Efficient CTL Research Articles

Efficient Photoinduced Thermocatalytic Chemiluminescence System Based on the Z-Scheme Heterojunction Ag3PO4/Ag/Bi4Ti3O12 for H2S Sensing.

Cataluminescence as a highly efficient gas transduction principle has attracted wide attention among research in environmental monitoring and clinical diagnosis with increasing awareness of human safety. Nowadays, the development of innovation sensing systems and the construction of the sensing mechanism to improve the analytical performance of compounds remain a major challenge. Herein, we construct an advanced photoinduced thermocatalytic chemiluminescence (PI-TC-CL) gas-sensing system via the introduction of a Z-scheme heterojunction Ag3PO4/Ag/Bi4Ti3O12 to achieve higher efficient detection of H2S. The unique electron transport path of the Z-scheme heterojunction and the LSPR effect of Ag nanoparticles fascinate the generation of the photoinduced electron-hole pair on the surface of catalysts when stimulated by LED lamps and slow down the recombination of electron-hole pairs under thermal conditions. Thus, based on the cooperative effect of the Z-scheme heterojunction AgPO/Ag/BTO and PI-TC-CL system, we have successfully established an efficient H2S CTL detection system, which has a response three times higher than that on the traditional CTL system and even 45 times higher than that on BTO and ranges among the best of the state-of-the-art CTL performance in H2S detection with the linear range of 0.095-8.87 μg mL-1 and a limit of detection of 0.0065 μg mL-1. Besides, to explore the gas-sensing mechanism, the synergetic effects of photoinduction and thermal catalysis are investigated thoroughly via conductivity and electrochemical experiments. This research provides a new perspective of engineering highly efficient catalysts and ingenious sensor systems through designing the nanostructure of materials and synergism catalytic mechanism.

Sub-clonal analysis of the murine C1498 acute myeloid leukaemia cell line reveals genomic and immunogenic diversity

BackgroundIn acute myeloid leukaemia (AML)-affected patients, the presence of heterogeneous sub-clones at diagnosis has been shown to be responsible for minimal residual disease and relapses. The role played by the immune system in this leukaemic sub-clonal hierarchy and maintenance remains unknown. As leukaemic sub-clone immunogenicity could not be evaluated in human AML xenograft models, we assessed the sub-clonal diversity of the murine C1498 AML cell line and the immunogenicity of its sub-clones in immune-competent syngeneic mice. MethodologyThe murine C1498 cell line was cultured in vitro and sub-clonal cells were generated after limiting dilution. The genomic profiles of 6 different sub-clones were analysed by comparative genomic hybridization arrays (CGH). The sub-clones were then injected into immune-deficient and – competent syngeneic mice. The immunogenicities of the sub-clones was evaluated through 1) assessment of mouse survival, 2) determination of leukaemic cell infiltration into organs by flow cytometry and the expression of a fluorescent reporter gene, 3) assessment of the CTL response ex vivo and 4) detection of residual leukaemic cells in the organs via amplification of the genomic reporter gene by real-time PCR (qPCR). ResultsGenomic analyses revealed heterogeneity among the parental cell line and its derived sub-clones. When injected individually into immune-deficient mice, all sub-clones induced cases of AML with different kinetics. However, when administered into immune-competent animals, some sub-clones triggered AML in which no mice survived, whereas others elicited reduced lethality rates. The AML-surviving mice presented efficient anti-leukaemia CTL activity ex vivo and eliminated the leukaemic cells in vivo. ConclusionWe showed that C1498 cell sub-clones presented genomic heterogeneity and differential immunogenicity resulting either in immune escape or elimination. Such findings could have potent implications for new immunotherapeutic strategies in patients with AML.

