title: Enterocarpam-III induces human liver and breast cancer cell apoptosis via mitochondrial and caspase-9 activation.
abstract: An aristolactam-type alkaloid, isolated from Orophea enterocarpa, is enterocarpam-III (10-amino-2,3,4,6- tetramethoxyphenanthrene-1-carboxylic acid lactam). It is cytotoxic to various human and murine cancer cell lines; however, the molecular mechanisms remain unclear. The aims of this study were to investigate cytotoxic effects on and mechanism (s) of human cancer cell death in human hepatocellular carcinoma HepG2 and human invasive breast cancer MDA-MB-231 cells compared to normal murine fibroblast NIH3T3 cells. Cell viability was determined by MTT assay to determine IC10, IC20 and IC50 levels, reactive oxygen species (ROS) production with 2',7'-dichlorohydrofluorescein diacetate and the caspase-3, -8 and -9 activities using specific chromogenic (p-nitroaniline) tetrapeptide substrates, viz., DEVD-NA, IETD-NA and LEHD-NA and employing a microplate reader. Mitochondrial transmembrane potential (MTP) was measured by staining with 3, 3'-dihexyloxacarbocyanine iodide (DiOC6) and using flow cytometry. The compound was cytotoxic to HepG2 and MDA-MB-231 cells with the IC50 levels of 26.0±4.45 and 51.3±2.05 μM, respectively. For murine normal fibroblast NIH3T3 cells, the IC50 concentration was 81.3±10.1 μM. ROS production was reduced in a dose-response manner in HepG2 cells. The caspase-9 and -3 activities increased in a concentration-dependent manner, whereas caspase-8 activity did not alter, indicating the intrinsic pathway activation. Enterocarpam-III decreased the mitochondrial transmembrane potential (MTP) dose-dependently in HepG2 cells, suggesting that the compound induced HepG2 cell apoptosis via the mitochondrial pathway. In conclusion, enterocarpam-III inhibited HepG2 and MDA-MB-231 cell proliferation and induced human HepG2 cells to undergo apoptosis via the intrinsic (mitochondrial) pathway and induction of caspase-9 activity.
authors.affiliation: Department of Biochemistry, Faculty of Medicine, Chiang Mai University, Chiang Mai, Thailand E-mail : rbanjerd@gmail.com.
authors.forename: Ratana
authors.initials: R
authors.lastname: Banjerdpongchai
authors.forename: Benjawan
authors.initials: B
authors.lastname: Wudtiwai
authors.forename: Wilart
authors.initials: W
authors.lastname: Pompimon
mesh_terms: D016475:3T3 Cells; D000470:Alkaloids / Q000494:pharmacology*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001943:Breast Neoplasms / Q000188:drug therapy*; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy*; D053148:Caspase 3 / Q000378:metabolism; D053181:Caspase 8 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D056945:Hep G2 Cells; D006801:Humans; D007769:Lactams / Q000494:pharmacology*; D008113:Liver Neoplasms / Q000188:drug therapy*; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D051379:Mice; D008928:Mitochondria / Q000378:metabolism; D010616:Phenanthrenes / Q000494:pharmacology*; D010936:Plant Extracts / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000470:Alkaloids; D007769:Lactams; D010616:Phenanthrenes; D010936:Plant Extracts; D017382:Reactive Oxygen Species; C000604570:enterocarpam-III; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9