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Research Paper

DNA damage signaling is activated during cancer progression in human colorectal carcinoma

Kazuhito Oka, Toshiki Tanaka, Tadahiko Enoki, Koichi Yoshimura, Mako Ohshima, Masayuki Kubo, Tomoyuki Murakami, Toshikazu Gondou, Yoshihide Minami, Yoshihiro Takemoto, Eijirou Harada, Takaaki Tsushimi, Tao-Sheng Li, Frank Traganos, Zbigniew Darzynkiewicz and Kimikazu Hamano

Purpose: Recent studies have shown that the DNA damage response (DDR) is activated in precancerous lesions, suggesting that neoplastic cells may avoid apoptosis by impairing the DDR which acts as a barrier against tumor progression. To define the role of the DDR pathway in human colorectal carcinoma, we investigated the level of phosphorylated proteins of the DDR pathway.

Experimental Design: Colorectal tissue samples were obtained at the time of surgery, from 55 patients at two hospitals. The tissues were classified into four groups according to pathology: normal mucosa, adenoma, early carcinoma and advanced carcinoma. We evaluated phosphorylated ataxia telangiectasia mutated (pATM), phosphorylated H2AX (γH2AX) and Chk2 (pChk2) protein levels by immunohistochemistry and Western blot analysis. We also evaluated apoptosis by the TUNEL assay.

Results: Immunostaining for pATM, γH2AX and pChk2 revealed that all were significantly expressed during tumor progression in advanced carcinoma (vs. normal tissue for pATM [p<0.05]; vs. normal and adenoma for γH2AX [p<0.05]; and vs. normal tissue for pChk2 [p<0.05). Western blot analysis of γH2AX and pChk2 revealed that their level increased gradually during tumor progression and was maximal in advanced carcinoma (vs. normal tissue; p<0.05). No apoptotic cells were found in any tissue sample.

Conclusions: The DDR pathway was activated during cancer progression, but no apoptosis was detected, even among the cells with activated DDR. It is likely that activation of DDR was induced by stress signaling as a consequence of oxidative, replication and mechanical stresses occurring during growth and expansion of the colorectal cancer.

Authors

Kazuhito Oka

Department of Surgery and Clinical Science Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan

Toshiki Tanaka Corresponding author: toshik@yamaguchi-u.ac.jp

Department of Surgery and Clinical Science Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan

Tadahiko Enoki

Department of Surgery and Clinical Science Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan

Koichi Yoshimura

Department of Surgery and Clinical Science Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan

Mako Ohshima

Department of Surgery and Clinical Science Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan

Masayuki Kubo

Department of Pathology, Kanmon Medical Center, Shimonoseki, Japan

Tomoyuki Murakami

Department of Pathology, Kanmon Medical Center, Shimonoseki, Japan

Toshikazu Gondou

Department of Pathology, Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan

Yoshihide Minami

Department of Surgery, Saiseikai Shimonoseki General Hospital, Shimonoseki, Japan

Yoshihiro Takemoto

Department of Surgery and Clinical Science Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan

Eijirou Harada

Department of Surgery and Clinical Science Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan

Takaaki Tsushimi

Department of Surgery and Clinical Science Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan

Tao-Sheng Li

Department of Surgery and Clinical Science Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan

Frank Traganos

Brander Cancer Research Institute, Department of Pathology, New York Medical College, NY, USA

Zbigniew Darzynkiewicz

Brander Cancer Research Institute, Department of Pathology, New York Medical College, NY, USA

Kimikazu Hamano

Department of Surgery and Clinical Science Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan