A Novel Zebrafish Liver-Specific Metastasis Model Reveals c-Met as a Driver of Liver Tropism

Abstract

Background and aims: Intrahepatic metastasis negatively impacts the prognosis of several cancers, including hepatocellular and colorectal carcinoma. Zebrafish larval xenografts serve as a robust vertebrate platform that allows direct visualisation of tumour behaviour within a living organism. However, organ-specific metastasis models in zebrafish remain limited, and liver metastasis has not yet been demonstrated. This study aimed to establish a zebrafish larval intrahepatic metastasis model and to determine the role of c-Met activation in mediating liver tropism of cancer cells.

Methods: An intravenous injection-based zebrafish model (zLiverMet) was developed using a liver-specific fluorescent reporter line to visualise tumour colonisation in vivo. Liver cancer cell lines with distinct c-Met expression and activation levels were injected into 2-day post-fertilisation larvae. The effects of c-Met overexpression and pharmacological inhibition on intrahepatic metastasis were analysed through confocal imaging and quantitative image measurements. The model's applicability was further tested using colorectal cancer (CRC) cell lines.

Results: Intravenous injection facilitated efficient intrahepatic colonisation, whereas yolk-sac injection failed to reproduce vascular dissemination. Liver cancer cell lines with high c-Met expression, SNU-449, Mahlavu and SK-HEP-1, exhibited strong liver tropism, while cell lines with no/low c-Met expression, HuH-7 and SNU-398, showed minimal hepatic metastasis. Overexpressing c-Met increased liver colonisation of HCC cells, and inhibiting c-Met activation with the c-Met inhibitor SU11274 reduced this effect.

Conclusions: The zLiverMet model successfully mimics intrahepatic metastasis and highlights c-Met as a driver of liver tropism. This zebrafish-based model offers an 'organism-on-a-chip' platform that is rapid, imageable and scalable-bridging in vitro assays and in vivo models for mechanistic and therapeutic studies of liver metastasis.

Keywords: CTC model; IV injection; hepatocellular carcinoma; liver‐specific metastasis; zLiverMet; zebrafish.