Article

Structure and Activation Mechanism of the CHK2 DNA Damage Checkpoint Kinase

Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
Molecular cell (Impact Factor: 14.02). 09/2009; 35(6):818-29. DOI: 10.1016/j.molcel.2009.09.007
Source: PubMed

ABSTRACT

The CHK2 protein kinase is an important transducer of DNA damage checkpoint signals, and its mutation contributes to hereditary and sporadic cancer. CHK2 activation is triggered by the phosphorylation of Thr68 by the DNA damage-activated ATM kinase. This leads to transient CHK2 dimerization, in part through intermolecular phosphoThr68-FHA domain interactions. Dimerization promotes kinase activation through activation-loop autophosphorylation, but the mechanism of this process has not been clear. The dimeric crystal structure of CHK2, described here, in conjunction with biochemical and mutational data reveals that productive CHK2 dimerization additionally involves intermolecular FHA-kinase domain and FHA-FHA interactions. Ile157, mutated in the Li-Fraumeni cancer-predisposition syndrome, plays a central role in the FHA-kinase domain interface, explaining the lack of dimerization and autophosphorylation of this mutant. In the dimer, the kinase active sites face each other in close proximity, indicating that dimerization may also serve to optimally position the kinase active sites for efficient activation loop transphosphorylation.

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    • "Phosphorylation of Chk1 leads to checkpoint activation and cell cycle arrest at the G1/S, intra S or G2/M phases. Despite their similarity in name, Chk1 and Chk2 differ substantially in the structure of their kinase pocket [18, 19] and in their cellular function with Chk1 suggested to be the major component responsible for responses to DNA damage. Inhibiting Chk1 following genotoxic stress (such as that induced by cytotoxic chemotherapy) results in checkpoint abrogation, inhibition of DNA repair and induction of cell death in cells with a defective p53 response [20, 21]. "
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    ABSTRACT: Background Chk1 inhibitors are currently in clinical trials as putative potentiators of cytotoxic chemotherapy drugs. Chk1 inhibitors may exhibit single agent anti-tumor activity in cancers with underlying DNA repair, DNA damage response or DNA replication defects. Methods Here we describe the cellular effects of the pharmacological inhibition of the checkpoint kinase Chk1 by the novel inhibitor V158411 in triple-negative breast cancer and ovarian cancer. Cytotoxicity, the effect on DNA damage response and cell cycle along with the ability to potentiate gemcitabine and cisplatin cytotoxicity in cultured cells was investigated. Western blotting of proteins involved in DNA repair, checkpoint activation, cell cycle and apoptosis was used to identify potential predictive biomarkers of Chk1 inhibitor sensitivity. Results The Chk1 inhibitors V158411, PF-477736 and AZD7762 potently inhibited the proliferation of triple-negative breast cancer cells as well as ovarian cancer cells, and these cell lines were sensitive compared to ER positive breast and other solid cancer cells lines. Inhibition of Chk1 in these sensitive cell lines induced DNA damage and caspase-3/7 dependent apoptosis. Western blot profiling identified pChk1 (S296) as a predictive biomarker of Chk1 inhibitor sensitivity in ovarian and triple-negative breast cancer and pH2AX (S139) in luminal breast cancer. Conclusions This finding suggests that Chk1 inhibitors either as single agents or in combination chemotherapy represents a viable therapeutic option for the treatment of triple-negative breast cancer. pChk1 (S296) tumor expression levels could serve as a useful biomarker to stratify patients who might benefit from Chk1 inhibitor therapy.
    Full-text · Article · Aug 2014 · BMC Cancer
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    • "The DDR can be activated by a range of endogenous and external insults including therapies currently used for the treatment of cancer such as ionizing radiation and cytotoxic chemotherapeutic agents such as gemcitabine, irinotecan and cisplatin [2,3]. Despite their similarity in name, Chk1 and Chk2 differ substantially in the structure of their kinase pocket [4,5] and in their cellular function with Chk1 suggested to be the major component responsible for responses to DNA damage [3,6,7]. Inhibiting Chk1 following genotoxic stress (such as that induced by cytotoxic chemotherapy) results in checkpoint abrogation, inhibition of DNA repair and induction of cell death in cells with a defective p53 response [8,9]. "
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    ABSTRACT: Background Chk1 forms a core component of the DNA damage response and small molecule inhibitors are currently being investigated in the clinic as cytotoxic chemotherapy potentiators. Recent evidence suggests that Chk1 inhibitors may demonstrate significant single agent activity in tumors with specific DNA repair defects, a constitutively activated DNA damage response or oncogene induced replicative stress. Methods Growth inhibition induced by the small molecule Chk1 inhibitor V158411 was assessed in a panel of human leukemia and lymphoma cell lines and compared to cancer cell lines derived from solid tumors. The effects on cell cycle and DNA damage response markers were further evaluated. Results Leukemia and lymphoma cell lines were identified as particularly sensitive to the Chk1 inhibitor V158411 (mean GI50 0.17 μM) compared to colon (2.8 μM) or lung (6.9 μM) cancer cell lines. Chk1 inhibition by V158411 in the leukemia and lymphoma cell lines induced DNA fragmentation and cell death that was both caspase dependent and independent, and prevented cells undergoing mitosis. An analysis of in vitro pharmacodynamic markers identified a dose dependent decrease in Chk1 and cyclin B1 protein levels and Cdc2 Thr15 phosphorylation along with a concomitant increase in H2AX phosphorylation at Ser139 following V158411 treatment. Conclusions These data support the further evaluation of Chk1 inhibitors in hematopoietic cancers as single agents as well as in combination with standard of care cytotoxic drugs.
    Full-text · Article · Jun 2014 · Molecular Cancer
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    • "Moreover, Chk2 induces transautophosphorylation of residues Thr383 and Thr387 and then cis-phosphorylation of residue Ser516 [5–8, 10–14]. After that, Chk2 will phosphorylate several downstream substrates, such as BRCA1 (breast cancer 1, early onset [15] [16]), Cdc25A (cell division cycle 25 homolog A), Cdc25C, and p53 [7] [8] [10]. Several researches indicated that Chk2 phosphorylates Cdc25A which is considered an oncogene on the residue Ser123 in S phase of cell cycle, and it also phosphorylates Cdc25C on the residue Ser216 in G2 phase helping prevent mitotic entry in cells with damaged DNA [5]. "
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    ABSTRACT: Checkpoint kinase 2 (Chk2) has a great effect on DNA-damage and plays an important role in response to DNA double-strand breaks and related lesions. In this study, we will concentrate on Chk2 and the purpose is to find the potential inhibitors by the pharmacophore hypotheses (PhModels), combinatorial fusion, and virtual screening techniques. Applying combinatorial fusion into PhModels and virtual screening techniques is a novel design strategy for drug design. We used combinatorial fusion to analyze the prediction results and then obtained the best correlation coefficient of the testing set (r test) with the value 0.816 by combining the BesttrainBesttest and FasttrainFasttest prediction results. The potential inhibitors were selected from NCI database by screening according to BesttrainBesttest + FasttrainFasttest prediction results and molecular docking with CDOCKER docking program. Finally, the selected compounds have high interaction energy between a ligand and a receptor. Through these approaches, 23 potential inhibitors for Chk2 are retrieved for further study.
    Full-text · Article · Apr 2014 · BioMed Research International
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