell cloning technology to longitudinally track the human B cell response to the yellow fever virus 17D (YFV-17D) vaccine. The early memory B cell (MBC) response was mediated by both classical immunoglobulin M (IgM) (IgM+CD27+) and switched immunoglobulin (swIg+) MBC populations; however, classical IgM MBCs waned rapidly, whereas swIg+ and atypical IgM+ and IgD+ MBCs were stable over time. Affinity maturation continued for 6 to 9 mo following vaccination, providing evidence for the persistence of germinal center activity long after the period of active viral replication in peripheral blood. Finally, a substantial fraction of the neutralizing antibody response was mediated by public clones that recognize a fusion loop-proximal antigenic site within domain II of the viral envelope glycoprotein. Overall, our findings provide a framework for understanding the dynamics and complexity of human B cell responses elicited by infection and vaccination.

Induction of Transient Virus Replication Facilitates Antigen-Independent Isolation of SIV-Specific Monoclonal Antibodies.

Structural characterization of the HIV-1 Envelope (Env) glycoprotein has facilitated the development of Env probes to isolate HIV-specific monoclonal antibodies (mAbs). However, preclinical studies have largely evaluated these virus-specific mAbs against chimeric viruses, which do not naturally infect non-human primates, in contrast to the unconstrained simian immunodeficiency virus (SIV)mac239 clone. Given the paucity of native-like reagents for the isolation of SIV-specific B cells, we examined a method to isolate SIVmac239-specific mAbs without using Env probes. We first activated virus-specific B cells by inducing viral replication after the infusion of a CD8ß-depleting mAb or withdrawal of antiretroviral therapy in SIVmac239-infected rhesus macaques. Following the rise in viremia, we observed 2- to 4-fold increases in the number of SIVmac239 Env-reactive plasmablasts in circulation. We then sorted these activated B cells and obtained 206 paired Ab sequences. After expressing 122 mAbs, we identified 14 Env-specific mAbs. While these Env-specific mAbs bound to both the SIVmac239 SOSIP.664 trimer and to infected primary rhesus CD4+ T cells, five also neutralized SIVmac316. Unfortunately, none of these mAbs neutralized SIVmac239. Our data show that this method can be used to isolate virus-specific mAbs without antigenic probes by inducing bursts of contemporary replicating viruses in vivo.

Vaccine protection against rectal acquisition of SIVmac239 in rhesus macaques.

A prophylactic vaccine against human immunodeficiency virus (HIV) remains a top priority in biomedical research. Given the failure of conventional immunization protocols to confer robust protection against HIV, new and unconventional approaches may be needed to generate protective anti-HIV immunity. Here we vaccinated rhesus macaques (RMs) with a recombinant (r)DNA prime (without any exogenous adjuvant), followed by a booster with rhesus monkey rhadinovirus (RRV)-a herpesvirus that establishes persistent infection in RMs (Group 1). Both the rDNA and rRRV vectors encoded a near-full-length simian immunodeficiency virus (SIVnfl) genome that assembles noninfectious SIV particles and expresses all nine SIV gene products. This rDNA/rRRV-SIVnfl vaccine regimen induced persistent anti-Env antibodies and CD8+ T-cell responses against the entire SIV proteome. Vaccine efficacy was assessed by repeated, marginal-dose, intrarectal challenges with SIVmac239. Encouragingly, vaccinees in Group 1 acquired SIVmac239 infection at a significantly delayed rate compared to unvaccinated controls (Group 3). In an attempt to improve upon this outcome, a separate group of rDNA/rRRV-SIVnfl-vaccinated RMs (Group 2) was treated with a cytotoxic T-lymphocyte antigen-4 (CTLA-4)-blocking monoclonal antibody during the vaccine phase and then challenged in parallel with Groups 1 and 3. Surprisingly, Group 2 was not significantly protected against SIVmac239 infection. In sum, SIVnfl vaccination can protect RMs against rigorous mucosal challenges with SIVmac239, a feat that until now had only been accomplished by live-attenuated strains of SIV. Further work is needed to identify the minimal requirements for this protection and whether SIVnfl vaccine efficacy can be improved by means other than anti-CTLA-4 adjuvant therapy.

The Frequency of Vaccine-Induced T-Cell Responses Does Not Predict the Rate of Acquisition after Repeated Intrarectal SIVmac239 Challenges in Mamu-B*08+ Rhesus Macaques.

