SC-560, a strucutral analogue of celecoxib, induces growth inhibition in a wide range of human cancer cells in a cyclooxygenase (COX)-independent manner. Since SC-560 suppresses the growth of cancer cells mainly by inducing cell cycle arrest, we sought to examine the role of p21CIP1, a cell cycle regulator protein, in the cellular response against SC-560 by using p21(+/+)and p21(-/-)isogenic HCT116 colon carcinoma cells. In HCT116 (p21(+/+)) cells, SC-560 dose-dependently induced growth inhibition and cell cycle arrest at the G1 phase without significant apoptosis induction. SC-560-induced cell cycle arrest was accompanied by upregulation of p21CIP1. However, the extent of SC-560-induced accumulation at the G1 phase was approximately equal in the p21(+/+)and the p21(-/-)cells. Nonetheless, the growth inhibition by SC-560 was increased in p21(-/-)cells than p21(+/+)cells. SC-560-induced reactive oxygen species (ROS) generation did not differ between p21(+/+)and p21(-/-)cells but the subsequent activaton of apoptotic caspase cascade was more pronounced in p21(-/-)cells compared with p21(+/+)cells. These results suggest that p21CIP1 blocks the SC-560-induced apoptotic response of HCT116 cells. SC-560 combined with other therapy that can block p21 CIP1 expression or function may contribute to the effective treatment of colon cancer.
Reactive Oxygen Species/metabolism ; Pyrazoles/*pharmacology ; Mutation ; Immunoblotting ; Humans ; HCT116 Cells ; Genotype ; Flow Cytometry ; Dose-Response Relationship, Drug ; Cyclooxygenase Inhibitors/pharmacology ; Cyclin-Dependent Kinase Inhibitor p21/genetics/metabolism/*physiology ; Colonic Neoplasms/genetics/metabolism/pathology ; Cell Survival/drug effects ; Cell Proliferation/drug effects ; Cell Differentiation/drug effects ; Cell Cycle Proteins/metabolism ; Cell Cycle/drug effects ; Apoptosis/drug effects ; Antineoplastic Agents/pharmacology

The inhibitory Smad6 and Smad7 are responsible for cross-talk between TGF-beta/bone morphogenic protein (BMP) signaling and other cellular signaling pathways, as well as negative feedback on their own signaling functions. Although inhibitory Smads are induced by various stimuli, little is known about the stimuli that increase Smad6 transcription, in contrast to Smad7. Here we demonstrate that etoposide, which induces double strand breaks during DNA replication, significantly up-regulates the transcription of the Smad6 gene in CMT-93 mouse intestinal cells by increasing specific DNA binding proteins. In addition, endogenous inhibition of the Smad6 gene by RNAi interference led to transient accumulation of G1 phase cells and reduction in incorporation of bromodeoxyuridine (BrdU). These findings strongly suggest that Smad6 plays a distinct role in the signaling of etoposide-induced DNA damage.
Animals ; Base Sequence ; Cell Line ; DNA-Binding Proteins/metabolism ; Enterocytes/*cytology/drug effects/*metabolism ; Etoposide/*pharmacology ; G1 Phase/*drug effects ; Mice ; Molecular Sequence Data ; Promoter Regions, Genetic/genetics ; RNA, Small Interfering/metabolism ; S Phase/*drug effects ; Smad6 Protein/*genetics ; Transcriptional Activation/drug effects