Related Experiment Videos
UCN-01 suppresses thymidylate synthase gene expression and enhances 5-fluorouracil-induced apoptosis in a
C T Hsueh1, D Kelsen, G K Schwartz
1Department of Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Abstract:
UCN-01, a protein kinase C/cyclin-dependent kinase inhibitor, suppressed thymidylate synthase (TS) protein expression in a dose-dependent manner with near complete suppression at 1 microM after a 24-h exposure in human gastric cancer cell line SK-GT5. Other protein kinase C/cyclin-dependent kinase inhibitors, including flavopiridol and safingol, had a similar effect on TS protein expression, but to a lesser degree. Moreover, UCN-01 repressed the induction of TS after 5-fluorouracil (FU) exposure by 90-95% and significantly enhanced the induction of apoptosis by FU from 4-8% with either FU or UCN-01 alone to 46+/-1% (P < 0.005 versus either single drug, reverse sequence, or the combination) when UCN-01 was given after FU. The effect of UCN-01 on TS was associated with a dose-dependent suppression of the E2F-1 protein, a transcriptional activator of TS. Northern blot analysis revealed that TS mRNA levels decreased gradually as the concentration of UCN-01 increased, but that E2F-1 mRNA levels remained relatively unchanged. UCN-01 may provide a novel way to enhance cellular sensitivity toward FU by means of suppressing TS expression mediated mainly by down-regulation of E2F-1.
Insights
UCN-01, a kinase inhibitor, suppresses thymidylate synthase (TS) protein expression and enhances 5-fluorouracil (FU) induced apoptosis in gastric cancer cells. This novel approach targets E2F-1 to improve FU sensitivity.
Area of Science:
- Oncology
- Molecular Biology
- Pharmacology
Background:
- Thymidylate synthase (TS) is a key enzyme in DNA synthesis and a target for cancer chemotherapy.
- 5-fluorouracil (FU) is a widely used chemotherapeutic agent that inhibits TS.
- Gastric cancer remains a significant health challenge with a need for improved therapeutic strategies.
Purpose of the Study:
- To investigate the effect of UCN-01, a protein kinase C/cyclin-dependent kinase inhibitor, on TS expression and its potential to enhance FU efficacy in human gastric cancer cells.
- To elucidate the molecular mechanisms underlying UCN-01's action on TS, including its effect on E2F-1.
Main Methods:
- Human gastric cancer cell line SK-GT5 was treated with UCN-01 and other kinase inhibitors.
- TS protein and mRNA levels were assessed using Western blot and Northern blot analyses.
- The expression of E2F-1 protein and mRNA was also evaluated.
- Apoptosis induction was measured in cells treated with FU alone, UCN-01 alone, or in combination.
Main Results:
- UCN-01 suppressed TS protein expression in a dose-dependent manner in SK-GT5 cells.
- Other kinase inhibitors showed a similar but less potent effect on TS.
- UCN-01 significantly repressed the induction of TS by FU and enhanced FU-induced apoptosis when administered after FU.
- UCN-01's effect on TS was linked to dose-dependent suppression of E2F-1 protein, a TS transcriptional activator, while E2F-1 mRNA levels remained largely unchanged.
Conclusions:
- UCN-01 effectively suppresses TS protein expression in gastric cancer cells.
- UCN-01 enhances the anti-cancer effects of FU by down-regulating E2F-1 and subsequently TS expression.
- UCN-01 represents a potential novel therapeutic strategy to improve cellular sensitivity to FU in gastric cancer treatment.