Replicating viral vector platform exploits alarmin signals for potent CD8+ T cell-mediated tumour immunotherapy

Viral infections lead to alarmin release and elicit potent cytotoxic effector T lymphocyte (CTLeff) responses. Conversely, the induction of protective tumour-specific CTLeff and their recruitment into the tumour remain challenging tasks. Here we show that lymphocytic choriomeningitis virus (LCMV) can be engineered to serve as a replication competent, stably-attenuated immunotherapy vector (artLCMV). artLCMV delivers tumour-associated antigens to dendritic cells for efficient CTL priming. Unlike replication-deficient vectors, artLCMV targets also lymphoid tissue stroma cells expressing the alarmin interleukin-33. By triggering interleukin-33 signals, artLCMV elicits CTLeff responses of higher magnitude and functionality than those induced by replication-deficient vectors. Superior anti-tumour efficacy of artLCMV immunotherapy depends on interleukin-33 signalling, and a massive CTLeff influx triggers an inflammatory conversion of the tumour microenvironment. Our observations suggest that replicating viral delivery systems can release alarmins for improved anti-tumour efficacy. These mechanistic insights may outweigh safety concerns around replicating viral vectors in cancer immunotherapy.

Efficient CTL Verification via Horn Constraints Solving

The use of temporal logics has long been recognised as a fundamental approach to the formal specification and verification of reactive systems. In this paper, we take on the problem of automatically verifying a temporal property, given by a CTL formula, for a given (possibly infinite-state) program. We propose a method based on encoding the problem as a set of Horn constraints. The method takes a program, modeled as a transition system, and a property given by a CTL formula as input. It first generates a set of forall-exists quantified Horn constraints and well-foundedness constraints by exploiting the syntactic structure of the CTL formula. Then, the generated set of constraints are solved by applying an off-the-shelf Horn constraints solving engine. The program is said to satisfy the property if and only if the generated set of constraints has a solution. We demonstrate the practical promises of the method by applying it on a set of challenging examples. Although our method is based on a generic Horn constraint solving engine, it is able to outperform state-of-art methods specialised for CTL verification.

Abstract 3216: Immunogenicity and genomic profiling reveal sub-clonal diversity of a murine acute myeloid leukemia (AML) cell line

Abstract In acute myeloid leukemia (AML), clonal heterogeneity at diagnosis and the presence of minor sub-clones were shown to be responsible for minimal residual disease and relapses in patients. Several studies have proposed that their persistence could be due to their dormant state (i.e. quiescence and/or control of their growth by immune cells). As contribution of immunity to clonal persistence could not be evaluated in xenografts models, we assessed for the clonal diversity of a murine AML cell line. Genomic analyses revealed heterogeneity among the parental cell line and its derived clones. Injected individually into immune-deficient mice, all clones induced AML with different kinetics. However, when administered into immune-competent animals, some clones triggered AML with no mice survival, whereas others elicited reduced lethality rates. The leukemia-surviving mice presented an efficient anti-tumoral CTL activity and reduced lymphopenia. Investigating for residual leukemic cells in the organs of these mice, elimination rather than persistence was observed. Thus, we showed that genetic heterogeneity among the clones led to differential immunogenicity resulting either in their immune escape or elimination. Such findings could have potent implications for new immunotherapeutic strategies in patients. Citation Format: Carine Brinster, Virginie DRISS, Bruno Quesnel, Martin Figeac, Frédéric Leprêtre, Céline Villenet, Isabelle Briche. Immunogenicity and genomic profiling reveal sub-clonal diversity of a murine acute myeloid leukemia (AML) cell line. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3216. doi:10.1158/1538-7445.AM2015-3216

Regulation of CTL trafficking into tumors and anti-tumor immunity by chemoattractant receptors BLT1 and CXCR3 (TUM7P.1015)

