PT - JOURNAL ARTICLE AU - Prabodh K Kandala AU - Sanjay K. Srivastava TI - Activation of Chk2 by 3,3' Diinolylmethane is required for causing G2/M cell cycle arrest in human ovarian cancer cells AID - 10.1124/mol.110.063750 DP - 2010 May 05 TA - Molecular Pharmacology PG - mol.110.063750 4099 - http://molpharm.aspetjournals.org/content/early/2010/05/05/mol.110.063750.short 4100 - http://molpharm.aspetjournals.org/content/early/2010/05/05/mol.110.063750.full AB - We evaluated the effect of 3,3'-diindolylmethane (DIM) in ovarian cancer cells. DIM treatment inhibited the growth of SKOV-3, TOV-21G and OVCAR-3 ovarian cancer cells in both dose and time-dependent manner with effective concentrations ranging from 40μM to 100μM. Growth inhibitory effects of DIM were mediated by cell cycle arrest in G2/M phase in all the three cell lines. G2/M arrest was associated with DNA damage as indicated by phosphorylation of H2A.X at Ser 139 and activation of Chk2 in all the three cell lines. Other G2/M regulatory molecules such as Cdc25C, Cdk1, Cyclin B1 were downregulated by DIM. Cycloheximide or Chk2 inhibitor pretreatment abrogated not only activation of Chk2 but also G2/M arrest and apoptosis mediated by DIM. To further establish the involvement of Chk2 in DIM-mediated G2/M arrest, cells were transfected with dominant negative Chk2 (DN-Chk2). Blocking Chk2 activation by DN-Chk2 completely protected cells from DIM- mediated G2/M arrest. These results were further confirmed in Chk2 knock out DT40 lymphoma cells where DIM failed to cause cell cycle arrest. These results clearly indicate the requirement of Chk2 activation to cause G2/M arrest by DIM in ovarian cancer cells. Moreover, blocking Chk2 activation also abrogates the apoptosis-inducing effects of DIM. Further our results show that DIM treatment cause ROS generation. Blocking ROS generation by NAC protect the cells from DIM- mediated G2/M arrest and apoptosis. Our results establish Chk2 as a potent molecular target of DIM in ovarian cancer cells and provide the rationale for further clinical investigation of DIM.The American Society for Pharmacology and Experimental Therapeutics