Virus-specific T-cell Responses Research Articles

Induction of Virus‐Specific Cytotoxic T Lymphocytes as a Basis for the Development of Broadly Protective Influenza Vaccines

There is considerable interest in the development of broadly protective influenza vaccines because of the continuous emergence of antigenic drift variants of seasonal influenza viruses and the threat posed by the emergence of antigenically distinct pandemic influenza viruses. It has been recognized more than three decades ago that influenza A virus-specific cytotoxic T lymphocytes recognize epitopes located in the relatively conserved proteins like the nucleoprotein and that they cross-react with various subtypes of influenza A viruses. This implies that these CD8+ T lymphocytes may contribute to protective heterosubtypic immunity induced by antecedent influenza A virus infections. In the present paper, we review the evidence for the role of virus-specific CD8+ T lymphocytes in protective immunity against influenza virus infections and discuss vaccination strategies that aim at the induction of cross-reactive virus-specific T-cell responses.

2009 Pandemic Influenza A H1N1 Vaccination In the Patients with Hematologic Malignancies: Requirement for Repeated Dosing to Optimize Seroprotection

AbstractAbstract 677In 2009 the spread of influenza A (H1N1) satisfied the World Health Organization (WHO) criteria for a global pandemic and led to the initiation of a vaccination campaign to ensure protection for the most vulnerable patients. However, the immunogenicity of the 2009 H1N1 vaccine in immunocompromised patients has not been specifically evaluated. Furthermore, the number of doses of vaccine required for effective immunization against H1N1 has not been established. Whereas the European Medicines Agency (EMEA) and the UK Department of Health (DoH) recommended the injection of two doses of inactivated H1N1 vaccine 3 weeks apart in immunocompromised individuals, the Centers for Disease Control and Prevention recommended immunization with one dose of inactivated H1N1 vaccine for patients with cancer receiving chemotherapy, followed by a booster vaccine after completion of treatment if the pandemic continued. The aim of this study was to determine the safety and efficacy of the 2009 H1N1 vaccine in patients with hematologic malignancies. We prospectively evaluated the humoral and cellular immune responses to monovalent influenza A/California/2009(H1N1)v-like strain surface antigen vaccine in 97 adults with hematologic malignancies and 25 adult controls. Patients received two intramuscular injections of the vaccine 21 days apart and controls received one dose. Antibody titers, expressed as geometric mean, were measured using a hemagglutination-inhibition assay on days 0, 21 and 49 after injection of the first dose. The induction of virus-specific T-cell responses by H1N1 vaccination was assessed directly ex-vivo by flow cytometric enumeration of antigen-specific CD8+ and CD4+ T-lymphocytes using an intracellular cytokine assay for IFN-γ and TNF-α production on days 0 and 49. Of the 97 patients, 32 had chronic myeloid leukemia (CML) in chronic phase in complete cytogenetic response on the tyrosine kinase inhibitors imatinib or dasatinib, 39 had a B-cell malignancy in complete remission (CR) or untreated, and 26 were recipients of allogeneic hematopoietic stem cell transplantation (allo-SCT) in CR at least 6 months beyond transplant and without evidence of graft versus host disease. The vaccine was well tolerated, with no obvious difference in side effects for patients and controls. By day 21 post-vaccination, protective antibody titers of 1:32 or more were seen in 100% of controls compared to 39% of patients with B-cell malignancies (p<0.001), 46% of allo-SCT recipients (p<0.001) and 85% of CML patients (p=0.086). The effect of a booster dose was assessed with a paired sample analysis. After a second vaccine dose, the seroprotection rates increased to 68% (p=0.008), 73% (p=0.031), and 95% (p=0.5) in patients with B-cell malignancies, allo-SCT recipients and CML patients respectively. Patients vaccinated within 6 months of rituximab-based chemotherapy failed to mount a seroprotective antibody response. We also assessed the cellular response to H1N1 vaccine. Prior to vaccination, pre-existing T-cells against H1N1 could be detected in 10/23 controls compared to 2/25 allo-SCT recipients (p=0.007), 2/28 patients with B-cell malignancies (p=0.003) and 6/28 of CML patients (p=0.131). These pre-existing H1N1 T-cell responses may be related to previous exposure to 2009 H1N1 virus but more likely are due to the presence of cross-reactive seasonal and pandemic H1N1 specific T-cells. Following vaccination, H1N1-specific T-cells were induced in a significant proportion of allo-SCT recipient (10/25, p=0.008) and patients with B-cell malignancies (10/28; p=0.008), but not in CML patients or healthy controls. The limited ability of vaccines to significantly increase pre-existing influenza-specific T-cells has been previously reported although the mechanism for this phenomenon is not fully elucidated. These data demonstrate efficacy of H1N1 vaccine in the majority of patients with hematologic malignancies and unequivocally support the EMEA and the UK DoH official guidelines for the administration of 2 vaccine doses in immunocompromised patients to induce protective immune response against 2009 H1N1 influenza. Based on the WHO analyses, it is expected that the pandemic 2009 H1N1 virus will remain globally predominant in 2010–2011. These results may contribute towards the development of evidence-based guidelines for influenza vaccination in patients with hematologic malignancies. [Display omitted] Disclosures:Marin:Novartis: Consultancy, Honoraria, Research Funding; Bristol Myers Squibb: Consultancy, Honoraria, Research Funding.

