Human Hepatocyte and Discovery Core

PROJECT SUMMARY/ABSTRACT Gene therapy for hemophilia A holds promise to accomplish a lasting cure. Adeno-associated viral (AAV) gene transfer of clotting factor FVIII to the livers of males can achieve therapeutic correction but this is not sustained. The reasons for hepatotoxicity and the decline in FVIII expression in clinical trials are unclear. The AAV therapies currently under investigation target hepatocytes in the liver. There is, however, a limited understanding of the interactions between the vector and hepatocytes as well as effect of FVIII expression and immune responses. Core B will provide primary human hepatocyte models to address these basic and mechanistic questions related to the biology of AAV and FVIII. The central hypothesis of this proposal is that multiple interconnected features of AAV and FVIII biology limit durability of therapeutic expression and pose serious safety concerns. The objectives of Core B, the Human Hepatocyte and Discovery Core, is to provide three primary human hepatocytes models to study AAV vector and FVIII biology. The first model is primary hepatocyte cultures that retain stable hepatocyte functions in 2-dimensional format. These will be used for in vitro studies. The second model is chimeric mice whose livers are highly repopulated with primary human hepatocytes. These will be used for in vivo AAV studies. And the third model is mice doubly engrafted with hematopoietic stem cells and livers for in vivo studies of human immune cells in the liver. Core B will provide these services to investigators to address major unanswered questions in FVIII biology, gene therapy for hemophilia, liver-directed gene transfer, and molecular and immunobiology of AAV vectors.