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Zebularine suppresses the apoptotic potential of 5-fluorouracil via cAMP/PKA/CREB pathway against human oral squamous cell carcinoma cells

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Abstract

Purpose

During tumorigenesis, tumor suppressor and tumor-related genes are commonly silenced by aberrant DNA methylation in their promoter regions, which is one of the important determinants of susceptibility to 5-fluorouracil (5-FU) in oral squamous cell carcinoma (OSCC) cells. Here, we examine the chemotherapeutic efficacy of epigenetic agents on 5-FU cytotoxicity.

Method

We investigated the effect of a DNA methyltransferase (DNMT) inhibitor, zebularine (Zeb), on the chemosensitivity of 5-FU and cisplatin (CDDP) by MTT and TUNEL methods, and compared the molecular mechanism of action with those of a GSK3β inhibitor, LiCl, and an Hsp90 inhibitor, 17-AAG.

Results

A significant apoptotic effect by a combination of Zeb or 17-AAG was found in CDDP treatment; however, considerable suppression of 5-FU-induced apoptosis was observed after incubation with Zeb, 17-AAG, or LiCl. Zeb’s suppressive effects were associated with activation of the cAMP/PKA/CREB pathway, differing from mechanisms of 17-AAG and LiCl. Suppression of 5-FU-induced apoptosis by Zeb was not associated with increased Bcl-2 and Bcl-xL expressions dependent on transcription factor CREB, and with the expression level of thymidylate synthase.

Conclusions

In the present study, we identified a more detailed mechanism of action by which Zeb suppresses 5-FU-induced apoptosis. These results indicate that combination therapies have to be carefully investigated due to potential harmful effects in the clinical application of DNMT inhibitors.

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Acknowledgments

We thank Mr. D. Mrozek for editing the manuscript. This work was supported in part by Grant-in-Aids for Scientific Research (20659309) from the Japan Society for the Promotion of Science and (19791480 and 19791482) from the Ministry of Education, Culture, Sports, Science, and Technology, Japan.

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Authors and Affiliations

  1. Department of Microbiology and Immunology, Tohoku University Graduate School of Dentistry, 4-1 Seiryo-machi, Aoba-ku, Sendai, 980-8575, Japan

    Maiko Suzuki, Manabu Endo & Hidemi Rikiishi

  2. Department of Oral Surgery, Tohoku University Graduate School of Dentistry, Sendai, 980-8575, Japan

    Fumiaki Shinohara, Manabu Endo, Masaki Sugazaki & Seishi Echigo

Authors
  1. Maiko Suzuki
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  2. Fumiaki Shinohara
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  3. Manabu Endo
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  4. Masaki Sugazaki
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  5. Seishi Echigo
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  6. Hidemi Rikiishi
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Corresponding author

Correspondence to Hidemi Rikiishi.

Electronic supplementary material

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280_2008_833_MOESM1_ESM.tif

Fig. S1 Effects of death ligands on cell growth. HSC-3 and Jurkat cells (as a positive control) were treated with increasing concentrations of a cross-linking antibody to Fas receptor, CH11 (A) or recombinant human TRAIL (B) for 48 h, followed by MTT assay. Results represent the percentages (means ± SE) of cell growth to the untreated control (TIF 1077 kb)

280_2008_833_MOESM2_ESM.tif

Fig. S2 p-CREB activity induced by Zeb/5-FU combination. HSC-3 cells pretreated with Zeb (220 μM) for 48 h, were treated with or without 5-FU (250 μg/ml) for 4 h. Nuclear proteins were extracted, and each sample (5 μg/well) was assayed by ELISA. Results show representative OD450 values of triplicate experiments(TIF 1047 kb)

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Suzuki, M., Shinohara, F., Endo, M. et al. Zebularine suppresses the apoptotic potential of 5-fluorouracil via cAMP/PKA/CREB pathway against human oral squamous cell carcinoma cells. Cancer Chemother Pharmacol 64, 223–232 (2009). https://doi.org/10.1007/s00280-008-0833-4

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  • DOI: https://doi.org/10.1007/s00280-008-0833-4

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