Attenuated type II TGF-B receptor signalling in human malignant oral keratinocytes induces a less differentiated and more aggressive phenotype that is associated with metastatic dissemination

We examined the effect of stable transfection of dominant negative TbetaR-II (din TbetaR-II) cDNA in a human oral carcinoma cell line that contained normal Ras and was growth inhibited by TGF-betaI. Two clonal cell lines containing dn TbetaR-II were isolated and compared to the vector-only control and parent cell line. The treatment of cells with exogenous TGF-betaI resulted in a decrease in ligand-induced growth inhibition and loss of c-myc downregulation in test cells compared to controls; transcriptional activation of certain genes including fra-I and collagenase was retained. Cells containing dn TbetaR-II grew faster in monolayer culture, expressed less keratin 10 and exhibited increased motility and invasion in vitro compared to control cell lines. Endogenous TGF-betaI production and the regulation of MMP-2 and MMP-9 by TGF-betaI remained unchanged. After orthotopic transplantation to the floor of the mouth in athymic mice, cells containing dn TbetaR-II formed comparable numbers of primary tumours at the site of inoculation as controls but the tumours were less differentiated as demonstrated by the absence of keratin 10 immunostaining. Further, metastatic dissemination to the lungs and lymphatics was more evident in grafts of cells containing dn TbetaR-II than controls. Taken together, the results demonstrate that attenuation of TGF-beta signalling through transfection of dn TbetaR-II cDNA leads to an enhanced growth rate, a loss of tumour cell differentiation and an increase in migration and invasion, characteristics that corresponded to the development of the metastatic phenotype. (C) 2004 Wiley-Liss, Inc.