HIV-specific Immune Responses Research Articles

Amplified antigen-specific immune responses in HIV-1 infected individuals in a double blind DNA immunization and therapy interruption trial

Immunotherapy in patients with HIV-1 infection aims to restore and broaden immunological competence, reduce viral load and thereby permit longer periods without combined antiretroviral treatment (cART). Twelve HIV-1-infected patients on cART were immunized on the skin with DNA plasmids containing genes of several HIV-1 subtypes with or without the addition of hydroxyurea (HU), or with placebo. The mean net gain of HIV-specific CD8+ T cell responses were higher and broader in the HIV DNA vaccine groups compared to non-vaccinated individuals (p<0.05). The vaccine-induced immune responses per se had no direct effect on viral replication. In all patients combined, including placebo, the viral set point after a final structured therapy interruption (STI) was lower than prior to initiation of cART (p=0.003). Nadir CD4 levels appeared to strongly influence the post-STI viral titers. After the sixth immunization or placebo, patients could stay off cART for a median time of 15 months. The study shows that HIV DNA immunization induces broader and higher magnitudes of HIV-specific immune responses compared to structured therapy interruptions alone. Although compromised by small numbers of patients, the study also demonstrates that well-monitored STI may safely function as an immunological read out of HIV vaccine efficacy.

Performance of V3-based HIV-1 sero subtyping in HIV endemic areas.

HIV-1 serosubtyping based on reactivity to peptides from the V3 region of gp120 is a low-cost and easy to perform procedure often used in geographical areas with high prevalence and incidence of HIV infection. We evaluated the performance of V3-based serotyping on 148 sera from 118 HIV-1-infected individuals living in Uganda, with estimated dates of seroconversion. Of the 148 tested samples, 68 (46.0%) specifically reacted with only one of the V3 peptides included in the test (SP), 64 (43.2%) did not react with any peptide (NR) and 16 (10.8%) reacted with two or more peptides (CR). According to the estimated seroconversion date, the large majority of samples collected early after infection belonged to the NR group. These samples had also a low Avidity Index. In contrast, samples collected later after infection belonged mainly to CR and SP groups and had also a higher avidity index. These results indicate that the performance of V3-based assays depends on maturation of HIV-specific immune response and can be significantly lowered when these tests are carried out on specimens collected from recently infected individuals.

Current and future assays for identifying recent HIV infections at the population level

SummaryThe precise diagnosis of recent human immunodeficiency virus (HIV) infection is crucial for estimating HIV incidence, defined as the number of new infections in a population, per person at risk, during a specified time period. Incidence assessment is considered to be a tool for surveillance, public health and research. Differentiating recent from long-term HIV infections is possible thanks to the evaluation of HIV-specific immune response development or viral markers measurement. Several methods that enable the recognition of recent HIV-1 infection with the use of a single blood specimen have been developed, and their value for use in population level studies has been demonstrated. However, they are still inadequate due to a variable window period and false recent rates among HIV clades and across populations. Application of these assays at an individual level is far more questionable because of person-to-person variability in the antibody response and the course of HIV infection, and because of the prospective regulatory approval requirements.In this article we review the principles and the limitations of the currently available major laboratory techniques that allow detection of recent HIV infection. The assays based on the alteration of serological parameters, as well as the newest method based on an increase of HIV genetic diversity with the progress of infection, are described.

Robust Vaccine-Elicited Cellular Immune Responses in Breast Milk following Systemic Simian Immunodeficiency Virus DNA Prime and Live Virus Vector Boost Vaccination of Lactating Rhesus Monkeys

