Adaptive immune responses to hepatitis C virus: from viral immunobiology to a vaccine

Hepatitis C virus (HCV) causes chronic infection in approximately two-thirds of cases, leading to chronic hepatitis, liver cirrhosis, liver disease, liver failure, and hepatocellular carcinoma in a substantial proportion of the 170 million HCV-infected individuals worldwide. It is generally accepted that the cellular immune response plays the most important role in determining the outcome of HCV infection. First, vigorous, multispecific and sustained CD4+ and CD8+ T-cell responses are associated with viral clearance. Second, depletion studies in chimpanzees, the only other host of HCV besides humans, have shown that both CD4+ and CD8+ T-cells are required for virus elimination. Third, the host's human leukocyte antigen alleles, which restrict the repertoire of CD4+ and CD8+ T-cell responses, influence the outcome of infection. Of note, protective immunity has been demonstrated in population-based studies, as well as in experimentally infected chimpanzees. Thus, a detailed understanding of the mechanisms contributing to the failure of the antiviral immune response should allow successful development of prophylactic and therapeutic vaccination strategies.     

High,Broad, Polyfunctional,and Durable T Cell Immune Responses Induced in Mice by a Novel Hepatitis C Virus (HCV) Vaccine Candidate (MVA-HCV) Based on Modified Vaccinia Virus Ankara Expressing the Nearly Full-Length HCV Genome

A major goal in the control of hepatitis C infection is the development of a vaccine. Here, we have developed a novel HCV vaccine candidate based on the highly attenuated poxvirus vector MVA (referred to as MVA-HCV) expressing the nearly full-length (7.9-kbp) HCV sequence, with the aim to target almost all of the T and B cell determinants described for HCV. In infected cells, MVA-HCV produces a polyprotein that is subsequently processed into the structural and nonstructural HCV proteins, triggering the cytoplasmic accumulation of dense membrane aggregates. In both C57BL/6 and transgenic HLA-A2-vaccinated mice, MVA-HCV induced high, broad, polyfunctional, and long-lasting HCV-specific T cell immune responses. The vaccine-induced T cell response was mainly mediated by CD8 T cells; however, although lower in magnitude, the CD4⁺ T cells were highly polyfunctional. In homologous protocol (MVA-HCV/MVA-HCV) the main CD8⁺ T cell target was p7+NS2, whereas in heterologous combination (DNA-HCV/MVA-HCV) the main target was NS3. Antigenic responses were also detected against other HCV proteins (Core, E1-E2, and NS4), but the magnitude of the responses was dependent on the protocol used. The majority of the HCV-induced CD8⁺ T cells were triple or quadruple cytokine producers. The MVA-HCV vaccine induced memory CD8⁺ T cell responses with an effector memory phenotype. Overall, our data showed that MVA-HCV induced broad, highly polyfunctional, and durable T cell responses of a magnitude and quality that might be associated with protective immunity and open the path for future considerations of MVA-HCV as a prophylactic and/or therapeutic vaccine candidate against HCV.     

Control of heterologous hepatitis C virus infection in chimpanzees is associated with the quality of vaccine-induced peripheral T-helper immune response

Prophylactic hepatitis C virus (HCV) vaccine trials with human volunteers are pending. There is an important need for immunological end points which correlate with vaccine efficacy and which do not involve invasive procedures, such as liver biopsies. By using a multicomponent DNA priming-protein boosting vaccine strategy, na?ve chimpanzees were immunized against HCV structural proteins (core, E1, and E2) as well as a nonstructural (NS3) protein. Following immunization, exposure to the heterologous HCV 1b J4 subtype resulted in a peak of plasma viremia which was lower in both immunized animals. Compared to the na?ve infection control and nine additional historical controls which became chronic, vaccinee 2 (Vac2) rapidly resolved the infection, while the other (Vac1) clearly controlled HCV infection. Immunization induced antibodies, peptide-specific gamma interferon (IFN-gamma), protein-specific lymphoproliferative responses, IFN-gamma, interleukin-2 (IL-2), and IL-4 T-helper responses in both vaccinees. However, the specificities were markedly different: Vac2 developed responses which were lower in magnitude than those of Vac1 but which were biased towards Th1-type cytokine responses for E1 and NS3. This proof-of-principle study in chimpanzees revealed that immunization with a combination of nonstructural and structural antigens elicited T-cell responses associated with an alteration of the course of infection. Our findings provide data to support the concept that the quality of the response to conserved epitopes and the specific nature of the peripheral T-helper immune response are likely pivotal factors influencing the control and clearance of HCV infection.     

