RESUMEN
The recepteur d'origine nantais (RON) receptor tyrosine kinase is overexpressed and stimulates invasive growth in pancreatic cancer cells, yet the mechanisms that underlie RON-mediated phenotypes remain poorly characterized. To better understand RON function in pancreatic cancer cells, we sought to identify novel RON interactants using multidimensional protein identification analysis. These studies revealed plectin, a large protein of the spectrin superfamily, to be a novel RON interactant. Plectin is a multifunctional protein that complexes with integrin-ß4 (ITGB4) to form hemidesmosomes, serves as a scaffolding platform crucial to the function of numerous protein signaling pathways and was recently described as an overexpressed protein in pancreatic cancer (Bausch D et al., Clin Cancer Res 2010; Kelly et al., PLoS Med 2008;5:e85). In this study, we demonstrate that on exposure to its ligand, macrophage-stimulating protein, RON binds to plectin and ITGB4, which results in disruption of the plectin-ITGB4 interaction and enhanced cell migration, a phenotype that can be recapitulated by small hairpin ribosomal nucleic acid (shRNA)-mediated suppression of plectin expression. We demonstrate that disruption of plectin-ITGB4 is dependent on RON and phosphoinositide-3 (PI3) kinase, but not mitogen-activated protein kinase (MEK), activity. Thus, in pancreatic cancer cells, plectin and ITGB4 form hemidesmosomes which serve to anchor cells to the extracellular matrix (ECM) and restrain migration. The current study defines a novel interaction between RON and plectin, provides new insight into RON-mediated migration and further supports efforts to target RON kinase activity in pancreatic cancer.
Asunto(s)
Movimiento Celular , Hemidesmosomas/metabolismo , Integrina beta4/metabolismo , Neoplasias Pancreáticas/patología , Plectina/metabolismo , Proteínas Tirosina Quinasas Receptoras/metabolismo , Western Blotting , Proliferación Celular , Células Cultivadas , Cromatografía Liquida , Técnica del Anticuerpo Fluorescente , Humanos , Riñón/citología , Riñón/metabolismo , Proteínas Quinasas Activadas por Mitógenos , Neoplasias Pancreáticas/metabolismo , Fosforilación , Transducción de Señal , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Cicatrización de HeridasRESUMEN
PURPOSE: The most common preclinical models of pancreatic adenocarcinoma utilize human cells or tissues that are xenografted into immunodeficient hosts. Several immunocompetent, genetically engineered mouse models of pancreatic cancer exist; however, tumor latency and disease progression in these models are highly variable. We sought to develop an immunocompetent, orthotopic mouse model of pancreatic cancer with rapid and predictable growth kinetics. EXPERIMENTAL DESIGN: Cell lines with epithelial morphology were derived from liver metastases obtained from Kras(G12D/+);LSL-Trp53(R172H/+);Pdx-1-Cre mice. Tumor cells were implanted in the pancreas of immunocompetent, histocompatible B6/129 mice, and the mice were monitored for disease progression. Relevant tissues were harvested for histologic, genomic, and immunophenotypic analysis. RESULTS: All mice developed pancreatic tumors by two weeks. Invasive disease and liver metastases were noted by six to eight weeks. Histologic examination of tumors showed cytokeratin-19-positive adenocarcinoma with regions of desmoplasia. Genomic analysis revealed broad chromosomal changes along with focal gains and losses. Pancreatic tumors were infiltrated with dendritic cells, myeloid-derived suppressor cells, macrophages, and T lymphocytes. Survival was decreased in RAG(-/-) mice, which are deficient in T cells, suggesting that an adaptive immune response alters the course of disease in wild-type mice. CONCLUSIONS: We have developed a rapid, predictable orthotopic model of pancreatic adenocarcinoma in immunocompetent mice that mimics human pancreatic cancer with regard to genetic mutations, histologic appearance, and pattern of disease progression. This model highlights both the complexity and relevance of the immune response to invasive pancreatic cancer and may be useful for the preclinical evaluation of new therapeutic agents.