title: Epidermal growth factor induces HCCR expression via PI3K/Akt/mTOR signaling in PANC-1 pancreatic cancer cells.
abstract: BACKGROUND
  Human cervical cancer oncoprotein 1 (HCCR-1), reported as a negative regulator of p53, is over-expressed in a variety of human cancers. However, it is yet unknown whether HCCR-1 plays any role in pancreatic cancer development. The aim of this study was to investigate the effect of epidermal growth factor on the expression of HCCR in pancreatic cancer cells, and to explore if PI3K/Akt/mTOR signaling pathway mediated this expression.
  
  
  METHODS
  A polyclonal antibody against HCCR protein was raised by immunizing Balb/c mice with the purified recombinant protein pMBPc-HCCR. Tissue samples were constructed on a tissue chip, and the expression of HCCR was investigated by immunohistochemistry assay and Western blotting. Pancreatic cell line, PANC-1 cells were stably transfected with plasmids containing sense-HCCR-1 fragment and HCCR siRNA fragment. MTT and transwell assay were used to investigate the proliferation and invasion of stable tansfectants. The specific inhibitor of PI3K and mTOR was used to see if PI3K/mTOR signal transduction was involved in the induction of HCCR gene expression. A Luciferase assay was used to see if Akt can enhance the HCCR promoter activity.
  
  
  RESULTS
  HCCR was up-regulated in pancreatic tumor tissues (mean Allred score 4.51+/-1.549 vs. 2.87+/-2.193, P<0.01), especially with high expression in poorly differentiated pancreatic cancer. The growth of cells decreased in HCCR-1 siRNA transfected cells compared with vector transfectants. The number of invasion cells was significantly lower in HCCR-1 siRNA transfected cells (24.4+/-9.9) than that in vector transfectants (49.1+/-15.4). Treatment of PANC-1 cells with epidermal growth factor increased HCCR protein level in a dose- and time-dependent manner. However, application of LY294002 and rapamycin caused a dramatic reduction of epidermal growth factor-induced HCCR expression. Over-expression of exogenous constitutively active Akt increased the HCCR promoter activity; in contrast, dominant negative Akt decreased the promoter activity.
  
  
  CONCLUSIONS
  EGF-induced HCCR-1 over-expression is mediated by PI3K/AKT/mTOR signaling which plays a pivotal role in pancreatic tumor progression, suggesting that HCCR-1 could be a potential target for cancer therapeutics.
authors.affiliation: Department of Gastroenterology, the First Affiliated Hospital of Nanjing Medical University, Nanjing 210029, China.
authors.forename: Zekuan
authors.initials: Z
authors.lastname: Xu
authors.forename: Yi
authors.initials: Y
authors.lastname: Zhang
authors.forename: Jiakai
authors.initials: J
authors.lastname: Jiang
authors.forename: Yang
authors.initials: Y
authors.lastname: Yang
authors.forename: Ruihua
authors.initials: R
authors.lastname: Shi
authors.forename: Bo
authors.initials: B
authors.lastname: Hao
authors.forename: Zhihong
authors.initials: Z
authors.lastname: Zhang
authors.forename: Zuhu
authors.initials: Z
authors.lastname: Huang
authors.forename: Jin W
authors.initials: JW
authors.lastname: Kim
authors.forename: Guoxin
authors.initials: G
authors.lastname: Zhang
mesh_terms: D000818:Animals; D000906:Antibodies; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D002867:Chromones / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D009025:Morpholines / Q000494:pharmacology; D009361:Neoplasm Invasiveness; D010190:Pancreatic Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011401:Promoter Regions, Genetic; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases; D013997:Time Factors; D046888:Tissue Array Analysis; D014162:Transfection; D015854:Up-Regulation
chemical_list: D000906:Antibodies; D002867:Chromones; D004815:Epidermal Growth Factor; D047908:Intracellular Signaling Peptides and Proteins; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D011994:Recombinant Proteins; D020123:Sirolimus; D058570:TOR Serine-Threonine Kinases; C476732:LETMD1 protein, human; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C546842:MTOR protein, human; C546843:mTOR protein, mouse