Cyclic AMP/PKA-dependent paradoxical activation of Raf/MEK/ERK signaling in polycystin-2 defective mice treated with sorafenib

Hepatology. 2012 Dec;56(6):2363-74. doi: 10.1002/hep.25872. Epub 2012 Aug 27.


Mutations in polycystins are a cause of polycystic liver disease. In polycystin-2 (PC2)-defective mice, cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA)-dependent activation of the Rat Sarcoma (Ras)/rapidly accelerated fibrosarcoma (Raf)/mitogen signal-regulated kinase-extracellular signal-regulated kinase (ERK) 1/2 pathway stimulates the growth of liver cysts. To test the hypothesis that sorafenib, a Raf inhibitor used for the treatment of liver and kidney cancers, inhibits liver cyst growth in PC2-defective mice, we treated PC2 (i.e., Pkd2(flox/-) :pCxCreER(TM) [Pkd2cKO]) mice with sorafenib-tosylate for 8 weeks (20-60 mg/kg/day). Sorafenib caused an unexpected increase in liver cyst area, cell proliferation (Ki67), and expression of phosphorylated ERK (pERK) compared with Pkd2cKO mice treated with vehicle. When given to epithelial cells isolated from liver cysts of Pkd2cKO mice (Pkd2cKO-cells), sorafenib progressively stimulated pERK1/2 and cell proliferation [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium and bromodeoxyuridine assay (MTS)] at doses between 0.001 and 1 μM; however, both pERK1/2 and cell proliferation significantly decreased at the dose of 10 μM. Raf kinase activity assay showed that whereas B-Raf is inhibited by sorafenib in both wild-type (WT) and Pkd2cKO cells, Raf-1 is inhibited in WT cells but is significantly stimulated in Pkd2cKO cells. In Pkd2cKO cells pretreated with the PKA inhibitor 14-22 amide, myristolated (1 μM) and in mice treated with octreotide in combination with sorafenib, the paradoxical activation of Raf/ERK1/2 was abolished, and cyst growth was inhibited.

Conclusion: In PC2-defective cells, sorafenib inhibits B-Raf but paradoxically activates Raf-1, resulting in increased ERK1/2 phosphorylation, cell proliferation, and cyst growth in vivo. These effects are consistent with the ability of Raf inhibitors to transactivate Raf-1 when a PKA-activated Ras promotes Raf-1/B-Raf heterodimerization, and are inhibited by interfering with cAMP/PKA signaling both in vitro and in vivo, as shown by the reduction of liver cysts in mice treated with combined octreotide and sorafenib.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antineoplastic Agents, Hormonal / pharmacology
  • Antineoplastic Agents, Hormonal / therapeutic use
  • Benzenesulfonates / pharmacology*
  • Benzenesulfonates / therapeutic use
  • Bile Ducts / cytology
  • Bile Ducts / metabolism
  • Caspase 3 / metabolism
  • Cell Proliferation / drug effects
  • Cells, Cultured
  • Cyclic AMP-Dependent Protein Kinases / metabolism*
  • Cysts / drug therapy
  • Cysts / enzymology*
  • Cysts / pathology
  • Drug Therapy, Combination
  • Epithelial Cells / drug effects
  • Epithelial Cells / enzymology
  • Ki-67 Antigen / metabolism
  • Liver Diseases / drug therapy
  • Liver Diseases / enzymology*
  • Liver Diseases / pathology
  • MAP Kinase Signaling System / drug effects*
  • Mice
  • Mice, Knockout
  • Niacinamide / analogs & derivatives
  • Octreotide / pharmacology
  • Octreotide / therapeutic use
  • Phenylurea Compounds
  • Phosphorylation
  • Protein Kinase Inhibitors / pharmacology*
  • Protein Kinase Inhibitors / therapeutic use
  • Proto-Oncogene Proteins B-raf / metabolism
  • Proto-Oncogene Proteins c-raf / metabolism
  • Pyridines / pharmacology*
  • Pyridines / therapeutic use
  • Sorafenib
  • TRPP Cation Channels / deficiency*
  • TRPP Cation Channels / genetics


  • Antineoplastic Agents, Hormonal
  • Benzenesulfonates
  • Ki-67 Antigen
  • Phenylurea Compounds
  • Protein Kinase Inhibitors
  • Pyridines
  • TRPP Cation Channels
  • polycystic kidney disease 2 protein
  • Niacinamide
  • Sorafenib
  • Proto-Oncogene Proteins B-raf
  • Proto-Oncogene Proteins c-raf
  • Cyclic AMP-Dependent Protein Kinases
  • Caspase 3
  • Octreotide