Abstract
BackgroundWe have previously shown in pancreatic ductal adenocarcinoma (PDAC) cells that the SRC inhibitors PP2 and PP1 effectively inhibited TGF-β1-mediated cellular responses by blocking the kinase function of the TGF-β type I receptor ALK5 rather than SRC. Here, we investigated the ability of the clinically utilised SRC/ABL inhibitor dasatinib to mimic the PP2/PP1 effect.MethodsThe effect of dasatinib on TGF-β1-dependent Smad2/3 phosphorylation, general transcriptional activity, gene expression, cell motility, and the generation of tumour stem cells was measured in Panc-1 and Colo-357 cells using immunoblotting, reporter gene assays, RT-PCR, impedance-based real-time measurement of cell migration, and colony formation assays, respectively.ResultsIn both PDAC cell lines, dasatinib effectively blocked TGF-β1-induced Smad phosphorylation, activity of 3TPlux and pCAGA(12)-luc reporter genes, cell migration, and expression of individual TGF-β1 target genes associated with epithelial-mesenchymal transition and invasion. Moreover, dasatinib strongly interfered with the TGF-β1-induced generation of tumour stem cells as demonstrated by gene expression analysis and single cell colony formation. Dasatinib also inhibited the high constitutive migratory activity conferred on Panc-1 cells by ectopic expression of kinase-active ALK5.ConclusionsOur data suggest that the clinical efficiency of dasatinib may in part be due to cross-inhibition of tumour-promoting TGF-β signalling. Dasatinib may be useful as a dual TGF-β/SRC inhibitor in experimental and clinical therapeutics to prevent metastatic spread in late-stage PDAC and other tumours.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-015-0468-0) contains supplementary material, which is available to authorized users.
Highlights
We have previously shown in pancreatic ductal adenocarcinoma (PDAC) cells that the SRC inhibitors PP2 and PP1 effectively inhibited TGF-β1-mediated cellular responses by blocking the kinase function of the transforming growth factor-β (TGF-β) type I receptor activin receptor-like kinase 5 (ALK5) rather than SRC
Dasatinib blocks TGF-β1-induced Smad2/3 activation in PDAC cells In order to test whether dasatinib can inhibit ALK5 function, we measured its effect on TGF-β1-induced C-terminal phosphorylation of Smad2 and Smad3 (p-Smad2C/3C), reflecting their state of activation
Dasatinib blocks TGF-β1-induced reporter gene activity The above data showed that TGF-β1-induced Smad2 and Smad3 activation was sensitive to dasatinib inhibition
Summary
We have previously shown in pancreatic ductal adenocarcinoma (PDAC) cells that the SRC inhibitors PP2 and PP1 effectively inhibited TGF-β1-mediated cellular responses by blocking the kinase function of the TGF-β type I receptor ALK5 rather than SRC. Due to its overexpression in PDAC and its various functions in driving tumour suppression, the non-receptor tyrosine kinase SRC represents a promising biological target in experimental and clinical approaches to treat PDAC [1,2,3]. Dasatinib (N-(2-chloro-6-methyl- phenyl)2-(6-(4-(2-hydroxyethyl)- piperazin-1-yl)-2-methylpyrimidin-4- ylamino)thiazole-5-carboxamide; BMS-354825, Sprycel), a tyrosine kinase inhibitor originally developed against BCR-ABL and SRC [4] and currently used in the treatment of CML [5] and Philadelphia chromosomepositive acute lymphoblastic leukemia (reviewed in [6]), has shown promise in the treatment of various epithelial tumours [6] including pancreatic cancer. A phase II study found that dasatinib as a single-agent did not have clinical activity as first-line therapy in patients with metastatic PDAC [10]
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