Breast milk transmission of HIV remains an important mode of infant HIV acquisition. Enhancement of mucosal HIV-specific immune responses in milk of HIV-infected mothers through vaccination may reduce milk virus load or protect against virus transmission in the infant gastrointestinal tract. However, the ability of HIV/SIV strategies to induce virus-specific immune responses in milk has not been studied. In this study, five uninfected, hormone-induced lactating, Mamu A*01(+) female rhesus monkey were systemically primed and boosted with rDNA and the attenuated poxvirus vector, NYVAC, containing the SIVmac239 gag-pol and envelope genes. The monkeys were boosted a second time with a recombinant Adenovirus serotype 5 vector containing matching immunogens. The vaccine-elicited immunodominant epitope-specific CD8(+) T lymphocyte response in milk was of similar or greater magnitude than that in blood and the vaginal tract but higher than that in the colon. Furthermore, the vaccine-elicited SIV Gag-specific CD4(+) and CD8(+) T lymphocyte polyfunctional cytokine responses were more robust in milk than in blood after each virus vector boost. Finally, SIV envelope-specific IgG responses were detected in milk of all monkeys after vaccination, whereas an SIV envelope-specific IgA response was only detected in one vaccinated monkey. Importantly, only limited and transient increases in the proportion of activated or CCR5-expressing CD4(+) T lymphocytes in milk occurred after vaccination. Therefore, systemic DNA prime and virus vector boost of lactating rhesus monkeys elicits potent virus-specific cellular and humoral immune responses in milk and may warrant further investigation as a strategy to impede breast milk transmission of HIV.

Evaluation of a Quantitative Real-Time PCR Assay to Measure HIV-Specific Mucosal CD8+ T Cell Responses in the Cervix

Several candidate HIV vaccines aim to induce virus-specific cellular immunity particularly in the genital tract, typically the initial site of HIV acquisition. However, standardized and sensitive methods for evaluating HIV-specific immune responses at the genital level are lacking. Therefore we evaluated real-time quantitative PCR (qPCR) as a potential platform to measure these responses. β-Actin and GAPDH were identified as the most stable housekeeping reference genes in peripheral blood mononuclear cells (PBMCs) and cervical mononuclear cells (CMCs) respectively and were used for normalizing transcript mRNA expression. HIV-specific cellular T cell immune responses to a pool of optimized CD8+ HIV epitopes (HIV epitope pool) and Staphylococcal enterotoxin B (SEB) superantigen control were assayed in HIV infected PBMC by qPCR, with parallel assessment of cytokine protein production. Peak HIV-specific mRNA expression of IFNγ, IL-2 and TNFα occurred after 3, 5 and 12 hours respectively. PBMCs were titrated to cervical appropriate cell numbers to determine minimum required assay input cell numbers; qPCR retained sensitivity with input of at least 2.5×104 PBMCs. This optimized qPCR assay was then used to assess HIV-specific cellular T cell responses in cytobrush-derived cervical T cells from HIV positive individuals. SEB induced IFNγ mRNA transcription was detected in CMCs and correlated positively with IFNγ protein production. However, qPCR was unable to detect HIV-induced cytokine mRNA production in the cervix of HIV-infected women despite robust detection of gene induction in PBMCs. In conclusion, although qPCR can be used to measure ex vivo cellular immune responses to HIV in blood, HIV-specific responses in the cervix may fall below the threshold of qPCR detection. Nonetheless, this platform may have a potential role in measuring mitogen-induced immune responses in the genital tract.

IL-15 increases the frequency of effector memory CD8+ T cells in rhesus monkeys immunized with HIV vaccine

Several studies have suggested that interleukin (IL)-15 is a promising adjuvant that promotes cellular immunity when administered with human immunodeficiency virus (HIV) vaccine. Here we evaluated the effect of IL-15 plasmid on HIV-specific immune responses, especially cellular immunity, in eight rhesus monkeys. These monkeys were immunized three times with HIV DNA vaccine with or without IL-15 plasmid and boosted with recombinant Tiantan strain vaccinia virus-based HIV vaccine (rTV) 22 weeks after the first immunization. Although we did not detect any significant differences in the HIV-specific CD8(+) T-cell response between monkeys with IL-15 coimmunization and monkeys with HIV vaccine alone, our results showed that the frequency of effector CD8(+) memory T cells in the peripheral blood was significantly higher in monkeys with IL-15 coimmunization than those with HIV vaccine alone at almost all of the time points examined. Furthermore, the titers of anti-HIV antibodies were higher in Group T than those in Group C after rTV boosting. These findings in rhesus monkeys suggest that IL-15 may be useful as a cytokine adjuvant for HIV vaccine.

