Cytotoxicity of epunctanone and four other phytochemicals isolated from the medicinal plants Garcinia epunctata and Ptycholobium contortum towards multi-factorial drug resistant cancer cells

Introduction: Resistance of cancer cells is a serious impediment to chemotherapy and several phytochemicals are active against multi-drug resistant (MDR) phenotypes. The cytotoxicity of five naturally occurring compounds: betulin (1), mundulea lactone (2), seputhecarpan A (3), seputheisoflavone (4) and epunctanone (5) was evaluated on a panel of 9 cancer cell lines including various sensitive and drug-resistant cell lines. The modes of action of compound 5 were further investigated. Methods: The resazurin reduction assay was used to evaluate cytotoxicity of samples and ferroptotic cell death induced by compound 5; caspase-Glo assay was used to detect the activation of caspases in CCRF-CEM leukemia cells treated with compound 5. Flow cytometry was used for cell cycle analysis in CCRF-CEM cells treated with compound 5, as well as detection of apoptotic cells by annexin V/PI staining, analysis of mitochondrial membrane potential (MMP) and measurement of reactive oxygen species (ROS). Results: Compounds 1-5 displayed cytotoxic effects in the 9 studied cancer cell lines with IC50 values below 70 mu M. The IC50 values varied from 8.20 mu M (in HCT116 (p53(-/-)) colon cancer cells) to 35.10 mu M (against HepG2 hepatocarcinoma cells) for 1, from 8.84 mu M (in CEM/ADR5000 leukemia cells) to 48.99 mu M (in MDA-MB-231 breast adenocarcinoma cells) for 2, from 12.17 mu M (in CEM/ADR5000 cells) to 65.08 mu M (in MDA-MB-231 cells) for 3, from 23.80 mu M (in U87MG.Delta EGFR glioblastoma cells) to 68.66 mu M (in HCT116 (p53(-/-)) cells) for 4, from 4.84 mu M (in HCT116 (p53(-/-)) cells) to 13.12 mu M (in HepG2 cells) for 5 and from 0.02 mu M (against CCRF-CEM cells) to 122.96 mu M (in CEM/ADR5000 cells) for doxorubicin. Compound 5 induced apoptosis in CCRF-CEM cells through alteration of MMP and increase in ROS production. In addition to apoptosis, ferroptosis was also identified as another mode of cell death induced by epunctanone. Conclusions: Compounds 1-5 are valuable cytotoxic compounds that could be used to combat MDR cancer cells. Benzophenoe 5 is the most active molecule and deserve more investigations to develop new anticancer drugs.