Functional delivery of xenogeneic p53 via extracellular vesicles induces apoptosis in human cancer cells

TP53 (encoding for the p53 tumor suppressor protein) is the most frequently mutated or deleted gene in human cancers, undermining p53's critical role in regulating cell cycle arrest and apoptosis. Here, we present a novel p53 replacement strategy using extracellular vesicles derived from chicken corneal epithelial cells, which naturally express high levels of cytoplasmic wild type p53. We demonstrate that corneal epithelium cells-derived extracellular vesicles encapsulate xenogeneic p53 and are readily internalized by human cancer cells, including those with mutant or null p53 backgrounds. Upon uptake, extracellular vesicles-delivered p53 translocates to the nucleus and activates canonical p53-dependent apoptotic pathways, evidenced by transcriptional upregulation of target genes such as p21, BAX, and PIDD, and induction of apoptosis across a range of human cancer cell lines. Proteomic and phosphoproteomic analyses of extracellular vesicles’ cargo revealed enrichment of apoptotic regulators and phosphorylated p53-associated proteins. In addition, p53-containing vesicles exhibited minimal cytotoxicity in primary human fibroblasts and human peripheral blood mononuclear cells, suggesting a selective anti-tumor activity. We also show that pre-treatment with chemotherapeutic agents sensitized cancer cells to vesicles-delivered p53, manifested by enhanced apoptosis. This study introduces an extracellular vesicles-based approach for functional p53 replacement, offering a promising therapeutic platform for targeting tumors with p53 loss or mutation.