Identification of a ligand for glucocorticoid-induced tumor necrosis factor receptor constitutively expressed in dendritic cells

Glucocorticoid-induced tumor necrosis receptor (GITR) has been implicated in regulation of T cell suppression by CD25(+)CD4(+) regulatory T cells (Tregs). We isolated a cDNA encoding GITR ligand (GITRL) from mouse endothelioma cells. When stably expressed in HEK293 cells, its specific interaction with GITR was confirmed by flow cytometry with the use of GITR-Fc. The interaction was greatly diminished by the addition of soluble GITRL. Consistent with this, soluble GITRL bound to the cell surface of the GITR-expressing HEK293 cells. Coexpression of GITR with GITRL or stimulation of the GITR-expressing cells with soluble GITRL led to activation of NF-kappaB, which was significantly reduced by anti-GITR. More importantly, GITRL was expressed by both immature and mature dendritic cells, suggesting that the interaction between GITR and GITRL may contribute to immune regulation of Tregs by dendritic cells. This isolated TNFRL represents a bona fide GITRL whose presence has been elusive until this time.     

Cutting edge: the natural ligand for glucocorticoid-induced TNF receptor-related protein abrogates regulatory T cell suppression

CD4(+)25(+) regulatory T (Treg) cells maintain immunological self-tolerance through mechanisms that are only in part understood. Previous studies suggest that the glucocorticoid-induced TNFR-related protein (GITR), which is preferentially expressed on the surface of Treg cells, potentially provides a signal that abrogates Treg suppression. In this study, we show that a soluble form of mouse GITR ligand (sGITR-L) induces GITR-dependent NF-kappaB activation and blocks in vitro suppression mediated by both resting and preactivated polyclonal and Ag-specific Treg cells. Since sGITR-L along with rIL-2 induces proliferation of CD4(+)25(+) cells, it appears that sGITR-L can break the anergic state of Treg cells. Because sGITR-L also up-regulates IL-2 secretion by activated CD4(+)25 (-)T cells, these two sGITR-L induced signals synergize to interfere with suppressor activity by CD4(+)25(+) Treg cells.     

SIV DNA vaccine co-administered with IL-12 expression plasmid enhances CD8 SIV cellular immune responses in cynomolgus macaques

Current evidence suggests that a strong induced CD8 human immunodeficiency virus type 1 (HIV-1)-specific cell mediated immune response may be an important aspect of an HIV vaccine. The response rates and the magnitude of the CTL responses induced by current DNA vaccines in humans need to be improved and cellular immune responses to DNA vaccines can be enhanced in mice by co-delivering DNA plasmids expressing immune modulators. Two reported to work well in the mouse systems are interleukin (IL)-12 and CD40L. We sought to compare these molecular adjuvants in a primate model system. The cDNA for macaque IL-12 and CD40L were cloned into DNA vectors. Groups of cynomolgus macaques were immunized with 2 mg of plasmid expressing SIVgag alone or in combination with either IL-12 or CD40L. CD40L did not appear to enhance the cellular immune response to SIVgag antigen. However, more robust results were observed in animals co-injected with the IL-12 molecular adjuvant. The IL-12 expanded antigen-specific IFN-gamma positive effector cells as well as granzyme B production. The vaccine immune responses contained both a CD8 component as well a CD4 component. The adjuvanted DNA vaccines illustrate that IL-12 enhances a CD8 vaccine immune response, however, different cellular profiles.     

