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Research Articles: Therapeutics, Targets, and Development

Chk1 and Chk2 are differentially involved in homologous recombination repair and cell cycle arrest in response to DNA double-strand breaks induced by camptothecins

Divisions of ¹ Anti-tumor Pharmacology and ² Chemistry, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, Peoples' Republic of China

Requests for reprints: Jian Ding, Division of Anti-tumor Pharmacology, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Zhangjiang Hi-Tech Park, Shanghai 201203, Peoples' Republic of China. Phone: 86-21-50806722; Fax: 86-21-50806722. E-mail: jding{at}mail.shcnc.ac.cn

Abstract

Camptothecins (CPT) activate S or G₂-M arrest and the homologous recombination (HR) repair pathway in tumor cells. In this process, both checkpoint kinases 1 and 2 (Chk1 and Chk2, respectively) are activated, but their differential roles, especially in the coordination of checkpoint and repair control, and potential clinic relevance remain to be fully elucidated. In this study, the repairable double-strand breaks were induced in human colon cancer HCT116 cells by 1-h exposure to 25 or 100 nmol/L CPT and its novel derivative chimmitecan. The cellular disposal of double-strand breaks was reflected as the progressive dispersal of -H2AX foci, reduction of "comet" tails, dynamic activation of RAD51-mediated HR repair, and reversible G₂-M arrest. In this model, the differential kinetics of Chk1 and Chk2 activation was characterized by the progressively increased phosphorylation of Chk2 until 72 h, the degradation of Chk1, and the disappearance of phosphorylated Chk1 48 h after drug removal. Using RNA interference, we further showed that Chk2 was essential to G₂-M arrest, whereas Chk1 was mainly required for HR repair in CPT-treated HCT116 cells. Moreover, Chk2, rather than Chk1, predominated over the control of cell survival in this model. The differential roles of Chk1 and Chk2 in regulating HR repair and G₂-M phase arrest were also confirmed in HT-29 colon cancer cells. Together, these findings systematically dissect the differential roles of Chk1 and Chk2 in a favorable model pursuing CPT-driven DNA damage responses, providing critical evidence to further explore checkpoint modulation, especially Chk2 inhibition as a therapeutic strategy in combination with CPT. [Mol Cancer Ther 2008;7(6):1440–9]

Grant support: National Natural Science Foundation of China grant 30772588.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

³ Supplementary material for this article is available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/).

Received 9/19/07; revised 3/13/08; accepted 3/15/08.