Multi-epitope Construct Research Articles

Immunogenicity assessment of in silico-selected T cell epitopes for a burkholderia biodefense vaccine (P4282)

Abstract Background: Burkholderia pseudomallei (BPM) and Burkholderia mallei (BM), the causative agents of Melioidosis and Glanders respectively, are Category B pathogens. Burkholderia cepacia (BC) causes severe disease in Cystic Fibrosis patients. No vaccines are available for these pathogens. We are developing a single Burkholderia vaccine that is effective for all three pathogens. Previously, we used informatics tools and HLA binding assays to select 70 promiscuous Class II T epitopes that are highly conserved in BPM, BM and/or BC. Here, an epitope-based vaccine is produced and its immunogenicity is evaluated in a mouse model. Methods: Epitopes were concatenated to form 3 multi-epitope gene constructs (PB1, PB2 and PB3) to produce DNA and protein vaccines. The VaxCAD algorithm ordered epitopes to minimize potential immunogenicity at epitope junctions. HLA DR3 transgenic mice were immunized by a DNA-prime/peptide- or protein-boost strategy. Immunogenicity was determined by IFNγ production to individual peptides by ELISpot assay. Results and Conclusion: Following DNA/peptide immunization, significant IFNγ production (p<0.01) was found in PB3- but not PB1-and PB2-immunized mice. Notably, only PB3 concatameric protein successfully expressed. DNA/PB3 protein immunization studies are ongoing. We describe the step-wise process of developing genome-derived vaccines and identify solutions to the problem of engineering epitope-rich constructs for a broadly reactive Burkholderia vaccine.

Priming and Activation of Human Ovarian and Breast Cancer-specific CD8+ T Cells by Polyvalent Listeria monocytogenes-based Vaccines

Immunotherapeutic vaccine is potentially an effective strategy to combat cancer. Essential components of an effective vaccine must include antigens that are processed by the major histocompatibility complex class I pathway, presented by the tumor major histocompatibility complex molecules, and an effective antigen delivery platform that is capable of breaking self-tolerance. In this study, we characterized a set of ovarian cancer-specific T-cell epitopes delivered by live-attenuated recombinant Listeria monocytogenes (Lm DeltaactADeltainlB) as a vaccine vector. We present data that peptide-specific T cells recognize the human monocytic cell line THP-1 infected with recombinant Lm DeltaactADeltainlB encoding the epitopes. Furthermore, we demonstrate that recombinant L. monocytogenes (Lm)-infected antigen-presenting cells can prime and expand epitope-specific CD8 T cells in vitro and such CD8 T cells recognize not only peptide-loaded targets but also ovarian and breast tumor cells presenting endogenous epitopes. Finally, peptide-specific T cells generated using peripheral blood mononuclear cell from ovarian cancer patients recognize target cells infected with recombinant Lm DeltaactADeltainlB encoding the epitopes. Our results demonstrate that live-attenuated recombinant Lm can be used effectively as a vehicle to deliver cancer peptide antigens singly or as a multiepitope construct. Thus, the use of recombinant live-attenuated Lm strains encoding endogenously processed and presented tumor epitopes/antigens represents an attractive strategy for active cancer immunotherapy in a clinical setting.

Multi-epitope DNA vaccine linked to the A 2/B subunit of cholera toxin protect mice against Toxoplasma gondii

The search for an effective vaccine against toxoplasmosis remains a challenging and elusive goal. Combination of epitopes from different stages of Toxoplasma gondii life cycle is an optimal strategy to overcome the antigen complexity of the parasite. Based on published epitope derived from several promising candidate vaccine antigens, we construct a DNA vaccine encoding multi-epitope of T. gondii and CpG motif, with or without the A 2/B subunit of cholera toxin as a genetic adjuvant. The immunity induced by this vaccine in BALB/c mice and the protection afforded against challenge with the highly virulent RH strain of T. gondii is assessed. This vaccine was able to elicit a significant humoral and cellular immune response in vaccinated mice. Furthermore, CTXA 2/B as a genetic adjuvant could enhance the magnitude of immune responses as well as increased survival rate in mice infected with the lethal RH tachyzoites. This study is the first report of a multi-epitope DNA construct strategy as a potential DNA vaccine against toxoplasmosis.

