Search

The contribution of host T-cell immunity and HLA class I alleles to the control of human immunodeficiency virus (HIV-1) replication in natural infection is widely recognized. We assessed whether vaccine-induced T-cell immunity, or expression of certain HLA alleles, impacted HIV-1 control after infection in the Step MRKAd5/HIV-1 gag/pol/nef study. Vaccine-induced T cells were associated with reduced plasma viremia, with subjects targeting ≥3 gag peptides presenting with half-log lower mean viral loads than subjects without Gag responses. This effect was stronger in participants infected proximal to vaccination and was independent of our observed association of HLA-B*27, –B*57 and –B*58:01 alleles with lower HIV-1 viremia. These findings support the ability of vaccine-induced T-cell responses to influence postinfection outcome and provide a rationale for the generation of T-cell responses by vaccination to reduce viremia if protection from acquisition is not achieved. Clinical trials identifier: NCT00095576. [ABSTRACT FROM PUBLISHER]

Several recent large clinical trials evaluated HIV vaccine candidates that were based on recombinant adenovirus serotype 5 (rAd-5) vectors expressing HIV-derived antigens. These vaccines primarily elicited T-cell responses, which are known to be critical for controlling HIV infection. In the current study, we present a meta-analysis of epitope mapping data from 177 participants in three clinical trials that tested two different HIV vaccines: MRKAd-5 HIV and VRC-HIVAD014-00VP. We characterized the population-level epitope responses in these trials by generating population-based epitope maps, and also designed such maps using a large cohort of 372 naturally infected individuals. We used these maps to address several questions: (1) Are vaccine-induced responses randomly distributed across vaccine inserts, or do they cluster into immunodominant epitope hotspots? (2) Are the immunodominance patterns observed for these two vaccines in three vaccine trials different from one another? (3) Do vaccine-induced hotspots overlap with epitope hotspots induced by chronic natural infection with HIV-1? (4) Do immunodominant hotspots target evolutionarily conserved regions of the HIV genome? (5) Can epitope prediction methods be used to identify these hotspots? We found that vaccine responses clustered into epitope hotspots in all three vaccine trials and some of these hotspots were not observed in chronic natural infection. We also found significant differences between the immunodominance patterns generated in each trial, even comparing two trials that tested the same vaccine in different populations. Some of the vaccine-induced immunodominant hotspots were located in highly variable regions of the HIV genome, and this was more evident for the MRKAd-5 HIV vaccine. Finally, we found that epitope prediction methods can partially predict the location of vaccine-induced epitope hotspots. Our findings have implications for vaccine design and suggest a framework by which different vaccine candidates can be compared in early phases of evaluation. [ABSTRACT FROM AUTHOR]

Functional T-cell epitope discovery is a key process for the development of novel immunotherapies, particularly for cancer immunology. In silico epitope prediction is a common strategy to try to achieve this objective. However, this approach suffers from a significant rate of false-negative results and epitope ranking lists that often are not validated by practical experience. A high-throughput platform for the identification and prioritization of potential T-cell epitopes is the iTopia™ Epitope Discovery System™, which allows measuring binding and stability of selected peptides to MHC Class I molecules. So far, the value of iTopia combined with in silico epitope prediction has not been investigated systematically. In this study, we have developed a novel in silico selection strategy based on three criteria: (1) predicted binding to one out of five common MHC Class I alleles; (2) uniqueness to the antigen of interest; and (3) increased likelihood of natural processing. We predicted in silico and characterized by iTopia 225 candidate T-cell epitopes and fixed-anchor analogs from three human tumor-associated antigens: CEA , HE R2 and TERT. HLA-A2-restricted fragments were further screened for their ability to induce cell-mediated responses in HLA-A2 transgenic mice. The iTopia binding assay was only marginally informative while the stability assay proved to be a valuable experimental screening method complementary to in silico prediction. Thirteen novel T-cell epitopes and analogs were characterized and additional potential epitopes identified, providing the basis for novel anticancer immunotherapies. In conclusion, we show that combination of in silico prediction and an iTopia-based assay may be an accurate and efficient method for MHC Class I epitope discovery among tumor-associated antigens. [ABSTRACT FROM AUTHOR]

