Induction of endoplasmic reticulum stress and mitochondrial dysfunction dependent apoptosis signaling pathway in human renal cancer cells by norcantharidin

// Min-Hua Wu 1, 2 , Hui-Ling Chiou 3 , Chu-Liang Lin 4 , Ching-Yi Lin 5 , Shun-Fa Yang 1, 6 and Yi-Hsien Hsieh 4, 7, 8 1 Institute of Medicine, Chung Shan Medical University, Taichung, Taiwan 2 Department of Laboratory, Chung-Kang Branch, Cheng-Ching General Hospital, Taichung, Taiwan 3 School of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung, Taiwan 4 Institute of Biochemistry, Microbiology and Immunology, Chung Shan Medical University, Taichung, Taiwan 5 Division Of Chest Medicine, Department of Internal Medicine, Taichung Veterans General Hospital, Taichung, Taiwan 6 Department of Medical Research, Chung Shan Medical University Hospital, Taichung, Taiwan 7 Department of Biochemistry, School of Medicine, Chung Shan Medical University, Taichung, Taiwan 8 Clinical Laboratory, Chung Shan Medical University Hospital, Taichung, Taiwan Correspondence to: Shun-Fa Yang, email: ysf@csmu.edu.tw Yi-Hsien Hsieh, email: hyhsien@csmu.edu.tw Keywords: norcantharidin; apoptosis; renal cancer cells; mitochondrial depolarization; endoplasmic reticulum Received: September 07, 2017      Accepted: November 16, 2017      Published: December 19, 2017 ABSTRACT Previous studies reported that norcantharidin (NCTD) has anti-tumor effects. We investigated the antitumor effects and underlying mechanism of NCTD on human renal cancer in vitro and in vivo . NCTD significantly decreased renal cancer cell viability by induction of apoptosis, as determined by the MTT assay and annexin V/PI staining. NCTD treatment of 786-O and A-498 cells altered the expression of caspase family proteins and PARP. Moreover, NCTD induced mitochondrial depolarization, which was accompanied by an increased level of Bax and decreased levels of Bcl-2 and Mcl-1. NCTD induced endoplasmic reticulum (ER) stress by increasing the expression of Grp78, p-elF2α, ATF4, and CHOP. Pretreatment with an ER stress inhibitor (salubrinal) significantly attenuated the effect of NCTD. NCTD also induced activation of the AKT pathway in 786-O and A-498 cells. Overexpression of AKT partly reversed the effect of NCTD on apoptosis. NCTD treatment led to decreased expression of Bcl-2 and Mcl-1, and increased expression of Bax, cleaved-caspase-9, cleaved-PARP, and p-elF2α. Our in vivo studies demonstrated that NCTD significantly inhibited tumor growth in a nude mouse xenograft model. Taken together, our results suggest that NCTD is a potential anti-tumor agent for treatment of renal carcinoma.