Approximately 50% of rhesus macaques (RMs) expressing the major histocompatibility complex class I (MHC-I) allele Mamu-B*08 spontaneously control chronic-phase viremia after infection with the pathogenic simian immunodeficiency virus mac239 (SIVmac239) clone. CD8+ T-cell responses in these animals are focused on immunodominant Mamu-B*08-restricted SIV epitopes in Vif and Nef, and prophylactic vaccination with these epitopes increases the incidence of elite control in SIVmac239-infected Mamu-B*08-positive (Mamu-B*08+ ) RMs. Here we evaluated if robust vaccine-elicited CD8+ T-cell responses against Vif and Nef can prevent systemic infection in Mamu-B*08+ RMs following mucosal SIV challenges. Ten Mamu-B*08+ RMs were vaccinated with a heterologous prime/boost/boost regimen encoding Vif and Nef, while six sham-vaccinated MHC-I-matched RMs served as the controls for this experiment. Vaccine-induced CD8+ T cells against Mamu-B*08-restricted SIV epitopes reached high frequencies in blood but were present at lower levels in lymph node and gut biopsy specimens. Following repeated intrarectal challenges with SIVmac239, all control RMs became infected by the sixth SIV exposure. By comparison, four vaccinees were still uninfected after six challenges, and three of them remained aviremic after 3 or 4 additional challenges. The rate of SIV acquisition in the vaccinees was numerically lower (albeit not statistically significantly) than that in the controls. However, peak viremia was significantly reduced in infected vaccinees compared to control animals. We found no T-cell markers that distinguished vaccinees that acquired SIV infection from those that did not. Additional studies will be needed to validate these findings and determine if cellular immunity can be harnessed to prevent the establishment of productive immunodeficiency virus infection.IMPORTANCE It is generally accepted that the antiviral effects of vaccine-induced classical CD8+ T-cell responses against human immunodeficiency virus (HIV) are limited to partial reductions in viremia after the establishment of productive infection. Here we show that rhesus macaques (RMs) vaccinated with Vif and Nef acquired simian immunodeficiency virus (SIV) infection at a lower (albeit not statistically significant) rate than control RMs following repeated intrarectal challenges with a pathogenic SIV clone. All animals in the present experiment expressed the elite control-associated major histocompatibility complex class I (MHC-I) molecule Mamu-B*08 that binds immunodominant epitopes in Vif and Nef. Though preliminary, these results provide tantalizing evidence that the protective efficacy of vaccine-elicited CD8+ T cells may be greater than previously thought. Future studies should examine if vaccine-induced cellular immunity can prevent systemic viral replication in RMs that do not express MHC-I alleles associated with elite control of SIV infection.

Mamu-B*17+ Rhesus Macaques Vaccinated with env, vif, and nef Manifest Early Control of SIVmac239 Replication.

Certain major histocompatibility complex class I (MHC-I) alleles are associated with spontaneous control of viral replication in human immunodeficiency virus (HIV)-infected people and simian immunodeficiency virus (SIV)-infected rhesus macaques (RMs). These cases of "elite" control of HIV/SIV replication are often immune-mediated, thereby providing a framework for studying anti-lentiviral immunity. In this study, we examined how vaccination impacts SIV replication in RMs expressing the MHC-I allele Mamu-B*17 Approximately 21% of Mamu-B*17+ and 50% of Mamu-B*08+ RMs control chronic-phase viremia after SIVmac239 infection. Because CD8+ T cells targeting Mamu-B*08-restricted SIV epitopes have been implicated in virologic suppression in Mamu-B*08+ RMs, we investigated whether this might also be true for Mamu-B*17+ RMs. Two groups of Mamu-B*17+ RMs were vaccinated with genes encoding Mamu-B*17-restricted epitopes in Vif and Nef. These genes were delivered by themselves (group 1) or together with env (group 2). Group 3 included MHC-I-matched RMs and served as the control group. Surprisingly, the group 1 vaccine regimen had little effect on viral replication compared to group 3, suggesting that unlike Mamu-B*08+ RMs, preexisting SIV-specific CD8+ T cells alone do not facilitate long-term virologic suppression in Mamu-B*17+ RMs. Remarkably, however, 5/8 group 2 vaccinees controlled viremia to <15 viral RNA copies/ml soon after infection. No serological neutralizing activity against SIVmac239 was detected in group 2, although vaccine-elicited gp140-binding antibodies correlated inversely with nadir viral loads. Collectively, these data shed new light on the unique mechanism of elite control in Mamu-B*17+ RMs and implicate vaccine-induced, nonneutralizing anti-Env antibodies in the containment of immunodeficiency virus infection.IMPORTANCE A better understanding of the immune correlates of protection against HIV might facilitate the development of a prophylactic vaccine. Therefore, we investigated simian immunodeficiency virus (SIV) infection outcomes in rhesus macaques expressing the major histocompatibility complex class I allele Mamu-B*17 Approximately 21% of Mamu-B*17+ macaques spontaneously controlled chronic phase viremia after SIV infection, an effect that may involve CD8+ T cells targeting Mamu-B*17-restricted SIV epitopes. We vaccinated Mamu-B*17+ macaques with genes encoding immunodominant epitopes in Vif and Nef alone (group 1) or together with env (group 2). Although neither vaccine regimen prevented SIV infection, 5/8 group 2 vaccinees controlled viremia to below detection limits shortly after infection. This outcome, which was not observed in group 1, was associated with vaccine-induced, nonneutralizing Env-binding antibodies. Together, these findings suggest a limited contribution of Vif- and Nef-specific CD8+ T cells for virologic control in Mamu-B*17+ macaques and implicate anti-Env antibodies in containment of SIV infection.