Knockdown of NAPA using short-hairpin RNA sensitizes cancer cells to cisplatin: implications to overcome chemoresistance.
Cisplatin is a widely used anti-cancer drug which targets DNA in replicating cells. In the present study, we found that NAPA--a protein found in the endoplasmic reticulum (ER) and implicated in protein trafficking--protects cells against cisplatin. Accordingly, knockdown of NAPA using lentivirus-encoded shRNA (shNAPA) induced ER stress similar to cisplatin treatment in HEK293 cells. A low dose of cisplatin also elicited a mild ER stress response associated with the accumulation of the protective proteins BiP and NAPA. Remarkably, knockdown of NAPA induced apoptosis and enhanced cisplatin-induced cytotoxicity/apoptosis, thereby sensitizing cancer cells to cisplatin. On the other hand, overexpression of NAPA increased resistance to cisplatin by reducing cisplatin-induced ER stress and apoptosis. The modulatory effects of shNAPA required the tumor suppressor p53 since the effects of NAPA knockdown were reduced by the p53 inhibitor PFT-alpha and in H1299 cells which are p53-null. A partial reversal of cisplatin resistance was also observed in cisplatin-resistant HeLa cells following knockdown of NAPA. Our results also indicated that calpain is required for ER-mediated apoptosis. Importantly, combined cisplatin/shNAPA treatment suppressed tumor growth in vivo in xenograph experiments performed in nude mice. Taken together, these observations suggest that NAPA represents a target of cisplatin, and that knockdown of NAPA may improve cisplatin-based cancer therapy.
Wu
Zchong-Zcho
ZZ
Department of Biochemistry and Molecular Biology, Graduate Institute of Biomedical Sciences, Chang Gung University, 259 Wen-Hua, 1st Road, Gueishan, Taoyuan 333, Taiwan, ROC. D9601101@stmail.cgu.edu.tw
Chao
Chuck C-K
CC
D000818:Animals; D000970:Antineoplastic Agents / Q000627:therapeutic use; D002460:Cell Line; D002945:Cisplatin / Q000627:therapeutic use; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D005260:Female; D055785:Gene Knockdown Techniques / Q000379:methods; D006367:HeLa Cells; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D051379:Mice; D008819:Mice, Nude; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000737:chemistry / Q000235:genetics*; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics; D023041:Xenograft Model Antitumor Assays / Q000379:methods
D000970:Antineoplastic Agents; D034741:RNA, Small Interfering; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D002945:Cisplatin