LGH00031, a novel ortho-quinonoid inhibitor of cell division cycle 25B, inhibits human cancer cells via ROS generation.
OBJECTIVE
To discover novel cell division cycle 25 (CDC25) B inhibitors and elucidate the mechanisms of inhibition in cancer cells.


METHODS
Cell growth inhibition was detected by MTT assay, the cell cycle was analyzed by flow cytometry, and protein expression and phosphorylation was examined by Western blot analysis.


RESULTS
LGH00031 inhibited CDC25B irreversibly in vitro in a dose-dependent manner, and impaired the proliferation of tumor cell lines. In synchronized HeLa cells, LGH00031 delayed the cell cycle progression at the G(2)/M phase. LGH00031 increased cyclin-dependent kinase 1 (CDK1) tyrosine 15 phosphorylation and cyclin B1 protein level. The activity of LGH00031 against CDC25B in vitro relied on the existence of 1,4-dithiothreitol (DTT) or dihydrolipoic acid and oxygen. The oxygen free radical scavenger catalase and superoxide dismutase reduced the inactivation of CDC25 by LGH00031, confirming that reactive oxygen species (ROS) mediate the inactivation process in vitro. LGH00031 accelerated cellular ROS production in a dose-dependent manner, and N-acetyl cysteine (NAC) markedly decreased the ROS production induced by LGH00031. Correspondingly, the LGH00031-induced decrease in cell viability and cell cycle arrest, cyclin B1 protein level, and phosphorylation of CDK1 tyrosine 15 were also rescued by NAC that decreased ROS production.


CONCLUSIONS
The activity of LGH00031 at the molecular and cellular level is mediated by ROS.
Zhou
Yu-bo
YB
National Center for Drug Screening, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
Feng
Xu
X
Wang
Li-na
LN
Du
Jun-qing
JQ
Zhou
Yue-yang
YY
Yu
Hai-ping
HP
Zang
Yi
Y
Li
Jing-ya
JY
Li
Jia
J
D000970:Antineoplastic Agents / Q000494:pharmacology*; D015153:Blotting, Western; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D006367:HeLa Cells / Q000187:drug effects; D006801:Humans; D011809:Quinones / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D020687:cdc25 Phosphatases / Q000037:antagonists & inhibitors*
D000970:Antineoplastic Agents; C544872:LGH00031; D011809:Quinones; D017382:Reactive Oxygen Species; C517008:CDC25B protein, human; D020687:cdc25 Phosphatases