Enhancement of p53-mutant human colorectal cancer cells radiosensitivity by flavonoid fisetin.
OBJECTIVE
The aim of this study was to investigate whether fisetin is a potential radiosensitizer for human colorectal cancer cells, which are relatively resistant to radiotherapy.


METHODS
Cell survival was examined by clonogenic survival assay, and DNA fragmentation was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay. The effects of treatments on cell cycle distribution and apoptosis were examined by flow cytometry. Western blot analysis was performed to ascertain the protein levels of gamma-H2AX, phospho-Chk2, active caspase-3, PARP cleavage, phospho-p38, phospho-AKT, and phospho-ERK1/2.


RESULTS
Fisetin pretreatment enhanced the radiosensitivity of p53-mutant HT-29 human colorectal cancer cells but not human keratocyte HaCaT cells; it also prolonged radiation-induced G(2)/M arrest, enhanced radiation-induced cell growth arrest in HT-29 cells, and suppressed radiation-induced phospho-H2AX (Ser-139) and phospho-Chk2 (Thr-68) in p53-mutant HT-29 cells. Pretreatment with fisetin enhanced radiation-induced caspase-dependent apoptosis in HT-29 cells. Fisetin pretreatment augmented radiation-induced phosphorylation of p38 mitogen-activated protein kinase, which is involved in caspase-mediated apoptosis, and SB202190 significantly reduced apoptosis and radiosensitivity in fisetin-pretreated HT-29 cells. By contrast, both phospho-AKT and phospho-ERK1/2, which are involved in cell proliferation and antiapoptotic pathways, were suppressed after irradiation combined with fisetin pretreatment.


CONCLUSIONS
To our knowledge, this study is the first to provide evidence that fisetin exerts a radiosensitizing effect in p53-mutant HT-29 cells. Fisetin could potentially be developed as a novel radiosensitizer against radioresistant human cancer cells.
Chen
Wen-Shu
WS
Department of Life Science, Tzu Chi University, Hualien, Taiwan.
Lee
Yi-Jang
YJ
Yu
Yi-Chu
YC
Hsaio
Ching-Hui
CH
Yen
Jui-Hung
JH
Yu
Sz-Hsien
SH
Tsai
Yu-Jia
YJ
Chiu
Shu-Jun
SJ
D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects; D002470:Cell Survival; D064447:Checkpoint Kinase 2; D015179:Colorectal Neoplasms / Q000235:genetics* / Q000378:metabolism; D053938:DNA Fragmentation; D005419:Flavonoids / Q000494:pharmacology*; D044948:Flavonols; D016158:Genes, p53; D045325:HCT116 Cells / Q000187:drug effects / Q000528:radiation effects; D019073:HT29 Cells / Q000187:drug effects / Q000528:radiation effects; D006657:Histones / Q000378:metabolism; D006801:Humans; D020287:In Situ Nick-End Labeling / Q000379:methods; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D011836:Radiation Tolerance / Q000187:drug effects* / Q000235:genetics; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D014410:Tumor Stem Cell Assay
D005419:Flavonoids; D044948:Flavonols; C468783:H2AX protein, human; D006657:Histones; D011838:Radiation-Sensitizing Agents; D011065:Poly(ADP-ribose) Polymerases; D064447:Checkpoint Kinase 2; C578634:CHEK2 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D020928:Mitogen-Activated Protein Kinases; D053148:Caspase 3; C017875:fisetin