Synergistic effect of exogenous P53 and sodium butyrate on tumor cell survival

BACKGROUND: The P53 protein is a transcription factor that regulates the expression of genes involved in numerous cellular processes, including cell cycle arrest, apoptosis, cell proliferation, and DNA repair. Its role as a multifunctional tumor suppressor makes P53 an attractive and promising target for cancer therapy.

AIM: This work aimed to analyze the combined effect of transfection with a plasmid encoding the p53 gene (a genetic approach), the histone deacetylase inhibitor sodium butyrate (epigenetic regulation), and co-cultivation with exosomes secreted by cells expressing wild-type P53 (modeling intercellular communication) on the survival of various human tumor cell lines.

METHODS: The study was conducted using four transplantable cell lines: HeLa (epithelioid cervical carcinoma) and HT-1080 (fibrosarcoma) cells harboring the wild-type p53 gene, as well as K562 (chronic myelogenous leukemia) and Gl-V (primary glioma cell culture) cells deficient in p53 (P53^–/–). Cell transfection was performed using a P53-GFP plasmid constructed in the Laboratory of Cell Biology, National Research Center “Kurchatov Institute.” This plasmid encodes the P53 protein fused to green fluorescent protein (GFP) at its N-terminus. Successful transfection was confirmed by detecting P53-GFP expression using confocal microscopy. The level of P53 protein in the cells was determined by Western blotting. To quantitatively assess proliferation and cell cycle parameters under conditions of histone deacetylase inhibition, sodium butyrate (NaBu) was added to the culture medium at a final concentration of 2.5 mM. The analysis was performed using an automated cell counter, flow cytometry, or colony formation assays. Exosomes were isolated from the collected conditioned medium by ultracentrifugation.

RESULTS: When only one of the above approaches was applied, the outcome largely depended on the P53 status of the analyzed tumor cells. The combination of epigenetic modulation through inhibition of histone deacetylase activity with genetic regulation or exposure to exosomes derived from wild-type P53–expressing cells produced a pronounced synergistic effect and several-fold increase in the efficiency of tumor cell growth suppression compared with monotherapy.

CONCLUSION: It appears that a strategy combining genetic methods, epigenetic modulation, and intercellular communication mechanisms that affect different components of the P53 regulatory network may substantially enhance the efficacy of P53-targeted anticancer therapy.