title: Inhibition of p38 MAPK-dependent MutS homologue-2 (MSH2) expression by metformin enhances gefitinib-induced cytotoxicity in human squamous lung cancer cells.
abstract: OBJECTIVE
  Gefitinib, a quinazoline-derived tyrosine kinase inhibitor, has anti-tumor activity in vivo and in vitro. Human MutS homologue-2 (MSH2) plays a central role in promoting genetic stability by correcting DNA replication errors. The present study investigated the effects of p38 mitogen-activated protein kinase (MAPK) signal on gefitinib-induced MSH2 expression in two human non-small cell lung squamous cancer cell lines.
  
  
  METHODS
  After the gefitinib treatment, the expressions of MSH2 mRNA were determined by real-time PCR and RT-PCR analysis. Protein levels of MSH2, phospho-MKK3/6, phospho-p38 MAPK were determined by Western blot analysis. We used specific MSH2, and p38 MAPK small interfering RNA to examine the role of p38 MAPK-MSH2 signal in regulating the chemosensitivity of gefitinib. Cell viability was assessed by MTS assay, trypan blue exclusion, and colony-forming ability assay.
  
  
  RESULTS
  Exposure of gefitinib increased MSH2 protein and mRNA levels, which was accompanied by MKK3/6-p38 MAPK activation in H520 and H1703 cells. Moreover, blocking p38 MAPK activation by SB202190 significantly decreased gefitinib-induced MSH2 expression by increasing mRNA and protein instability. In contrast, enhancing p38 activation using constitutively active MKK6 (MKK6E) increased MSH2 protein and mRNA levels. Specific inhibition of MSH2 expression by siRNA enhanced gefitinib-induced cytotoxicity. Metformin, an anti-diabetic drug, might reduce cancer risk. In human lung squamous cancer cells, metformin decreased gefitinib-induced MSH2 expression and augmented the cytotoxic effect and growth inhibition by gefitinib. Transient expression of MKK6E or HA-p38 MAPK vector could abrogate metformin and gefitinib-induced synergistic cytotoxic effect in H520 and H1703 cells.
  
  
  CONCLUSIONS
  Together, down-regulation of MSH2 expression can be a possible strategy to enhance the sensitivity of gefitinib to human lung squamous cancer cells.
authors.affiliation: Department of Internal Medicine, National Taiwan University Hospital, Hsin-Chu Branch, Taiwan; Department of Nursing, Yuanpei University, Hsinchu, Taiwan; Institute of Technology Law, National Chiao Tung University, Hsinchu, Taiwan.
authors.forename: Jen-Chung
authors.initials: JC
authors.lastname: Ko
authors.forename: Hsien-Chun
authors.initials: HC
authors.lastname: Chiu
authors.forename: Ting-Yu
authors.initials: TY
authors.lastname: Wo
authors.forename: Yi-Jhen
authors.initials: YJ
authors.lastname: Huang
authors.forename: Sheng-Chieh
authors.initials: SC
authors.lastname: Tseng
authors.forename: Yu-Ching
authors.initials: YC
authors.lastname: Huang
authors.forename: Huang-Jen
authors.initials: HJ
authors.lastname: Chen
authors.forename: Jhan-Jhang
authors.initials: JJ
authors.lastname: Syu
authors.forename: Chien-Yu
authors.initials: CY
authors.lastname: Chen
authors.forename: Yun-Ting
authors.initials: YT
authors.lastname: Jian
authors.forename: Yi-Jun
authors.initials: YJ
authors.lastname: Jian
authors.forename: Yun-Wei
authors.initials: YW
authors.lastname: Lin
mesh_terms: D002294:Carcinoma, Squamous Cell / Q000188:drug therapy* / Q000276:immunology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D017024:Chemotherapy, Adjuvant; D004357:Drug Synergism; D000077156:Gefitinib; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000276:immunology; D048371:MAP Kinase Kinase 3 / Q000378:metabolism; D048669:MAP Kinase Kinase 6 / Q000235:genetics / Q000378:metabolism; D008687:Metformin / Q000494:pharmacology*; D051718:MutS Homolog 2 Protein / Q000235:genetics / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D011725:Pyridines / Q000494:pharmacology; D011799:Quinazolines / Q000494:pharmacology*; D034741:RNA, Small Interfering / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects; D019076:Transgenes / Q000235:genetics; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D007093:Imidazoles; D011725:Pyridines; D011799:Quinazolines; D034741:RNA, Small Interfering; D008687:Metformin; D048051:p38 Mitogen-Activated Protein Kinases; D048371:MAP Kinase Kinase 3; D048669:MAP Kinase Kinase 6; C482566:MAP2K6 protein, human; C497172:MSH2 protein, human; D051718:MutS Homolog 2 Protein; C090942:4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)imidazole; D000077156:Gefitinib
keywords: Cytotoxicity; Gefitinib; MSH2; Metformin; Non-small cell lung cancer; Tyrosine kinase inhibitor; p38 MAPK