Abstract A major hurdle limiting the success of tumor immunotherapies in the clinic is poor recruitment of CD8+ T cells (Cytotoxic T Lymphocytes-CTL) into tumors despite efficient CTL responses in the periphery. However, little is known about the mechanisms that regulate their infiltration into tumors. We investigated the role of leukotriene B4 receptor, BLT1 and CXCR3, the receptor for CXCL9 and CXCL10 in anti-tumor immunity using a syngeneic B16 melanoma mice model. Significant acceleration in tumor growth and reduced survival was observed in both BLT1-/- and CXCR3-/- mice as compared to WT mice. Tumor immune infiltration studies revealed significant reduction of CTLs in BLT1-/- and CXCR3-/- tumors as compared to WT tumors, their frequencies being similar in the periphery. Adoptive transfer of WT but not BLT1-/- or CXCR3-/- CTLs reduced tumor growth in Rag2-/- animals due to enhanced WT CTL infiltration to tumors. Furthermore, adoptive transfer of BLT1-/-, CXCR3-/- or BLT1-/-CXCR3-/- CTLs together with WT CTLs showed that initial migration of BLT1+/+ CTLs to tumors facilitates additional CTL recruitment in a CXCR3 dependent manner. An anti-PD-1 antibody based vaccine completely lost its efficacy in both BLT1-/- and CXCR3-/- mice suggesting a requirement for these receptors in generating sustained anti-tumor immune response. These results demonstrate a critical role for BLT1 and CXCR3 in CTL migration to tumors and thus can be targeted to elicit effective anti-tumor responses.

Heat shock protein 90 targets a chaperoned peptide to the static early endosome for efficient cross-presentation by human dendritic cells.

The presentation of an exogenous antigen in a major histocompatibility complex class-I-restricted fashion to CD8+ T cells is called cross-presentation. Heat shock proteins (HSPs) such as Hsp70, gp96, and Hsp90 have been shown to elicit efficient CTL responses by cross-presentation through an as-yet entirely unknown mechanism. Hsp90 is the most abundant cytosolic HSP and is known to act as a molecular chaperone. We have shown that a tumor antigen peptide complexed with Hsp90 could be cross-presented by dendritic cells (DCs) through an endosomal pathway in a murine system. However, it has not been determined whether human DCs also cross-present an Hsp90–peptide complex and induce peptide-specific CTLs. In this study, we found that an Hsp90–cancer antigen peptide complex was efficiently cross-presented by human monocyte-derived DCs and induced peptide-specific CTLs. Furthermore, we observed that the internalized Hsp90–peptide complex was strictly sorted to the Rab5+, EEA1+ static early endosome and the Hsp90-chaperoned peptide was processed and bound to MHC class I molecules through an endosome-recycling pathway. Our data indicate that targeting of the antigen to a “static” early endosome by Hsp90 is essential for efficient cross-presentation.

Efficient CTL model-checking for pushdown systems

Pushdown systems (PDS) are well adapted to model sequential programs with (possibly recursive) procedure calls. Therefore, it is important to have efficient model checking algorithms for PDSs. We consider in this paper CTL model checking for PDSs. We consider the “standard” CTL model checking problem where whether a configuration of a PDS satisfies an atomic proposition or not depends only on the control state of the configuration. We consider also CTL model checking with regular valuations, where the set of configurations in which an atomic proposition holds is a regular language. We reduce these problems to the emptiness problem in Alternating Büchi Pushdown Systems, and we give an algorithm to solve this emptiness problem. Our algorithms are more efficient than the other existing algorithms for CTL model checking for PDSs in the literature. We implemented our techniques in a tool, and we applied it to different case studies. Our results are encouraging. In particular, we were able to confirm the existence of known bugs in Linux source code.

HIV-1 gag specific exosome-targeted CD8+ T cell based vaccine stimulates specific CTL responses leading to long-term immunity (P4493)

