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




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BMC Cancer

, 12:198

First Online: 28 May 2012Received: 20 December 2011Accepted: 13 April 2012DOI: 10.1186-1471-2407-12-198

Cite this article as: Dai, Y. & Siemann, D.W. BMC Cancer 2012 12: 198. doi:10.1186-1471-2407-12-198

Abstract

BackgroundThe 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.

MethodsCell 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.

ResultsAlthough 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.

ConclusionsThese 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.

KeywordsBMS-777607 c-Met HGF Neutralizing antibody Prostate cancer AbbreviationsHGFHepatocyte growth factor

DMSODimethyl sulfoxide

ERKExtracellular signal-regulated kinases

FAKFocal adhesion kinase

S6KS6 kinase

FBSFetal bovine serum

CMConditioned medium

poly-HEMA2-hydroxyethyl methacrylate

RIPARadioimmunoprecipitation assay

HRPHorseradish peroxidase

mTORMammalian target of rapamycin.

Electronic supplementary materialThe online version of this article doi:10.1186-1471-2407-12-198 contains supplementary material, which is available to authorized users.

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Autor: Yao Dai - Dietmar W Siemann

Fuente: https://link.springer.com/







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