Pei-Yu Yang1, Dan-Ning Hu2, Ying-Hsien Kao3, I-Ching Lin4, Chih-Yuan Chou5, Yang-Chang Wu6. 1. Department of Medical Research, Show Chwan Memorial Hospital, Changhua, Taiwan, ROC. Electronic address: peyyuh2900@gmail.com. 2. Tissue Culture Center, New York Eye and Ear Infirmary of Mount Sinai, New York, NY, USA. Electronic address: hu2095@yahoo.com. 3. Department of Medical Research, E-DA Hospital, Kaohsiung, Taiwan, ROC. Electronic address: danyhkao@gmail.com. 4. Department of Family Medicine, Changhua Christian Hospital, Changhua, Taiwan, ROC; School of Medicine, Chung Shan Medical University, Taichung, Taiwan, ROC; School of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan, ROC. Electronic address: licypy01@gmail.com. 5. Division of Urology, Department of Surgery, Show-Chwan Memorial Hospital, Changhua, Taiwan, ROC. Electronic address: yuan117@gmail.com. 6. School of Pharmacy, College of Pharmacy, China Medical University, Taichung, Taiwan, ROC. Electronic address: yachwu@mail.cmu.edu.tw.
Abstract
BACKGROUND: Norcantharidin, a modified pure compound from blister beetles, was previously demonstrated to induce apoptosis of cancer cells. This study investigated its anti-cancer activity in prostate cancer cells and the mechanisms involved. METHODS: Two human prostate cancer cell lines, 22Rv1 and Du145, were treated with norcantharidin at concentrations ranging from 3 to 30μg/ml. Cytotoxic effect of norcantharidin was determined by use of the 3-(4,5-dimethylthiazol-yl)-diphenyl tetrazoliumbromide (MTT) assay. The effects of apoptosis were evaluated by cell death assay, Caspase-3, -8, -9 activity and cytochrome c release. The apoptotic related protein expressions (Bcl-2 family and inhibitor of apoptosis proteins) were determined using western blotting. RESULTS: An MTT assay revealed that norcantharidin induced cytotoxicity against both prostate cancer cells in dose- and time-dependent manners. Treatment with norcantharidin at 3μg/ml or higher significantly increased oligonucleosomal formation with concomitant appearance of PARP cleavage, implicating the induction of apoptosis. Norcantharidin intrinsically elevated cytosolic cytochrome c levels and activated caspase-3, -8, and -9. Extrinsically, it upregulated the expression of not only the death receptors Fas and DR5 in 22Rv1 cells, but also of RIP and TRADD adaptor proteins in Du145 cells. Mechanistically, norcantharidin increased ratios of pro-/anti-apoptotic proteins and decreased expression of IAP family member proteins, including cIAP1 and survivin, regardless of the distinct status of androgen receptor expression in both cells. CONCLUSIONS: Norcantharidin exhibited cytotoxicity against 22Rv1 and Du145 prostate cancer cells by inducing both intrinsic and extrinsic apoptotic pathways and could thus potentially be a remedy for prostate cancer.
BACKGROUND:Norcantharidin, a modified pure compound from blister beetles, was previously demonstrated to induce apoptosis of cancer cells. This study investigated its anti-cancer activity in prostate cancer cells and the mechanisms involved. METHODS: Two humanprostate cancer cell lines, 22Rv1 and Du145, were treated with norcantharidin at concentrations ranging from 3 to 30μg/ml. Cytotoxic effect of norcantharidin was determined by use of the 3-(4,5-dimethylthiazol-yl)-diphenyl tetrazoliumbromide (MTT) assay. The effects of apoptosis were evaluated by cell death assay, Caspase-3, -8, -9 activity and cytochrome c release. The apoptotic related protein expressions (Bcl-2 family and inhibitor of apoptosis proteins) were determined using western blotting. RESULTS: An MTT assay revealed that norcantharidin induced cytotoxicity against both prostate cancer cells in dose- and time-dependent manners. Treatment with norcantharidin at 3μg/ml or higher significantly increased oligonucleosomal formation with concomitant appearance of PARP cleavage, implicating the induction of apoptosis. Norcantharidin intrinsically elevated cytosolic cytochrome c levels and activated caspase-3, -8, and -9. Extrinsically, it upregulated the expression of not only the death receptors Fas and DR5 in 22Rv1 cells, but also of RIP and TRADD adaptor proteins in Du145 cells. Mechanistically, norcantharidin increased ratios of pro-/anti-apoptotic proteins and decreased expression of IAP family member proteins, including cIAP1 and survivin, regardless of the distinct status of androgen receptor expression in both cells. CONCLUSIONS:Norcantharidin exhibited cytotoxicity against 22Rv1 and Du145 prostate cancer cells by inducing both intrinsic and extrinsic apoptotic pathways and could thus potentially be a remedy for prostate cancer.