Abstract The limitations of highly active anti-retroviral therapy (HAART) have necessitated the development of alternative therapeutics for human immunodeficiency virus type-1 (HIV-1)-infected patients with dysfunctional dendritic cells (DCs) and CD4+T cell deficiency. We have recently demonstrated that Con A stimulated non-specific CD8+T cells could uptake OVA pulsed DC-derived exosome (EXO), act as CD8+Tc antigen presenting cells (Th-APCs), and capable of stimulating DC- and CD4+T cell-independent OVA-specific CTL responses leading to long-term immunity. This formed the new concept novel EXO-targeted CD8+T cell vaccine. The extreme conservation and intolerance of escape variants in HIV-1 Gag p24 make it a potential target for vaccine design. In this study, we constructed adenoviral vector AdV-gag and expression vector pEGFP-gag both expressing Gag. We generated Gag specific EXO-targeted T (Gag-Texo) cell vaccine by using C57BL/6 mouse CD8+ ConA T cells with uptake of AdV-gag transfected bone marrow DC (DCgag) released EXOgag. We then generated BL6-10gag cell lines by transfection of B16 melanoma cell line (BL6-10) with pEGFP-gag. We demonstrated that Gag-Texo vaccine is capable of stimulating efficient Gag-specific CD8+ CTL responses in vivo, leading to antitumor immunity against Gag expressing B16 melanoma (BL6-10 gag) in animal studies. Therefore, this novel HIV-1-specific EXO-targeted Gag-Texo vaccine may be useful in induction of efficient CTL responses in AIDS patients.

Engineering of papaya mosaic virus (PapMV) nanoparticles with a CTL epitope derived from influenza NP

BackgroundThe ever-present threat of infectious disease, e.g. influenza pandemics, and the increasing need for new and effective treatments in immunotherapy are the driving forces that motivate research into new and innovative vaccine platforms. Ideally, such platforms should trigger an efficient CTL response, be safe, and easy to manufacture. We recently developed a novel nanoparticle adjuvant comprised of papaya mosaic virus (PapMV) coat protein (CP) assembled around an RNA. The PapMV nanoparticle is an efficient vaccine platform in which the peptide antigen is fused to the C-terminus of the PapMV CP, leading to nanoparticles presenting surface-exposed epitope. The fusion stabilizes the epitope and improves its immunogenicity. We found recently that C-terminal fusions are not always efficient, depending on the nature of the peptide fused to the platform.ResultsWe chose a CTL epitope derived from the nucleocapsid (NP) of influenza virus (NP147-155) for this proof-of-concept demonstration. Recombinant nanoparticles harbouring a fusion at the N-terminus were more efficient in triggering a CTL response. Efficacy appeared to be linked to the stability of the nanoparticles at 37°C. We also showed that discs—smaller than nanoparticles—made of 20 subunits of PapMV CP are less efficient for induction of a CTL response in mice, revealing that assembly of the recombinant PapMV CP into nanoparticles is crucial to triggering an efficient CTL response.ConclusionThe point of fusion on the PapMV vaccine platform is critical to triggering an efficient CTL response. Efficacy is linked to nanoparticle stability; nanoparticles must be stable at 37°C but remain susceptible to cellular proteases to ensure efficient processing of the CTL epitope by cells of the immune system. The results of this study improve our understanding of the PapMV vaccine platform, which will facilitate the design of efficient vaccines to various infectious threats.

The outcome of cross-priming during virus infection is not directly linked to the ability of the antigen to be cross-presented

The initiation of CD8(+) T cell (CTL) immune responses can occur via cross-priming. Recent data suggested a relationship between cross-presentation and immunodominance of epitope-specific T cells. To test this association, we evaluated the efficacy of cross-presentation for several virus epitopes in vitro and examined if this can be extrapolated in vivo. Employing lymphocytic choriomeningitis virus (LCMV), we demonstrate that the cross-presentation and cross-priming of LCMV antigens were dominated by NP396, but not NP205 when analyzing the LCMV-NP. Although with LCMV-GP, cross-presentation was dominated by GP276, and cross-priming was dominated by GP33. Importantly, although NP396 was significantly more efficient than GP33 in cross-presentation, cross-priming of their specific CTL was comparable. In a subsequent virus challenge after cross-priming, GP33-specific CTL dominated the response. Accordingly, based on our data, the ability of viral epitopes to be cross-presented in vitro does not entirely reflect what would occur in cross-priming. Thus, weak cross-presenting antigens may still cross-prime an efficient CTL response depending on other in vivo elements such as the naïve T-cell precursor frequencies.

