Abrogation of Chk1-mediated S/G2 checkpoint by UCN-01 enhances ara-C-induced cytotoxicity in human colon cancer cells.
OBJECTIVE
To investigate whether 7-hydroxystaurosporine (UCN-01) affects cell cycle progression in arabinosylcytosine (ara-C) treated human colon carcinoma HT-29 cells.


METHODS
Cytotoxicity, DNA synthesis, cell cycle distribution, protein level, and kinase activity were determined by clonogenic assay, flow cytometry, DNA synthesis assay, immunoblotting, and kinase assays, respectively.


RESULTS
UCN-01 abrogated an S/G2-phase checkpoint in HT-29 cells treated with ara-C. When UCN-01 was added after treatment with ara-C, the rate of recovery of DNA synthesis was enhanced and colony-forming ability diminished. Thus, premature recovery of DNA synthesis was associated with increased cytotoxicity. Measurements of cyclin A and B protein levels, Cdk2 and Cdc2 kinase activities, Cdc25C phosphorylation, and Chk1 kinase activity were consistent with UCN-01-induced abrogation of the S/G2-phase checkpoint in ara-C treated cells.


CONCLUSIONS
The abrogation of the S/G2 checkpoint may be due to inhibition of Chk1 kinase by UCN-01. The enhanced cytotoxicity produced when UCN-01 was combined with ara-C suggested a rationale for the use of this drug combination for tumors that might be susceptible to cell cycle checkpoint abrogation.
Shao
Rong-Guang
RG
Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences, Peking Union Medical College, Beijing 100050, China. shaor@public3.bta.net.cn
Cao
Chun-Xia
CX
Pommier
Yves
Y
D000970:Antineoplastic Agents / Q000494:pharmacology; D042846:CDC2-CDC28 Kinases / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism; D000071877:Checkpoint Kinase 1; D019925:Cyclin A / Q000378:metabolism; D019926:Cyclin B / Q000378:metabolism; D051357:Cyclin-Dependent Kinase 2; D003561:Cytarabine / Q000494:pharmacology*; D004273:DNA, Neoplasm / Q000096:biosynthesis; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016195:G2 Phase / Q000187:drug effects*; D019073:HT29 Cells; D006801:Humans; D011494:Protein Kinases / Q000378:metabolism*; D016196:S Phase / Q000187:drug effects*; D019311:Staurosporine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D020687:cdc25 Phosphatases / Q000378:metabolism
D000970:Antineoplastic Agents; D018797:Cell Cycle Proteins; D019925:Cyclin A; D019926:Cyclin B; D004273:DNA, Neoplasm; D004791:Enzyme Inhibitors; D003561:Cytarabine; C054852:7-hydroxystaurosporine; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases; D019311:Staurosporine