Antigen-specific CD4+ T cells produce sufficient IFN-gamma to mediate robust protective immunity against genital Chlamydia muridarum infection

Chlamydia has been shown to evade host-specific IFN-gamma-mediated bacterial killing; however, IFN-gamma-deficient mice exhibit suboptimal late phase vaginal Chlamydia muridarum clearance, greater dissemination, and oviduct pathology. These findings introduce constraints in understanding results from murine chlamydial vaccination studies in context of potential implications to humans. In this study, we used mice deficient in either IFN-gamma or the IFN-gamma receptor for intranasal vaccination with a defined secreted chlamydial Ag, chlamydial protease-like activity factor (CPAF), plus CpG and examined the role of IFN-gamma derived from adoptively transferred Ag-specific CD4+ T cells in protective immunity against genital C. muridarum infection. We found that early Ag-specific IFN-gamma induction and CD4+ T cell infiltration correlates with the onset of genital chlamydial clearance. Adoptively transferred IFN-gamma competent CPAF-specific CD4+ T cells failed to enhance the resolution of genital chlamydial infection within recipient IFN-gamma receptor-deficient mice. Conversely, IFN-gamma production from adoptively transferred CPAF-specific CD4+ T cells was sufficient in IFN-gamma-deficient mice to induce early resolution of infection and reduction of subsequent pathology. These results provide the first direct evidence that enhanced anti-C. muridarum protective immunity induced by Ag-specific CD4+ T cells is dependent upon IFN-gamma signaling and that such cells produce sufficient IFN-gamma to mediate the protective effects. Additionally, MHC class II pathway was sufficient for induction of robust protective anti-C. muridarum immunity. Thus, targeting soluble candidate Ags via MHC class II to CD4+ T cells may be a viable vaccine strategy to induce optimal IFN-gamma production for effective protective immunity against human genital chlamydial infection.     

Optimized vaccination regimen linked to exhaustive screening approaches identifies 2 novel HLA-B7 restricted epitopes within hepatitis C virus NS3 protein

Broad immune responses, in particular specific for the NS3 protein and mediated by both CD8+ and CD4+T lymphocytes, are thought to play a critical role in the control of hepatitis C virus (HCV) infection. In this study, we searched for novel HLA-B*0702 NS3 restricted epitopes following an optimized NS3NS4 immunization protocol in transgenic mice expressing HLA-B*0702 molecule. Combining predicted and overlapping peptides, we identified two novel epitopes, WPA10 (aa 1111-1120) and LSP10 (aa 1153-1162), which triggered significant IFN-gamma-producing T cell frequencies and high CTL responses. Both epitopes were shown to be immunogenic when used as synthetic peptides to immunize mice. The relevance of these epitopes to humans was demonstrated, as both were able in vitro to recall specific IFN-gamma and IL10-producing cells from peripheral blood mononuclear cells of HCV infected patients. Such epitopes enlarge the pool of NS3-specific CD8+T cell epitopes available to perform immunomonitoring of HCV infection and to develop vaccines.     