Complement as an Endogenous Adjuvant for Dendritic Cell-Mediated Induction of Retrovirus-Specific CTLs

Previous studies have demonstrated the involvement of complement (C) in induction of efficient CTL responses against different viral infections, but the exact role of complement in this process has not been determined. We now show that C opsonization of retroviral particles enhances the ability of dendritic cells (DCs) to induce CTL responses both in vitro and in vivo. DCs exposed to C-opsonized HIV in vitro were able to stimulate CTLs to elicit antiviral activity significantly better than non-opsonized HIV. Furthermore, experiments using the Friend virus (FV) mouse model illustrated that the enhancing role of complement on DC-mediated CTL induction also occurred in vivo. Our results indicate that complement serves as natural adjuvant for DC-induced expansion and differentiation of specific CTLs against retroviruses.

Cutting Edge: Priming of CD8 T Cell Immunity to Herpes Simplex Virus Type 1 Requires Cognate TLR3 Expression InVivo

Despite its potential for involvement in viral immunity, little evidence links TLR3 to adaptive antiviral responses. Here we show that TLR3 is required for the generation of CD8 T cell immunity to HSV-1. The magnitude of the gB-specific CD8 T cell response after flank infection by HSV-1 was significantly reduced in mice lacking TIR domain-containing adaptor-inducing IFN-beta or TLR3, but not MyD88. Impaired CTL induction was evident in chimeric mice lacking TLR3 in bone marrow (BM)-derived cells. Among the dendritic cell subsets, TLR3 was expressed by CD8alpha(+) dendritic cells, known to be involved in priming HSV-1-specific CD8 T cells. Use of mixed BM chimeras revealed that TLR3 and the MHC class I-restriction element must be expressed by the same BM-derived cell for effective priming. These data imply that a cognate linkage between TLR3 and MHC class I is required for efficient CTL priming to HSV-1.

Targeting to Static Endosome Is Required for Efficient Cross-Presentation of Endoplasmic Reticulum-Resident Oxygen-Regulated Protein 150-Peptide Complexes

Heat shock proteins (HSPs) such as Hsp70, gp96, and Hsp90 have been shown to elicit intriguing, efficient CTL responses by cross-presentation via an as yet entirely unknown mechanism. Oxygen-regulated protein 150 (ORP150), also known as grp170, is an endoplasmic reticulum-resident HSP and is up-regulated by hypoxia. It has been demonstrated that ORP150 binds tumor-associated Ag peptides within cancer cells. Immunization with an ORP150-tumor Ag complex has been shown to generate tumor-specific CTLs. Most recently, it has been shown that exogenous ORP150 induces cross-presentation of a chaperoned Ag, thereby stimulating Ag-specific CTLs. However, the mechanism underlying this efficient cross-presentation is still unsolved. In this study, we show that the ORP150-precursor peptide complex can elicit CTL response through cross-presentation as well as the CD4(+) T cell response by dendritic cells. Furthermore, we observed that the internalized ORP150-peptide complex, but not OVA protein, which was not cross-presented, was sorted to the Rab5(+), EEA1(+) static early endosome, followed by translocation to a recycling endosome, where the ORP150-chaperoned peptide was processed and bound to MHC class I molecules. Moreover, we observed that immunization of mice with ORP150-peptide complexes elicited strong peptide-specific CTLs and antitumor effects in vivo. Our data indicate that targeting of the Ag to a "static" early endosme by ORP150 is required for the efficient cross-presentation.

