Reversal of cisplatin and multidrug resistance by ribozyme-mediated glutathione suppression.
gamma-Glutamylcysteine synthetase (gamma-GCS) is a key enzyme in glutathione (GSH) synthesis, and is thought to play a significant role in intracellular detoxification, especially of anticancer drugs. Increased levels of GSH are commonly found in the drug-resistant human cancer cells. We designed a hammerhead ribozyme against gamma-GCS mRNA (anti-gamma-GCS Rz), which specifically down-regulated gamma-GCS gene expression in the HCT-8 human colon cancer cell line. The aim of this study was to reverse the cisplatin and multidrug resistance for anticancer drugs. The cisplatin-resistant HCT-8 cells (HCT-8DDP cells) overexpressed MRP and MDR1 genes, and showed resistance to not only cisplatin (CDDP), but also doxorubicin (DOX) and etoposide (VP-16). We transfected a vector expressing anti-gamma-GCS Rz into the HCT-8DDP cells (HCT-8DDP/Rz). The anti-gamma-GCS Rz significantly suppressed MRP and MDR, and altered anticancer drug resistance. The HCT-8DDP/Rz cells were more sensitive to CDDP, DOX and VP-16 by 1.8-, 4.9-, and 1.5-fold, respectively, compared to HCT-8DDP cells. The anti-gamma-GCS Rz significantly down-regulated gamma-GCS gene expression as well as MRP/MDR1 expression, and reversed resistance to CDDP, DOX and VP-16. These results suggested that gamma-GCS plays an important role in both cisplatin and multidrug resistance in human cancer cells.
Nagata
J
J
Department of Pathology, Tokai University School of Medicine, Bohseidai, Kanagawa, 259-1193, Japan.
Kijima
H
H
Hatanaka
H
H
Asai
S
S
Miyachi
H
H
Takagi
A
A
Miwa
T
T
Mine
T
T
Yamazaki
H
H
Nakamura
M
M
Kondo
T
T
Scanlon
K J
KJ
Ueyama
Y
Y
D000970:Antineoplastic Agents / Q000627:therapeutic use*; D002945:Cisplatin / Q000627:therapeutic use*; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism / Q000628:therapy*; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D005721:Glutamate-Cysteine Ligase / Q000235:genetics; D005978:Glutathione / Q000037:antagonists & inhibitors*; D006801:Humans; D016337:RNA, Catalytic / Q000627:therapeutic use*; D012334:RNA, Neoplasm / Q000096:biosynthesis / Q000235:genetics; D013778:Tetrazolium Salts / Q000737:chemistry; D013844:Thiazoles / Q000737:chemistry; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D016337:RNA, Catalytic; D012334:RNA, Neoplasm; D013778:Tetrazolium Salts; D013844:Thiazoles; D005721:Glutamate-Cysteine Ligase; C022616:thiazolyl blue; D005978:Glutathione; D002945:Cisplatin