Chlorpromazine activates p21Waf1/Cip1 gene transcription via early growth response-1 (Egr-1) in C6 glioma cells

Exp Mol Med. 2010 May 31;42(5):395-405. doi: 10.3858/emm.2010.42.5.041.


2-Chloro-10-[3(-dimethylamino)propyl]phenothiazinemonohydrochloride (chlorpromazine) is a phenothiazine derivative used clinically to control psychotic disorders. It also exhibits an anticancer activity. Treatment with chlorpromazine (CPZ) results in cell-cycle arrest at the G2/M phase in rat C6 glioma cells. CPZ reduces the expression of cell cycle-related proteins, such as cyclin D1, cyclin A, and cyclin B1, but causes an increase in the p21(Waf1/Cip1) level. The molecular mechanism by which CPZ regulates p21(Waf1/Cip1) expression is unknown. Here, we provide evidence that CPZ activates the p21(Waf1/Cip1) gene promoter via induction of the transcription factor early growth response-1 (Egr-1) independently of p53 in C6 cells. A point mutation in the Egr-1-binding motif within the p21(Waf1/Cip1) promoter abrogated promoter inducibility due to CPZ. Forced expression of Egr-1 enhanced p21(Waf1/Cip1) promoter activity. In contrast, knockdown of endogenous Egr-1 by small interference RNA attenuated CPZ-induced p21(Waf1/Cip1) promoter activity. A chromatin immunoprecipitation assay demonstrated that Egr-1 binds to the p21(Waf1/Cip1) gene promoter. Further analysis showed that the ERK and JNK MAP kinases are required for induction of Egr-1 by CPZ. Finally, stable silencing of Egr-1 expression lead to attenuated CPZ-inducible p21(Waf1/Cip1) expression and inhibited G2/M phase cell-cycle arrest. These results demonstrate that a functional link between ERK and JNK MAP kinase pathways and p21(Waf1/Cip1) induction via Egr-1 contributes to CPZ-induced anticancer activity in C6 glioma cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antineoplastic Agents / pharmacology
  • Cell Line, Tumor
  • Cell Proliferation / drug effects
  • Chlorpromazine / pharmacology*
  • Cyclin-Dependent Kinase Inhibitor p21 / metabolism*
  • Early Growth Response Protein 1 / genetics
  • Early Growth Response Protein 1 / metabolism*
  • Extracellular Signal-Regulated MAP Kinases / metabolism*
  • Gene Expression Regulation, Neoplastic / drug effects
  • Glioma / drug therapy
  • Glioma / metabolism*
  • JNK Mitogen-Activated Protein Kinases / metabolism*
  • RNA, Small Interfering
  • Rats
  • Transcription, Genetic / drug effects
  • Tumor Suppressor Protein p53 / metabolism


  • Antineoplastic Agents
  • Cyclin-Dependent Kinase Inhibitor p21
  • EGR1 protein, human
  • Early Growth Response Protein 1
  • RNA, Small Interfering
  • Tumor Suppressor Protein p53
  • Extracellular Signal-Regulated MAP Kinases
  • JNK Mitogen-Activated Protein Kinases
  • Chlorpromazine