Development\nof a Versatile Cancer Vaccine Format Targeting\nAntigen-Presenting Cells Using Proximity-Based Sortase A-Mediated\nLigation of T‑Cell Epitopes

Cancer vaccines are a promising strategy to increase\ntumor-specific\nimmune responses in patients who do not adequately respond to checkpoint\ninhibitors. Cancer vaccines that contain patient-specific tumor antigens\nare most effective but also necessitate the production of patient-specific\nvaccines. This study aims to develop a versatile cancer vaccine format\nin which patient-specific tumor antigens can be site-specifically\nconjugated by a proximity-based Sortase A (SrtA)-mediated ligation\n(PBSL) approach to antibodies that specifically bind to antigen-presenting\ncells to stimulate immune responses. DEC205 and CD169 are both receptors\nexpressed on antigen-presenting cells that can be targeted to deliver\nantigens and stimulate T-cell responses. We used the CRISPR/HDR platform\nto produce mouse heavy chain IgG2a antibodies with DEC205 or CD169\nspecificity containing an SrtA recognition motif followed by a SpyTag\nat the C-terminus. Using a recombinant protein of SrtA linked to SpyCatcher,\nwe applied proximity-based SrtA-mediated ligation to ligate fluorescein isothiocyanate\n(FITC)-labeled or antigenic peptides to the antibodies. Ligated antibodies\nbound to DEC205-expressing dendritic cells or CD169-expressing macrophages\nboth <i>in vitro</i> and <i>in vivo</i>. More\nimportantly, immunization with DEC205- or CD169-specific Abs linked\nto T-cell epitopes efficiently stimulated T-cell responses <i>in vivo</i>. To conclude, we have developed a cancer vaccine\nformat using PBSL that enables the rapid incorporation of tumor antigens\nand could potentially be implemented for the synthesis of personalized\ncancer vaccines.