T cell directed HIV vaccines are based upon the induction of CD8+ T cell memory responses that would be effective in inhibiting infection and subsequent replication of an infecting HIV-1 strain, a process that requires a match or near-match between the epitope induced by vaccination and the infecting viral strain. We compared the frequency and specificity of the CTL epitope responses elicited by the replication-defective Ad5 gag/pol/nef vaccine used in the Step trial with the likelihood of encountering those epitopes among recently sequenced Clade B isolates of HIV-1. Among vaccinees with detectable 15-mer peptide pool ELISpot responses, there was a median of four (one Gag, one Nef and two Pol) CD8 epitopes per vaccinee detected by 9-mer peptide ELISpot assay. Importantly, frequency analysis of the mapped epitopes indicated that there was a significant skewing of the T cell response; variable epitopes were detected more frequently than would be expected from an unbiased sampling of the vaccine sequences. Correspondingly, the most highly conserved epitopes in Gag, Pol, and Nef (defined by presence in .80% of sequences currently in the Los Alamos database www.hiv. lanl.gov) were detected at a lower frequency than unbiased sampling, similar to the frequency reported for responses to natural infection, suggesting potential epitope masking of these responses. This may be a generic mechanism used by the virus in both contexts to escape effective T cell immune surveillance. The disappointing results of the Step trial raise the bar for future HIV vaccine candidates. This report highlights the bias towards less-conserved epitopes present in the same vaccine used in the Step trial. Development of vaccine strategies that can elicit a greater breadth of responses, and towards conserved regions of the genome in particular, are critical requirements for effective T-cell based vaccines against HIV-1. [ABSTRACT FROM AUTHOR]

Background: Adenoviral (Ad) vaccine vectors represent both a vehicle to present a novel antigen to the immune system as well as restimulation of immune responses against the Ad vector itself. To what degree Ad-specific CD8+ T cells are restimulated by Ad vector vaccination is unclear, although such knowledge would be important as vector-specific CD8+ T cell expansion could potentially further limit Ad vaccine efficacy beyond Ad-specific neutralizing antibody alone. Methodology/Principal Findings: Here we addressed this issue by measuring human Adenovirus serotype 5 (Ad5)-specific CD8+ T cells in recipients of the Merck Ad5 HIV-1 vaccine vector before, during, and after vaccination by multicolor flow cytometry. Ad5-specific CD8+ T-cells were detectable in 95% of subjects prior to vaccination, and displayed primarily an effector-type functional profile and phenotype. Peripheral blood Ad5-specific CD8+ T-cell numbers expanded after Ad5-HIV vaccination in all subjects, but differential expansion kinetics were noted in some baseline Ad5-neutralizing antibody (Ad5 nAb) seronegative subjects compared to baseline Ad5 nAb seropositive subjects. However, in neither group did vaccination alter polyfunctionality, mucosal targeting marker expression, or memory phenotype of Ad5-specific CD8+ T-cells. Conclusions: These data indicate that repeat Ad5-vector administration in humans expands Ad5-specific CD8+ T-cells without overtly affecting their functional capacity or phenotypic properties. This is a secondary analysis of samples collected during the 016 trial. Results of the Merck 016 trial safety and immunogenicity have been previously published in the journal of clinical infectious diseases. [ABSTRACT FROM AUTHOR]

Background. We report composite results from the Merck phase I program of near-consensus clade B human immunodeficiency virus (HIV) type 1 gag vaccines. Methods. Healthy HIV-uninfected adults were enrolled in 6 blinded placebo-controlled studies evaluating the immunogenicity of (1) a 4-dose regimen of a DNA vaccine, (2) a 3-dose priming regimen of the DNA vaccine with a booster dose of an adenovirus type 5 (Ad5)-vectored vaccine, or (3) a 3-dose regimen of the Ad5 vaccine. The DNA plasmid was provided with or without an aluminum phosphate or CRL1005 adjuvant. The primary end point was the unfractionated HIV-1 gag-specific interferon γ enzyme-linked immunospot (ELISpot) response 4 weeks after the final dose. Results. Overall, 254 (83%) of 307 subjects randomized to the vaccine groups were evaluable. Adjuvants did not enhance immunogenicity of the DNA vaccine. Postboost ELISpot responder frequencies were higher for Ad5-containing regimens than for the DNA/DNA regimen (33%) but were similar for DNA/Ad5 (55%) and Ad5/Ad5 (50%). DNA/DNA elicited mainly a CD4 response, whereas Ad5/Ad5 elicited mainly a CD8 response; DNA/Ad5 generated CD4 and CD8 responses comparable to those of DNA/DNA and Ad5/Ad5, respectively. Conclusions. The DNA vaccine alone or as a priming regimen for the Ad5 vaccine did not increase unfractionated ELISpot responses compared with the Ad5 vaccine alone. Qualitative T cell responses to different vaccine regimens deserve further study. [ABSTRACT FROM AUTHOR]