Immunogenic Profiling in Mice of a HIV/AIDS Vaccine Candidate (MVA-B) Expressing Four HIV-1 Antigens and Potentiation by Specific Gene Deletions

BackgroundThe immune parameters of HIV/AIDS vaccine candidates that might be relevant in protection against HIV-1 infection are still undefined. The highly attenuated poxvirus strain MVA is one of the most promising vectors to be use as HIV-1 vaccine. We have previously described a recombinant MVA expressing HIV-1 Env, Gag, Pol and Nef antigens from clade B (referred as MVA-B), that induced HIV-1-specific immune responses in different animal models and gene signatures in human dendritic cells (DCs) with immunoregulatory function.Methodology/Principal FindingsIn an effort to characterize in more detail the immunogenic profile of MVA-B and to improve its immunogenicity we have generated a new vector lacking two genes (A41L and B16R), known to counteract host immune responses by blocking the action of CC-chemokines and of interleukin 1β, respectively (referred as MVA-B ΔA41L/ΔB16R). A DNA prime/MVA boost immunization protocol was used to compare the adaptive and memory HIV-1 specific immune responses induced in mice by the parental MVA-B and by the double deletion mutant MVA-B ΔA41L/ΔB16R. Flow cytometry analysis revealed that both vectors triggered HIV-1-specific CD4+ and CD8+ T cells, with the CD8+ T-cell compartment responsible for >91.9% of the total HIV-1 responses in both immunization groups. However, MVA-B ΔA41L/ΔB16R enhanced the magnitude and polyfunctionality of the HIV-1-specific CD4+ and CD8+ T-cell immune responses. HIV-1-specific CD4+ T-cell responses were polyfunctional and preferentially Env-specific in both immunization groups. Significantly, while MVA-B induced preferentially Env-specific CD8+ T-cell responses, MVA-B ΔA41L/ΔB16R induced more GPN-specific CD8+ T-cell responses, with an enhanced polyfunctional pattern. Both vectors were capable of producing similar levels of antibodies against Env.Conclusions/SignificanceThese findings revealed that MVA-B and MVA-B ΔA41L/ΔB16R induced in mice robust, polyfunctional and durable T-cell responses to HIV-1 antigens, but the double deletion mutant showed enhanced magnitude and quality of HIV-1 adaptive and memory responses. Our observations are relevant in the immune evaluation of MVA-B and on improvements of MVA vectors as HIV-1 vaccines.

Persistence of Genital Tract T Cell Responses in HIV-Infected Women on Highly Active Antiretroviral Therapy

Initiation of highly active antiretroviral therapy (HAART) for HIV-infected individuals is associated with control of viremia, improved CD4 counts, and declining systemic HIV-specific immune responses. While HAART effectively reduces plasma viremia, it remains unclear how effectively antiretroviral drugs reach mucosal surfaces, such as those of the genital tract. The aim of this study was to determine the effect of HAART on genital tract CD4 T cell reconstitution, HIV shedding, and HIV-specific T cell responses. Cervical cytobrush and blood specimens were obtained from 35 HIV-infected, HAART-naïve women and 27 women on HAART in order to investigate HIV Gag-specific T cell responses by intracellular gamma interferon (IFN-γ) staining. Interleukin 1β (IL-1β), IL-6, and IL-8 concentrations were measured by enzyme-linked immunosorbent assays (ELISA). We show that for HIV-infected women, HAART is associated with significantly improved CD4 T cell counts both in blood and at the cervix. While HAART effectively suppressed both blood and cervical viremia, HIV-specific CD8 T cell responses in blood were lost, while those at the cervix were preserved.

