Effect of hammerhead ribozyme against human thymidylate synthase on the cytotoxicity of thymidylate synthase inhibitors.
One of the resistance mechanisms to folate-based thymidylate synthase (TS) inhibitors is the increase in TS activity in tumor cells. Human B lymphoblastoid cell line (W1L2) was made resistant to a lipophilic non-polyglutamatable TS inhibitor (ZM249148), and the subline (W1L2:R179) showed a 20-fold increase in TS enzyme activity with concomitant overexpression of TS mRNA. To overcome the resistance, we designed a ribozyme that can cleave the CUC sequences in a triple tandemly repeated sequence of TS mRNA. Expression of this ribozyme in W1L2:R179 cells transfected with Epstein Barr virus-based expression vector resulted in sensitization to TS inhibitors concomitantly with a decrease of TS expression. The ribozyme expressed in transfectants was shown to be functional in cleaving artificial TS RNA in vitro.
Kobayashi
H
H
Department of Laboratory Medicine, National Defense Medical College, Saitama.
Takemura
Y
Y
Miyachi
H
H
Skelton
L
L
Jackman
A L
AL
D001402:B-Lymphocytes / Q000187:drug effects / Q000201:enzymology; D001483:Base Sequence; D004351:Drug Resistance; D004357:Drug Synergism; D004790:Enzyme Induction / Q000187:drug effects; D005492:Folic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D005493:Folic Acid Antagonists / Q000494:pharmacology; D006801:Humans; D008727:Methotrexate / Q000494:pharmacology; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011799:Quinazolines / Q000494:pharmacology*; D016337:RNA, Catalytic / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism*; D012091:Repetitive Sequences, Nucleic Acid; D013876:Thiophenes / Q000494:pharmacology; D013940:Thymidylate Synthase / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection; D016597:Trimetrexate / Q000494:pharmacology
D005493:Folic Acid Antagonists; D011799:Quinazolines; D016337:RNA, Catalytic; D012333:RNA, Messenger; D013876:Thiophenes; C094299:ZD 9331; C097903:ZM 249148; C031662:CB 3717; D005492:Folic Acid; D013940:Thymidylate Synthase; C068874:raltitrexed; D016597:Trimetrexate; D008727:Methotrexate