The clearance of hepatitis C virus infection in chimpanzees may not necessarily correlate with the appearance of acquired immunity

Clearance of hepatitis C virus (HCV) infection in humans and chimpanzees is thought to be associated with the induction of strong T-cell responses. We studied four chimpanzees infected with HCV derived from an infectious full-length HCV genotype 1b cDNA. Two of the chimpanzees cleared the infection to undetectable levels for more than 12 months of follow-up; the other two became persistently infected. Detailed analyses of HCV-specific immune responses were performed during the courses of infection in these chimpanzees. Only weak and transient T helper responses were detected during the acute phase in all four chimpanzees. A comparison of the frequency of gamma interferon (IFN-gamma)-producing CD4(+) and CD8(+) T cells in peripheral blood by ELISpot assay did not reveal any correlation between viral clearance and T-cell responses. In addition, analyses of IFN-gamma, IFN-alpha, and interleukin-4 mRNA levels in liver biopsies, presumably indicative of intrahepatic T-cell responses, revealed no distinct pattern in these chimpanzees with respect to infection outcome. The present study suggests that the outcome of HCV infection in chimpanzees is not necessarily attributable to HCV sequence variation and that chimpanzees may recover from HCV infection by mechanisms other than the induction of readily detectable HCV-specific T-cell responses.     

Immune-based novel therapies for chronic hepatitis C virus infection

Hepatitis C virus (HCV) infection is a great public health problem, with an estimated 200 million chronically infected patients worldwide. No vaccines are currently available for HCV, and only a subset of HCV patients responds to interferon-alpha (IFN-alpha) and Ribavirin treatment. Substantial evidence has emerged recently to support the role of the host immune response in the outcome and pathogenesis of HCV infection. Our aims of this article are to present the immune-based novel therapeutic options for HCV infection and the evidence supporting their use in patients with chronic hepatitis C. There is a growing consensus that acute control of HCV infection is associated with a vigorous intrahepatic antiviral CD4+ and CD8+ T cell response. IFN-gamma was detectable in the livers of the chimpanzees that cleared or controlled the virus, raising the possibility that IFN-gamma might perform antiviral effector functions during HCV infection. Based on these observations, therapeutic induction of intrahepatic IFN-gamma by adoptive immunotherapy might be able to control chronic HCV infection. Immune-based novel therapies appear to hold great promise in treating chronic HCV infection.     

Protective vaccination with hepatitis C virus NS3 but not core antigen in a novel mouse challenge model

BACKGROUND: Efficient vaccines against hepatitis C virus (HCV) infection are urgently needed. Vaccine development has been hampered by the lack of suitable small animal models to reliably test the protective capacity of immmunization. METHODS: We used recombinant murine gammaherpesvirus 68 (MHV-68) as a novel challenge virus in mice and tested the efficacy of heterologous candidate human vaccines based on modified vaccinia virus Ankara or adenovirus, both delivering HCV non-structural NS3 or core proteins. RESULTS: Recombinant MHV-68 expressing NS3 (MHV-68-NS3) or core (MHV-68-core) were constructed and characterized in vitro and in vivo. Mice immunized with NS3-specific vector vaccines and challenged with MHV-68-NS3 were infected but showed significantly reduced viral loads in the acute and latent phase of infection. NS3-specific CD8+ T cells were amplified in immunized mice after challenge with MHV-68-NS3. By contrast, we did neither detect a reduction of viral load nor an induction of core-specific CD8+ T cells after core-specific immunization. CONCLUSIONS: Our data suggest that the challenge system using recombinant MHV-68 is a highly suitable model to test the immunogenicity and protective capacity of HCV candidate vaccine antigens. Using this system, we demonstrated the usefulness of NS3-specific immunization. By contrast, our analysis rather discarded core as a vaccine antigen.     

