Extracellular vesicles from IL-18-overexpressing and/or TGF-β1-deprived tumor cells as an immunogenic antigen source for next-generation DC vaccines

Dendritic cell (DC)-based antitumor immunotherapies have demonstrated significant therapeutic potential. Nonetheless, their broader clinical application is limited for several reasons, including the lack of comprehensive sources of tumor antigens. Tumor-derived extracellular vesicles (TEVs) provide a rich source of tumor-associated antigens. However, unmodified TEVs, due to the high levels of immunosuppressive molecules, inhibit processes related to antigen processing and their presentation to naïve T lymphocytes. This study aimed to engineer TEVs via lentiviral modification of parental colon carcinoma MC38 cells to overexpress IL-18 and/or shRNA targeting IL-10 or TGF-β1 to enhance the immunogenic potential of TEVs. The modified TEVs were characterized with TEM, Western blotting, ELISA, and RT-qPCR. Functional activity was tested in vitro using flow cytometry and confocal imaging. In vivo studies used a C57BL/6 syngeneic MC38 tumor model. mTEVs from MC38 cells engineered to overexpress IL-18 and/or silence TGF-β1 strongly stimulated DCs, triggering a potent, multifaceted antitumor response in vitro and in vivo . The combination of both modifications exerted an additive effect, enhancing the therapeutic efficacy of DCs and significantly inhibiting tumor growth. In addition to altering cytokine levels, genetic modifications also changed the vesicle's miRNA and protein cargo, potentially boosting their immunostimulatory role. Our findings provide compelling evidence that rational genetic engineering of tumor cells can effectively transform the extracellular vesicles they produce, shifting their immunosuppressive, tumor-supporting potential into an immunogenic one. Interleukin-18-loaded and TGF-β1-deprived TEVs can serve as a highly versatile and naturally immunogenic platform for the next-generation DC-based anticancer vaccines. • Modified TEVs constitute an effective antigen source for dendritic cell activation. • mTEV-stimulated DCs increase cytotoxic activity of T lymphocytes. • IL-18 overexpression in mTEVs exhibits the highest immunostimulatory potential. • MC38/shTGFβ1/IL18 TEVs in combination with CY decreased tumor growth by 95%. • Employed modifications resulted in significant miRNA and protein changes in mTEVs.