Cell-derived vesicles modified with pH-sensitive polymers for intracellular delivery of autologous antigen

Abstract

Event Abstract Back to Event Cell-derived vesicles modified with pH-sensitive polymers for intracellular delivery of autologous antigen Eiji Yuba1*, Takanori Sakano1*, Takumi Kato1*, Takuma Urasaki1, Atsushi Harada1* and Kenji Kono1* 1 Osaka Prefecture University, Graduate School of Engineering, Japan Introduction: Efficient antigen carriers to dendritic cells (DCs) are needed for establishment of cancer immunotherapy. We previously reported cytoplasmic delivery using liposomes modified with pH-sensitive polymers, such as carboxylated poly(glycidol)s or dextrans[1],[2]. These liposomes delivered model antigen, ovalbumin (OVA), into cytosol of DCs and induced OVA-specific antitumor immunity. However, tumor antigen of each patient is different because of the difference of genetic mutation in each patient. Therefore, cancer cell-derived vesicles, such as plasma membrane vesicles (PMVs) or exosomes, were used as resources of tumor antigen and modified with pH-sensitive polymers (Figure 1). In this study, the effects of polymer modification on antigen delivery performance of cell-derived vesicles were examined. Materials and Methods: PMVs or exosomes were extracted from cancer cells, such as murine T lymphoma E.G7-OVA cells or their culture medium using repeated centrifugation, respectively. They were modified with 3-methylglutarylated or 2-carboxycyclohexane-1-carboxylated poly(glycidol)s (MGluHPG or CHexHPG, respectively) and characterized using DLS, AFM and TEM. pH-sensitivity of vesicles were measured by change in optical density or fusion assay based on FRET. Fluorescence-labeled vesicles were added to DC2.4 cells, a murine dendritic cell line, and then, cellular association and intracellular distribution of vesicles were evaluated by FCM and CLSM. These vesicles or vesicle-treated DCs were administered to tumor-bearing mice and their antitumor effects were investigated. Results and Discussion: Polymer-modified PMVs showed around 300 nm of diameters, while PMV formed micrometer-sized aggregates, indicating that polymer-modification increased the colloidal stability of PMVs (Figure 2A). Exosomes showed almost same size distribution irrespective of polymer modification. Polymer-modified vesicles showed increase of optical density and induced fusion with liposomes at acidic pH, which suggests that polymers became hydrophobic at acidic pH and induced fusion with other membranes. Polymer modification increased cellular association of PMVs and exosomes to DCs, indicating that carboxyl groups in polymers modified onto vesicles were recognized by scavenger receptors of DCs. Polymer-modified vesicles delivered their contents into cytosol of DCs (Figure 2B). Antitumor effects induced by administration of these vesicles or vesicle-treated DCs were promoted by polymer-modification probably because of their efficient delivery of cancer cell derived-antigen into cytosol of DCs. Conclusion: In this study, cancer cell-derived vesicles modified with pH-sensitive polymers were developed. Polymer modification to PMVs or exosomes enhanced their cellular association, intracellular delivery performance and antitumor effects. Therefore, cancer cell-derived vesicles modified with pH-sensitive polymers are promising as delivery carriers of autologous cancer antigen for establishment of cancer immunotherapy. This work was supported by grant from The Naito Foundation.