title: Application of the granzyme B ELISPOT assay for monitoring cancer vaccine trials.
abstract: Granzyme B (GrB) is present in the granules of cytolytic lymphocytes and is a key mediator of cell-mediated target cell death via the granule-mediated pathway. The release of GrB can be used as an indicator of a cytotoxic T lymphocyte response. Herein, we report that the GrB enzyme-linked immunospot assay (ELISPOT) can be used to measure ex vivo antigen-specific cytotoxicity of peripheral blood mononuclear cells from cancer patients vaccinated with a peptide-based cancer vaccine. We compare the reactivity of patients' peripheral blood mononuclear cells in the GrB ELISPOT with reactivity in the tetramer, interferon (IFN)-gamma ELISPOT, and Cr-release assays. Differences in immune response over all assays tested were found between patients and 4 response patterns were observed. Reactivity in the GrB ELISPOT was more closely associated with cytotoxicity in the Cr-release assay than the tetramer or IFN-gamma ELISPOT assays. Moreover, the higher affinity g209-2M peptide (used for vaccination) elicited greater GrB secretion than the native g209 peptide, although this difference was not observed with IFN-gamma secretion. Taken together with the fact that GrB is a specific mediator released by cytotoxic T lymphocytes, these results show that simultaneous use of the GrB ELISPOT assay with other immunologic assays may provide important additional immunologic insight into patient responses to cancer vaccines.
authors.affiliation: Laboratory of Cell-mediated Immunity, SAIC-Frederick, Inc, National Cancer Institute at Frederick, Frederick, MD 21702, USA.
authors.forename: Kimberly
authors.initials: K
authors.lastname: Shafer-Weaver
authors.forename: Steven
authors.initials: S
authors.lastname: Rosenberg
authors.forename: Susan
authors.initials: S
authors.lastname: Strobl
authors.forename: W
authors.initials: W
authors.lastname: Gregory Alvord
authors.forename: Michael
authors.initials: M
authors.lastname: Baseler
authors.forename: Anatoli
authors.initials: A
authors.lastname: Malyguine
mesh_terms: D000941:Antigens / Q000737:chemistry; D019496:Cancer Vaccines / Q000737:chemistry; D002986:Clinical Trials as Topic / Q000379:methods*; D003601:Cytotoxicity Tests, Immunologic / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D053804:Granzymes; D006801:Humans; D007107:Immune System; D007371:Interferon-gamma / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D008545:Melanoma / Q000097:blood* / Q000628:therapy; D008562:Membrane Glycoproteins / Q000737:chemistry; D015233:Models, Statistical; D010455:Peptides / Q000737:chemistry; D012697:Serine Endopeptidases / Q000737:chemistry*; D058951:gp100 Melanoma Antigen
chemical_list: D000941:Antigens; D019496:Cancer Vaccines; D008562:Membrane Glycoproteins; C548637:PMEL protein, human; D010455:Peptides; C548638:g209-2M melanoma peptide; D058951:gp100 Melanoma Antigen; D007371:Interferon-gamma; C509213:GZMB protein, human; D053804:Granzymes; D012697:Serine Endopeptidases