Type II cGMP-dependent protein kinase inhibits activation of key members of the RTK family in gastric cancer cells.

Receptor tyrosine kinases (RTKs) are key to the regulation of biological cell activities, particularly of tumor cells. In a previous study, we demonstated that type II cGMP-dependent protein kinase (PKG II) may inhibit the activation of epidermal growth factor receptor (EGFR), a key member of the RTK family, and inhibit the consequent signal transduction in gastric cancer cells. Since RTKs exhibit a high level of conservation in their molecular structure, we hypothesized that PKG II may exert a similar inhibitory effect on the activation of other members of the RTK family. The aim of the present study was to investigate the potential inhibitory effect of PKG II on the activation of vascular endothelial growth factor receptor (VEGFR), platelet-derived growth factor receptor (PDGFR) and insulin-like growth factor-1 receptor (IGF-1R). The AGS gastric cancer cell line was transfected with adenoviral vector encoding PKG II cDNA (Ad-PKG II) to increase the expression of PKG II and incubated with 8-pCPT-cGMP to activate the kinase. The cells were then stimulated with VEGF, PDGF and IGF-1, and the phosphorylation/activation of relative RTKs was detected by western blot analysis. The results demonstrated that stimulating cells with VEGF-C (100 ng/ml), PDGF-BB (100 ng/ml) and IGF-1 (100 ng/ml) for 5 min led to a clear increase of phosphorylation of VEGFR2 (Tyr 951), PDGFRβ (Tyr 751) and IGF-1R (Tyr 1161), respectively. Pre-infection of the cells with Ad-PKG II with a multiplicity of infection (MOI) of 100% overnight and pre-incubation of cells with 8-pCPT-cGMP (100 and 250 μM) for 1 h efficiently inhibited the ligand-binding-induced phosphorylation/activation of the RTKs. PKG II also inhibited the MAPK/ERK- and PI3K-mediated signal transductions induced by VEGF-C, PDGF-BB and IGF-1. The results demonstrated that PKG II may exert a wide range of inhibitory effects on the activation of RTKs and provided further evidence to confirm PKG II as a tumor suppressor.