Intracellular Activation of Bioorthogonal Nanozymes through Endosomal Proteolysis of the Protein Corona.

Bioorthogonal activation of prodrugs provides a strategy for on-demand on-site production of therapeutics. Intracellular activation provides a strategy to localize therapeutics, poten-tially minimizing off-target effects. To this end, nanoparticles embedded with transition metal catalysts (nanozymes) were engineered to generate either 'hard' irreversible or 'soft' re-versible coronas in serum. The hard corona induced nanozyme aggregation, effectively inhibiting nanozyme activi-ty, whereas only modest loss of activity was observed with the non-aggregating soft corona nanozymes. In both cases complete activity was restored by treatment with proteases. Intracellular activity mirrored this reactivation: endogenous proteases in the endosome provided intracellular activation of both nanozymes. The role of intracellular proteases in nanozyme reactivation was verified through treatment of the cells with protease inhibitors, which prevented reactivation. This study demonstrates the use of intracellular proteolysis as a strategy for localization of therapeutic generation to within cells.