Mutational landscape of hepatoblastoma goes beyond the Wnt-β-catenin pathway
Authors: Ze-Guang Han
Journal: Hepatology (2014)

Hepatoblastoma (HB) is a common pediatric liver cancer that could originate from embryonal and fetal liver cells.1 The etiology of HB is mostly unclear; however, it is well known that certain inherited conditions including familial adenomatous polyposis (FAP) and Beckwith-Wiedemann Syndrome, as well as congenital anomalies and low birth weight, may dramatically raise the risk of HB. The unique clinical features of HB suggest that developmental factors as well as the interactions between genetic and environmental elements may affect the self-renewal and differentiation of hepatic stem/progenitor cells, and lead to the malignant transformation during embryogenesis. However, the exact mechanism by which normal hepatic development is malignantly transformed to HB remains obscure. Previous investigations have uncovered a number of genetic aberrations in HB.2 FAP cases with HB are caused by heritable germline mutation of tumor suppressor gene adenomatous polyposis coli (APC). However, the unique germline APC mutations are rare in sporadic HB cases. In sporadic HB cases, the most prevalent somatic mutations involve the genes encoding components of the Wnt-β-catenin pathway which play a critical role in embryonic and fetal liver development.3 In addition to the low APC mutations found in HB cases, the most common recurrent mutations occur in CTNNB1, which encodes β-catenin, a key component of the Wnt-β-catenin pathway. The occurrence of genetic aberrations, including point mutations and in-frame deletion of the third exon in CTNNB1, has been reported as to be high as 75% of HB tumors. Moreover, the loss-of-function somatic mutations of tumor suppressor genes AXIN1 and AXIN2 whose products affect the β-catenin degradation have also been reported in some HB (Fig. 1). In normal cells, the N-terminal serine and threonine residues of β-catenin encoded by exon 3 of CTNNB1 are phosphorylated by the APC/AXIN/GSK3b complex, and result in β-catenin ubiquitination and subsequent degradation. The constitutive activity of Wnt-β-catenin pathway, which is caused by the gain-of-function mutations of CTNNB1 or loss-of-function mutations of APC and AXIN1/2, leads to the cytoplasmic and nuclear accumulation of β-catenin, and then activates the expression of many target genes, including cyclin D1 for cell proliferation, by the binding of β-catenin to the transcription factors TCF/LEF. In addition to the Wnt-β-catenin pathway, based on the conventional Sanger sequencing of selected genes, the acquired somatic mutations of the components in other signaling pathways are infrequently found in sporadic HB cases. A hypothetical schematic showing the aberrant signaling pathways triggered by newly found somatic mutations in hepatoblastoma. The candidate oncogenes or cancer-promoting genes are shown in red boxes; potential tumor suppressors are shown in green boxes. The predicted gain-of-function and loss-of-function mutations are labeled by red and green lightning symbols with new mutational sites, respectively. Whole genome and exome sequencing have been successfully applied in uncovering the somatic mutations in malignancies, including hepatocellular carcinoma (HCC)4 and intrahepatic cholangiocarcinoma (ICC).5 Significantly, in this issue of Hepatology, Jia et al.6 performed exome sequencing on six sporadic HB cases, and exhibited the first mutational landscape of pediatric liver cancer. They found 144 somatic mutations, including 28 nonsynonymous mutations, with an average of 4.67 nonsilenced mutations per case. The numbers of somatic mutations in the pediatric HB are far less than those in adult HCC and ICC, implying that a few driver mutations may initiate the malignant transformation of HB in embryonal and fetal livers. In this study, they confirmed 24 nonsynonymous mutations within 21 genes by Sanger sequencing. As expected, the point mutations of CTNNB1 exist in two of the six HB tumors. These point mutations include a previous uncharacterized mutation (G512V), located near the armadillo repeat domain of β-catenin. The domain of β-catenin can interact with many binding partners that affect its stability and localization. Like other known CTNNB1 mutations, the G512V is documented as a gain-of-function mutation that significantly promotes proliferation of HB cells by activating β-catenin (Fig. 1). Interestingly, although