Quantiferon-CMV Test in Prediction of Cytomegalovirus Infection After Kidney Transplantation

Infection with cytomegalovirus (CMV) is a major cause of morbidity and mortality in immunosuppressed patients, including organ and bone marrow transplant recipients. The majority of CMV disease is caused by reactivation of alatent infection rather that by newly acquired virus. Many techniques have been currently available to aid in the diagnostics of CMV disease. In this report we performed a prospective evaluation of Quantiferon-CMV assay (Cellestis) to determine whether the test is predictive of CMV disease. CD8+ T-cell CMV-specific immunity was assessed in a longitudinal cohort of 14 kidney transplant recipients. According to our data, subjects with higher cellular immune response measured with Quantiferon test had a lower risk of manifestation of CMV infection than subjects with lower responses. Despite the small number of patients and large intra- and interindividual variability of the data in the study, we observed the Quantiferon-CMV assay to be a sensitive specific test to detect a virus-specific T-cell response. We propose that this assay in combination with viral DNA load estimates may prove to be useful to stratify patients at risk of CMV disease.

Multistrain influenza protection induced by a nanoparticulate mucosal immunotherapeutic.

All commercial influenza vaccines elicit antibody responses that protect against seasonal infection, but this approach is limited by the need for annual vaccine reformulation that precludes efficient responses against epidemic and pandemic disease. In this study we describe a novel vaccination approach in which a nanoparticulate, liposome-based agent containing short, highly conserved influenza-derived peptides is delivered to the respiratory tract to elicit potent innate and selective T cell-based adaptive immune responses. Prepared without virus-specific peptides, mucosal immunostimulatory therapeutic (MIT) provided robust, but short-lived, protection against multiple, highly lethal strains of influenza in mice of diverse genetic backgrounds. MIT prepared with three highly conserved epitopes that elicited virus-specific memory T-cell responses but not neutralizing antibodies, termed MITpep, provided equivalent, but more durable, protection relative to MIT. Alveolar macrophages were more important than dendritic cells in determining the protective efficacy of MIT, which induced both canonical and non-canonical antiviral immune pathways. Through activation of airway mucosal innate and highly specific T-cell responses, MIT and MITpep represent novel approaches to antiviral protection that offer the possibility of universal protection against epidemic and pandemic influenza.

