Development of citrinin-loaded liposomes and in vitro toxicogenetic evaluation on breast cells

This study aimed to synthesize a liposomal nanoformulation containing citrinin (LP-CIT) to improve its availability and to evaluate its toxicogenomic profile, focusing on cytotoxic and genotoxic effects in breast cancer (MDA-MB-231) and non-tumorigenic mammary epithelial (MCF-10A) cell lines. The formulation was characterized by macroscopic appearance, pH, particle size, polydispersity index (PDI), zeta potential, encapsulation efficiency, and estability of nanoparticles. Citrinin content, encapsulation efficiency, and liposome stability were evaluated. Cytotoxicity was assessed using MTT, trypan blue exclusion, and flow cytometry. Cell death mechanisms were investigated by mitochondrial membrane potential analysis and expression of apoptotic markers (caspase-9 and BAX). Genotoxicity was evaluated using the comet assay. The liposomal nanosystem consisted of small unilamellar vesicles with a mean diameter of 143.3 nm, a PDI of 0.3, and a positive surface charge (+ 28.5 mV). The average was 63.69 ± 0.18% of citrinin encapsulated in liposomes with stability more than 21 days. Both free citrinin (CIT) and LP-CIT showed cytotoxic effects in MDA-MB-231 and MCF-10A cells, with lower LC₅₀ values observed for LP-CIT. Although LP-CIT induced more pronounced cytotoxic and genotoxic effects in the cancer cell line, toxic effects were also observed in non-tumorigenic cells, indicating limited selectivity. Cell death was associated with mitochondrial membrane depolarization and increased caspase-9 and BAX expression. Both formulations induced DNA damage as detected by the comet assay. Liposomal encapsulation enhanced citrinin biological activity but did not indicate tumor selectivity. Thus, this study represents a toxicogenomic assessment of citrinin and its liposomal formulation for potential therapeutic applicability.