title: Pancreatic tumor suppression by benzyl isothiocyanate is associated with inhibition of PI3K/AKT/FOXO pathway.
abstract: OBJECTIVE
  Our previous studies have shown that benzyl isothiocyanate (BITC) suppress pancreatic cancer growth by inducing apoptosis but the molecular mechanism was unclear. In this study we hypothesized the involvement of PI3K/AKT/FOXO pathway in BITC-induced apoptosis.
  
  
  METHODS
  Mice were implanted BxPC-3 tumor xenografts and orally gavaged with 12 μmol BITC. Plasma and tumor BITC concentration was estimated by liquid chromatography/tandem mass spectrometry. BxPC-3 and PanC-1 cells were used to elucidate PI3K/AKT/FOXO pathway. Electrophoretic mobility shift assay (EMSA), DNA binding activity, immunofluorescence, and gene transfection were used to delineate the mechanism.
  
  
  RESULTS
  BITC-treated mice showed 43% less tumor growth as compared with control mice and correlated well with the therapeutic concentrations of 6.5 μmol/L BITC achieved in plasma and 7.5 μmol/g BITC in tumor tissue. Western blot analyses and immunohistochemistry revealed that tumors from BITC-treated mice showed reduced phosphorylation of PI3K, AKT, PDK1, mTOR, FOXO1, and FOXO3a and increased apoptosis. Complementing our in vivo results, we made similar observations in a dose- and time-dependent manner in BITC-treated BxPC-3 and Panc-1 cells. Binding of FOXO1 with 14-3-3 proteins was also reduced drastically by BITC treatment indicating nuclear retention of FOXO1 and this observation was further confirmed with EMSA, immunofluorescence, DNA binding, and upregulation of FOXO-responsive proteins Bim, p27, and p21 in BxPC-3 cells. Overexpression of AKT by transient transfection significantly blocked the modulation of FOXO proteins and protected the cells from BITC-mediated apoptosis and growth suppression.
  
  
  CONCLUSIONS
  Our results provide convincing evidence on the involvement of PI3K/AKT/FOXO pathway in BITC-mediated pancreatic tumor growth suppression.
authors.affiliation: Department of Biomedical Sciences and Cancer Biology Center, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas 79106, USA.
authors.forename: Srinivas Reddy
authors.initials: SR
authors.lastname: Boreddy
authors.forename: Kartick C
authors.initials: KC
authors.lastname: Pramanik
authors.forename: Sanjay K
authors.initials: SK
authors.lastname: Srivastava
mesh_terms: D000107:Acetylation; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D000072224:Bcl-2-Like Protein 11; D050882:CREB-Binding Protein / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000378:metabolism; D015536:Down-Regulation; D000071161:Forkhead Box Protein O1; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors / Q000378:metabolism*; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism; D017879:Isothiocyanates / Q000494:pharmacology*; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D010190:Pancreatic Neoplasms / Q000188:drug therapy* / Q000473:pathology; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000502:physiology*; D010766:Phosphorylation; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D021381:Protein Transport / Q000187:drug effects; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000502:physiology*; D015398:Signal Transduction / Q000187:drug effects; D037761:Sirtuins / Q000378:metabolism; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism; D014183:Transplantation, Heterologous; D047368:Tumor Burden / Q000187:drug effects; D050880:p300-CBP Transcription Factors / Q000378:metabolism; D000099224:p300-CBP-Associated Factor
chemical_list: D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; D000072224:Bcl-2-Like Protein 11; D050882:CREB-Binding Protein; D053148:Caspase 3; D050760:Cyclin-Dependent Kinase Inhibitor p27; D000071161:Forkhead Box Protein O1; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; D051550:I-kappa B Kinase; D017879:Isothiocyanates; D008565:Membrane Proteins; D019869:Phosphatidylinositol 3-Kinases; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D037761:Sirtuins; D058570:TOR Serine-Threonine Kinases; D050880:p300-CBP Transcription Factors; D000099224:p300-CBP-Associated Factor; C000606775:BCL2L11 protein, human; C000606776:Bcl2l11 protein, mouse; C085126:FOXO1 protein, human; C499517:FOXO3 protein, human; C031403:benzyl isothiocyanate; C494363:CREBBP protein, human; C499320:PARP1 protein, human; C546842:MTOR protein, human