The mechanisms underlying possible increased HIV-1 acquisition in adenovirus 5 (Ad5)-seropositive subjects vaccinated with Ad5–HIV-1 vectors in the Merck STEP trial remain unclear. We find that Ad5 serostatus does not predict Ad5-specific CD4+ T cell frequency, and we did not observe durable significant differences in Ad5-specific CD4+ T cells between Ad5-seropositive and Ad5-seronegative subjects after vaccination. These findings indicate no causative role for Ad5-specific CD4+ T cells in increasing HIV-1 susceptibility in the STEP trial. [ABSTRACT FROM AUTHOR]

Sub-unit vaccines are primarily designed to include antigens required to elicit protective immune responses and to be safer than whole-inactivated or live-attenuated vaccines. But their purity and inability to self-adjuvant often result in weaker immunogenicity. Emerging evidence suggests that bio-engineered nanoparticles can be used as immunomodulatory adjuvants. Therefore, in this study we explored the potential of novel Merck-proprietary lipid nanoparticle (LNP) formulations to enhance immune responses to sub-unit viral antigens. Immunization of BALB/c and C57BL/6 mice revealed that LNPs alone or in combination with a synthetic TLR9 agonist, immune-modulatory oligonucleotides, IMO-2125 (IMO), significantly enhanced immune responses to hepatitis B virus surface antigen (HBsAg) and ovalbumin (OVA). LNPs enhanced total B-cell responses to both antigens tested, to levels comparable to known vaccine adjuvants including aluminum based adjuvant, IMO alone and a TLR4 agonist, 3-O-deactytaled monophosphoryl lipid A (MPL). Investigation of the quality of B-cell responses demonstrated that the combination of LNP with IMO agonist elicited a stronger Th1-type response (based on the IgG2a:IgG1 ratio) than levels achieved with IMO alone. Furthermore, the LNP adjuvant significantly enhanced antigen specific cell-mediated immune responses. In ELISPOT assays, depletion of specific subsets of T cells revealed that the LNPs elicited potent antigen-specific CD4 + and CD8 + T cell responses. Intracellular FACS analyses revealed that LNP and LNP + IMO formulated antigens led to higher frequency of antigen-specific IFNγ + TNFα + IL-2 + , multi-functional CD8 + T cell responses, than unadjuvanted vaccine or vaccine with IMO only. Overall, our results demonstrate that lipid nanoparticles can serve as future sub-unit vaccine adjuvants to boost both B-cell and T-cell responses in vivo , and that addition of IMO can be used to manipulate the quality of immune responses. [ABSTRACT FROM AUTHOR]

Abstract: Maternal immunity to human cytomegalovirus (HCMV) prior to conception is ∼70% protective against congenital transmission and in utero infection of HCMV. Both functional antibodies capable of neutralizing virus and effective T-cells are believed to be important for the protection. Previous HCMV vaccines have rarely been shown able to induce neutralizing antibody titers comparable to those seen in naturally infected HCMV seropositive subjects. Recent studies link a glycoprotein H (gH) complex to receptor-mediated viral entry of endothelial/epithelial cells and leukocytes. This pentameric gH complex, composed of five proteins (gH, gL, UL128, UL130 and UL131 proteins), is notably missing in all HCMV vaccine previously evaluated in clinic. Here we showed that a HCMV virus, with restored expression of the pentameric gH complex, can induce 10-fold higher neutralizing antibody titers than an attenuated AD169 virus or a recombinant glycoprotein B vaccine in multiple animal species in which viral replication is not expected. Encouragingly, the peak neutralizing titers post vaccination in rabbits and monkeys were within 2–4-fold of the levels determined in HCMV seropositive subjects. Functional antibodies by vaccination could further be improved when formulated with a novel adjuvant, and the titers of the antiviral antibodies were sustained in rabbits for over a year after vaccination. These results indicate that the pentameric gH complex is associated with greatly improved functional antibodies following vaccination, and support a vaccine concept based on a nonreplicating whole HCMV with the pentameric gH-associated epithelial tropism restored. [Copyright &y& Elsevier]

