title: Loss of Smad4 in colorectal cancer induces resistance to 5-fluorouracil through activating Akt pathway.
abstract: BACKGROUND
  Higher frequency of Smad4 inactivation or loss of expression is observed in metastasis of colorectal cancer (CRC) leading to unfavourable survival and contributes to chemoresistance. However, the molecular mechanism of how Smad4 regulates chemosensitivity of CRC is unknown.
  
  
  METHODS
  We evaluated how the loss of Smad4 in CRC enhanced chemoresistance to 5-fluorouracil (5-FU) using two CRC cell lines in vitro and in vivo. Immunoblotting with cell and tumour lysates and immunohistochemical analyses with tissue microarray were performed.
  
  
  RESULTS
  Knockdown or loss of Smad4 induced tumorigenicity, migration, invasion, angiogenesis, metastasis, and 5-FU resistance. Smad4 expression in mouse tumours regulated cell-cycle regulatory proteins leading to Rb phosphorylation. Loss of Smad4 activated Akt pathway that resulted in upregulation of anti-apoptotic proteins, Bcl-2 and Bcl-w, and Survivin. Suppression of phosphatidylinositol-3-kinase (PI3K)/Akt pathway by LY294002 restored chemosensitivity of Smad4-deficient cells to 5-FU. Vascular endothelial growth factor-induced angiogenesis in Smad4-deficient cells might also lead to chemoresistance. Low levels of Smad4 expression in CRC tissues correlated with higher levels of Bcl-2 and Bcl-w and with poor overall survival as observed in immunohistochemical staining of tissue microarrays.
  
  
  CONCLUSIONS
  Loss of Smad4 in CRC patients induces resistance to 5-FU-based therapy through activation of Akt pathway and inhibitors of this pathway may sensitise these patients to 5-FU.
authors.affiliation: 1] Birmingham Veterans Affairs Medical Center, Birmingham, AL, USA [2] Department of Medicine, University of Alabama at Birmingham, 1824 6th Avenue South, WTI 520C, Birmingham, AL 35294, USA [3] Department of Surgery, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, USA.
authors.forename: B
authors.initials: B
authors.lastname: Zhang
authors.affiliation: Department of Surgery, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, USA.
authors.forename: B
authors.initials: B
authors.lastname: Zhang
authors.affiliation: Department of Surgery, Hepatic Surgery Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1095 Jiefang Avenue, Wuhan 430030, China.
authors.forename: X
authors.initials: X
authors.lastname: Chen
authors.affiliation: Department of Medicine, University of Alabama at Birmingham, 1824 6th Avenue South, WTI 520C, Birmingham, AL 35294, USA.
authors.forename: S
authors.initials: S
authors.lastname: Bae
authors.affiliation: Department of Medicine, University of Alabama at Birmingham, 1824 6th Avenue South, WTI 520C, Birmingham, AL 35294, USA.
authors.forename: K
authors.initials: K
authors.lastname: Singh
authors.affiliation: Department of Pathology, Vanderbilt University School of Medicine, Nashville, TN, USA.
authors.forename: M K
authors.initials: MK
authors.lastname: Washington
authors.affiliation: 1] Birmingham Veterans Affairs Medical Center, Birmingham, AL, USA [2] Department of Medicine, University of Alabama at Birmingham, 1824 6th Avenue South, WTI 520C, Birmingham, AL 35294, USA [3] Department of Surgery, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, USA.
authors.forename: P K
authors.initials: PK
authors.lastname: Datta
mesh_terms: D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D051017:Apoptosis Regulatory Proteins; D018797:Cell Cycle Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics; D049109:Cell Proliferation; D002470:Cell Survival / Q000235:genetics; D002867:Chromones / Q000494:pharmacology; D015179:Colorectal Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology*; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005472:Fluorouracil / Q000494:pharmacology*; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000096:biosynthesis; D051379:Mice; D008807:Mice, Inbred BALB C; D009025:Morpholines / Q000494:pharmacology; D009361:Neoplasm Invasiveness; D009362:Neoplasm Metastasis / Q000235:genetics; D009368:Neoplasm Transplantation; D009389:Neovascularization, Pathologic / Q000235:genetics; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D012097:Repressor Proteins / Q000096:biosynthesis; D016160:Retinoblastoma Protein / Q000378:metabolism; D051901:Smad4 Protein / Q000172:deficiency / Q000235:genetics*; D000077022:Survivin; D015854:Up-Regulation; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D014945:Wound Healing; D023041:Xenograft Model Antitumor Assays
chemical_list: D000964:Antimetabolites, Antineoplastic; D051017:Apoptosis Regulatory Proteins; D018797:Cell Cycle Proteins; D002867:Chromones; D004791:Enzyme Inhibitors; D005472:Fluorouracil; D051034:Inhibitor of Apoptosis Proteins; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011506:Proteins; D051057:Proto-Oncogene Proteins c-akt; D019253:Proto-Oncogene Proteins c-bcl-2; D012097:Repressor Proteins; D016160:Retinoblastoma Protein; D051901:Smad4 Protein; D000077022:Survivin; D042461:Vascular Endothelial Growth Factor A; C101155:Bcl2l2 protein, mouse; C117749:Birc5 protein, mouse; C490810:Smad4 protein, mouse; C467480:vascular endothelial growth factor A, mouse; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one