Pre-occupation of Empty Carriers Decreases Endo/Lysosome Escape and Reduces the Protein Delivery Efficiency of Mesoporous Silica Nanoparticles.

Endo/lysosome escape is a major challenge in nanoparticles-based protein delivery for cancer therapy. To enhance endo/lysosomal escape and increase the efficacy of proteins delivery, current strategies mainly focuses on destroying endo/lysosomes by employing modified nanoparticles, such as pH-sensitive polyplexes, cell-penetrating peptides, and photosensitive molecules. Herein, we hypothesize that pre-treatment with empty nanocarriers might make endo/lysosomes occupied and affect the endo/lysosomal escape of subsequent protein delivery by nanocarriers. We first treated breast carcinoma MDA-MB-231 cells with a high concentration of empty nanocarriers mesoporous silica nanoparticles (MSN), to occupy endo/lysosome. After 2 hours, we treated the cells with a lower concentration of fluorescein isothiocyanate-labeled MSN (MSN-FITC) and investigated the intracellular spatial and temporal distribution of MSN-FITC and its colocalization with endo/lysosomes. We discovered the pre-occupation of endo/lysosomes by the empty nanocarriers did exist, mainly through changing the spatial distribution of the subsequently introduced nanocarriers. Furthermore, for the protein delivery, we observed reduced MSN-saporin delivery after endo/lysosome pre-occupation by MSN empty carriers. Similar result is observed for delivery of Cytochrome C by MSN, but not for small molecule anti-cancer drug doxorubicin. The results show that the empty nanocarriers inhibit the endo/lysosome intracellular trafficking process and decrease the endo/lysosome escape of proteins subsequently delivered by the nanocarriers. This new discovered phenomenon of declined endo/lysosome escape after endo/lysosome pre-occupation indicates that repeated treatment by nanomaterials with low protein loading capacity may not yield good cancer therapeutic effect. Therefore, it provides a new insightful perspective on the role of nanomaterial carriers in intracellular protein delivery.