Induction of Intrahepatic HCV NS4B,NS5A and NS5B-Specific Cellular Immune Responses following Peripheral Immunization

Numerous studies have suggested that an effective Hepatitis C Virus (HCV) vaccine must induce strong cytotoxic and IFN-γ+ T cell responses targeting the non-structural region of the virus. Most importantly, these responses must be able to migrate into and remain functional within the liver, an organ known to cause T cell tolerance. Using three novel HCV DNA vaccines encoding non-structural proteins NS4B, NS5A and NS5B, we assessed the ability of peripheral immunization to induce functional intrahepatic immunity both in the presence and absence of cognate HCV antigen expression within the liver. We have shown that these constructs induced potent HCV-specific CD4+ and CD8+ T cell responses in the spleen of C57BL/6 mice and that these responses were detected within the liver following peripheral immunization. Additionally, using a transfection method to express HCV antigen within the liver, we showed that intrahepatic HCV-specific T cells remained highly functional within the liver and retained the ability to become highly activated as evidenced by upregulation of IFN-γ and clearance of HCV protein expressing hepatocytes. Taken together, these findings suggest that peripheral immunization can induce potent HCV-specific T cell responses able to traffic to and function within the tolerant environment of the liver.     

Conserved hierarchy of helper T cell responses in a chimpanzee during primary and secondary hepatitis C virus infections

Control of hepatitis C virus (HCV) infection could be influenced by the timing and magnitude of CD4+ T cell responses against individual epitopes. We characterized CD4+ T cells targeting seven Pan troglodytes (Patr) class II-restricted epitopes during primary and secondary HCV infections of a chimpanzee. All Patr-DR-restricted HCV epitopes bound multiple human HLA-DR molecules, indicating the potential for overlap in epitopes targeted by both species. Some human MHC class II molecules efficiently stimulated IL-2 production by chimpanzee virus-specific T cell clones. Moreover, one conserved epitope designated NS3(1248) (GYKVLVLNPSV) overlapped a helper epitope that is presented by multiple HLA-DR molecules in humans who spontaneously resolved HCV infection. Resolution of primary infection in the chimpanzee was associated with an initial wave of CD4+ T cells targeting a limited set of dominant epitopes including NS3(1248.) A second wave of low-frequency CD4+ T cells targeting other subdominant epitopes appeared in blood several weeks later after virus replication was mostly contained. During a second infection 7 years later, CD4+ T cells against all epitopes appeared in blood sooner and at higher frequencies but the pattern of dominance was conserved. In summary, primary HCV infection in this individual was characterized by T cell populations targeting two groups of MHC class II-restricted epitopes that differed in frequency and kinetics of appearance in blood. The hierarchial nature of the CD4+ T cell response, if broadly applicable to other HCV-infected chimpanzees and humans, could be a factor governing the outcome of HCV infection.     

Immunopathogenesis of viral hepatitis C. Immunological markers of the disease progression

Results of studies of immune response during hepatitis C virus (HCV) infection were reviewed in order to reveal immunologic markers of the disease progression. Genetic heterogeneity of HCV and immunogenetic features of the host determine heterogeneity of immune response to the virus and differences in the course of the disease and outcomes. Spontaneous elimination of HCV-infection in acute phase occurs due to vigorous and sustained multispecific Th1-response toviral antigens. During such response proliferation of virus-specific CD4+ T-cells and secretion of IFN-gamma by them are observed, otherwise chronic hepatitis develops. Great importance in persistence of HCV as well as in quantitative and functional suppression of HCV-specific CD8+ T-cells has increased number of CD4+ CD25+ regulatory T-cells. Cellular immune response plays a key role not only in the elimination of HCV, but also in liver pathology associated with HCV-infection. Progression of the process and shift to its chronic form are also associated with decrease of production of IFN-gamma, alpha, IL-2 by peripheral blood mononuclear cells and increase of TNF-alpha, IFN-gamma, IL-4, IL-2r levels in blood serum.     

