Abstract
Recent advances in cell cycle regulation have led to a suggestion of therapeutically targeting cell cycle checkpoint pathways in cancer cells to increase the toxicity of DNA-damaging agents. In this study, we investigate whether knockdowns of checkpoint kinases Chk1 and Chk2 by RNA interfering potentiate the cytotoxicity and abrogate G(2)/M checkpoint induced by DNA-damaging agent lidamycin (LDM) in HCT116 cells with different p53 status. Our results showed that Chk1 knockdown enhanced the cytotoxicity of LDM through abrogating G(2)/M arrest and increasing apoptosis to a greater extent in HCT116 p53(-/-) cells than in p53(wt) cells. Abrogation of LDM-induced G(2)/M arrest by Chk1 knockdown was associated with reducing the inactivated phosphorylations of Cdc25C and Cdc2. LDM-induced gamma-H2AX was increased in cells with Chk1 knockdown, indicating that DNA double-strand breaks (DSBs) were enhanced. Furthermore, knockdown of Chk1 also increased LDM-mediated apoptotic cell death in p53 knockout cells with activation of caspase-2 and caspase-3. On the contrary, knockdown of Chk2 had no impact on G(2)/M arrest or apoptosis induced by LDM. Moreover, dual knockdown of Chk1 and Chk2 failed to achieve better efficacy than Chk1 alone. Taken together, we suggest that Chk1 is a potential therapeutic target to sensitize human p53 deficient cancer cells to LDM.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Aminoglycosides / pharmacology*
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Antibiotics, Antineoplastic / pharmacology*
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Apoptosis / genetics
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Blotting, Western
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Caspase 2 / metabolism
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Caspase 3 / metabolism
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / metabolism
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Cell Division / drug effects
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Cell Division / genetics
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Cell Growth Processes / drug effects
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Cell Growth Processes / genetics
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Checkpoint Kinase 1
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Checkpoint Kinase 2
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Colonic Neoplasms / drug therapy*
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Colonic Neoplasms / enzymology*
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Colonic Neoplasms / genetics
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Colonic Neoplasms / pathology
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Cysteine Endopeptidases / metabolism
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Enediynes / pharmacology*
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G2 Phase / drug effects
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G2 Phase / genetics
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Gene Knockdown Techniques
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HCT116 Cells
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Humans
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Phosphorylation
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Protein Kinase Inhibitors / pharmacology
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Protein Kinases / deficiency*
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Protein Kinases / genetics
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Protein Kinases / metabolism
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Protein Serine-Threonine Kinases / deficiency
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / metabolism
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RNA, Small Interfering / genetics
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Transfection
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Tumor Suppressor Protein p53 / metabolism*
Substances
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Aminoglycosides
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Antibiotics, Antineoplastic
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Cell Cycle Proteins
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Enediynes
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Protein Kinase Inhibitors
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RNA, Small Interfering
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TP53 protein, human
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Tumor Suppressor Protein p53
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C 1027
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Protein Kinases
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Checkpoint Kinase 2
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CHEK1 protein, human
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CHEK2 protein, human
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Checkpoint Kinase 1
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Protein Serine-Threonine Kinases
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CASP2 protein, human
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Caspase 2
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Caspase 3
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Cysteine Endopeptidases