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. 2013 Mar;1(2):373-378.
doi: 10.3892/mco.2012.53. Epub 2012 Dec 18.

S-adenosyl Methionine Specifically Protects the Anticancer Effect of 5-FU via DNMTs Expression in Human A549 Lung Cancer Cells

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Free PMC article

S-adenosyl Methionine Specifically Protects the Anticancer Effect of 5-FU via DNMTs Expression in Human A549 Lung Cancer Cells

Myeong-Sun Ham et al. Mol Clin Oncol. .
Free PMC article

Abstract

Cellular methylation is associated with stabilization of the chromatin structure. S-adenosyl methionine (SAM), a metabolite of methionine metabolism, is the methyl donor of essential cellular methyltransferase reactions. Using 3-(4,5-dimethylthiazol-2-yl)-2,5-dephenyl tetrazolium bromide (MTT) assay, we found that combination treatment of SAM and 5-fluorouracil (5-FU) specifically protected the anticancer effect of 5-FU, whereas the combination of SAM and cisplatin had no effect. This result was confirmed by FACS analysis. The combination treatment of SAM and 5-FU significantly decreased the dead cell population, while the G1 cell population was slightly increased, suggesting that protection of SAM is not associated with the cell cycle arrest of DNA-damaging drugs. We also analyzed which cellular methylation-related proteins were involved in the protective effect. Results showed the expression of DNA methyltransferases (DNMTs) was decreased with 5-FU alone but was increased with the combination treatment of SAM and 5-FU, suggesting that SAM protects the anticancer effect of 5-FU by regulating the expression of DNMTs. Taken together, the results indicated that SAM specifically modulates the anti-cancer effect of the DNA damage agent 5-FU and this may be modulated by aberrant DNA methylation.

Keywords: 5-fluorouracil; DNA methyltransferases; S-adenosyl methionine; cisplatin; protection.

Figures

Figure 1
Figure 1
Effect on the cell growth of S-adenosyl methionine (SAM), 5-fluorouracil (5-FU) and cisplatin. Cells (1,000 cells/well) were seeded and treated with various concentrations of the indicated drugs for 2 days and this was followed by 3-(4,5-dimethylthiazol-2-yl)-2,5-dephenyl tetrazolium bromide (MTT) assay. SAM exhibit no specific cytotoxic effect in A549 cells. However, 5-FU and cisplatin significantly reduced cell growth. Data are shown as the mean ± standard deviation of triplicate assays. *P<0.05, compared to the control group.
Figure 2
Figure 2
Combination effect of S-adenosyl methionine (SAM) and 5-fluorouracil (5-FU) or SAM and cisplatin. Cells were seeded in 96-well plates, and treated using the indicated drugs for 4 days. Combined treatment of SAM and 5-FU showed a little protection of the 5-FU anticancer effect. This phenomenon was not detected in the combination treatment of SAM and cisplatin. Data are shown as the mean ± standard deviation of triplicate assays. *P<0.05, compared to the control group.
Figure 3
Figure 3
FACS analysis in the combination of 5-fluorouracil (5-FU) or cisplatin with S-adenosyl methionine (SAM). Cells (5×105 cells/well) were seeded and treated with the indicated drugs for 24 h and fixed with ethanol. Cells were stained with propidium iodide and analyzed with FACScan. 5-FU treatment induced cell death without any relation to a specific cell cycle, but the combined treatment of SAM and 5-FU significantly decreased the dead cell population. The combined treatment of SAM and cisplatin has shown as muh G2/M arrest as with cisplatin alone. *P<0.05, compared to the G0 population treated with 5-FU alone.
Figure 4
Figure 4
Expression analyses of methylation related proteins in combination of 5-fluorouracil (5-FU) or cisplatin with S-adenosyl methionine (SAM). (A) 5-FU decreased induced DNMT1 and DNMT3A proteins but not in cisplatin. However, combination treatment of SAM and 5-FU restored DNA methyltransferases (DNMTs) protein level. (B) 5-FU treatment decreased most DNMTs expression at the RNA level. SAM restored the effect 5-FU on DNMTs expression.

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