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

Abstract

Camptothecins (CPT) activate S or G 2 -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 G 2 -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 G 2 -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 G 2 -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.

Trifluridine Induces p53-Dependent Sustained G2 Phase Arrest with Its Massive Misincorporation into DNA and Few DNA Strand Breaks

“…HCT-116 cells were cultured in the presence of FdUrd or FTD at IC 50 concentrations for 24 and 72 hours, and ssDNA and dsDNA breaks were evaluated by alkaline-and neutral-comet assays. As reported previously (28,29), a significant increase in the mean score of the tail moment was observed in alkaline-and neutral-comet assays after camptothecin treatment ( Supplementary Fig. S7).…”

Activation of ATM/Chk1 by curcumin causes cell cycle arrest and apoptosis in human pancreatic cancer cells

“…Curcumin treatment modulates expression of G2/M cell cycle regulatory proteins DNA damage generally leads to the activation of the ATM/ATR pathway (Molinari, 2000;Abraham, 2001;Lukas et al, 2001;Huang et al, 2008). To further delineate the molecular mechanism of curcumin-mediated G2/M arrest, we determine its effect on the key-signaling proteins of this pathway.…”

Eupatilin: A flavonoid compound isolated from the artemisia plant, induces apoptosis and G2/M phase cell cycle arrest in human melanoma A375 cells

“…This study also demonstrated that Eupatilin arrests the cell cycle as a result of the DNA damage and indicated that Eupatilin induced growth inhibition is likely to involve the modulation of cell cycle progression in A375 cells. During DNA damage, cells are blocked in G2/M phase to provide time to repair damaged DNA (Molinari 2000;Abraham, 2001), or lead to apoptotic cell death in case of severe DNA damage (Huang et al, 2008).…”