Development of an MHC class I L(d)-restricted PSA peptide-loaded tetramer for detection of PSA-specific CD8(+) T cells in the mouse

OBJECTIVES: We set out to develop a prostate specific antigen (PSA) peptide-loaded tetramer for enumeration of PSA-specific CD8(+) T cells in the Balb/c mouse model. METHODS: A candidate MHC class I PSA peptide (HPQKVTKFML(188–197)) was selected based on its ability to restimulate PSA-specific CD8(+) T cells to secrete IFN-γ in our assays. Next, H-2L(d)-restricted peptide-loaded and fluorescently labeled tetramers were produced in conjunction with the NIH Tetramer Core Facility. This tetramer was then tested for staining specificity and optimized for detection of PSA-specific CD8(+) T cells induced by our PSA-encoding adenovirus tumor vaccine. RESULTS: The MHC class I PSA peptide demonstrated successful restimulation of CD8(+) T cells isolated from mice previously vaccinated with a PSA-encoding adenovirus tumor vaccine, with no restimulation observed in control vaccinated mice. The peptide-loaded H-2L(d) tetramer exhibited the desired binding specificity and allowed for detection and frequency determination of PSA-specific CD8(+) T cells by flow cytometry. CONCLUSIONS: We have successfully designed and validated a PSA peptide tetramer for use in the Balb/c mouse model that can be used to test PSA-based prostate cancer vaccines. Until now, PSA-specific CD8(+) T cells in the mouse have only been detectable via cytotoxic T lymphocyte (CTL) assays or intracellular cytokine staining, which primarily assess Ag-specific functional activity, not their absolute number. This research tool provides laboratories the ability to directly quantitate CD8(+) T cells elicited by PSA-specific immunotherapies and cancer vaccines that are tested in mouse models.