Trans-splicing repair of CD40 ligand deficiency results in naturally regulated correction of a mouse model of hyper-IgM X-linked immunodeficiency

X-linked immunodeficiency with hyper-IgM (HIGM1), characterized by failure of immunoglobulin isotype switching, is caused by mutations of the CD40 ligand (CD40L), which is normally expressed on activated CD4(+) T cells. As constitutive expression of CD40L induces lymphomas, we corrected the mutation while preserving the natural regulation of CD40L using pre-mRNA trans-splicing. Bone marrow from mice lacking CD40L was modified with a lentivirus trans-splicer encoding the normal CD40L exons 2-5 and was administered to syngenic CD40L-knockout mice. Recipient mice had corrected CD40L mRNA, antigen-specific IgG1 responses to keyhole limpet hemocyanin immunization, regulated CD4(+) T-cell CD40L expression after CD3 stimulation in primary and secondary transplanted mice, attenuation of Pneumocystis carinii pneumonia, and no evidence of lymphoproliferative disease over 1 year. Thus, HIGM1 can be corrected by CD40L trans-splicing, leading to functional correction of the genetic defect without the adverse consequences of unregulated expression of the CD40L gene.     

Soluble CD40 ligand induces beta-chemokine production by macrophages and resistance to HIV-1 entry

CD40 ligand (CD40L) is a cell surface molecule of CD4(+)T cells that interacts with its receptor CD40 on antigen presenting cells to mediate thymus-dependent humoral immunity and inflammatory reactions. We report here that treating monocyte-derived macrophages (MDM) with a trimeric soluble form of CD40L (CD40LT) induced them to secrete high levels of the beta-chemokines RANTES, MIP-1alpha and MIP-1beta that are ligands for CCR5 and able to inhibit HIV-1 entry. CD40LT inhibited the entry of M-tropic HIV-1 reporter viruses. Furthermore, supernatants obtained from CD40LT-stimulated macrophages protected CEMx174-CCR5 cells from infection by HIV-1(JRFL)reporter virus. The inhibitory activity appeared to be due to beta-chemokines present in the supernatant, since pretreating them with a cocktail of antibodies to RANTES, MIP-1alpha and MIP-1beta neutralized the inhibitory activity of the supernatants. In addition, treating monocytes with CD40LT caused CCR5 and CD4 to be downregulated from the cell surface. In vivo, macrophages activated through CD40 could interfere with HIV replication.     

CD40 ligand mutants responsible for X-linked hyper-IgM syndrome associate with wild type CD40 ligand

CD40 ligand (CD40L) is a 33-kDa type II membrane glycoprotein mainly expressed on activated CD4(+) T cells in trimeric form. When it is mutated, the clinical consequences are X-linked hyper-IgM syndrome (XHIM), a primary immunodeficiency disorder characterized by low levels of IgG, IgA, and elevated or normal levels of IgM. Mutated CD40L can no longer bind CD40 nor provide signals for B cells to proliferate and to switch from IgM to other immunoglobulin isotypes. When considering gene therapy for XHIM, it is important to address the possibility that the mutated CD40L associates with transduced wild type CD40L, and as a consequence, immune reconstitution is not attained. In this study, we demonstrate that the various mutated CD40L species we have identified in patients with XHIM, including both full-length and truncated mutants, associate with wild type CD40L on the cell surface of co-transfected COS cells. The association between wild type and mutated CD40L was also observed in CD4(+) T cell lines established from XHIM patients with leaky splice site mutations. The clinical phenotype of these patients suggests that this association between wild type and mutated CD40L species may result in less efficient cross-linking of CD40.     

Recombinant monomeric CD40 ligand for delivering polymer particles to dendritic cells

Dendritic cells (DCs) are considered the most efficient antigen-presenting cells and are therefore ideal targets for in vivo delivery of antigen for vaccines. We are investigating the strategy of using CD40 ligand (CD40L) as a targeting moiety because this protein has the potential to not only target DCs, but also stimulate cell maturation, leading to more potent immune responses. We have shown that a recombinant, monomeric CD40 ligand fusion protein conjugated to polystyrene micro- and nanoparticles led to significantly enhanced uptake by DCs in vitro. This enhancement was observed for particles of both sizes and in both a murine DC cell line and primary DCs. The uptake appeared to be specifically mediated by CD40L binding to CD40 expressed on DCs. Enhanced uptake of nanoparticles in draining lymph nodes of mice was not observed, however, 48 hours after subcutaneous injection. These findings suggest that CD40 ligand may be a potentially useful targeting moiety for delivery of particulate vaccines to DCs, and that further optimization of both CD40L and the polymer carriers is necessary to achieve efficacy in vivo.     