Impaired effector function of hepatitis C virus-specific CD8+ T cells in chronic hepatitis C virus infection

The cellular immune response contributes to clearance of hepatitis C virus (HCV) and persists for decades after recovery from infection. The immunological basis for the inefficiency of the cellular immune response in chronically infected persons is not known. Here, we used four HLA-A2 tetramers, specific for two HCV core and two HCV NS3 epitopes, to investigate at the single-cell level effector function and phenotype of HCV-specific CD8+ T cells in 20 chronically infected and 12 long-term recovered patients. Overall, HCV-specific, tetramer+ T cells were more frequently found in PBMCs of chronically infected patients than in those of recovered patients. However, when compared with HCV-tetramer+ T cells of recovered patients, they displayed an impaired proliferative capacity. As a result of the impaired proliferative capacity, HCV-specific T cell lines derived from chronically infected patients displayed less peptide-specific cytotoxicity than those from recovered patients. In addition, proliferation and ex vivo IFN-gamma production of HCV-tetramer+ cells, but not influenza-virus-specific T cells, were defective in chronically infected patients and could not be restored by in vitro stimulation with peptide and IL-2. At least three distinct phenotypes of HCV-specific CD8+ T cells were identified and associated with certain functional characteristics. In addition, impairment of proliferative, cytokine, and cytotoxic effector functions of tetramer+ T cells in viremic patients was associated with weak ex vivo HCV-specific CD4+ T cell responses. Thus, the defective functions of HCV-specific CD8+ T cells might contribute to viral persistence in chronically infected patients, and knowledge on their reversibility may facilitate the development of immunotherapeutic vaccines.     

A novel adenovirus type 6 (Ad6)-based hepatitis C virus vector that overcomes preexisting anti-ad5 immunity and induces potent and broad cellular immune responses in rhesus macaques

Success in resolving hepatitis C virus (HCV) infection has been correlated to vigorous, multispecific, and sustained CD8(+) T-cell response in humans and chimpanzees. The efficacy of inducing T-cell-mediated immunity by recombinant serotype 5 adenovirus vector has been proven in many animal models of infectious diseases, but its immunogenicity can be negatively influenced by preexisting immunity against the vector itself. To evaluate the less prevalent adenovirus serotype 6 (Ad6) as an alternative vector for and HCV vaccine development, we have generated serotype 5 and 6 adenoviral vectors directing expression of the nonstructural region of HCV (MRKAd5-NSmut and MRKAd6-NSmut). Immunogenicity studies in mice showed that the two vectors induced comparable T-cell responses but that only MRKAd6-NSmut was not suppressed in the presence of anti-Ad5 immunity. In contrast, preexisting anti-Ad5 immunity dramatically blunted the immunogenicity of the serotype 5-based HCV vector. Furthermore, MRKAd6-NSmut showed equivalent potency, breadth, and longevity of HCV-specific T-cell responses in rhesus macaques as the corresponding Ad5-based vector over a wide range of doses and was capable of boosting DNA-primed animals even if administered at low doses. These data support the use of the MRKAd6-NSmut for anti-HCV immunotherapy and, more generally, for the Ad6 serotype as a better genetic vaccine vehicle than Ad5.     

IL-15 prolongs the duration of CD8+ T cell-mediated immunity in mice infected with a vaccine strain of Toxoplasma gondii.

Immunization of mice with a vaccine (ts-4) strain of Toxoplasma gondii is known to induce complete protection against subsequent lethal infection. Ts-4-mediated protection has been reported to be primarily dependent on IFN-gamma-producing CD8+ T cells. However, duration of CD8+ T cell-mediated immunity in the ts-4-vaccinated animals is not known. In the present study, the kinetics of the CD8+ T cell response in mice immunized with the ts-4 strain of T. gondii was evaluated. Optimal CD8+ T cell immunity persisted at least 6 mo after vaccination, and mice at this time point continued to overcome lethal challenge with a more virulent strain. However, at 9 mo postimmunization, CD8+ T cell immunity was severely diminished and the mice succumbed to Toxoplasma challenge. Pretreatment of animals, vaccinated 9 mo earlier, with rIL-15 prevented the mortality induced by Toxoplasma challenge. The protective effect of IL-15 treatment was due to a rise in the frequency of Ag-specific CD8+ T cells. CD8+ T cells from IL-15-administered animals showed increased proliferation and IFN-gamma production in response to antigenic restimulation. These findings suggest that rIL-15 can reverse the decline in the long-term CD8+ T cell immune response in mice immunized with vaccine strain of T. gondii.     

