Chemically Tuning the Antigen Release Kinetics from Spherical Nucleic Acids Maximizes Immune Stimulation

[Image: see text] Cancer vaccine structure is emerging as an important design factor that offers tunable parameters to enhance the targeted immune response. We report the impact of altering the antigen release rate from spherical nucleic acid (SNA) vaccines—nanoparticles with a liposomal core and surface-anchored adjuvant DNA—on immune stimulation. Peptide antigens were incorporated into SNAs using either a nonreducible linker or one of a series of reduction-triggered traceless linkers that release the native peptide at rates controlled by their substitution pattern. Compared with a nonreducible linkage, the traceless attachment of antigens resulted in lower EC(50) of T cell proliferation in vitro and greater dendritic cell (DC) activation and higher T cell killing ability in vivo. Traceless linker fragmentation rates affected the rates of antigen presentation by DCs and were correlated with the in vitro potencies of SNAs. Antigen release was correlated with the ex vivo −log(EC(50)), and more rapid antigen release resulted in an order of magnitude improvement in the EC(50) and earlier and greater antigen presentation over the same time-period. In vivo, increasing the rate of antigen release resulted in higher T cell activation and target killing. These findings provide fundamental insights into and underscore the importance of vaccine structure.