Relationship between Regulatory T Cells and Immune Activation in Human Immunodeficiency Virus-Infected Patients Interrupting Antiretroviral Therapy

Persistent immune activation plays a central role in driving Human Immunodeficiency Virus (HIV) disease progression. Whether CD4+CD25+ regulatory T cells (Tregs) are harmful by suppressing HIV-specific immune responses and/or beneficial through a decrease in immune activation remains debatable. We analysed the relationship between proportion and number of regulatory T cells (Tregs) and immune activation in HIV-infected patients interrupting an effective antiretroviral therapy (ART). Twenty-five patients were included in a substudy of a prospective multicenter trial of treatment interruption (TI) (ANRS 116). Proportions and numbers of Tregs and the proportion of activated CD4 and CD8 T cells were assessed at baseline and month 12 (M12) of TI. Specific anti-HIV CD4 and CD8 responses were investigated at baseline and M12. Non parametric univariate analyses and multivariate linear regression models were conducted. At baseline, the proportion of Tregs negatively correlated with the proportion of HLA-DR+CD8+T cells (r = −0.519). Following TI, the proportion of Tregs increased from 6.3% to 7.2% (p = 0.029); absolute numbers of Tregs decreased. The increase in the proportion of HLA-DR+CD38+CD8+T cells was significantly related to the increase in proportion of Tregs (p = 0.031). At M12, the proportion of Tregs did not negatively correlate with CD8 T-cell activation. Nevertheless, Tregs retain a suppressive function since depletion of Treg-containing CD4+CD25+ cells led to an increase in lymphoproliferative responses in most patients studied. Our data suggest that Tregs are efficient in controlling residual immune activation in patients with ART-mediated viral suppression. However, the insufficient increase in the proportion and/or the decrease in the absolute number of Tregs result in a failure to control immune activation following TI.Trial RegistrationClinicalTrials.gov NCT00118677

HIV+ elite controllers have low HIV-specific T-cell activation yet maintain strong, polyfunctional T-cell responses

HIV elite controllers are a unique group of rare individuals who maintain undetectable viral loads in the absence of antiretroviral therapy. We studied immune responses in these individuals to inform vaccine development, with the goal of identifying the immune correlates of protection from HIV. We compared markers of cellular activation, HIV-specific immune responses and regulatory T (Treg) cell frequencies in four groups of individuals: HIV-negative healthy controls, elite controllers (HIV RNA level <75 copies/ml), individuals on HAART and individuals with HIV RNA level more than 10,000 copies/ml (noncontrollers). Elite controllers possessed significantly lower levels of activated HIV-specific CD8 T cells and of recently divided HIV-specific CD4 T cells than noncontrollers, whereas these differences were not seen in the respective cytomegalovirus-specific T-cell populations. Elite controllers also mounted a stronger and broader cytokine and chemokine response following HIV-specific stimulation than individuals on HAART and noncontrollers. Finally, we found that HAART-suppressed individuals had elevated Treg cell frequencies, whereas elite controllers and noncontrollers maintained normal percentages of Treg cells. Elite controllers maintain high levels of HIV-specific immune responses with low levels of HIV-specific T-cell activation and do not have elevated Treg cell levels. Based on these data an ideal HIV vaccine would induce strong HIV-specific immune responses whereas minimizing HIV-specific T-cell activation.

NSG-hu mice generate HIV specific human immune responses after gp96 vaccination (46.14)

Abstract NOD.SCID common gamma chain (NSG) knockout mice have no functional adaptive immune system of their own; they readily accept human CD34+ hematopoietic stem cell xenografts (NSG-hu). Therefore, NSG-hu mice present a model system in which to study human immunology. Previously, we developed a cell based vaccine combining secreted heat shock protein gp96 and HIV gag protein (293-gp96Ig-HIVgag). Here we use NSG-hu mice vaccinated with 293-gp96Ig-HIVgag to study human immunological responses. Flow cytometry analysis of peripheral blood in pre-vaccination (8-9 week old) animals revealed human engraftment (mCD45 vs. hCD45) ranging from 25%-50%. In addition, human CD45+ cells were detected in specific tissues (spleen, PEC, IEL and MLN); CD4+, CD8+ T cells and NK cells were found within secondary murine lymphoid organs. Post vaccination (12-13 week old) human engraftment of peripheral blood was observed ranging from 15%-79%. In an ELISPOT for IFN-γ, PECs from NSG-hu vaccinated with 293-gp96Ig-HIVgag i.p. generated an HIV specific immune response when stimulated with SLYNTVATL peptide, producing an average of 778 spots per 2x105 cells plated whereas mice vaccinated with 293 cells alone generated an average of 88.5 spots per 2x105 cells plated (p&amp;lt;0.05) We conclude that NSG-hu mice are able to establish and maintain stable hematopoiesis from human derived cells CD34+ cells. Furthermore, NSG-hu mice are capable of mounting a human HIV-specific immune response after gp96 vaccination.

