Scalable manufacturing of human iPSC-derived functional liver organoids for acute liver failure treatment

Rationale: Acute liver failure (ALF) is a life-threatening disease associated with high mortality rates. Human induced pluripotent stem cell (hiPSC)-derived liver organoids (LOs) hold considerable promise as cell-based bridging therapies for ALF. However, the scalable production of highly consistent and mature LOs remains a major barrier to their clinical translation. In this study, we aimed to establish a scalable manufacturing platform for the reproducible generation of highly uniform and functionally mature liver organoids. Methods: We generated LOs from hiPSCs and established a differentiation platform designed for high-density LO production using low-attachment culture vessels combined with spin culture. The hepatic functions of conventionally cultured LOs (Con-LOs) and spin-cultured LOs (Spin-LOs) were evaluated using qPCR, albumin secretion assay, and detoxification assay. The molecular effects of spin culture on LO development were further analyzed using RNA sequencing. Furthermore, microwell bags were introduced to enhance manufacturing scalability. Finally, the LOs were transplanted into the intraperitoneal spaces of acetaminophen (APAP)-induced ALF mice to assess their therapeutic potential. Results: Compared to Con-LOs, Spin-LOs exhibited markedly elevated hepatic gene expression and significantly enhanced liver-specific functions. Mechanistically, spin culture enabled a metabolic transition from glycolysis to oxidative phosphorylation, thereby driving further functional maturation of liver organoids, resulting in improved synthetic, metabolic, and detoxification capacities. Moreover, integrating a microwell bag-based LO formation system with spin-culture-driven LO differentiation further promoted the efficiency and scalability of functional LO manufacturing. Importantly, these LOs provided short-term metabolic support following intraperitoneal transplantation into immunocompetent mice and significantly ameliorated APAP-induced liver failure. Conclusion: This study establishes a scalable manufacturing platform for the reproducible generation of highly uniform and functionally mature liver organoids. Our findings provide an evidence-supported foundation for their future clinical translation as cell-based bridging therapies for ALF.