Design, Synthesis and Antitumor Activity Evaluation of Novel Ergosterol Peroxide Derivatives Based on Carbamate and Urea/Thiourea Dual Pharmacophores

Ergosterol peroxide (EP) is a natural endoperoxide with a widely range of antitumor activities. However, its low bioavailability and moderate activity have limited its further application. Through the structural modification of the C-3 position of EP, a series of EP derivatives coupled with carbamate and urea/thiourea moieties were successfully synthesized. The cytotoxicity of these derivatives against three human cancer cell lines (human non-small cell lung cancer A549 cells; human hepatocellular cancer HepG2 cells; human breast cancer MCF-7 cells) was assessed through the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Compound 4n showed the most significant cytotoxicity against the A549 cell line, with the half-maximal inhibitory concentration (IC50) value of 2.86 μM, which is 6.8 times higher than that of EP (IC50 = 19.42 μM). In contrast, compound 4n showed low toxicity in human embryonic lung fibroblasts (MRC-5, IC50 = 80.64 μM), and had a selectivity index of 28.19 (IC50 MRC-5/IC50 A549). Apoptosis assays demonstrated that compound 4n induced substantial early apoptosis (22.9%) and late apoptosis (8.7%) in the A549 cell line, whereas cells in the control condition exhibited only 4.0% early and 1.7% late apoptosis. Further mechanistic studies revealed that, in the A549 cell line, compound 4n increased the intracellular reactive oxygen species (ROS) levels and decreased the mitochondrial membrane potential (MMP, 30.1% change) compared to the control and ultimately led to apoptosis. These results suggest that compound 4n has the potential to be developed as a clinical drug.