Authors' Reply to Maintenance Immunosuppressive Agents as Risk Factors for BK Virus Nephropathy

We appreciate the comments by Kuypers et al. regarding our study on the association of maintenance immunosuppressive agents and BK nephropathy (BKN) (1). We are in an agreement with the authors that mycophenolate mofetil (MMF) dose is not a reliable surrogate for drug exposure, and multiple factors, such as renal dysfunction, serum albumin level, and concomitant medications, could significantly affect the concentration of its metabolites (2–6). However, the clinical value of mycophenolic acid (MPA) therapeutic drug monitoring remains a controversial and supportive evidence regarding its added benefit is conflicting (7). Although, many retrospective studies found an association between the risk of acute rejection and total MPA area under the curve (AUC0–12 h) (7–9), the relationship between adverse effects of MMF, such as anemia, leukopenia, gastrointestinal symptoms, and infection, and the AUC0–12 h is less well established (7, 9, 10). Some studies have found better correlation between the incidence of infection or leucopenia with free rather than total MPA level (2–4). Others have suggested that adverse events are better predicted by MMF dose, rather than blood concentration (8, 11). We agree with the authors that the wide interindividual and intraindividual variability in MMF pharmacokinetics makes its dose a less than optimal indicator of its effect at the cellular level. For the same reason, a single MPA trough level suffers from similar shortcomings and is the reason that there is no consensus on its use for drug monitoring or its optimal target level. Therefore, acknowledging the pitfall of using the drug dose, we are not sure that MPA trough level is a better surrogate for drug exposure and effect. Moreover, the use of complete or abbreviated AUC for comparing the drug exposure between the cases and controls in this setting is not practically feasible, because a single testing after the diagnosis may not reflect the exposure over time, and retrospective AUC measurements at different time points would not be possible. To better study this matter, a prospective cohort study would be a much better design, but the relatively low incidence of the disease would require enrolment of a large cohort, which would make it an expensive study and impair its feasibility. Published guidelines advocate reduction in immunosuppression as the first step after a diagnosis of BKN (12). However, no specific approach has been established regarding whether calcineurin inhibitors (CNI) or antiproliferative agents should be reduced first. Clinical trials to validate different strategies are lacking and published reports on different immunosuppression reduction protocols contain mixed results (13). Therefore, the question on how to manage immunosuppressive agents in this patient population remains uncertain. In our study, we suggested reduction in tacrolimus level as the first step, while maintaining the MMF dose, based on our observation of a significant association between BKN and tacrolimus level (1). Even though, similar to other strategies, there is no clinical trial to support this recommendation, evidence continue to accumulate supporting the benefit of significant reduction in CNI. Recently, Egli et al. (14) explored BK virus-specific T-cell response as a marker of antiviral immune protection. The study found that BK virus-specific T-cell response in vitro inversely correlated with tacrolimus concentration, although there was no relationship with MMF concentration. This further supports the idea that reduction in CNI level would be a better first step for the management of BKN. The risk of rejection after reduction in immunosuppression remains a valid concern, as acute rejection rates as high as 75% have been reported after reduction in immunosuppression with various protocols (13). However, we believe that with our suggested strategy, this risk will not be higher than the traditional reduction in MPA and maintenance of CNI as the first step. In this setting, we agree that the therapeutic drug monitoring for MPA level might have a role in the prevention of acute rejection (7–9). In conclusion, we admit that well-designed clinical trials would be the best way of studying the role of differential reduction in immunosuppressive agents in patients with BKN and use of standard pharmacokinetic methods to examine the drug exposure provides more accurate data; however, in the absence of such studies, the use of observational clinical data such as ours and in vitro studies would provide a better rationale for a therapeutic approach than following practices that are not based on tangible evidence. Wana Manitpisitkul1 Abdolreza Haririan2 1Department of Pharmacy, University of Maryland Medical Center Baltimore, Maryland 2 Division of Nephrology University of Maryland Medical Center Baltimore, Maryland