Abstract: It has been suggested that poor immunogenicity may explain the lack of vaccine efficacy in preventing or controlling HIV infection in the Step trial. To investigate this issue we vaccinated eight Indian rhesus macaques with a trivalent replication-incompetent adenovirus serotype 5 vaccine expressing SIV Gag, Pol, and Nef using a regimen similar to that employed in the Step trial. We detected broad vaccine-induced CD8+ (2–7 pool-specific responses) and CD4+ (5–19 pool-specific responses) T-cell responses in IFN-γ ELISPOT assays at one week post-boost using fresh PBMC. However, using cryopreserved cells at one and four weeks post-boost we observed a reduction in both the number and magnitude of most vaccine-induced responses. This demonstrates that the time points and conditions chosen to perform immune assays may influence the observed breadth and frequency of vaccine-induced T-cell responses. To evaluate protective efficacy, we challenged the immunized macaques, along with naïve controls, with repeated, limiting doses of the heterologous swarm isolate SIVsmE660. Vaccination did not significantly affect acquisition or control of virus replication in vaccinees compared to naïve controls. Post-infection we observed an average of only two anamnestic CD8+ T-cell responses per animal, which may not have been sufficiently broad to control heterologous virus replication. While the trivalent vaccine regimen induced relatively broad T-cell responses in rhesus macaques, it failed to protect against infection or control viral replication. Our results are consistent with those observed in the Step trial and indicate that SIV immunization and challenge studies in macaque models of HIV infection can be informative in assessing pre-clinical HIV vaccines. [Copyright &y& Elsevier]

Recombinant viruses hold promise as vectors for vaccines to prevent infectious diseases with significant global health impacts. One of their major limitations is that preexisting anti-vector neutralizing antibodies can reduce T cell responses to the insert antigens; however, the impact of vector-specific cellular immunity on subsequent insert-specific T cell responses has not been assessed in humans. Here, we have identified and compared adenovirus-specific and HIV-specific T cell responses in subjects participating in two HIV-1 vaccine trials using a vaccine vectored by adenovirus serotype 5 (Ad5). Higher frequencies of pre-immunization adenovirus-specific CD4+ T cells were associated with substantially decreased magnitude of HIV-specific CD4+ T cell responses and decreased breadth of HIV-specific CD8+ T cell responses in vaccine recipients, independent of type-specific preexisting Ad5-specific neutralizing antibody titers. Further, epitopes recognized by adenovirus-specific T cells were commonly conserved across many adenovirus serotypes, suggesting that cross-reactivity of preexisting adenovirus-specific T cells can extend to adenovirus vectors derived from rare serotypes. These findings provide what we believe to be a new understanding of how preexisting viral immunity may impact the efficacy of vaccines under current evaluation for prevention of HIV, tuberculosis, and malaria. [ABSTRACT FROM AUTHOR]

Abstract: Following the disappointing outcome of the phase IIb test-of-concept step study in which Merck''s adenovirus type 5 (Ad5) HIV-1 clade B gag/pol/nef vaccine failed to demonstrate efficacy in HIV high-risk individuals, an extensive review of the trial and preclinical studies which supported the trial is ongoing. One point of interest is how well preclinical nonhuman primate immunogenicity studies predicted what was observed in humans. Here we compare the HIV-1-specific cellular immune responses elicited in nonhuman primates and human clinical trial subjects to several HIV-1 vaccine candidates. We find that although rhesus macaques are immunologically more responsive to vaccination than humans, the hierarchy in potency of single-modality prime–boost regimens using several vector approaches (adenovirus, DNA, and pox vectors) was well predicted. Vaccine approaches using complex formulations such as novel adjuvants (DNA+CRL1005) or mixed-modality prime–boost (DNA/Ad5; Ad5/ALVAC) did not correlate as well between rhesus macaques and humans. Although the immunogenicity of the vaccines and vaccine regimens evaluated were not all accurately predicted, testing in rhesus macaques generally offers an indispensable tool for ranking the immunological potential of HIV-1 vaccine candidates. [Copyright &y& Elsevier]

