Development of a Versatile Cancer Vaccine Format Targeting Antigen-Presenting Cells Using Proximity-Based Sortase A-Mediated Ligation of T-Cell Epitopes

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