Inhibition by 5-fluorouracil of ERCC1 and gamma-glutamylcysteine synthetase messenger RNA expression in a cisplatin-resistant HST-1 human squamous carcinoma cell line.
Pretreatment of 5-fluorouracil (5-FU), but not posttreatment, has been shown to augment the cytotoxicity of cisplatin (CDDP) or even circumvent CDDP resistance by inhibiting repair of platinum-DNA interstrand crosslinks as well as by reducing the cellular glutathione (GSH) contents in CDDP-resistant HST-1/CP0.2 human squamous carcinoma cells. Because exogenous thymidine, which compensates for 5-FU-mediated inhibition of de novo DNA synthesis via salvage pathway, did not affect this schedule-dependent synergism, the modulatory effect of 5-FU on CDDP resistance would be attributed to the 5-FU-induced RNA damage. We therefore examined the effect of 5-FU on the steady-state levels of messenger RNA (mRNA) of a human excision repair gene ERCC1 and gamma-glutamylcysteine synthetase (gamma-GCS) gene coding for a rate-limiting enzyme for GSH synthesis. The HST-1/ CP0.2 cells were found to have significantly more mRNA expression of these respective genes than do parental HST-1 cells. In these cells, 5-FU pretreatment progressively inhibited mRNA expression of both ERCC1 and gamma-GCS after removal of 5-FU, without affecting glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA. A maximal mRNA suppression was observed at 48 h posttreatment. Such 5-FU-induced suppression of mRNA transcripts of these genes seems to be consistent with its inhibitory activity on DNA repair capacity and cellular GSH contents. In contrast, 5-FU did not reduce the level of glutathione-S-transferase-pi (GST-pi) or DNA topoisomerase 1 mRNA. Although not convinced, our data suggest that 5-FU, when incorporated into RNA, may inhibit both GSH synthesis and repair of platinum-DNA adducts by downregulating the ERCC1 and gamma-GCS genes, thereby enhancing antitumor activity of CDDP and reversing resistance to CDDP in HST-1/CP0.2 cells.
Fujishima
H
H
First Department of Internal Medicine, Faculty of Medicine, Kyushu University, Fukuoka, Japan.
Nakano
S
S
Masumoto
N
N
Esaki
T
T
Tatsumoto
T
T
Kondo
T
T
Niho
Y
Y
D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D002294:Carcinoma, Squamous Cell / Q000473:pathology*; D002945:Cisplatin / Q000494:pharmacology; D004260:DNA Repair / Q000187:drug effects; D004264:DNA Topoisomerases, Type I / Q000235:genetics; D004268:DNA-Binding Proteins; D019008:Drug Resistance, Neoplasm; D004720:Endonucleases; D005472:Fluorouracil / Q000494:pharmacology*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D005721:Glutamate-Cysteine Ligase / Q000235:genetics*; D006801:Humans; D011506:Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D012334:RNA, Neoplasm / Q000235:genetics; D013936:Thymidine / Q000494:pharmacology
D000964:Antimetabolites, Antineoplastic; D004268:DNA-Binding Proteins; D011506:Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; C071447:ERCC1 protein, human; D004720:Endonucleases; D004264:DNA Topoisomerases, Type I; D005721:Glutamate-Cysteine Ligase; D002945:Cisplatin; D005472:Fluorouracil; D013936:Thymidine