The article presents a case-cohort analysis which assessed the MRKAd5 HIV-1 gag/pol/nef vaccine for vaccine-induced immunity and potential contributions to infection risk. The study was performed in response to the results of the Step Study which found higher HIV-1 incidence in vaccine-treated men than in placebo-treated men with pre-existing adenovirus serotype 5 immunity. Validated interferon-γ ELISPOT and intracellular cytokine staining assays were used to characterise HIV-specific T cells to assess for immunogenicity. Flow cytometric studies were conducted to establish the effects of vaccine and pre-existing Ad5 immunity on infection risk. The MRKAd5 HIV-1 gag/pol/nef vaccine was found to be highly immunogenic for inducing HIV-specific CD8+ T cells.

Abstract: We sought to determine how effectively a CD8+ T cell inducing vaccine controls SHIV-89.6P infection in rhesus macaques at a range of challenge times post-vaccination. To this end, twenty eight Mamu-A⁎01 + rhesus macaques were given replication incompetent human serotype 5 adenovirus vector expressing SIVmac239 gag DNA and boosted 24 weeks later. Groups of 4 monkeys were then challenged with SHIV-89.6P at 1, 3, 6, 12, and 24 weeks after the boost. We compared the kinetics of viral load, CD4+ and virus-specific CD8+ T cells in these macaques. Measurements of CD8+ T cells taken before challenge show an exponential decay between 1 and 12 weeks following vaccination (p <0.0001). After week 12, no further decay was observed. Twenty of 24 vaccinated animals maintained more CD4+ T cells and kept their viral load at least one order of magnitude lower than the control animals throughout the chronic phase of the study. All 24 vaccinated animals survived the duration of the study. The viral and T cell kinetics over the first two weeks differed between the vaccinated groups, with more recent vaccination improving the early control of virus (p-value=0.027). The rates of virus specific CD8+ T cell expansion were greater in animals having higher viral loads at one week (r =0.45, p =0.029), suggesting that the kinetics of early viral load may have a role in virus specific CD8+ T cell generation, although these early differences did not lead to different clinical outcomes within the vaccinated animals. [Copyright &y& Elsevier]

The article compares anti-HIV-1 T-cell immune responses among unvaccinated individuals from Botswana and Cameroon results infected with diverse HIV-1 clades. Response rates to clade B Gag and/or clade B Nef proteins were the same when compared with countries including Brazil, Thailand, South Africa, Malawi and the United States. Overall, 93 percent of individuals responded to either all 3 clade Gag or all 3 clade Nef proteins in Botswana and 98 percent responded to clade B Gag and/or Nef proteins in Cameroon.

Results from Merck's phase II adenovirus type 5 (Ad5) gag/pol/nef test-of-concept trial showed that the vaccine lacked efficacy against human immunodeficiency virus (HIV) infection in a high-risk population. Among the many questions to be explored following this outcome are whether (i) the Ad5 vaccine induced the quality of T-cell responses necessary for efficacy and (ii) the lack of efficacy in the Ad5 vaccine can be generalized to other vector approaches intended to induce HIV type 1 (HIV-1)-specific T-cell responses. Here we present a comprehensive evaluation of the T-cell response profiles from cohorts of clinical trial subjects who received the HIV CAM-1 gag insert delivered by either a regimen with DNA priming followed by Ad5 boosting (n = 50) or a homologous Ad5/Ad5 prime-boost regimen (n = 70). The samples were tested using a statistically qualified nine-color intracellular cytokine staining assay measuring interleukin-2 (IL-2), tumor necrosis factor alpha, macrophage inflammatory protein 1β, and gamma interferon production and expression of CD107a. Both vaccine regimens induced CD4+ and CD8+ HIV gag-specific T-cell responses which variably expressed several intracellular markers. Several trends were observed in which the frequencies of HIV-1-specific CD4+ T cells and IL-2 production from antigen-specific CD8+ T cells in the DNA/Ad5 cohort were more pronounced than in the Ad5/Ad5 cohort. Implications of these results for future vaccine development will be discussed. [ABSTRACT FROM AUTHOR]