Core-specific adaptive regulatory T-cells in different outcomes of hepatitis C

CD4+ Treg-cells (regulatory T-cells) probably contribute to the impaired virus-specific T-cell responses in chronic HCV (hepatitis C virus) infection; however, their antigen-specificity has remained elusive. In the present study, we analysed peripheral blood CD4+ Treg-cells in patients with chronic hepatitis C and subjects with self-limited HCV infection and characterized individual Treg-cell clones obtained from both groups at the phenotypic and functional level. Foxp3 (forkhead box p3)+CD25+CD4+ Treg-cells were detected more frequently in patients with chronic hepatitis C than self-limited HCV infection, which responded to HCV core stimulation and inhibited proliferation of reporter cells. Cloning under limiting dilution conditions resulted in 14 and six hypoproliferative Foxp3+CD25+CD127-CD4+ T-cell clones from patients with chronic hepatitis C and subjects with self-limited HCV infection respectively. All clones expressed Treg-cell markers and produced IL (interleukin)-10 upon mitogen stimulation. However, exclusively Treg-cell clones from chronic hepatitis C produced IL-10 in response to HCV core and inhibited proliferation of reporter T-cells. These core-specific Treg-cell clones recognized epitopes in two regions of HCV core (amino acids 1-44 and 79-113). Co-culture inhibition assays demonstrated Treg-cells to inhibit reporter T-cells via secretion of IL-10 and IL-35 rather than cell-contact-dependent mechanisms. Finally, the HCV-specific Treg-cell clones lost their functional capacity, along with Foxp3 expression, if kept in culture without HCV core exposure. In conclusion, we identified functionally active HCV core-specific Treg-cells in patients with chronic hepatitis C, which share their epitopes with conventional T-cells and require the continued presence of antigen to maintain their functional differentiation. Thus HCV core-specific Treg-cells may contribute to the immunoregulatory balance in chronic hepatitis C.

Occult cytomegalovirus in vivarium-housed mice may influence transplant allograft acceptance

We have recently shown that latent murine cytomegalovirus (MCMV) can influence murine transplant allograft acceptance. During these studies we became aware that vivarium-housed control mice can acquire occult MCMV infection. The purpose of this investigation was to confirm occult MCMV transmission and determine the timing, vehicle, and possible consequences of transmission. Mice arriving from a commercial vendor were negative for MCMV both by commercial serologic testing and by our nested PCR. Mice housed in our vivarium became positive for MCMV DNA 30–60 days after arrival, but remained negative for MCMV by commercial serologic testing. To confirm MCMV we sequenced PCR products for several genes and showed > 99% homology to MCMV. Further sequence analyses show that the occult MCMV is similar to a laboratory strain of MCMV, but the vehicle of transmission remains unclear. Control tissues from historical experiments with unexplained graft losses were evaluated for occult MCMV, and mice with unexplained allograft losses showed significantly higher incidence of occult MCMV than did allograft acceptors. Deliberate infection with very low titer MCMV confirmed that viral transmission can occur without measurable virus specific antibody or T-cell responses. These data suggest that vivarium-housed mice can develop occult MCMV that is missed by currently available commercial serologic testing, and that these infections may influence transplant allograft acceptance.

Cytotoxic T Cells Are the Predominant Players Providing Cross-Protective Immunity Induced by γ-Irradiated Influenza A Viruses

We previously demonstrated that a single dose of nonadjuvanted intranasal gamma-irradiated influenza A virus can provide robust protection in mice against both homologous and heterosubtypic challenges, including challenge with an H5N1 avian virus strain. We investigated the mechanism behind the observed cross-protection to define which arms of the adaptive immune response are involved in mediating this protection. Studies with gene knockout mice showed the cross-protective immunity to be mediated mainly by T cells and to be dependent on the cytolytic effector molecule perforin. Adoptive transfer of memory T cells from immunized mice, but not of memory B cells, protected naïve recipients against lethal heterosubtypic influenza virus challenge. Furthermore, gamma-irradiated influenza viruses induced cross-reactive Tc-cell responses but not cross-neutralizing or cross-protective antibodies. In addition, histological analysis showed reduced lung inflammation in vaccinated mice compared to that in unvaccinated controls following heterosubtypic challenge. This reduced inflammation was associated with enhanced early recruitment of T cells, both CD4(+) and CD8(+), and with early influenza virus-specific cytotoxic T-cell responses. Therefore, cross-protective immunity induced by vaccination with gamma-irradiated influenza A virus is mediated mainly by Tc-cell responses.