CD40 ligand (CD154) enhances the Th1 and antibody responses to respiratory syncytial virus in the BALB/c mouse

CD40 ligand (CD40L) is a cell surface costimulatory molecule expressed mainly by activated T cells. CD40L is critically important for T-B cell and T cell-dendritic cell interactions. CD40L expression promotes Th1 cytokine responses to protein Ags and is responsible for Ig isotype switching in B cells. Respiratory syncytial virus (RSV) is an important pathogen of young children and the elderly, which causes bronchiolitis and pneumonia. Studies of mice infected with RSV suggest that a Th2 cytokine response may be responsible for enhanced pulmonary disease. To investigate the effect CD40L has on RSV immunity, mice were infected simultaneously with RSV and either an empty control adenovirus vector or one expressing CD40L or were coimmunized with plasmid DNA vectors expressing CD40L and RSV F and/or G proteins and subsequently challenged with RSV. The kinetics of the intracellular and secreted cytokine responses, the cytotoxic T lymphocyte precursor frequency, NO levels in lung lavage, rates of virus clearance, and anti-RSV Ab titers were determined. These studies show that coincident expression of CD40L enhances the Th1 (IL-2 and IFN-gamma) cytokine responses, increases the expression of TNF-alpha and NO, accelerates virus clearance, and increases the anti-F and anti-G Ab responses. These data suggest that CD40L may have the adjuvant properties needed to optimize the safety and efficacy of RSV vaccines.     

ISCOMATRIX vaccines mediate CD8+ T-cell cross-priming by a MyD88-dependent signaling pathway

Generating a cytotoxic CD8(+) T-cell response that can eradicate malignant cells is the primary objective of cancer vaccine strategies. In this study we have characterized the innate and adaptive immune response to the ISCOMATRIX adjuvant, and the ability of vaccine antigens formulated with this adjuvant to promote antitumor immunity. ISCOMATRIX adjuvant led to a rapid innate immune cell response at the injection site, followed by the activation of natural killer and dendritic cells (DC) in regional draining lymph nodes. Strikingly, major histocompatibility complex (MHC) class I cross-presentation by CD8α(+) and CD8α(-) DCs was enhanced by up to 100-fold when antigen was formulated with ISCOMATRIX adjuvant. These coordinated features enabled efficient CD8(+) T-cell cross-priming, which exhibited prophylactic and therapeutic tumoricidal activity. The therapeutic efficacy of an ISCOMATRIX vaccine was further improved when co-administered with an anti-CD40 agonist antibody, suggesting that ISCOMATRIX-based vaccines may combine favorably with other immune modifiers in clinical development to treat cancer. Finally, we identified a requirement for the myeloid differentiation primary response gene 88 (MyD88) adapter protein for both innate and adaptive immune responses to ISCOMATRIX vaccines in vivo. Taken together, our findings support the utility of the ISCOMATRIX adjuvant for use in the development of novel vaccines, particularly those requiring strong CD8(+) T-cell immune responses, such as therapeutic cancer vaccines.     

Cooperation of TNF family members CD40 ligand,receptor activator of NF-kappa B ligand,and TNF-alpha in the activation of dendritic cells and the expansion of viral specific CD8+ T cell memory responses in HIV-1-infected and HIV-1-uninfected individuals

