title: Imiquimod activates p53-dependent apoptosis in a human basal cell carcinoma cell line.
abstract: BACKGROUND
  The tumor suppressor p53 controls DNA repair, cell cycle, apoptosis, autophagy and numerous other cellular processes. Imiquimod (IMQ), a synthetic toll-like receptor (TLR) 7 ligand for the treatment of superficial basal cell carcinoma (BCC), eliminates cancer cells by activating cell-mediated immunity and directly inducing apoptosis and autophagy in cancer cells.
  
  
  OBJECTIVE
  To evaluate the role of p53 in IMQ-induced cell death in skin cancer cells.
  
  
  METHODS
  The expression, phosphorylation and subcellular localization of p53 were detected by real-time PCR, luciferase reporter assay, cycloheximide chase analysis, immunoblotting and immunocytochemistry. Using BCC/KMC1 cell line as a model, the upstream signaling of p53 activation was dissected by over-expression of TLR7/8, the addition of ROS scavenger, ATM/ATR inhibitors and pan-caspase inhibitor. The role of p53 in IMQ-induced apoptosis and autophagy was assessed by genetically silencing p53 and evaluated by a DNA content assay, immunoblotting, LC3 puncta detection and acridine orange staining.
  
  
  RESULTS
  IMQ induced p53 mRNA expression and protein accumulation, increased Ser15 phosphorylation, promoted nuclear translocation and up-regulated its target genes in skin cancer cells in a TLR7/8-independent manner. In BCC/KMC1 cells, the induction of p53 by IMQ was achieved through increased ROS production to stimulate the ATM/ATR-Chk1/Chk2 axis but was not mediated by inducing DNA damage. The pharmacological inhibition of ATM/ATR significantly suppressed IMQ-induced p53 activation and apoptosis. Silencing of p53 significantly decreased the IMQ-induced caspase cascade activation and apoptosis but enhanced autophagy. Mutant p53 skin cancer cell lines were more resistant to IMQ-induced apoptosis than wildtype p53 skin cancer cell lines.
  
  
  CONCLUSIONS
  IMQ induced ROS production to stimulate ATM/ATR pathways and contributed to p53-dependent apoptosis in a skin basal cell carcinoma cell line BCC/KMC1.
authors.affiliation: Department of Education and Research, Taichung Veterans General Hospital, Taichung, Taiwan.
authors.forename: Shi-Wei
authors.initials: SW
authors.lastname: Huang
authors.affiliation: Institute of Clinical Medicine, National Yang Ming University, Taipei, Taiwan.
authors.forename: Shu-Hao
authors.initials: SH
authors.lastname: Chang
authors.affiliation: Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan.
authors.forename: Szu-Wei
authors.initials: SW
authors.lastname: Mu
authors.affiliation: Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan.
authors.forename: Hsin-Yi
authors.initials: HY
authors.lastname: Jiang
authors.affiliation: Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan.
authors.forename: Sin-Ting
authors.initials: ST
authors.lastname: Wang
authors.affiliation: Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan; Department of Pediatrics, Children's Hospital, Changhua Christian Hospital, Changhua, Taiwan.
authors.forename: Jun-Kai
authors.initials: JK
authors.lastname: Kao
authors.affiliation: Department of Bioscience Technology, Chang Jung Christian University, Tainan, Taiwan.
authors.forename: Jau-Ling
authors.initials: JL
authors.lastname: Huang
authors.affiliation: Division of Gastroenterology and Hepatology, Taichung Veterans General Hospital, Taichung, Taiwan.
authors.forename: Chun-Ying
authors.initials: CY
authors.lastname: Wu
authors.affiliation: Department of Dermatology, Taichung Veterans General Hospital, Taichung, Taiwan.
authors.forename: Yi-Ju
authors.initials: YJ
authors.lastname: Chen
authors.affiliation: Department of Education and Research, Taichung Veterans General Hospital, Taichung, Taiwan; Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan; Rong Hsing Research Center for Translational Medicine, National Chung Hsing University, Taichung, Taiwan. Electronic address: shiehjj@vghtc.gov.tw.
authors.forename: Jeng-Jer
authors.initials: JJ
authors.lastname: Shieh
mesh_terms: D021581:Active Transport, Cell Nucleus; D000634:Aminoquinolines / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D064007:Ataxia Telangiectasia Mutated Proteins / Q000235:genetics / Q000378:metabolism; D001343:Autophagy / Q000187:drug effects; D002280:Basal Cell Carcinoma / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D000077271:Imiquimod; D009154:Mutation; D010766:Phosphorylation; D034622:RNA Interference; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D012878:Skin Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D013997:Time Factors; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*
chemical_list: D000634:Aminoquinolines; D000970:Antineoplastic Agents; D017382:Reactive Oxygen Species; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; C576324:ATM protein, human; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D000077271:Imiquimod
keywords: Apoptosis; Autophagy; Imiquimod; p53