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Technology Developmental Project 1

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A Novel ORFeome Approach to Identify CD8+ T Cell Responses to S. Typhi Proteins In Humans

Project Leader: Marcelo B. Sztein, MD 

This project is directed towards the development of novel technology to identify antigens recognized by pathogen-specific class-I MHC restricted T cells by systematic screening of all pathogen protein coding genes, its ORFeome in humans. The system is based on the observation that E. coli expressing a pathogen-derived CD8+ T-cell epitope together with cytoplasmic listeriolysin (cytoLLO) is able to stimulate epitope-specific class I restricted CD8+ T-cells following processing of whole bacteria by antigen presenting cells (APCs).

We will focus these studies on immunity to S. Typhi, a pathogen of great public health importance. As discussed above, the determinants of protection from typhoid fever remain largely undefined. Thus, the identification of putative S. Typhi proteins that might be involved in protection, by screening of the S. Typhi ORFeome, will allow us to explore the hypothesis that the increased efficacy of four doses of the oral Ty21a typhoid vaccine as compared to a single dose is in part due to the repertoire of S. Typhi specific CD8+ T cells in vaccinees.

One of the novel features of the proposed technology is an ability to confirm that S. Typhi proteins are expressed in E. coli and can be presented by APCs in the context of class-I molecules by fusing them with positive epitope controls. This proposal brings together collaborators with expertise in cellular immunology, genomics and high-throughput assays systems with the express purpose of tackling an important global health problem via improved vaccine development.