Characterization of a thymidylate synthase (TS)-inducible cell line: a model system for studying sensitivity to TS- and non-TS-targeted chemotherapies.
Thymidylate synthase (TS) is responsible for the de novo synthesis of thymidylate, which is required for DNA synthesis and repair and which is an important target for fluoropyrimidines such as 5-fluorouracil (5-FU), and antifolates such as Tomudex (TDX), ZD9331, and multitargeted antifolate (MTA). To study the importance of TS expression in determining resistance to these agents, we have developed an MDA435 breast cancer-derived cell line with tetracycline-regulated expression of TS termed MTS-5. We have demonstrated that inducible expression of TS increased the IC(50) dose of the TS-targeted therapeutic agents 5-FU, TDX, and ZD9331 by 2-, 9- and 24-fold respectively. An IC(50) dose for MTA was unobtainable when TS was overexpressed in these cells, which indicated that MTA toxicity is highly sensitive to increased TS expression levels. The growth inhibitory effects of the chemotherapeutic agents CPT-11, cisplatin, oxaliplatin, and Taxol were unaffected by TS up-regulation. Cell cycle analyses revealed that IC(50) doses of 5-FU, TDX and MTA caused an S-phase arrest in cells that did not overexpress TS, and this arrest was overcome when TS was up-regulated. Furthermore, the S-phase arrest was accompanied by 2- to 4-fold increased expression of the cell cycle regulatory genes cyclin E, cyclin A, and cyclin dependent kinase 2 (cdk2). These results indicate that acute increases in TS expression levels play a key role in determining cellular sensitivity to TS-directed chemotherapeutic drugs by modulating the degree of S-phase arrest caused by these agents. Moreover, CPT-11, cisplatin, oxaliplatin, and Taxol remain highly cytotoxic in cells that overexpress TS.
Longley
D B
DB
Department of Oncology, Cancer Research Centre, Queen's University of Belfast, Belfast City Hospital, Lisburn Road, Belfast BT9 7AB, Northern Ireland.
Ferguson
P R
PR
Boyer
J
J
Latif
T
T
Lynch
M
M
Maxwell
P
P
Harkin
D P
DP
Johnston
P G
PG
D015152:Blotting, Northern; D015153:Blotting, Western; D042846:CDC2-CDC28 Kinases; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000494:pharmacology; D002453:Cell Cycle / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002945:Cisplatin / Q000494:pharmacology; D019925:Cyclin A / Q000187:drug effects / Q000378:metabolism; D019927:Cyclin E / Q000187:drug effects / Q000378:metabolism; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases / Q000187:drug effects / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005472:Fluorouracil / Q000494:pharmacology; D005493:Folic Acid Antagonists / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D000077146:Irinotecan; D009369:Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology; D009944:Organoplatinum Compounds / Q000494:pharmacology; D000077150:Oxaliplatin; D017239:Paclitaxel / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000187:drug effects / Q000378:metabolism; D011799:Quinazolines / Q000494:pharmacology; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D013752:Tetracycline / Q000494:pharmacology*; D013876:Thiophenes / Q000494:pharmacology; D013940:Thymidylate Synthase / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D014407:Tumor Cells, Cultured
D019925:Cyclin A; D019927:Cyclin E; D005493:Folic Acid Antagonists; D009944:Organoplatinum Compounds; D011799:Quinazolines; D012333:RNA, Messenger; D013876:Thiophenes; C094299:ZD 9331; D000077150:Oxaliplatin; D000077146:Irinotecan; D013940:Thymidylate Synthase; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases; D013752:Tetracycline; C068874:raltitrexed; D017239:Paclitaxel; D002945:Cisplatin; D005472:Fluorouracil; D002166:Camptothecin