Members of the TNF superfamily have been shown to be instrumental in enhancing cell-mediated immune responses, primarily through their interactions with dendritic cells (DCs). We systematically evaluated the ability of three TNF superfamily molecules, CD40 ligand (CD40L), receptor activator of NF-kappaB ligand (RANKL), and TNF-alpha, to expand ex vivo EBV-specific CTL responses in healthy human individuals and ex vivo HIV-1-specific CTL responses in HIV-1-infected individuals. In both groups of individuals, we found that all three TNF family molecules could expand CTL responses, albeit at differing degrees. CD40L treatment alone was better than RANKL or TNF-alpha alone to mature DCs and to expand CTL. In healthy volunteers, TNF-alpha or RANKL could cooperate with CD40L to maximize the ability of DCs to expand virus-specific CTL responses. In HIV-1 infection, cooperative effects between TNF-alpha or RANKL in combination with CD40L were variable. TNF-alpha and RANKL cooperated with CD40L via differing mechanisms, i.e., TNF-alpha enhanced IL-12 production, whereas RANKL enhanced survival of CD40L-stimulated DCs. These findings demonstrate that optimal maturation of DCs requires multiple signals by TNF superfamily members that include CD40L. In HIV-1 infection, DCs may only require CD40L to maximally expand CTL. Finally, CTL responses were higher in CD4(+) T cell-containing conditions even in the presence of TNF family molecules, suggesting that CD4(+) T cells can provide help to CD8(+) T cells independently of CD40L, RANKL, or TNF-alpha.     

The adjuvant effects of co-stimulatory molecules on cellular and memory responses to HBsAg DNA vaccination

Because DNA vaccines on their own tend to induce weak immune responses in humans, adjuvant methods are needed in order to improve their efficacy. The co-stimulatory molecules 4-1BBL, OX40L, and CD70 have been shown to induce strong T cell activities; therefore, in this study, we investigated whether they may be used as molecular adjuvants for a hepatitis B surface antigen (HBsAg) DNA vaccine (pcDS2) in eliciting strong cellular and memory responses. Compared to mice immunized with pcDS2 alone, addition of the co-stimulatory molecules increased T cell proliferation and an HBsAg-specific antibody response that was marked with a higher ratio of IgG2a/IgG1. Importantly, pcDS2 plus these co-stimulatory molecules elicited a higher level of IFN-gamma and IL-4 in CD4(+) T cells and a higher level of IFN-gamma in CD8(+) T cells. In addition, a significantly robust antigen-specific cytotoxic T lymphocyte (CTL) response and the production of long-term memory CD8(+) T cells were also observed in the groups immunized with pcDS2 plus 4-1BBL, OX40L, or CD70. Consistently, as late as 100 days after immunization, upregulated expressions of BCL-2, Spi2A, IL-7Ra, and IL-15Ra were still observed in mice immunized with pcDS2 plus these co-stimulatory molecules, suggesting the generation of memory T cells in these groups. Together, these results suggest that the co-stimulatory molecules 4-1BBL, OX40L, or CD70 can enhance the immunogenicity of HBsAg DNA vaccines, resulting in strong humoral, cellular, and memory responses. This approach may lead to an effective therapeutic vaccine for chronic hepatitis B virus (HBV) infection.     

Glucocorticoid-induced TNF receptor-triggered T cells are key modulators for survival/death of neural stem/progenitor cells induced by ischemic stroke

Increasing evidences show that immune response affects the reparative mechanisms in injured brain. Recently, we have demonstrated that CD4(+)T cells serve as negative modulators in neurogenesis after stroke, but the mechanistic detail remains unclear. Glucocorticoid-induced tumor necrosis factor (TNF) receptor (GITR), a multifaceted regulator of immunity belonging to the TNF receptor superfamily, is expressed on activated CD4(+)T cells. Herein, we show, by using a murine model of cortical infarction, that GITR triggering on CD4(+)T cells increases poststroke inflammation and decreases the number of neural stem/progenitor cells induced by ischemia (iNSPCs). CD4(+)GITR(+)T cells were preferentially accumulated at the postischemic cortex, and mice treated with GITR-stimulating antibody augmented poststroke inflammatory responses with enhanced apoptosis of iNSPCs. In contrast, blocking the GITR-GITR ligand (GITRL) interaction by GITR-Fc fusion protein abrogated inflammation and suppressed apoptosis of iNSPCs. Moreover, GITR-stimulated T cells caused apoptosis of the iNSPCs, and administration of GITR-stimulated T cells to poststroke severe combined immunodeficient mice significantly reduced iNSPC number compared with that of non-stimulated T cells. These observations indicate that among the CD4(+)T cells, GITR(+)CD4(+)T cells are major deteriorating modulators of poststroke neurogenesis. This suggests that blockade of the GITR-GITRL interaction may be a novel immune-based therapy in stroke.     

