Genome engineering of primary and pluripotent stem cell-derived hepatocytes for modeling liver tumor formation

ABSTRACT: Genome editing has demonstrated its utility in generating isogenic cell-based disease models, enabling the precise introduction of genetic alterations into wildtype cells to mimic disease phenotypes and explore underlying mechanisms. However, its application in liver-related diseases has been limited by challenges in genetic modification of mature hepatocytes in a dish. Here we conducted a systematic comparison of various methods for primary hepatocyte culture and gene delivery to achieve robust genome editing of hepatocytes ex vivo. Our efforts yielded editing efficiencies of up to 80% in primary murine hepatocytes cultured in monolayer and 20% in organoids. To model human hepatic tumorigenesis, we utilized hepatocytes differentiated from human pluripotent stem cells (hPSCs) as an alternative human hepatocyte source. We developed a series of cellular models by introducing various single or combined oncogenic alterations into hPSC-derived hepatocytes. Our findings demonstrated that distinct mutational patterns led to phenotypic variances, affecting both overgrowth and transcriptional profiles. Notably, we discovered that the PI3KCA E542K mutant, whether alone or in combination with exogenous c-MYC, significantly impaired hepatocyte functions and facilitated cancer metabolic reprogramming. In conclusion, our study demonstrates genome-engineered hepatocytes as valuable cellular models for understanding hepatocarcinoma (HCC), especially highlighting PIK3CA and c-MYC—key oncogenes often amplified or upregulated in HCC—and providing insights into early tumorigenesis mechanisms.

ORGANISM(S): Homo sapiens

PROVIDER: GSE275241 | GEO | 2024/08/25

REPOSITORIES: GEO