Repeated DNA therapeutic vaccination of chronically SIV-infected macaques provides additional virological benefit

We have previously reported that therapeutic immunization by intramuscular injection of optimized plasmid DNAs encoding SIV antigens effectively induces immune responses able to reduce viremia in antiretroviral therapy (ART)-treated SIVmac251-infected Indian rhesus macaques. We subjected such therapeutically immunized macaques to a second round of therapeutic vaccination using a combination of plasmids expressing SIV genes and the IL-15/IL-15 receptor alpha as molecular adjuvant, which were delivered by the more efficacious in vivo constant-current electroporation. A very strong induction of antigen-specific responses to Gag, Env, Nef, and Pol, during ART (1.2–1.6% of SIV-specific T cells in the circulating T lymphocytes) was obtained with the improved vaccination method. Immunological responses were characterized by the production of IFN-γ, IL-2, and TNF-α either alone, or in combination as double or triple cytokine positive multifunctional T cells. A significant induction of CD4 + T cell responses, mainly targeting Gag, Nef, and Pol, as well as of CD8 + T cells, mainly targeting Env, was found in both T cells with central memory and effector memory markers. After release from ART, the animals showed a virological benefit with a further ∼1 log reduction in viremia. Vaccination with plasmid DNAs has several advantages over other vaccine modalities, including the possibility for repeated administration, and was shown to induce potent, efficacious, and long-lasting recall immune responses. Therefore, these data support the concept of adding DNA vaccination to the HAART regimen to boost the HIV-specific immune responses.

Polymorphisms in innate immunity genes and patients response to dendritic cell-based HIV immuno-treatment

Dendritic cells (DCs)-based vaccine was demonstrated to increase HIV specific cellular immune response; however, in some HIV-infected patients, the response to the vaccine resulted to be not effective. In order to understand if the outcome of the vaccination may be influenced by the host's genome and natural immunity, we studied the innate immune genome of HIV-infected patients previously vaccinated with DCs. We identified 15 SNPs potentially associated with the response to the immuno-treatment and two SNPs significantly associated with the modulation of the response to the DC vaccine: MBL2 rs10824792 and NOS1 rs693534. These two SNPs were also studied in different ethnic groups (Brazilians, African and Caucasian) of HIV-infected, exposed uninfected and unexposed uninfected subjects. The HIV positive Caucasian patients were also characterized by different disease progressions.Our findings suggest that, independently and/or in addition to other variables, the host's genome could significantly contribute to the modulation of the response to the DC vaccine.

Influence of a Vaccination Schedule on Viral Load Rebound and Immune Responses in Successfully Treated HIV-Infected Patients

Vaccination is recommended for HIV-infected patients. Transient increases of viral load (VL) and risk of developing resistance to HAART have been described. In addition, VL rebounds could increase HIV-specific immune responses. Twenty-six successfully treated HIV-infected adults were randomized to receive a vaccination schedule or placebo during 12 months. Afterward, HAART was discontinued. Influences of vaccination over VL, genotypic mutations, different T cell subsets, and HIV-1-specific immune responses were evaluated. Patients did not present any secondary effect. No differences in incidence of detectable VL determinations were detected between groups [relative risk 0.54 (95% CI 0.23-1.26)]. No relevant resistance mutations were detected. The vaccinated group showed a significant drop in CD4(+) T cells (p = 0.046) associated with increases in activated T cells. HIV-1-specific lymphoproliferative responses increased more in the vaccinated group during the vaccination period. Viral rebound dynamics after interrupting HAART were similar in both groups. A vaccination schedule in successfully treated HIV patients was safe, was not associated with an increase in detectable VL, and did not increase the risk of developing resistance mutations. However, it induced an increase in T cell activation and a drop in CD4(+) T cells, although these changes did not influence the VL rebound dynamics after HAART interruption.

