Constitutively active c-Met kinase in PC-3 cells is autocrine-independent and can be blocked by the Met kinase inhibitor BMS-777607.
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.
Department of Radiation Oncology, University of Florida, 2033 Mowry Road, Cancer Genetic Research Complex, Room 485E, Gainesville, FL 32610, USA. daiyao@ufl.edu
Yao
Y
Dai
Dietmar W
DW
Siemann
D000631:Aminopyridines / Q000494:pharmacology*; D057134:Antibodies, Neutralizing / Q000276:immunology / Q000494:pharmacology; D019898:Autocrine Communication / Q000187:drug effects*; D045744:Cell Line, Tumor; D015972:Gene Expression Regulation, Neoplastic; D017228:Hepatocyte Growth Factor / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D006801:Humans; D008297:Male; D011471:Prostatic Neoplasms / Q000235:genetics / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D019859:Proto-Oncogene Proteins c-met / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D011728:Pyridones / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects
D000631:Aminopyridines; D057134:Antibodies, Neutralizing; C066855:HGF protein, human; C550356:N-(4-(2-amino-3-chloropyridin-4-yloxy)-3-fluorophenyl)-4-ethoxy-1-(4-fluorophenyl)-2-oxo-1,2-dihydropyridine-3-carboxamide; D047428:Protein Kinase Inhibitors; D011728:Pyridones; D012333:RNA, Messenger; D017228:Hepatocyte Growth Factor; D019859:Proto-Oncogene Proteins c-met