Constitutive OX40/OX40 ligand interaction induces autoimmune-like diseases

The interaction between OX40 and OX40 ligand (OX40L) is suggested to provide T cells with an effective costimulatory signals during T cell-APC interaction. To examine the in vivo effect of constitutive OX40/OX40L interaction during immune regulation, we report the establishment of OX40L-transgenic (OX40L-Tg) mice that constitutively express OX40L on T cells. Markedly elevated numbers of effector memory CD4(+) T cells, but not CD8(+) T cells, were observed in the secondary lymphoid organs of OX40L-Tg mice. Upon immunization with keyhole limpet hemocyanin in the absence of adjuvant, profound T cell proliferative responses and cytokine productions were seen in the OX40L-Tg mice as compared with wild-type mice. Furthermore, in OX40L-Tg mice administrated with superantigen, this constitutive OX40/OX40L interaction on CD4(+) T cells completely prevented normal in vivo clonal T cell deletion. Interestingly, OX40L-Tg mice on the C57BL/6 background spontaneously developed interstitial pneumonia and inflammatory bowel disease that was accompanied with a significant production of anti-DNA Ab in the sera. Surprisingly, these diseases were not evident on the OX40L-Tg mice on the BALB/c strain. However, such inflammatory diseases were successfully reproducible in recombination-activating gene (RAG)2-deficient mice upon transfer of OX40L-Tg CD4(+) T cells. Blockade of OX40/OX40L interaction in the recipient RAG2-deficient mice completely prevented disease development. The present results orchestrated in this study indicate that OX40/OX40L interaction may be a vital link in our understanding of T cell-mediated organ-specific autoimmunity.     

Varied immunity generated in mice by DNA vaccines with large and small hepatitis delta antigens

Whether the hepatitis delta virus (HDV) DNA vaccine can induce anti-HDV antibodies has been debatable. The role of the isoprenylated motif of hepatitis delta antigens (HDAg) in the generation of immune responses following DNA-based immunization has never been studied. Plasmids p2577L, encoding large HDAg (L-HDAg), p2577S, expressing small HDAg (S-HDAg), and p25L-211S, encoding a mutant form of L-HDAg with a cysteine-to-serine mutation at codon 211, were constructed in this study. Mice were intramuscularly injected with the plasmids. The anti-HDV antibody titers, T-cell proliferation responses, T-helper responses, and HDV-specific, gamma interferon (IFN-gamma)-producing CD8(+) T cells were analyzed. Animals immunized with p2577S showed a strong anti-HDV antibody response. Conversely, only a low titer of anti-HDV antibodies was detected in mice immunized with p2577L. Epitope mapping revealed that the anti-HDV antibodies generated by p2577L vaccination hardly reacted with epitope amino acids 174 to 194, located at the C terminus of S-HDAg. All of the HDAg-encoding plasmids could induce significant T-cell proliferation responses and generate Th1 responses and HDV-specific, IFN-gamma-producing CD8(+) T cells. In conclusion, HDAg-specific antibodies definitely exist following DNA vaccination. The magnitudes of the humoral immune responses generated by L-HDAg- and S-HDAg-encoding DNA vaccines are different. The isoprenylated motif can mask epitope amino acids 174 to 195 of HDAg but does not interfere with cellular immunity following DNA-based immunization. These findings are important for the choice of a candidate HDV DNA vaccine in the future.     

