LAUSR

Our group has previously established several inducible hepatocellular carcinoma (HCC) models in zebrafish by transgenic expression of selected oncogenes. One common and important feature of these liver tumor models is the reversibility of tumor after suppression of oncogene expression by removing chemical inducer. This phenomenon could be useful to model the process of targeted cancer therapy in human. However, the question remains unanswered is whether the tumor cells have been eliminated or reverted to normal cells. For this, Cre/loxP based lineage tracing was carried out during tumor regression. Thus a new transgenic line, Tg(fabp10: loxP-EGFP-stop-loxP-DsRed; TRE: CreER) (abbreviated to CreER) was established and crossed the xmrk tumor line Tg(fabp10: rtTA; TRE: xmrk; krt8: EGFP). In the CreER/xmrk double transgenic fish under no induction, normal hepatocytes were GFP+. Doxycycline treatment activated xmrk oncogene and transformed hepatocytes into tumor cells that also expressed CreER; subsequent treatment of 4-hydroxytamoxifen activated Cre-mediated loxP recombination and irreversibly labeled tumor cells with RFP expression. In the established HCC of the CreER/xmrk fish, a labeling efficiency of N90% was achieved for the tumor cells. During tumor regression after removal of doxycycline, restoration of normal liver was contributed by both reversion of tumor cells (indicated by the presence of RFP+ hepatocytes) and differentiation of progenitor cells (indicated by the increase of GFP+ hepatocytes). Furthermore, transcriptome analysis of the fully restored liver suggested that the RFP+ hepatocytes from the tumor cell lineage and the GFP+ hepatocytes from the progenitor cells shared very similar gene expression profile. The deregulated genes in tumor cells including the cancer hallmarks genes and metabolism genes have returned to normal level of expression in the tumor-reverted hepatocytes. Thus, our lineage tracing studies demonstrated the possibility of transformed tumor cells to revert to normal cells after the suppression of expression of a primary oncogene.