Constitutively active c-Met kinase in PC-3 cells is autocrine-independent and can be blocked by the Met kinase inhibitor BMS-777607

BMC Cancer. 2012 May 28;12:198. doi: 10.1186/1471-2407-12-198.

Abstract

Background: The c-Met receptor tyrosine kinase is aberrantly activated in many solid tumors. In a prior study we showed that prostate cancer PC-3 cells exhibit constitutively activated c-Met without exogenous hepatocyte growth factor (HGF); however whether this characteristic is due to an endogenous HGF/c-Met autocrine loop remains controversial. In the current study we examined the response of PC-3 cells to an anti-HGF neutralizing antibody or a small molecule Met kinase inhibitor (BMS-777607).

Methods: Cell scattering was tested by monitoring cell morphology after HGF stimulation. Cell migration was examined by both "wound-healing" and transwell assasy and invasion was detected by Matrigel-coated transwell assay. Proliferation, survival and anoikis were determined by MTT, colony formation and trypan blue exclusion assay, respectively. Gene and protein expression were assessed by real-time PCR and Western blot, respectively.

Results: Although HGF mRNA could be detected in PC-3 cells, the molecular weight of secreted "HGF" protein was inconsistent with the functional recombinant HGF. Furthermore, conditioned medium from PC-3 cell cultures was ineffective at triggering either motogenic behavior or c-Met signaling in DU145, another prostate cancer cell line expressing c-Met but lacking basal c-Met activation. PC-3 cells also were not responsive to the anti-HGF neutralizing antibody in experiments assessing proliferation, migration, or c-Met signaling. BMS-777607 treatment with micromolar doses nonetheless led to significant inhibition of multiple PC-3 cell functions including proliferation, clonogenicity, migration and invasion. At the molecular level, BMS-777607 suppressed autophosphorylated c-Met and downstream c-Src and Akt pathways.

Conclusions: These results suggest that the constitutive c-Met activation in PC-3 is independent of autocrine stimulation. Because PC-3 cells were responsive to BMS-777607 but not the anti-HGF antibody, the findings also indicate that under circumstances where c-Met is constitutively hyperactive in the absence of functional HGF, targeting the c-Met receptor remains a viable therapeutic option to impede cancer progression.

MeSH terms

  • Aminopyridines / pharmacology*
  • Antibodies, Neutralizing / immunology
  • Antibodies, Neutralizing / pharmacology
  • Autocrine Communication / drug effects*
  • Cell Line, Tumor
  • Gene Expression Regulation, Neoplastic
  • Hepatocyte Growth Factor / genetics
  • Hepatocyte Growth Factor / immunology
  • Hepatocyte Growth Factor / metabolism
  • Humans
  • Male
  • Prostatic Neoplasms / genetics
  • Prostatic Neoplasms / metabolism
  • Protein Kinase Inhibitors / pharmacology*
  • Proto-Oncogene Proteins c-met / antagonists & inhibitors*
  • Proto-Oncogene Proteins c-met / metabolism*
  • Pyridones / pharmacology*
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Signal Transduction / drug effects

Substances

  • Aminopyridines
  • Antibodies, Neutralizing
  • HGF protein, human
  • N-(4-(2-amino-3-chloropyridin-4-yloxy)-3-fluorophenyl)-4-ethoxy-1-(4-fluorophenyl)-2-oxo-1,2-dihydropyridine-3-carboxamide
  • Protein Kinase Inhibitors
  • Pyridones
  • RNA, Messenger
  • Hepatocyte Growth Factor
  • Proto-Oncogene Proteins c-met