Reverse signaling through GITR ligand enables dexamethasone to activate IDO in allergy

Glucocorticoid-induced tumor necrosis factor receptor (GITR) on T cells and its natural ligand, GITRL, on accessory cells contribute to the control of immune homeostasis. Here we show that reverse signaling through GITRL after engagement by soluble GITR initiates the immunoregulatory pathway of tryptophan catabolism in mouse plasmacytoid dendritic cells, by means of noncanonical NF-kappaB-dependent induction of indoleamine 2,3-dioxygenase (IDO). The synthetic glucocorticoid dexamethasone administered in vivo activated IDO through the symmetric induction of GITR in CD4(+) T cells and GITRL in plasmacytoid dendritic cells. The drug exerted IDO-dependent protection in a model of allergic airway inflammation. Modulation of tryptophan catabolism via the GITR-GITRL coreceptor system might represent an effective therapeutic target in immune regulation. Induction of IDO could be an important mechanism underlying the anti-inflammatory action of corticosteroids.     

Sustained peptide-specific gamma interferon T-cell response in rhesus macaques immunized with human immunodeficiency virus gag DNA vaccines

We examined the influence of dose and method of antigen delivery on the dynamics and durability of T-cell responses to candidate human immunodeficiency virus (HIV) vaccines. Codon-optimized sequences from the HIV gag gene were inserted into alternative DNA vaccine vectors to express the coding sequence with or without the tissue plasminogen activator leader sequence. We delivered the vaccines by intramuscular injection as plasmid DNA without adjuvant or as plasmid DNA formulated with a novel block copolymer adjuvant (CRL8623) and then monitored the ensuing T-cell responses by using a gamma interferon enzyme-linked immunospot assay. We demonstrated persistence of the cell-mediated immune (CMI) response in rhesus macaques for at least 18 months following a four-dose vaccination regimen. The plasmid vaccine, with or without CRL8623, was immunogenic in macaques; however, the form coadministered with adjuvant exhibited improved T-cell responses, with a bias toward more antigen-specific CD8(+) T cells. Finally, we examined the fine specificity of the T-cell response to the gag vaccines by testing the response of 23 vaccinated macaques to individual Gag 20-mer peptides. Collectively, the monkeys responded to 25 epitopes, and, on average, each monkey recognized a minimum of 2.7 epitopes. The results indicate that a broad and durable CMI response to HIV DNA vaccines can be induced in a relevant nonhuman primate model.     

Molecular characterization of CD40 signaling intermediates

Signal transduction through the CD40 receptor is initiated by binding of its trimeric ligand and propagated by interactions of tumor necrosis factor receptor-associated factor (TRAF) proteins with the multimerized CD40 cytoplasmic domain. Using defined multimeric constructs of the CD40 cytoplasmic domain expressed as either soluble or myristoylated proteins, we have addressed the extent of receptor multimerization needed to initiate signal transduction and identified components of CD40 signaling complexes. Signal transduction in human embryonic kidney 293 cells, measured by nuclear factor kappaB activation, was observed in cells expressing soluble trimeric CD40 cytoplasmic domain and to a lesser extent in cells expressing dimeric CD40 cytoplasmic domain. Nuclear factor kappaB activation was strongest in cells expressing myristoylated trimeric CD40 cytoplasmic domain. Signal transduction through trimeric CD40 cytoplasmic domains with various point mutations in the TRAF binding sites was similar to signal transduction through analogous full-length receptors. Transiently expressed soluble trimeric CD40 cytoplasmic domain was isolated as complexes that contained TRAF2, TRAF3, TRAF5, TRAF6, and the inhibitor of apoptosis protein (c-IAP1). Association of c-IAP1 with the CD40 cytoplasmic domain complex was indirect and dependent on the presence of an intact TRAF1/2/3 binding site. These results suggest that extracellular ligation of CD40 can be bypassed and that soluble trimerized CD40 complexes can be isolated and used to identify components that link CD40 with signaling pathways.     