Immunotherapy for Epstein-Barr virus-associated malignancies

Epstein-Barr virus (EBV) is present in the malignant cells of several human cancers including post-transplant lymphoproliferative disease (PTLD), Hodgkin's lymphoma, nasopharyngeal carcinoma, natural killer/T lymphomas and Burkitt's lymphoma. Yet in > 90% of the world's adult population, who carry EBV as a lifelong asymptomatic infection, the oncogenic potential of this virus is controlled by a strong virus-specific T-cell response. Accordingly, EBV-associated malignancies represent good candidates for a T-cell-based therapy and provide an important model for developing such therapies for other human cancers. This review summarizes the impressive results seen with T-cell therapy for PTLD and discusses, in the light of recent technological advances, the prospects for developing more effective approaches for other EBV-associated cancers.

The regulatory T-cell response during acute retroviral infection is locally defined and controls the magnitude and duration of the virus-specific cytotoxic T-cell response

AbstractCytotoxic CD8+ T cells control acute viremia in many viral infections. However, most viruses that establish chronic infections evade destruction by CD8+ T cells, and regulatory T cells (Treg) are thought to be involved in this immune evasion. We have infected transgenic mice, in which Treg can be selectively depleted, with Friend retrovirus (FV) to investigate the influence of Treg on pathogen-specific CD8+ T-cell responses in vivo. We observed that Treg expansion during acute infection was locally defined to organs with high viral loads and massive activation of virus-specific effector CD8+ T cells. Experimental ablation of Treg resulted in a significant increase of peak cytotoxic CD8+ T-cell responses against FV. In addition, it prevented the development of functional exhaustion of CD8+ T cells and significantly reduced FV loads in lymphatic organs. Surprisingly, despite the massive virus-specific CD8+ T-cell response after temporary Treg depletion, no evidence of immunopathology was found. These results demonstrate the important role of Treg in controlling acute retrovirus-specific CD8+ T-cell responses, and suggest that temporary manipulation of Treg might be a possible therapeutic approach in chronic infectious diseases.

P16-32. Antigen exposure regulates the balance between proliferating and cytotoxic subsets of virus-specific CD8 T-cells

Methods Virus-specific CD8 T-cell responses (n = 74) including CMV, EBV, Flu and HIV-1 were characterized using tetramer complexes or peptide stimulation and analyzed for proliferation capacity, expression of CD127 and perforin. CCR7, CD45RA, PD-1 and CD57 were used to assess T-cell differentiation/exhaustion. Simultaneous expression of IFNγ, IL-2, TNFα and perforin was analysed by polychromatic flow cytometry.

Asymptomatic Reactivation of JC Virus in Patients Treated with Natalizumab

Progressive multifocal leukoencephalopathy (PML) occurs in a fraction of patients with multiple sclerosis who were treated with natalizumab. Most adults who are infected with the JC virus, the etiologic agent in PML, do not have symptoms. We sought to determine whether exposure to natalizumab causes subclinical reactivation and neurotropic transformation of JC virus. We followed 19 consecutive patients with multiple sclerosis who were treated with natalizumab over an 18-month period, performing quantitative polymerase-chain-reaction assays in blood and urine for JC virus reactivation; BK virus, a JC virus-related polyomavirus, was used as a control. We determined JC virus-specific T-cell responses by means of an enzyme-linked immunospot assay and antibody responses by means of an enzyme-linked immunosorbent assay and analyzed JC virus regulatory-region sequences. After 12 months of natalizumab therapy, the prevalence of JC virus in the urine of the 19 patients increased from a baseline value of 19% to 63% (P=0.02). After 18 months of treatment, JC virus was detectable in 3 of 15 available plasma samples (20%) and in 9 of 15 available samples of peripheral-blood mononuclear cells (60%) (P=0.02). JC virus regulatory-region sequences in blood samples and in most of the urine samples were similar to those usually found in PML. Conversely, BK virus remained stable in urine and was undetectable in blood. The JC virus-specific cellular immune response dropped significantly between 6 and 12 months of treatment, and variations in the cellular immune response over time tended to be greater in patients in whom JC viremia developed. None of the patients had clinical or radiologic signs of PML. Subclinical reactivation of JC virus occurs frequently in natalizumab-treated patients with multiple sclerosis. Viral shedding is associated with a transient drop in the JC virus-specific cellular immune response.

