title: PKI-587 and sorafenib targeting PI3K/AKT/mTOR and Ras/Raf/MAPK pathways synergistically inhibit HCC cell proliferation.
abstract: BACKGROUND
  Deregulated Ras/Raf/MAPK and PI3K/AKT/mTOR signaling pathways are found in hepatocellular carcinoma (HCC). This study aimed to test the inhibitory effects of PKI-587 and sorafenib as single agents or in combination on HCC (Huh7 cell line) proliferation.
  
  
  METHODS
  (3)H-thymidine incorporation and MTT assay were used to assess Huh7 cell proliferation. Phosphorylation of the key enzymes in the Ras/Raf/MAPK and PI3K/AKT/mTOR pathways was detected by Western blot.
  
  
  RESULTS
  We found that PKI-587 is a more potent PI3K/mTOR inhibitor than PI-103. Combination of PKI-587 and sorafenib was a more effective inhibitor of Huh7 proliferation than the combination of PI-103 and sorafenib. Combination of PKI-587 and sorafenib synergistically inhibited epidermal growth factor (EGF)-stimulated Huh7 proliferation compared with monodrug therapy. EGF increased phosphorylation of Ras/Raf downstream signaling proteins MEK and ERK; EGF-stimulated activation was inhibited by sorafenib. However, sorafenib, as a single agent, increased AKT (Ser473) phosphorylation. EGF-stimulated AKT (ser473) activation was inhibited by PKI-587. PKI-587 is a potent inhibitor of AKT (Ser473), mTOR (Ser2448), and S6K (Thr389) phosphorylation; in contrast, rapamycin stimulated mTOR complex 2 substrate AKT(Ser473) phosphorylation although it inhibited mTOR complex 1 substrate S6K phosphorylation. PKI-587, as a single agent, stimulated MEK and ERK phosphorylation. However, when PKI-587 and sorafenib were used in combination, they inhibited all the tested kinases in the Ras/Raf /MAPK and PI3K/AKT/mTOR pathways.
  
  
  CONCLUSIONS
  The combination of PKI-587 and sorafenib has the advantage over monodrug therapy on inhibition of HCC cell proliferation by blocking both PI3K/AKT/mTOR and Ras/Raf/MAPK signaling pathways.
authors.affiliation: Department of Surgery, University of Kentucky, College of Medicine, Lexington, Kentucky 40536-0293, USA. rgeda2@uky.edu
authors.forename: Roberto
authors.initials: R
authors.lastname: Gedaly
authors.forename: Paul
authors.initials: P
authors.lastname: Angulo
authors.forename: Jonathan
authors.initials: J
authors.lastname: Hundley
authors.forename: Michael F
authors.initials: MF
authors.lastname: Daily
authors.forename: Changguo
authors.initials: C
authors.lastname: Chen
authors.forename: B Mark
authors.initials: BM
authors.lastname: Evers
mesh_terms: D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology; D001557:Benzenesulfonates / Q000494:pharmacology*; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy* / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D004357:Drug Synergism; D025461:Feedback, Physiological / Q000187:drug effects; D005663:Furans / Q000494:pharmacology; D006801:Humans; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects* / Q000502:physiology; D000076222:Mechanistic Target of Rapamycin Complex 1; D009025:Morpholines / Q000494:pharmacology*; D046912:Multiprotein Complexes; D009536:Niacinamide / Q000031:analogs & derivatives; D010671:Phenylurea Compounds; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011506:Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology*; D011743:Pyrimidines / Q000494:pharmacology; D020123:Sirolimus / Q000494:pharmacology; D000077157:Sorafenib; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D014227:Triazines / Q000494:pharmacology*
chemical_list: D000903:Antibiotics, Antineoplastic; D000970:Antineoplastic Agents; D001557:Benzenesulfonates; D005663:Furans; D000076222:Mechanistic Target of Rapamycin Complex 1; D009025:Morpholines; D046912:Multiprotein Complexes; D009536:Niacinamide; D010671:Phenylurea Compounds; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011506:Proteins; D051057:Proto-Oncogene Proteins c-akt; D011725:Pyridines; D011743:Pyrimidines; D020123:Sirolimus; D000077157:Sorafenib; D058570:TOR Serine-Threonine Kinases; D014157:Transcription Factors; D014227:Triazines; C493245:CRTC2 protein, human; C522973:PI103; C549060:gedatolisib; C546842:MTOR protein, human; C494918:AKT1 protein, human