Zinc Fingers and Homeoboxes 2 is Required for Diethylnitrosamine-induced Liver Tumor Formation in C57BL/6 Mice

Liver cancer, comprised mainly of hepatocellular carcinoma (HCC), is the third leading cause of cancer deaths worldwide and increasing in Western countries. We previously identified the transcription factor Zinc Fingers and Homeoboxes 2 (Zhx2) as a regulator of hepatic gene expression, and many Zhx2 target genes are dysregulated in HCC. Here, we investigate HCC in Zhx2-deficient mice using the Diethylnitrosamine (DEN)-induced liver tumor model. Our study using whole-body Zhx2 knock-out ( Zhx2 KO ) mice revealed the complete absence of liver tumors 9 and 10 months after DEN exposure.Analysis soon after DEN treatment showed no differences in expression of the DEN bioactivating enzyme CYP2E1 and DNA polymerase delta 2, or in the numbers of γH2AX foci between Zhx2 KO and wild-type ( Zhx2 wt ) mice. The absence of Zhx2, therefore, did not alter DEN bioactivation or DNA damage. Zhx2 KO livers showed fewer positive foci for Ki67 staining and reduced IL-6 and AKT2 expression compared to Zhx2 wt livers, suggesting that Zhx2 loss reduces liver cell proliferation and may account for reduced tumor formation. Tumors were reduced but not absent in DEN-treated liver-specific Zhx2 knock-out mice, suggesting that Zhx2 acts in both hepatocytes and non-parenchymal cells to inhibit tumor formation. Analysis of data from The Cancer Genome Atlas and Clinical Proteomic Tumor Consortium indicated that ZHX2 mRNA and protein levels were significantly higher in HCC patients and associated with clinical pathological parameters. Conclusions In contrast to previous studies in human hepatoma cell lines and other HCC mouse models showing that Zhx2 acts as a tumor suppressor, our data indicate that Zhx2 acts as an oncogene in the DEN-induced HCC model and is consistent with the higher ZHX2 expression in HCC patients.