Immunologic activity and safety of autologous HIV RNA-electroporated dendritic cells in HIV-1 infected patients receiving antiretroviral therapy

Immunogenicity, manufacturing feasibility, and safety of a novel, autologous dendritic cell (DC)-based immunotherapy (AGS-004) was evaluated in ten human immunodeficiency virus type 1 (HIV-1)-infected adults successfully treated with antiretroviral therapy (ART). Personalized AGS-004 was produced from autologous monocyte-derived DCs electroporated with RNA encoding CD40L and HIV antigens (Gag, Vpr, Rev, and Nef) derived from each subjects' pre-ART plasma. Patients received monthly injections of AGS-004 in combination with ART. AGS-004 was produced within a mean of 6 weeks and yielded 4-12 doses/subject Full or partial HIV-specific proliferative immune responses occurred in 7 of 9 evaluable subjects. Responses were specific for the AGS-004 presented HIV antigens and preferentially targeted CD8(+) T cells. Mild adverse events included flu-like symptoms, fatigue, and injection site reactions. No evidence of autoimmunity, changes in viral load, or significant changes in absolute CD4(+) and CD8(+) T cell counts were observed. This pilot study supports the further clinical investigation of AGS-004.

P03-02. SCID-hu mice generate HIV specific human immune responses after gp96 vaccination

P03-02. SCID-hu mice generate HIV specific human immune responses after gp96 vaccination

P02-04. Multimeric soluble CD40 ligand efficiently enhances HIV specific cellular immune responses during DNA prime and boost with attenuated poxvirus strains

1 page.-- Poster presentation.-- This article is part of the supplement: AIDS Vaccine 2009.

P16-02. Induction of systemic HIV-1 specific cellular immune responses by oral exposure in exposed uninfected partner of discordant couples

Background No protective immune response has been identified so far and no HIV-vaccine has yet proven successful. However, some individuals seem to be naturally protected against HIV, or at least less susceptible for HIV transmission. These so called exposed uninfected individuals have been heavily studied in hope to revealing the mechanisms behind the protection. Such information would be highly relevant for any HIV-vaccine design. Our aim was to investigate whether oral exposure of HIV is enough to induce a systemic HIV-1 specific cellular immune response.

P18-08. Characterization of CD34+ derived dendritic cells generated in vitro and transfected with HIV gene as potential therapeutic vaccine in macaque

Background Antiretroviral therapies against HIV infection reduce the plasma viral load but can not eradicate the virus. Dendritic cells (DC) transfected with messenger ribonucleic acids (mRNA) encoding endogenous viral proteins are expected to enhance the HIV specific immune response and are considered as potent therapeutic vaccines. In this context we studied the feasibility and efficiency of mRNA loaded CD34+ derived DC as therapeutic vaccine in SIV infected macaques as a model of HIV infection and AIDS.

P19-05. Use of autologous apoptotic pseudovirus infected cells for vaccination against HIV, evaluation in macaques

Background Antigen-presenting dendritic cells (DC) present viral antigens to T-cells after uptake of apoptotic bodies derived from virus-infected cells. Besides, processing and presentation of ingested antigen, DC were recently shown to be able to incorporate part of the DNA from the apoptotic cells into their own chromosomes and express the encoded antigens. This mechanism could lead to a more efficient presentation of foreign antigens and activation of antigen specific naive T-cells. It was recently exploited for induction of HIV specific immune responses in a HIV-1/ MuLV mouse infection model.