Advanced Antigen Delivery of Murine Survivin: Chimeric Virus-Like Particles in Cancer Vaccine Research

Success in cancer immunotherapy depends on the identification and efficient targeting of specific tumor-associated antigens. Two pivotal strategies to prime patients' immune system against malignant cells are tumor-specific adoptive T-cell therapy and tumor-specific vaccination. Here, we will focus on immunotherapeutic vaccination and discuss the advantages and disadvantages of different strategies to deliver tumor-specific T-cell epitopes. A particular focus will be put on virus-like particles (VLPs) as vehicle to deliver tumor-specific epitopes in the context of full-length proteins, as multi-epitope constructs or as individual tumor-associated T-cell epitopes. VLPs represent non-infectious and non-replicating antigen delivery systems devoid of any nucleic acid. They constitute innovative immunotherapeutic agents against cancer due to their superior, adjuvant-like antigenicity. We will present various tumor-associated antigens currently in different stages of development including survivin, as promising candidates for targeted tumor therapies.

Protective efficacy of multiepitope human leukocyte antigen-A*0201 restricted cytotoxic T-lymphocyte peptide construct against challenge with human T-cell lymphotropic virus type 1 Tax recombinant vaccinia virus.

Human T-cell lymphotropic virus type 1 (HTLV-1) is the causative agent of adult T-cell leukemia. Multiepitope T-cell vaccines are more likely to generate a broad long-lasting immune response than those composed of single epitopes. We recently reported a novel multivalent cytotoxic T-lymphocyte peptide construct derived from the Tax protein of HTLV-1 separated by arginine spacers that elicited high cellular responses against individual epitopes simultaneously in human leukocyte antigen (HLA)-A*0201 transgenic mice. We now report the effect of epitope orientation on the processing of the multiepitope construct by 20s proteasomes and the effect of the processing rates on the immunogenicity of the intended epitopes. A positive correlation was found between processing rates and the immunogenicity of the intended epitopes. The construct with the highest immunogenicity for each epitope was tested for protective efficacy in a preclinical model of infection using HTLV-1 Tax recombinant vaccinia virus and HLA-A*0201 transgenic mice. Mice vaccinated with the multiepitope construct displayed a statistically significant reduction in viral replication that was dependent on CD8 T cells. Reduction in viral replication was also confirmed to be specific to Tax-vaccinia virus. These results demonstrate the activation of Tax-specific CD8+ T cells by vaccination and are supportive of a multivalent peptide vaccine approach against HTLV-1 infections.

Optimization of epitope processing enhances immunogenicity of multiepitope DNA vaccines

Experimental DNA vaccines comprised of multiple minimal cytotoxic T lymphocytes (CTL) epitopes can effectively induce broad CTL responses; however, such constructs frequently exhibit significant variation in epitope immunogenicity. Antigenicity assays utilizing human cells transfected with one such multiepitope construct revealed that the epitopes with poor immunogenicity were inefficiently processed in transfected cells. Compilation of a database of 94 epitope/flanking region combinations, for which immunogenicity was measured experimentally, revealed that the type of residue immediately following the carboxyl-terminus of the epitope exerted a prominent effect on immunogenicity. Experiments utilizing a variety of HBV-specific vaccine constructs demonstrated epitope immunogenicity could be modulated by the insertion of a single amino acid and the effect on immunogenicity could be ascribed to modulation of processing efficiency. These findings demonstrate that multiepitope DNA vaccines can be engineered to enhance CTL immunogenicity by increasing processing efficiency .