2-methoxyestradiol, an endogenous mammalian metabolite, radiosensitizes colon carcinoma cells through c-Jun NH2-terminal kinase activation.
OBJECTIVE
2-Methoxyestradiol (2ME), an estrogen metabolite, induces apoptosis in various cell types. We investigated whether 2ME pretreatment can radiosensitize colon adenocarcinoma cells.


METHODS
Radiosensitizing effects of 2ME were evaluated by cell death, clonogenic assay, nuclear fragmentation, and tumor progression of xenografts. Ionizing radiation-induced DNA damage was evaluated by histone H2AX phosphorylation and its foci. The c-Jun NH2-terminal kinase (JNK) activation was evaluated by anti-phosphorylated JNK antibody and inhibited by the JNK-specific inhibitor SP600125 or dominant-negative SEK1 expression.


RESULTS
Clonogenic assays revealed that 2ME, but not estradiol, radiosensitized three colon carcinoma cells, DLD-1, HCT-8, and HCT-15, and strongly suppressed tumor progression of DLD-1 xenografts. Gene transfer-mediated Bcl-xL overexpression largely abolished both augmented apoptosis and reduced survival fractions. Pretreatment with 2ME enhanced H2AX phosphorylation, its foci, and phosphorylation of ATM kinase and delayed re-entry of cell cycle progression after ionizing radiation. Augmentation of both radiosensitivity and H2AX phosphorylation was substantially reduced by SP600125 or overexpression of a dominant-negative mutant SEK1.


CONCLUSIONS
2ME radiosensitized colon carcinoma cells through enhanced DNA damage via JNK activation, thereby representing a novel radiosensitizing therapy against colon cancer.
Zou
HuiChao
H
Division of Molecular Oncology and Molecular Diagnosis, Graduate School of Medicine, First Department of Internal Medicine, Sapporo Medical University, Sapporo, Japan.
Adachi
Masaaki
M
Imai
Kohzoh
K
Hareyama
Masato
M
Yoshioka
Katsuji
K
Zhao
Shiguang
S
Shinomura
Yasuhisa
Y
D000077584:2-Methoxyestradiol; D000230:Adenocarcinoma / Q000628:therapy*; D000818:Animals; D017209:Apoptosis / Q000502:physiology; D015153:Blotting, Western; D002453:Cell Cycle / Q000528:radiation effects; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000628:therapy*; D003131:Combined Modality Therapy; D004249:DNA Damage / Q000528:radiation effects; D004958:Estradiol / Q000031:analogs & derivatives* / Q000378:metabolism / Q000493:pharmacokinetics; D006657:Histones / Q000378:metabolism; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D008819:Mice, Nude; D018613:Microscopy, Confocal; D010766:Phosphorylation; D011839:Radiation, Ionizing; D011838:Radiation-Sensitizing Agents / Q000493:pharmacokinetics*; D014162:Transfection; D023041:Xenograft Model Antitumor Assays
C468783:H2AX protein, human; D006657:Histones; D011838:Radiation-Sensitizing Agents; D004958:Estradiol; D000077584:2-Methoxyestradiol; D048031:JNK Mitogen-Activated Protein Kinases