Hepatoblastoma, the most common pediatric liver cancer, is tightly linked to excessive Wnt/beta-catenin signaling. Here, we used microarray analysis to identify two tumor subclasses resembling distinct phases of liver development and a discriminating 16-gene signature. beta-catenin activated different transcriptional programs in the two tumor types, with distinctive expression of hepatic stem/progenitor markers in immature tumors. This highly proliferating subclass was typified by gains of chromosomes 8q and 2p and upregulated Myc signaling. Myc-induced hepatoblastoma-like tumors in mice strikingly resembled the human immature subtype, and Myc downregulation in hepatoblastoma cells impaired tumorigenesis in vivo. Remarkably, the 16-gene signature discriminated invasive and metastatic hepatoblastomas and predicted prognosis with high accuracy.

Tbx3 encodes a transcriptional repressor that is important for diverse patterning events during development, and Tbx3 mutation in humans causes the ulnar-mammary syndrome. Here, we describe the identification of Tbx3 in array-based search for genes downstream Wnt/B-catenin that are implicated in liver tumorigenesis. Overexpression of Tbx3 is closely associated with the mutational status of B-catenin in murine liver tumors induced by Myc as well as in human hepatocellular carcinomas and hepatoblastomas. Moreover, Tbx3 transcription is activated by ectopic expression of B-catenin in mouse liver and in human tumor cell lines. Evidence that Tbx3 transcription is directly regulated by B-catenin is provided by chromatin immunoprecipitation and reporter assays. Although HepG2 cells stably transfected with Tbx3 display moderately enhanced growth rate, the dominant negative mutant Tbx3-Y149S drastically inhibits hepatoma cell growth in vitro and in vivo. Moreover, small interfering RNAs (siRNA) directed against Tbx3 inhibit anchorageindependent growth of liver and colon carcinoma cells. We further show that inhibition of Tbx3 expression by specific siRNAs blocks B-catenin-mediated cell survival and renders cells sensitive to doxorubicin-induced apoptosis. Conversely, ectopic expression of Tbx3 inhibits apoptosis induced by B-catenin depletion. Marked overexpression of Tbx3 in a subset of hepatoblastomas is associated with chemotherapyresistant phenotype and unfavorable patient outcome. These results reveal an unsuspected role of Tbx3 as a mediator of B-catenin activities on cell proliferation and survival and as an important player in liver tumorigenesis. [Cancer Res 2007;67(3):901-10]

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