We describe a method for discovery of new tumor antigens that uses dendritic cells (DCs) as antigen-presenting cells to prime autologous naïve CD4+ and CD8+ T cells from healthy donors against tumor proteins and peptides. For the identification of HLA class I-restricted tumor antigens, peptides were extracted from tumor HLA class I molecules, fractionated by reverse phase-high performance liquid chromatography, and loaded onto in vitro-generated DCs to prime naïve CD8+ T cells. Our results show that we were able to prime naïve CD8+ T cells in vitro to several peptide fractions and generate specificity for the tumor. Electrospray ionization mass spectrometry was used to confirm that these fractions contained peptides derived from MHC class I molecules, and the primed CD8+ T cells were used to further analyze the immunostimulatory peptide fractions. For the identification of HLA class II-restricted tumor antigens, we fractionated tumor protein extracts using reverse phase-high performance liquid chromatography and loaded individual fractions onto DCs to prime naïve CD4+ T cells. Our results show that we were also able to prime naïve CD4+ T cells to several protein fractions and generate specificity for the tumor. These results illustrate the potential of this method to identify new immunostimulatory MHC class I- and class II-restricted tumor antigens.
This work is supported by Department of Defense Grant DAMD 17-9-1-7057 (to H. K.) and NIH Grant 1PO1CA 73743 (to O. J. F.).