Vaccination with a DNA vaccine based on human PSCA and HSP70 adjuvant enhances the antigen-specific CD8+ T-cell response and inhibits the PSCA+ tumors growth in mice

BACKGROUND: DNA vaccines have been shown to be an effective approach to induce antigen-specific cellular and humoral immunity. However, the lower immune intensity in clinical trials limits the application of DNA vaccine. Here we intend to develop a new DNA vaccine based on prostate stem-cell antigen (PSCA), which has been suggested as a potential target for prostate cancer therapy, and enhance the DNA vaccine potency with heat shock proteins (HSPs) as adjuvant. METHODS: A series of DNA plasmids encoding human PSCA, human HSP70 and their conjugates was constructed and injected into male mice intramuscularly (i.m.). To evaluate the immune responses and therapeutic efficacy of these plasmids, major histocompatibility complex (MHC)-restricted PSCA and HSP70-specific epitopes were predicted and a mouse model with a human PSCA-expressing tumor was constructed. RESULTS: The result showed that mice vaccinated with PSCA-HSP plasmids generated the strongest PSCA-specific CD8+ T-cell immune response, but the CD4+ TH1 and TH2 cell immune responses were similar with those vaccinated with other HSP-adjuvant PSCA plasmids or only PSCA DNA. The immunity of HSP70 was also observed and the mice i.m. injected with PSCA+ HSP mixed plasmids generated the lowest anti-HSP antibodies. Furthermore, these vaccinations inhibited the growth of PSCA-expressing tumors and prolonged mouse survival. CONCLUSIONS: These observations emphasize and extend the potential of the human HSP70 gene as adjuvant for DNA vaccines, and the vaccine based on PSCA and HSP70 is of potential value for treating prostate cancer.     

Protection from bacterial infection by a single vaccination with replication-deficient mutant herpes simplex virus type 1

Adaptive immune responses in which CD8(+) T cells recognize pathogen-derived peptides in the context of major histocompatibility complex class I molecules play a major role in the host defense against infection with intracellular pathogens. Cells infected with intracellular bacteria such as Listeria monocytogenes, Salmonella enterica serovar Typhimurium, or Mycobacterium tuberculosis are directly lysed by cytotoxic CD8(+) T cells. For this reason, current vaccines for intracellular pathogens, such as subunit vaccines or viable bacterial vaccines, aim to generate robust cytotoxic T-cell responses. In order to investigate the capacity of a herpes simplex virus type 1 (HSV-1) vector to induce strong cytotoxic effector cell responses and protection from infection with intracellular pathogens, we developed a replication-deficient, recombinant HSV-1 (rHSV-1) vaccine. We demonstrate in side-by-side comparison with DNA vaccination that rHSV-1 vaccination induces very strong CD8(+) effector T-cell responses. While both vaccines provided protection from infection with L. monocytogenes at low, but lethal doses, only rHSV-1 vaccines could protect from higher infectious doses; HSV-1 induced potent memory cytotoxic T lymphocytes that, upon challenge by pathogens, efficiently protected the animals. Despite the stimulation of relatively low humoral and CD4-T-cell responses, rHSV-1 vectors are strong candidates for future vaccine strategies that confer efficient protection from subsequent infection with intracellular bacteria.     

Cutting edge: a potent adjuvant effect of ligand to receptor activator of NF-kappa B gene for inducing antigen-specific CD8+ T cell response by DNA and viral vector vaccination

The ligand to receptor activator of NF-kappaB (RANK-L)/RANK interaction has been implicated in CD40 ligand/CD40-independent T cell priming by dendritic cells. In this report, we show that the coadministration of the RANK-L gene with a Trypanosoma cruzi gene markedly enhances the induction of Trypanosoma Ag-specific CD8(+) T cells and improves the DNA vaccine efficacy. A similarly potent adjuvant effect of the RANK-L gene on the induction of Ag-specific CD8(+) T cells was also observed when recombinant influenza virus expressing murine malaria Ag was used as an immunogen. In contrast, the coadministration of the CD40L gene was not effective in these systems. Our results demonstrated, for the first time, the potent immunostimulatory effect of the RANK-L gene to improve the CD8(+) T cell-mediated immunity against infectious agents.