Sanguinarine-induced apoptosis is associated with an early and severe cellular glutathione depletion.
OBJECTIVE
The quaternary benzophenanthridine alkaloid sanguinarine exhibits a broad range of activity, including cytotoxicity against various human tumour and normal cell lines. Here, we examined its potency as an anticancer drug.


METHODS
The differential cytotoxicity against cancer versus normal cells was assessed in vitro by two fluorimetric assays (RRT and Hoechst 33342 dye DNA assays, respectively) in a panel of human solid cancer cell lines and a human fibroblast primary culture. The ability to induce apoptosis was demonstrated in PC3 human prostatic adenocarcinoma cells by analysis of morphological changes, internucleosomal DNA fragmentation, cellular poly(ADP-ribose) polymerase cleavage and caspase 3/7 activation. Production of reactive oxygen species was evaluated by the 2',7'-dichlorofluorescin diacetate assay. Depletion of cellular glutathione content was assessed with the monochlorobimane assay.


RESULTS
Sanguinarine markedly inhibited the growth of all tested cells (IC(50) 0.9-3.3 microM) without differential cytotoxicity against normal versus cancer cells. In PC3 cells, continuous treatment with 5 microM sanguinarine induced an early (within 10 min) cellular reduced glutathione depletion insensitive to dithiothreitol or N-acetylcysteine treatment, followed by a caspase 3/7-dependent apoptotic response within 2 h. Complementary assays suggested that the glutathione depletion was initiated by direct reactivity of sanguinarine with reduced glutathione.


CONCLUSIONS
Taken together, these results show that (1) sanguinarine exhibits no specificity for cancer cells, and (2) its strong cytotoxicity is probably due to a rapid apoptotic response induced by an early and severe glutathione-depleting effect. They also suggest that the clinical usefulness of this alkaloid as an anticancer drug is limited.
Debiton
Eric
E
UMR-INSERM U-484, Rue Montalembert, BP 184, 63005, Clermont-Ferrand, France.
Madelmont
Jean-Claude
JC
Legault
Jean
J
Barthomeuf
Chantal
C
D000470:Alkaloids / Q000494:pharmacology*; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001562:Benzimidazoles; D053119:Benzophenanthridines; D053148:Caspase 3; D053179:Caspase 7; D020169:Caspases / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D053938:DNA Fragmentation; D004587:Electrophoresis, Agar Gel; D005978:Glutathione / Q000502:physiology*; D006801:Humans; D007202:Indicators and Reagents; D007546:Isoquinolines; D009369:Neoplasms / Q000473:pathology; D010078:Oxazines / Q000737:chemistry; D017382:Reactive Oxygen Species / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014966:Xanthenes
D000470:Alkaloids; D000972:Antineoplastic Agents, Phytogenic; D001562:Benzimidazoles; D053119:Benzophenanthridines; D007202:Indicators and Reagents; D007546:Isoquinolines; D010078:Oxazines; D017382:Reactive Oxygen Species; D014966:Xanthenes; C005843:resazurin; C005705:sanguinarine; C505407:CASP3 protein, human; C505520:CASP7 protein, human; D053148:Caspase 3; D053179:Caspase 7; D020169:Caspases; D005978:Glutathione; C017807:bisbenzimide ethoxide trihydrochloride