The duration of TCR/pMHC interactions regulates CTL effector function and tumor‐killing capacity

Effector CTL contribute to tumoral immunity by killing tumor cells through secretion of cytotoxic granules and cytokines. Activation of CTL requires specific recognition of cognate peptide-MHC-I (pMHC) complexes on the tumor cell surface by the CTL TCR. It has been suggested that the half-life (t(1/2)) of the TCR/pMHC interaction modulates the activation of naïve CD8(+) T cells; however, it remains unknown whether CTL effector function can also be regulated by the TCR/pMHC t(1/2). Here, we have studied CTL activity in response to tumor cells loaded with pMHC that bind the TCR with different t(1/2). We observed that the TCR/pMHC t(1/2) can differentially regulate CTL effector function during the interaction with tumor cells and defines the nature of anti-tumoral CTL responses in vivo. Although prolonged TCR/pMHC t(1/2) promoted only partial expression of cytotoxic molecules, short t(1/2) induced partial polarization of lytic machinery toward target cells. In contrast, intermediate TCR/pMHC t(1/2) induced strong expression of cytotoxic molecules, efficient polarization of lytic machinery and subsequent release of toxic granules by CTL that killed tumor cells. Consistently, efficient in vivo CTL-mediated tumor clearance was only observed for tumors expressing intermediate t(1/2) pMHC ligands. These data suggest that there is an optimal TCR/pMHC t(1/2) for efficient CTL activity.

Quantitative expression and immunogenicity of MAGE‐3 and ‐6 in upper aerodigestive tract cancer

The MAGE antigens are frequently expressed cancer vaccine targets. However, quantitative analysis of MAGE expression in upper aerodigestive tract (UADT) tumor cells and its association with T-cell recognition has not been performed, hindering the selection of appropriate candidates for MAGE-specific immunotherapy. Using quantitative RT-PCR (QRT-PCR), we evaluated the expression of MAGE-3/6 in 65 UADT cancers, 48 normal samples from tumor matched sites and 7 HLA-A*0201+ squamous cell carcinoma of the head and neck (SCCHN) cell lines. Expression results were confirmed using Western blot. HLA-A*0201:MAGE-3- (271-279) specific cytotoxic T lymphocytes (MAGE-CTL) from SCCHN patients and healthy donors showed that MAGE-3/6 expression was highly associated with CTL recognition in vitro. On the basis of the MAGE-3/6 expression, we could identify 31 (47%) of the 65 UADT tumors, which appeared to express MAGE-3/6 at levels that correlated with efficient CTL recognition. To confirm that the level of MAGE-3 expression was responsible for CTL recognition, 2 MAGE-3/6 mRNA(high) SCCHN cell lines, PCI-13 and PCI-30, were subjected to MAGE-3/6-specific knockdown. RNAi-transfected cells showed that MAGE expression and MAGE-CTL recognition were significantly reduced. Furthermore, treatment of cells expressing low MAGE-3/6 mRNA with a demethylating agent, 5-aza-2'-deoxycytidine (DAC), increased the expression of MAGE-3/6 and CTL recognition. Thus, using QRT-PCR UADT cancers frequently express MAGE-3/6 at levels sufficient for CTL recognition, supporting the use of a QRT-PCR-based assay for the selection of candidates likely to respond to MAGE-3/6 immunotherapy. Demethylating agents could increase the number of patients amenable for targeting epigenetically modified tumor antigens in vaccine trials.

Impact of Intrinsic Cooperative Thermodynamics of Peptide-MHC Complexes on Antiviral Activity of HIV-Specific CTL

The antiviral activity of HIV-specific CTL is not equally potent but rather is dependent on their specificity. But what characteristic of targeted peptides influences CTL antiviral activity remains elusive. We addressed this issue based on HLA-B35-restricted CTLs specific for two overlapping immunodominant Nef epitopes, VY8 (VPLRPMTY) and RY11 (RPQVPLRPMTY). VY8-specific CTLs were more potently cytotoxic toward HIV-infected primary CD4(+) cells than RY11-specific CTLs. Reconstruction of their TCR revealed no substantial difference in their functional avidity toward cognate Ags. Instead, the decay analysis of the peptide-MHC complex (pMHC) revealed that the VY8/HLA-B35 complex could maintain its capacity to sensitize T cells much longer than its RY11 counterpart. Corroboratively, the introduction of a mutation in the epitopes that substantially delayed pMHC decay rendered Nef-expressing target cells more susceptible to CTL killing. Moreover, by using differential scanning calorimetry and circular dichroism analyses, we found that the susceptible pMHC ligands for CTL killing showed interdependent and cooperative, rather than separate or sequential, transitions within their heterotrimer components under the thermally induced unfolding process. Collectively, our results highlight the significant effects of intrinsic peptide factors that support cooperative thermodynamics within pMHC on the efficient CTL killing of HIV-infected cells, thus providing us better insight into vaccine design.

