Hepatoblastoma is the most common liver neoplasm in children and accounts for approximately 1% of all childhood malignancies. Five-year survival for hepatoblastoma is 59-74%; one of the lowest survival rates for childhood cancer and is dictated largely by surgical control. Therefore, an improved understanding of the mechanisms underlying this cancer will enable the development of improved and more targeted clinical therapies. Hepatoblastoma is an embryonal tumor with distinct phenotypes and whose classification and treatment is guided largely by both histology and immunohistochemical staining. The molecular defects present in hepatoblastoma have been identified but their testing and inclusion of targeted therapies into clinical therapy have not been adopted and consequently curative treatment largely consists of surgery alone. Our central hypothesis is that the most common mutation present in hepatoblastoma tumors, CTNNB1 cause differentiation defects, increased cellular proliferation, and tumorigenesis in a cell-specific manner. Therefore, the lack of robust models of hepatoblastoma in a relevant and appropriate cell type has significantly impaired our ability to better understand the molecular underpinnings underlying this cancer and to develop improved and effective clinical therapies. These tumors have relative genetic simplicity, consequently we can readily generate cell lines containing genetically similar and relevant mutations. We propose to use induced pluripotent stem cells (PSC) and PSC derived hepatoblasts and hepatocyte-like cells engineered using CrispR technology to generate mutations in ?-catenin, the most common gene that is mutated in hepatoblastoma. In our preliminary data, we have used CrispR/Cas9 technologies to generate human PSC lines with similar mutations in CTNNB1. These PSC lines have increased activation of WNT signaling but differentiate normally into hepatoblast-like cells, at which point, these cells proliferate and have differentiation defects. Mutant CTNNB1 containing hepatoblasts and hepatocyte-like cells form tumors in vivo. The proposed work will determine the role that mutant CTNNB1 plays in driving hepatoblastoma formation. We will study CTNNB1- mutant containing PSC derived hepatoblasts and hepatocyte-like cells and determine the effect of mutant CTNNB1 on cellular differentiation, proliferation, the transcriptomic and metabolomic signature, and tumorigenesis in vivo. We will then use inducible Cas9 platforms to dynamically generate mutations to determine whether the developmental stage at which CTNNB1 mutations form leads to further downstream alterations in cellular differentiation, phenotype, and tumorigenesis. Finally we will determine whether the timing and order of coincident mutations in addition to the specific types of cells in which the mutations are generated impacts hepatoblastoma formation. The dissection of key regulators of hepatoblastoma formation will enable us to identify and target pathways regulating hepatoblastoma development and growth and will lead to the identification of downstream regulators and clinical targets for hepatoblastoma treatment.
Cancer is the second most common cause of death in children. In particular, hepatoblastoma incidence has nearly doubled; faster than any other childhood cancer with only a five-year survival for hepatoblastoma of 59- 74%. We have developed a new human stem cell derived model to allow for the study of the underlying genetic mutations of hepatoblastoma in clinically relevant and developmentally appropriate human primary cell types and the dissection of key regulators of hepatoblastoma formation and progression will enable us to identify and target pathways regulating hepatoblastoma development and lead to the identification of downstream regulators and clinical targets for hepatoblastoma treatment.