Impact of Cytotoxic-T-Lymphocyte Memory Induction without Virus-Specific CD4 + T-Cell Help on Control of a Simian Immunodeficiency Virus Challenge in Rhesus Macaques

Despite many efforts to develop AIDS vaccines eliciting virus-specific T-cell responses, whether induction of these memory T cells by vaccination before human immunodeficiency virus (HIV) exposure can actually contribute to effective T-cell responses postinfection remains unclear. In particular, induction of HIV-specific memory CD4(+) T cells may increase the target cell pool for HIV infection because the virus preferentially infects HIV-specific CD4(+) T cells. However, virus-specific CD4(+) helper T-cell responses are thought to be important for functional CD8(+) cytotoxic-T-lymphocyte (CTL) induction in HIV infection, and it has remained unknown whether HIV-specific memory CD8(+) T cells induced by vaccination without HIV-specific CD4(+) T-cell help can exert effective responses after virus exposure. Here we show the impact of CD8(+) T-cell memory induction without virus-specific CD4(+) T-cell help on the control of a simian immunodeficiency virus (SIV) challenge in rhesus macaques. We developed a prophylactic vaccine by using a Sendai virus (SeV) vector expressing a single SIV Gag(241-249) CTL epitope fused with enhanced green fluorescent protein (EGFP). Vaccination resulted in induction of SeV-EGFP-specific CD4(+) T-cell and Gag(241-249)-specific CD8(+) T-cell responses. After a SIV challenge, the vaccinees showed dominant Gag(241-249)-specific CD8(+) T-cell responses with higher effector memory frequencies in the acute phase and exhibited significantly reduced viral loads. These results demonstrate that virus-specific memory CD8(+) T cells induced by vaccination without virus-specific CD4(+) T-cell help could indeed facilitate SIV control after virus exposure, indicating the benefit of prophylactic vaccination eliciting virus-specific CTL memory with non-virus-specific CD4(+) T-cell responses for HIV control.

44. Chronic social stress influences virus specific T-cell response to LCMV infection

44. Chronic social stress influences virus specific T-cell response to LCMV infection

Transient Nature of Long-Term Nonprogression and Broad Virus-Specific Proliferative T-Cell Responses with Sustained Thymic Output in HIV-1 Controllers

BackgroundHIV-1+ individuals who, without therapy, conserve cellular anti-HIV-1 responses, present with high, stable CD4+ T-cell numbers, and control viral replication, facilitate analysis of atypical viro-immunopathology. In the absence of universal definition, immune function in such HIV controllers remains an indication of non-progression.Methodology/Principal FindingsCD4 T-cell responses to a number of HIV-1 proteins and peptide pools were assessed by IFN-γ ELISpot and lymphoproliferative assays in HIV controllers and chronic progressors. Thymic output was assessed by sjTRECs levels. Follow-up of 41 HIV-1+ individuals originally identified as “Long-term non-progressors” in 1996 according to clinical criteria, and longitudinal analysis of two HIV controllers over 22 years, was also performed. HIV controllers exhibited substantial IFN-γ producing and proliferative HIV-1-specific CD4 T-cell responses to both recombinant proteins and peptide pools of Tat, Rev, Nef, Gag and Env, demonstrating functional processing and presentation. Conversely, HIV-specific T-cell responses were limited to IFN-γ production in chronic progressors. Additionally, thymic output was approximately 19 fold higher in HIV controllers than in age-matched chronic progressors. Follow-up of 41 HIV-1+ patients identified as LTNP in 1996 revealed the transitory characteristics of this status. IFN-γ production and proliferative T-cell function also declines in 2 HIV controllers over 22 years.ConclusionsAlthough increased thymic output and anti-HIV-1 T-cell responses are observed in HIV controllers compared to chronic progressors, the nature of nonprogressor/controller status appears to be transitory.

