II. Class I and class II restricted epitopes from a representative sample of the different arenavirus species pathogenic in humans.
This project addresses the discovery and validation of cytotoxic and helper T cell epitopes presented by HLA class I and class II MHC molecules, respectively, that are derived from a group of prevalent arenaviruses (Junin, Guanarito, Sabia, Machupo, Lassa, LCMV and Whitewater arroyo) with known potential for causing disease in humans, and representative of a diverse set of arenavirus phylogenetic groups. We also plan to investigate and determine epitopes presented by murine, non-human primate and human MHC molecules. To accomplish these goals we plan to utilize a multidisciplinary approach, based on bioinformatic scanning of viral genomes, high throughput binding assays utilizing purified MHC molecules, in vitro cellular assays and in vivo assays. These studies should lead to the definition of a broad range of epitopes, facilitate development of diagnostic reagents allowing a rigorous evaluation of T cell responses associated with infection in humans, and enable the evaluation of the performance of different vaccine candidates. Little is known about the degree to which cellular responses in humans are cross-reactive amongst various arenavirus species. Thus, for each virus, we will map the epitopes recognized within each viral antigen by class I and class II molecules and determine their degree of cross-reactivity with homologous sequences from other arenaviruses. We will also pursue the development of a multi-epitope vaccine construct, encompassing epitopes cross-reactive with multiple arenaviruses, as well as a collection of non-cross-reactive epitopes derived from each distinct virus, so that a single vaccine entity might be able to protect from, or ameliorate the disease course caused by, several major arenavirus species.
III. Identification of class I and class II restricted epitopes derived from variola and vaccinia viruses.
This study addresses the discovery and validation of cytotoxic and helper T cell epitopes derived from the vaccinia and variola viruses and presented by murine, non-human primate and human MHC molecules. To accomplish these goals we plan to utilize a multidisciplinary approach based on genome wide scanning of the viruses genome, high throughput binding assays utilizing purified MHC molecules, in vitro cellular assays and in vivo assays, as feasible and appropriate. In the first part of our studies, we will 1) determine the immunodominant antigens recognized by class I and class II restricted responses in humans immunized with vaccinia virus, 2) map the epitopes recognized within each antigen, and 3) determine their degree of cross-reactivity with homologous variola virus-derived sequences.
IV. Epitope-based Multi-peptide Vaccines for Expanded Coverage Against Inter-pandemic Influenza. (In collaboration with Dr. Mbawuike, Baylor College of Medicine, Houston, TX)
The CDC estimates that influenza epidemics kill about 20-40,000 persons annually in the U.S, most of them elderly. Currently licensed inactivated subunit influenza virus vaccines (IVVs) show reduced effectiveness against variant interpandemic drift influenza A viruses because of the inevitably poor match between vaccine-elicited antibodies and the virus. Influenza subunit vaccines are also ineffective in inducing influenza virus-specific CD8+ cytotoxic T lymphocytes (CTL) which appear to be critical in mediating the clearance of virus infection, thus promoting recovery and lowering rates of morbidity and mortality. The inadequacy of subunit influenza vaccine is obviously due to the narrow spectrum of response it elicits and frequent mutation of influenza virus. We hypothesize that a vaccine based on reproducing the breath of responses induced by the whole organism, including T cells, B cells and appropriate cytokines, should provide better protection against influenza disease. Such optimally effective vaccine should include epitopes derived from all viral antigens recognized by most of the genetically diverse human population.
The goal of this proposal is to identify and characterize immunodominant and subdominant CTL and helper T lymphocyte (HTL) epitopes contained in all ten influenza protein segments restricted by major human HLA supertypes.
V. Human T-cell Epitopes in SARS. (In collaboration with Dr. Buchmeier, The Scripps Research Institute, San Diego, CA).
The discovery and validation of HLA-restricted cytotoxic and helper T cell epitopes derived from the Severe Acute Respiratory Syndrome coronavirus (SARS CoV) represents a significant challenge because of the large genome size and because of the extreme polymorphism of HLA alleles. Herein, we propose to combine bioinformatic approaches and high throughput MHC-peptide binding assays to identify epitopes restricted by HLA class I and class II molecules representative of >95% of the general population, irrespective of ethnicity. These epitopes will be further validated with in vivo and in vitro immunogenicity studies utilizing HLA transgenic mice and human PBMC from healthy unexposed donors. Animal models are likely to play a vital role in the development of SARS specific vaccines and diagnostics. Accordingly, we also plan to identify epitopes presented by mouse and non-human primate MHC molecules.