Vanilloid receptor 1 agonists, capsaicin and resiniferatoxin, induce efficient anti-viral CTL responses by increasing MHC-restricted antigen presentation (78.29)

Abstract DCs, like the sensory neurons, express vanilloid receptor 1 (VR1). Here we demonstrate that VR1 agonists, capsaicin (CP) and resiniferatoxin (RTX), increase MHC-restricted antigen presentation in DCs. Treatment of DCs, that were infected with recombinant vaccinia virus (VV) expressing OVA (VV-OVA), with CP or RTX significantly increased MHC class I-restricted OVA presentation. In accordance with the in vitro results, oral administration of CP into VV-OVA-infected mice significantly enhanced MHC class I-restricted OVA presentation, and consequently elicited potent OVA-specific CTL activity compared to untreated controls. The effects of VR1 agonists on the exogenous antigen presentation were also examined in DCs. CP and RTX enhanced MHC class I-restricted, but not MHC class II-restricted, presentation of exogenous OVA. CP and RTX did not affect phagocytic activity, the expression level of total MHC molecules and co-stimulatory molecules on DC, suggesting that they enhance the intracellular processing events of phagocytosed antigens. These results demonstrate that VR1 agonists preferentially increase MHC class I-restricted antigen processing pathways. The present study also shows that oral consumption of CP, a common component of pungent foods, could enhance the efficacy of DC-based anti-viral and anti-tumor vaccines.

Viral load and clinical disease enhancement associated with a lentivirus cytotoxic T lymphocyte vaccine regimen

Effective DNA-based vaccines against lentiviruses will likely induce CTL against conserved viral proteins. Equine infectious anemia virus (EIAV) infects horses worldwide, and serves as a useful model for lentiviral immune control. Although attenuated live EIAV vaccines have induced protective immune responses, DNA-based vaccines have not. In particular, DNA-based vaccines have had limited success in inducing CTL responses against intracellular pathogens in the horse. We hypothesized that priming with a codon-optimized plasmid encoding EIAV Gag p15/p26 with co-administration of a plasmid encoding an equine IL-2/IgG fusion protein as a molecular adjuvant, followed by boosting with a vaccinia vector expressing Gag p15/p26, would induce protective Gag-specific CTL responses. Although the regimen induced Gag-specific CTL in four of seven vaccinated horses, CTL were not detected until after the vaccinia boost, and protective effects were not observed in EIAV challenged vaccinates. Unexpectedly, vaccinates had significantly higher viral loads and more severe clinical disease, associated with the presence of vaccine-induced CTL. It was concluded that (1) further optimization of the timing and route of DNA immunization was needed for efficient CTL priming in vivo, (2) co-administration of the IL-2/IgG plasmid did not enhance CTL priming by the Gag p15/p26 plasmid, (3) vaccinia vectors are useful for lentivirus-specific CTL induction in the horse, (4) Gag-specific CTL alone are either insufficient or a more robust Gag-specific CTL response is needed to limit EIAV viremia and clinical disease, and (5) CTL-inducing vaccines lacking envelope immunogens can result in lentiviral disease enhancement. Although the mechanisms for enhancement associated with this vaccine regimen remain to be elucidated, these results have important implications for development of lentivirus T cell vaccines.

713. Sendai Virus Vector-Mediated Efficient CTL Induction Against AIDS Even in the Presence of Anti-Vector Antibody

713. Sendai Virus Vector-Mediated Efficient CTL Induction Against AIDS Even in the Presence of Anti-Vector Antibody