Down-regulation of thymidylate synthase expression and its steady-state mRNA by oxaliplatin in colon cancer cells.
Recently, evidence has accumulated that weekly 24-h infusion of high-dose 5-fluorouracil (5-FU) with leucovorin (LV, folinic acid) biochemical modulation may improve the response rates compared with the bolus 5-FU regimens in colorectal cancer (CRC). Combining the infusional 5-FU/LV (iFL) regimens with oxaliplatin or irinotecan is widely adopted to further improve treatment efficacy. Either oxaliplatin-iFL or irinotecan-iFL may achieve an overall response rate of more than 50% in the first-line treatment. Intriguingly, in the salvage treatment for metastatic CRC patients who had failed iFL, only oxaliplatin-iFL may achieve a response rate of about 13-25%. In contrast, oxaliplatin alone or irinotecan-iFL had a very low response rate of 5% or less. To test if the oxaliplatin may reverse the iFL-related 5-FU resistance in CRC, we used DLD-1 colon adenocarcinoma cells as the in vitro study model. First, we revealed that oxaliplatin and 5-FU act synergistically on DLD-1 cells by MTT cytotoxicity assay and median drug effect analysis. Second, we treated the DLD-1 cells with serial concentrations of oxaliplatin (0.1-10 microM). Oxaliplatin treatment results in down-regulation of free thymidylate synthase (TS) protein expression by Western blotting. Further, we analyzed the TS mRNA level by reverse transcription and real-time quantitative polymerase chain reaction assay. Oxaliplatin treatment results in down-regulation of the TS mRNA level up to 40% (mean +/- SD of ratio to reference control = 0.60 +/- 0.21, range 0.42-0.84). In this study, our data provide important information explaining the reason why the combination of oxaliplatin and 5-FU results in a better objective response in 5-FU-resistant patients than oxaliplatin alone does. Our data also suggest that TS down-regulation happens at the transcriptional level. TS modulation and down-regulation had, thus, shed light on the useful potential strategy to achieve objective responses in 5-FU-resistant CRC patients.
Yeh
Kun-Huei
KH
Department of Internal Medicine, Far Eastern Memorial Hospital, Taiwan. kevinyeh@ha.mc.ntu.edu.tw
Cheng
Ann-Lii
AL
Wan
Joeu-Pei
JP
Lin
Chien-Shing
CS
Liu
Chih-Chun
CC
D000230:Adenocarcinoma / Q000201:enzymology; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology*; D015153:Blotting, Western; D002151:Calorimetry; D003110:Colonic Neoplasms / Q000201:enzymology; D015536:Down-Regulation; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D004357:Drug Synergism; D005472:Fluorouracil / Q000494:pharmacology; D006801:Humans; D020128:Inhibitory Concentration 50; D009944:Organoplatinum Compounds / Q000494:pharmacology*; D000077150:Oxaliplatin; D012333:RNA, Messenger / Q000096:biosynthesis*; D013940:Thymidylate Synthase / Q000096:biosynthesis* / Q000235:genetics; D014407:Tumor Cells, Cultured
D000964:Antimetabolites, Antineoplastic; D000970:Antineoplastic Agents; D009944:Organoplatinum Compounds; D012333:RNA, Messenger; D000077150:Oxaliplatin; D013940:Thymidylate Synthase; D005472:Fluorouracil