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Dissecting the role of TGF-beta type I receptor/ALK5 in pancreatic ductal adenocarcinoma: Smad activation is crucial for both the tumor suppressive and prometastatic function

Abstract

In the present study, we have analysed the effects of transforming growth factor-beta (TGF-β) signaling on the growth behavior of pancreatic carcinoma cells in vitro and on their tumorigenicity in vivo. Ectopic expression of dominant-negative mutants of the TGF-β type II receptor or type I receptor/activin receptor-like kinase 5 (ALK5) in TGF-β-sensitive pancreatic ductal adenocarcinoma PANC-1 cells prevented the TGF-β-induced activation of transfected Smad-responsive reporter genes and growth arrest. The growth-inhibitory effect was mimicked by stable expression of kinase-active ALK5 (ALK5-T204D), and was dependent on ALK5's ability to activate Smad signaling, as a ALK5-derived mutant with an intact kinase domain but deficient in its ability to activate Smads (RImL45) failed to suppress proliferation in the absence of added TGF-β. Moreover, this mutant often displayed opposite effects to those of ALK5-TD and blocked various ligand-induced responses in vitro, indicating that it acts in a dominant-negative fashion to inhibit endogenous wild-type receptors. ALK5-TD-, but not RImL45-TD-transduced cells underwent epithelial-to-mesenchymal transition, exhibited a higher ratio of thrombospondin-1 to vascular endothelial growth factor-A expression and upregulated various metastasis-associated genes. Upon orthotopic transplantation of PANC-1 clones into immunodeficient mice, ALK5-TD, but not RImL45-TD, greatly reduced tumor size and induced the formation of liver metastases in otherwise non-metastatic PANC-1 cells. These results suggest a causal, dominant role for the endogenous Smad2/3 signaling pathway in the tumor suppressor and prometastatic activities of TGF-β in pancreatic tumor cells.

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Abbreviations

ALK5:

activin receptor-like kinase 5

EMT:

epithelial-to-mesenchymal transition

ERK:

extracellular signal-regulated kinase

kd:

kinase-deficient

MAPK:

mitogen-activated protein kinase

MVD:

microvessel density

PAI-1:

plasminogen activator inhibitor-1

PDAC:

pancreatic ductal adenocarcinoma

TGF-β:

transforming growth factor-β

TSP-1:

thrombospondin-1

VEGF:

vascular endothelial growth factor

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Acknowledgements

We thank M Jansen and B Körtge for excellent technical assistance and Drs V Ellenrieder, P Knaus, SE Kern, J Massagué, K Miyazono and YE Zhang for generously providing expression plasmids. Part of this work was supported by a grant from the DFG (UN 128/1–2). Some data of this work are part of the doctoral thesis of SG.

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Correspondence to H Ungefroren.

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Schniewind, B., Groth, S., Sebens Müerköster, S. et al. Dissecting the role of TGF-beta type I receptor/ALK5 in pancreatic ductal adenocarcinoma: Smad activation is crucial for both the tumor suppressive and prometastatic function. Oncogene 26, 4850–4862 (2007). https://doi.org/10.1038/sj.onc.1210272

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