Dynamic decrease in PD-1 expression correlates with HBV-specific memory CD8 T-cell development in acute self-limited hepatitis B patients

Programmed death-1 (PD-1) upregulation can impair virus-specific CD8 T-cell responses during chronic viral infection. Whether and how PD-1 affects virus-specific memory CD8 T cells in humans with acute viral infection, however, remains largely undefined. The association between PD-1 expression and HBV-specific memory CD8 T-cell responses were longitudinally analyzed in eighteen patients with acute hepatitis B virus (HBV) infection, including ten patients with human leucocyte antigen (HLA)-A201 and eight with other HLA-A2 subtypes. At clinical onset, PD-1 was significantly up-regulated and subsequently led to the functional suppression of HBV-specific effector CD8 T cells, as blocking PD-1/PD-L1 interactions in vitro enhanced their proliferation and IFN-gamma production. Following disease resolution, HBV-specific effector CD8 T cells developed into memory T cells. During this period, the dynamic PD-1 decrease was numerically correlated with the reduction of HBV-specific CD8 T-cell frequency, phenotypically with an acquisition of CCR7, CD45RA and CD127 expressions, and functionally with the increase in proliferation and IFN-gamma production of the memory T cells. PD-1-mediated inhibitory signaling not only attenuates HBV-specific CD8 T-cell effector function during the acute phase of infection but also correlates with the development of HBV-specific memory CD8 T cells following disease resolution.

Polyfunctional CD4 + T-Cell Induction in Neutralizing Antibody-Triggered Control of Simian Immunodeficiency Virus Infection

Rapid depletion of memory CD4(+) T cells and delayed induction of neutralizing antibody (NAb) responses are characteristics of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) infections. Although it was speculated that postinfection NAb induction could have only a limited suppressive effect on primary HIV replication, a recent study has shown that a single passive NAb immunization of rhesus macaques 1 week after SIV challenge can result in reduction of viral loads at the set point, indicating a possible contribution of postinfection NAb responses to virus control. However, the mechanism accounting for this NAb-triggered SIV control has remained unclear. Here, we report rapid induction of virus-specific polyfunctional T-cell responses after the passive NAb immunization postinfection. Analysis of SIV Gag-specific responses of gamma interferon, tumor necrosis factor alpha, interleukin-2, macrophage inflammatory protein 1beta, and CD107a revealed that the polyfunctionality of Gag-specific CD4(+) T cells, as defined by the multiplicity of these responses, was markedly elevated in the acute phase in NAb-immunized animals. In the chronic phase, despite the absence of detectable NAbs, virus control was maintained, accompanied by polyfunctional Gag-specific T-cell responses. These results implicate virus-specific polyfunctional CD4(+) T-cell responses in this NAb-triggered virus control, suggesting possible synergism between NAbs and T cells for control of HIV/SIV replication.

