Guggulsterone-induced apoptosis in human prostate cancer cells is mediated by reactive oxygen species-dependent activation of c-Jun N-terminal kinase

Abstract

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Guggulsterone [4,17(20)-(cis)-pregnadiene-3,16-dione] is a plant sterolderived from the gum resin (guggulu) of the tree Commiphoramukul that has been used extensively in Indian Ayurvedic medicine for the treatment of different ailments including bone fracture,arthritis, inflammation, cardiovascular disease, and lipid disorders. Guggulsteroneis an antagonist of bile acid farnesoid X receptor and regulates cholesterol homeostasis by increasing the transcription of bile salt export pump. Recent studies have also shown that guggulsterone is a potent suppressor of nuclear factor-κB (NF-κB) in tumor cells. NF-κB, a transcription factor constitutively activated in a variety of hematological and solid tumor cells including prostate cancer cells, is involved in regulation of various genes including inflammatory cytokines, chemokines, cell adhesion molecules, growth factors and interferon. We have shown previously that guggulsterone causes Bax/Bak-dependent apoptosis in PC-3 human prostate cancer cell line, whereas a normal prostate epithelial cell line (PrEC) is significantly more resistant to guggulsterone-induced cell death compared with prostate cancer cells. We now demonstrate that guggulsterone-induced apoptosis in human prostate cancer cells is regulated by reactive oxygen species (ROS)-dependent activation of c-Jun N-terminal kinase (JNK). Treatment of PC-3 (androgen-independent cell line lacking functional p53) and LNCaP cells (androgen responsive cell line with wild-type p53) with growth suppressive concentrations of guggulsterone (20 or 40 µM) resulted in activation of JNK and p38 mitogen activated protein kinase (MAPK) in both cell lines, and activation of extracellular signal-regulated kinase (ERK) in LNCaP cells. The guggulsterone-induced apoptosis in both cell lines was significantly attenuated in the presence of JNK-specific inhibitor SP600125. On the other hand, pharmacological inhibition of either ERK (using MEK1 inhibitor PD98059) or p38 MAPK (using p38 inhibitor SB202190) did not have any appreciable effect on guggulsterone-induced cell death in PC-3 or LNCaP cells. The guggulsterone-mediated activation of JNK correlated with generation of ROS in both PC-3 and LNCaP cells as judged by flow cytometry following staining with 6-carboxy-2',7'-dichlorodihydrofluoresceindiacetate. The guggulsterone-mediated JNK activation as well as cytoplasmic histone-associated DNA fragmentation (apoptosis) was significantly blocked on pretreatment with anti-oxidant N-acetylcysteine and overexpression of catalase and superoxide dismutase. In conclusion, the present study provides novel insights into the mechanism of guggulsterone-induced apoptosis in human prostate cancer cells involving ROS-dependent activation of JNK. This study was supported in part by NCI grants CA101753, CA113363 and CA115498.