Clinical significance and functional role of nuclear met in hepatocellular carcinoma

Abstract

Hepatocellular carcinoma (HCC) is the fifth most prevalent cancer and the third most common cause of cancer mortality worldwide. Understanding the underlying mechanisms that contributed to hepatocarcinogenesis would certainly provide more effective therapeutic options as the prognosis of HCC remains poor. Met/HGF signaling, implicated in various carcinomas including HCC, is an attractive target for human cancers. Indeed, therapeutic strategies against Met surface receptor are currently in clinical trials. The presence of Met inside nucleus will undoubtedly challenge the old-school idea of receptor tyrosine kinase signaling always occurred at the membrane surface, suggesting a novel “signaling shortcut”.

In this study, we attempted to provide a comprehensive understanding regarding the involvement of nuclear Met in HCC by focusing on three major goals; first, the expression and clinical relevance of nuclear Met in HCC; second, the functions of nuclear Met in HCC tumourigenesis and progression; and finally, the underlying signaling network mediated by nuclear Met.

The expression of nuclear Met was examined in human HCC cell lines and tissue samples. Nuclear Met was shown to be overexpressed in HCC tumorous tissues than the corresponding non-tumorous tissues. Moreover, its expression increased progressively along HCC development, from non-tumorous liver, cirrhotic liver to early and advanced HCC. Besides, nuclear Met overexpression was associated with venous invasion and poorer overall survival in HCC patients. We then confirmed that nuclear Met, at a size of around 48 kDa, only consists of the cytoplasmic fragment of full length Met as it could only be detected by Met antibody that directed against the cytoplasmic domain but not the extracellular domain of Met. Subsequent cellular fractionation and immunofluorescence microscopy confirmed that this 48 kDa fragment of Met was present in the nucleus in a ligand-independent fashion. We also identified a region preceding the tyrosine kinase domain after juxtamembrane region (P1027-L1157) is necessary for the translocation of nuclear Met.

Nuclear Met overexpressing stable clones were established to functionally characterize the nuclear Met in HCC by subjecting stable clones to various functional studies. Collectively, we demonstrated the promoting effect of nuclear Met in in vitro cell proliferation, anchorage independent growth, motility and invasiveness. Moreover, nuclear Met overexpression also enhanced the in vivo tumourigenicity of HCC cells. Our data provided the first compelling evidences of an oncogenic function of nuclear Met in HCC.

To elucidate the underlying mechanism mediated by nuclear Met in promoting HCC tumourigenesis and aggressiveness, we performed luciferase reporter assay and showed the activation of NF-κB reporter by nuclear Met expression. Subsequent inhibition of IKKβ and p65 activity, two important effectors of NF-κB signaling, abolished the promoting effect of nuclear Met on HCC cells motility and invasiveness. Also, quantitative polymerase chain reaction analysis revealed that MMP-2 might be involved in this nuclear Met mediated NF-κB signaling to confer the aggressiveness to HCC cells.

Though we could not yet prove cleavage of Met, our study has provided novel insights into the functional role of nuclear Met in HCC and warranted further investigation into other potential role of nuclear Met in carcinogenesis.