Decreased expression of hMLH1 correlates with reduced 5-fluorouracil-mediated apoptosis in colon cancer cells

  • Authors:
    • Hideyuki Fujita
    • Jun Kato
    • Joichiro Horii
    • Keita Harada
    • Sakiko Hiraoka
    • Hidenori Shiraha
    • Kohsaku Sakaguchi
    • Yasushi Shiratori
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  • Published online on: November 1, 2007     https://doi.org/10.3892/or.18.5.1129
  • Pages: 1129-1137
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Abstract

Patients with sporadic microsatellite instable colorectal cancers, in most of which the function of the hMLH1 mismatch repair gene is impaired, do not gain a survival benefit from 5-fluorouracil (5-FU)-based chemotherapy. However, the effect of hMLH1 on the cytotoxicity induced by 5-FU has not yet been sufficiently confirmed. In this study, we assessed the effect of hMLH1 on cytotoxicity and apoptosis induced by 5-FU using newly developed cell lines. We constructed two cell lines: SW480 (originally hMLH1-proficient), in which the expression of hMLH1 was reduced using a small interfering RNA (siRNA) technique, and HCT116 (originally hMLH1-deficient), in which the expression of hMLH1 can be regulated by doxycycline. Using these cell lines, a clonogenic survival assay, 4',6-diamidino-2-phenylindole (DAPI) staining and an Annexin-V assay were performed. Moreover, the incorporation of 5-FU into DNA was determined using tritium-labeled 5-FU. In both of our two cell lines, hMLH1-deficient cells exhibited approximately 2.4-fold clonal surviving fraction compared to hMLH1-proficient cells for 10 days after the administration of 5-FU. Additionally, hMLH1-deficient cells treated with 5-FU exhibited 34-45% less apoptosis than hMLH1-proficient cells according to the results of DAPI staining and Annexin-V assay. Furthermore, hMLH1-deficient cells treated with 5-FU exhibited an approximately 2-fold greater incorporation of 5-FU into DNA than control cells, suggesting that the recognition of 5-FU-incorporated DNA is impaired in hMLH1-deficient cells, resulting in reduced apoptosis. Our conclusions were that decreased expression of hMLH1 in colon cancer cells reduced the apoptosis induced by 5-FU, suggesting that hMLH1 is a key determinant of 5-FU chemosensitivity.

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November 2007
Volume 18 Issue 5

Print ISSN: 1021-335X
Online ISSN:1791-2431

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Spandidos Publications style
Fujita H, Kato J, Horii J, Harada K, Hiraoka S, Shiraha H, Sakaguchi K and Shiratori Y: Decreased expression of hMLH1 correlates with reduced 5-fluorouracil-mediated apoptosis in colon cancer cells. Oncol Rep 18: 1129-1137, 2007.
APA
Fujita, H., Kato, J., Horii, J., Harada, K., Hiraoka, S., Shiraha, H. ... Shiratori, Y. (2007). Decreased expression of hMLH1 correlates with reduced 5-fluorouracil-mediated apoptosis in colon cancer cells. Oncology Reports, 18, 1129-1137. https://doi.org/10.3892/or.18.5.1129
MLA
Fujita, H., Kato, J., Horii, J., Harada, K., Hiraoka, S., Shiraha, H., Sakaguchi, K., Shiratori, Y."Decreased expression of hMLH1 correlates with reduced 5-fluorouracil-mediated apoptosis in colon cancer cells". Oncology Reports 18.5 (2007): 1129-1137.
Chicago
Fujita, H., Kato, J., Horii, J., Harada, K., Hiraoka, S., Shiraha, H., Sakaguchi, K., Shiratori, Y."Decreased expression of hMLH1 correlates with reduced 5-fluorouracil-mediated apoptosis in colon cancer cells". Oncology Reports 18, no. 5 (2007): 1129-1137. https://doi.org/10.3892/or.18.5.1129