The complexity of protective immunity against liver-stage malaria

Sterile protective immunity against challenge with Plasmodium spp. sporozoites can be induced in multiple model systems and humans by immunization with radiation-attenuated Plasmodium spp. sporozoites. The infected hepatocyte has been established as the primary target of this protection, but the underlying mechanisms have not been completely defined. Abs, CD8+ T cells, CD4+ T cells, cytokines (including IFN-gamma and IL-12), and NO have all been implicated as critical effectors. Here, we have investigated the mechanisms of protective immunity induced by immunization with different vaccine delivery systems (irradiated sporozoites, plasmid DNA, synthetic peptide/adjuvant, and multiple Ag peptide) in genetically distinct inbred strains, genetically modified mice, and outbred mice. We establish that there is a marked diversity of T cell-dependent immune responses that mediate sterile protective immunity against liver-stage malaria. Furthermore, we demonstrate that distinct mechanisms of protection are induced in different strains of inbred mice by a single method of immunization, and in the same strain by different methods of immunization. These data underscore the complexity of the murine host response to a parasitic infection and suggest that an outbred human population may behave similarly. Data nevertheless suggest that a pre-erythrocytic-stage vaccine should be designed to induce CD8+ T cell- and IFN-gamma-mediated immune responses and that IFN-gamma responses may represent an in vitro correlate of pre-erythrocytic-stage protective immunity.     

Effective DNA vaccination against listeriosis by prime/boost inoculation with the gene gun.

Protective immunity against Listeria monocytogenes strongly depends on CD8+ T lymphocytes, and both IFN-gamma secretion and target cell killing are considered relevant to protection. We analyzed whether we could induce a protective type 1 immune response by DNA vaccination with the gene gun using plasmids encoding for two immunodominant listerial Ags, listeriolysin and p60. To induce a Th1 response, we 1) coprecipitated a plasmid encoding for GM-CSF, 2) employed a prime/boost vaccination schedule with a 45-day interval, and 3) coinjected oligodeoxynucleotides (ODN) containing immunostimulatory CpG motifs. DNA immunization of BALB/c mice with plasmids encoding for listeriolysin (pChly) and p60 (pCiap) efficiently induced MHC class I-restricted, Ag-specific CD8+ T cells that produced IFN-gamma. Coinjection of CpG-ODN significantly increased the frequency of specific IFN-gamma-secreting T cells. Although pChly induced specific CD8+ T cells expressing CTL activity, it failed to stimulate CD4+ T cells. Only pCiap induced significant CD4+ T cell and humoral responses, which were predominantly of Th2 type. Vaccination with either plasmid induced protective immunity against listerial challenge, and coinjection of CpG ODN improved vaccine efficacy in some situations. This study demonstrates the feasibility of gene gun administration of plasmid DNA for inducing immunity against an intracellular pathogen for which protection primarily depends on type 1 CD8+ T cells.     

Induction of broad CD4+ and CD8+ T-cell responses and cross-neutralizing antibodies against hepatitis C virus by vaccination with Th1-adjuvanted polypeptides followed by defective alphaviral particles expressing envelope glycoproteins gpE1 and gpE2 and nonstructural proteins 3, 4, and 5

