Enzyme-Linked Immunosorbent Spot Assay for the Detection of Wilms’ Tumor 1-Specific T Cells Induced by Dendritic Cell Vaccination

BACKGROUND Despite recent advances in cancer immunotherapy and the development of various assays for T cell assessment, a lack of universal standards within immune monitoring remains. The objective of this study was to evaluate the enzyme-linked immunosorbent spot (ELISpot) assay in comparison with major histocompatibility complex-tetramer analysis in the context of dendritic cell (DC)-based cancer immunotherapy. METHODS The ELISpot assay was performed on peripheral blood mononuclear cells to assess reproducibility, daily precision, and linearity using HLA-A*24:02-restricted Cytomegalovirus peptide. Wilms' tumor 1 (WT1) antigen-specific cytotoxic T cells were then evaluated by both the ELISpot assay and WT1 tetramer analysis in peripheral blood from 46 cancer patients who received DC vaccinations pulsed with human leukocyte antigen (HLA)-A*24:02-restricted modified WT1 peptides. RESULTS The ELISpot assay was proven to have reproducibility (coefficient of variation (CV) ranged from 7.4% to 16.3%), daily precision (CV ranged from 5.0% to 17.3%), and linearity (r = 0.96-0.98). WT1-specific immune responses were detected by the ELISpot assay in 34 out of 46 patients (73.9%) post-vaccination. A Spearman's rank-correlation coefficient of 0.82 between the ELISpot assay and WT1 tetramer analysis was obtained. CONCLUSION This is the first report of a comparison of an ELISpot assay and tetramer analysis in the context of dendritic cell (DC)-based cancer immunotherapy. The ELISpot assay has reproducibility, linearity, and excellent correlation with the WT1 tetramer analysis. These findings suggest that the validated ELISpot assay is useful to monitor the acquired immunity by DC vaccination targeting WT1.