Impact of Epitope Escape on PD-1 Expression and CD8 T-Cell Exhaustion during Chronic Infection

During some persistent viral infections, virus-specific T-cell responses wane due to the antigen-specific deletion or functional inactivation (i.e., exhaustion) of responding CD8 T cells. T-cell exhaustion often correlates with high viral load and is associated with the expression of the inhibitory receptor PD-1. In other infections, functional T cells are observed despite high levels of pathogen persistence. The reasons for these different T-cell fates during chronic viral infections are not clear. Here, we tracked the fate of virus-specific CD8 T cells in lymphocytic choriomeningitis virus (LCMV)-infected mice during viral clearance, the persistence of wild-type virus, or the selection and persistence of a viral variant that abrogates the presentation of a single epitope. Viral clearance results in PD-1(lo) functional virus-specific CD8 T cells, while the persistence of wild-type LCMV results in high PD-1 levels and T-cell exhaustion. However, following the emergence of a GP35V-->A variant virus that abrogates the presentation of the GP33 epitope, GP33-specific CD8 T cells remained functional, continued to show low levels of PD-1, and reexpressed CD127, a marker of memory T-cell differentiation. In the same animals and under identical environmental conditions, CD8 T cells recognizing nonmutated viral epitopes became physically deleted or were PD-1(hi) and nonfunctional. Thus, the upregulation of PD-1 and the functional inactivation of virus-specific T cells during chronic viral infection is dependent upon continued epitope recognition. These data suggest that optimal strategies for vaccination should induce high-magnitude broadly specific T-cell responses that prevent cytotoxic T-lymphocyte escape and highlight the need to evaluate the function of vaccine-induced T cells in the context of antigens presented during virus persistence.

Primary Occult Hepadnavirus Infection Induces Virus-Specific T-Cell and Aberrant Cytokine Responses in the Absence of Antiviral Antibody Reactivity in the Woodchuck Model of Hepatitis B Virus Infection

Although the virological features of serologically silent hepadnaviral primary occult infection (POI) have been relatively well recognized in the woodchuck model of hepatitis B virus infection, the characteristics of accompanying immune responses remain unknown. In this study, the kinetics of woodchuck hepatitis virus (WHV)-specific and generalized (mitogen-induced) T-cell proliferative responses and cytokine expression profiles in circulating lymphoid cells and the liver, along with WHV-specific antibody responses, were investigated during experimentally induced POI and subsequent challenge with a liver-pathogenic dose (>10(3) virions) or liver-nonpathogenic dose (50 virions) of the same virus. The data revealed that POI, which does not prompt WHV surface antigenemia, antiviral antibody response, and hepatitis or protect from challenge with a liver-pathogenic virus dose, was accompanied by the appearance of a strong WHV-specific T-cell response directed against multiple viral epitopes that intermittently persisted at low levels for up to 10-months during follow-up. Furthermore, immediately after exposure to a liver-nonpathogenic dose of WHV, lymphocytes acquired a heightened capacity to proliferate in response to mitogenic stimuli and displayed augmented expression of alpha interferon, interleukin-12 (IL-12), and IL-2, but not tumor necrosis factor alpha. Overall, the kinetics of WHV-specific and mitogen-induced T-cell proliferative and cytokine responses in POI were closely comparable to those seen in infection induced by liver-pathogenic viral doses. The data demonstrated that virus-specific T-cell proliferative reactivity is a very sensitive indicator of exposure to hepadnavirus, even to small amounts inducing serologically mute infection. They also showed that hepadnaviral POI is not only a molecularly but also an immunologically identifiable and distinctive entity.

Evaluation of the immunogenicity of replication-competent V-knocked-out and replication-defective F-deleted Sendai virus vector-based vaccines in macaques

Viral vectors are promising vaccine tools for eliciting antigen-specific T-cell responses. We previously showed the potential of recombinant Sendai virus (SeV) vectors to induce virus-specific T-cell responses in macaque AIDS models. Here, we have evaluated the immunogenicity of replication-competent V-knocked-out and replication-defective F-deleted SeV vectors in macaques. Intranasal replication-competent and replication-defective SeV immunizations both elicited robust systemic antigen-specific T-cell responses, whereas the responses induced by the former were more durable than those by the latter. However, even the latter-induced T-cell responses remained detectable in a local, retropharyngeal lymph node two months after the immunization. These findings are useful for establishment of a vaccine protocol using SeV vectors.