Broad, multispecific CD4⁺ and CD8⁺ T-cell responses to the hepatitis C virus (HCV), as well as virus-cross-neutralizing antibodies, are associated with recovery from acute infection and may also be associated in chronic HCV patients with a favorable response to antiviral treatment. In order to recapitulate all of these responses in an ideal vaccine regimen, we have explored the use of recombinant HCV polypeptides combined with various Th1-type adjuvants and replication-defective alphaviral particles encoding HCV proteins in various prime/boost modalities in BALB/c mice. Defective chimeric alphaviral particles derived from the Sindbis and Venezuelan equine encephalitis viruses encoding either the HCV envelope glycoprotein gpE1/gpE2 heterodimer (E1E2) or nonstructural proteins 3, 4, and 5 (NS345) elicited strong CD8⁺ T-cell responses but low CD4⁺ T helper responses to these HCV gene products. In contrast, recombinant E1E2 glycoproteins adjuvanted with MF59 containing a CpG oligonucleotide elicited strong CD4⁺ T helper responses but no CD8⁺ T-cell responses. A recombinant NS345 polyprotein also stimulated strong CD4⁺ T helper responses but no CD8⁺ T-cell responses when adjuvanted with Iscomatrix containing CpG. Optimal elicitation of broad CD4⁺ and CD8⁺ T-cell responses to E1E2 and NS345 was obtained by first priming with Th1-adjuvanted proteins and then boosting with chimeric, defective alphaviruses expressing these HCV genes. In addition, this prime/boost regimen resulted in the induction of anti-E1E2 antibodies capable of cross-neutralizing heterologous HCV isolates in vitro. This vaccine formulation and regimen may therefore be optimal in humans for protection against this highly heterogeneous global pathogen.     

Augmentation of the CD8+ T cell response by IFN-gamma in IL-12-deficient mice during Toxoplasma gondii infection

The importance of IFN-gamma in regulating the host CD8+ T cell response during microbial infection has not been delineated. Mice deficient for the p40 chain of the IL-12 heterodimer have impaired IFN-gamma production and are susceptible to infection with the intracellular parasite Toxoplasma gondii. The administration of exogenous IFN-gamma to parasite-infected p40-/- mice increases survival and up-regulates the depressed CD8+ T cell response following infection. CD8+ T cells isolated from cytokine-treated p40-/- mice exhibit an increase in both precursor CTL frequency and IFN-gamma production compared with untreated controls. The enhancement of the CD8+ T cell response is independent of CD4+ T cell help. These CD8+ T cells induce protective immunity against a lethal challenge when adoptively transferred into naive p40-/- and IFN-gamma-/- mice. These observations indicate that IFN-gamma can regulate the CD8+ T cell response during T. gondii infection.     

IL-10 from regulatory T cells determines vaccine efficacy in murine Leishmania major infection

Leishmaniasis affects 12 million people, but there are no vaccines. Immunological correlates of vaccine efficacy are unclear. Polarized Th1 vs Th2 responses in Leishmania major-infected mice suggested that a shift in balance from IL-4 to IFN-gamma was the key to vaccine success. Recently, a role for IL-10 and regulatory T cells in parasite persistence was demonstrated, prompting re-evaluation of vaccine-induced immunity. We compared DNA/modified vaccinia virus Ankara heterologous prime-boost with Leishmania homolog of the receptor for activated C kinase (LACK) or tryparedoxin peroxidase (TRYP). Both induced low IL-4 and high IFN-gamma prechallenge. Strikingly, high prechallenge CD4 T cell-derived IL-10 predicted vaccine failure using LACK, whereas low IL-10 predicted protection with TRYP. The ratio of IFN-gamma:IL-10 was thus a clear prechallenge indicator of vaccine success. Challenge infection caused further polarization to high IL-10/low IFN-gamma with LACK and low IL-10/high IFN-gamma with TRYP. Ex vivo quantitative RT-PCR and in vitro depletion and suppression experiments demonstrated that Ag-driven CD4+ CD25+ T regulatory 1-like cells were the primary source of IL-10 in LACK-vaccinated mice. Anti-IL-10R treatment in vivo demonstrated that IL-10 was functional in determining vaccine failure, rendering LACK protective in the presence of high IFN-gamma/low IL-5 responses.