APC, AXIN1 and AXIN2 are not mutated, two hemizygous mutations (R968H/S969C) of CAPRIN2, whose product can enhance the canonical Wnt-β-catenin pathway by regulating LRP5/6 phosphorylation,7 were found in a single HB case. The R968H/S969C mutant of CAPRIN2 is evidently proven as gain-of-function mutations to activate the Wnt-β-catenin pathway (Fig. 1). Significantly, in addition to the Wnt-β-catenin pathway, the mutational landscape of HB also suggests that the ubiquitin ligase complex could be involved in HB pathogenesis. The acquired genetic mutations of SPOP, KLHL22, TRPC4AP, and RNF169, encoding the components of ubiquitin ligase system, were identified in three of the six HB cases. Among them, SPOP mutations have been found in prostate cancer8 and endometrial cancer.9 In this issue, Jia et al.6 showed that wild-type SPOP inhibits HB cell proliferation by arresting cell cycle progression at the G2 phase by way of up-regulating CDKN2B (p15), a well-known cell growth regulator and tumor suppressor, whereas the S119N mutant of SPOP looses the function. In addition to SPOP mutation in a single case not accompanied by CTNNB1 or CAPRIN2 mutations, Jia et al.6 also showed that SPOP mRNA and protein levels are significantly decreased in HB specimens of two additional cohorts. This suggests that SPOP functions as a tumor suppressor, and its loss-of-function mutations or decreased expression could be a critical event in HB carcinogenesis. SPOP encodes a component of the multisubunit cullin 3 (CUL3)-dependent ubiquitin ligase complex, and the MATH domain within SPOP acts as the substrate recognition for ubiquitin-mediated degradation of the targeted proteins. Known substrates of SPOP involve Gli2 and Gli3,10 which have been known to be the crucial transcription factors of Sonic hedgehog signaling responsible for embryonic development. The newly identified S119N mutant within the MATH domain of SPOP could disrupt SPOP-mediated ubiquitin-proteasomal degradation by affecting substrate recognition, and lead to the constitutive activation of multiple signaling pathways, including Sonic hedgehog signaling by way of Gli2 and Gli3 accumulation, which synthetically contribute to HB carcinogenesis (Fig. 1). KLHL22 also encodes an adaptor of the CUL3-dependent ubiquitin ligase complex. The complex targets the known cancer-promoting factor Polo-like kinase 1 (PLK1) that critically regulates mitosis through its dynamic localization to kinetochores and centrosomes.11 Interestingly, TRPC4AP can bind specifically to the oncoprotein Myc, and suppress Myc-mediated transactivation and transformation in a dose-dependent manner by enhancing Myc ubiquitination and degradation.12 As is known, Myc overexpression can induce HB-like tumors in mice, and plays a critical role in the aggressive phenotype of liver tumors originating from hepatic progenitor cells.3 Whether the TRPC4AP mutation leads to Myc overexpression is worthy of further investigation. It should be pointed out that, like SPOP, TRPC4AP was also mutated in a single HB case without CTNNB1 or CAPRIN2 mutations, implying that SPOP and TRPC4AP mutations may promote HB development by other mechanisms independent of the canonical Wnt-β-catenin pathway (Fig. 1). In general, the first exome sequencing analysis of the pediatric HB tumors identifies a number of novel mutant genes, and provides the platform for a comprehensive understanding of HB tumorigenesis beyond the Wnt-β-catenin pathway (Fig. 1). We should further distinguish the driver mutations enhancing the growth advantage of hepatic stem/progenitor cells from the passenger mutations not affecting the growth advantage of these cells. It has been generally recognized that HB initiation is closely associated with constitutive activation of the Wnt-β-catenin pathway, Myc overexpression, and reprogramming of genomic imprinted genes such as the IGF2-H19 and DLK1-GTL2 regions.2, 3 Therefore, further study should investigate the effect of these mutated genes on the Wnt-β-catenin pathway, Myc, and the imprinted genes. This may provide profound insights into the roles of the driver mutations within these key genes in HB tumorigenesis. Ze-Guang Han, M.D.1,2 1Key Laboratory of Systems Biomedicine (Ministry of Education), Shanghai Center for Systems Biomedicine, Shanghai Jiao Tong University, Shanghai, China; 2Shanghai-MOST Key Laboratory for Disease and Health Genomics, Chinese National Human Genome Center at Shanghai, Shanghai, China.