M1970 Knock Down of Signal Tranducer and Avtivator of Transcripton3 (STAT3) Enhance Gemcitabine Mediated Apoptosis in Chemoresistant Pancreatic Cancer Cells In Vitro and In Vivo

Abstract

Background: Protein kinase D1 (PKD1) is the founding member of a new protein kinase family, separate from the previously identified PKCs. PKD family members are rapidly activated by G protein-coupled receptor (GPCR) agonists which are increasingly implicated as autocrine/paracrine growth factors for multiple solid tumors, including pancreatic ductal adenocarcinoma, a devastating disease with overall 5 year survival of only 3-5%. A variety of GPCR agonists, including neurotensin (NT), stimulate DNA synthesis in human pancreatic adenocarcinoma cell lines. We used inducible stable expression of wild-type (WT) and kinase-dead (K618N) forms of PKD1 in PANC-1 cells to elucidate its function in the regulation of NT-induced MAP kinases, DNA synthesis and cell proliferation. Results: Neurotensin (NT) stimulates protein kinase D1 (PKD1), extracellular signal regulated kinase (ERK), c-Jun Nterminal Kinase (JNK) and DNA synthesis in the human pancreatic adenocarcinoma cell line PANC-1. To determine the effect of PKD1 overexpression on these biological responses, we generated inducible stable PANC-1 clones that express wild-type (WT) or kinase-dead (K618N) forms of PKD1 in response to the ecdysone analog ponasterone-A (PonA). NT potently stimulated c-Jun Ser63 phosphorylation (indicative of JNK activity) in both wild type and clonal derivatives of PANC-1 cells. PonA-induced expression of WT, but not K618N PKD1, rapidly blocked NT-mediated c-Jun Ser63 phosphorylation either at the level of or upstream of MKK4. This is the first demonstration that PKD1 suppresses GPCR-induced JNK/cJun activation in any cell type. In contrast, PKD1 overexpression markedly increased the duration of NT-induced ERK activation and stimulated DNA synthesis and proliferation of PANC-1 cells cultured in regular dishes or in polyhydroxyethylmethacrylate [Poly-(HEMA)]coated dishes to eliminate cell adhesion (anchorage-independent growth). Conclusion: The reciprocal influence of PKD1 signaling on pro-mitogenic ERK and pro-apoptotic JNK/c-Jun pathways provides a mechanism by which PKD1 overexpression promotes DNA synthesis and proliferation of PANC-1 cells. These results suggest that PKDs could be a novel target for the development of therapeutic drugs that restrict the proliferation of pancreatic cancer cells.