Characterization of the humanized FRG mouse model and development of an AAV-LK03 variant with improved liver lobular biodistribution
Marti Cabanes‐Creus, Renina Gale Navarro, Sophia H.Y. Liao, Suzanne Scott, Rodrigo Carlessi, Ramon Roca-Pinilla, Maddison Knight, Grober Baltazar, Erhua Zhu, Matthew Jones, Elena Denisenko, Alistair R. R. Forrest, Ian E. Alexander, Janina E. E. Tirnitz‐Parker, Leszek Lisowski
Abstract
Recent clinical successes have intensified interest in using adeno-associated virus (AAV) vectors for therapeutic gene delivery. The liver is a key clinical target, given its critical physiological functions and involvement in a wide range of genetic diseases. In the present study, we first investigated the validity of a liver xenograft mouse model repopulated with primary hepatocytes using single-nucleus RNA sequencing (sn-RNA-seq) by studying the transcriptomic profile of human hepatocytes pre- and post-engraftment. Complementary immunofluorescence analyses performed in highly engrafted animals confirmed that the human hepatocytes organize and present appropriate patterns of zone-dependent enzyme expression in this model. Next, we tested a set of rationally designed HSPG de-targeted AAV-LK03 variants for relative transduction performance in human hepatocytes. We used immunofluorescence, next-generation sequencing, and single-nucleus transcriptomics data from highly engrafted FRG mice to demonstrate that the optimally HSPG de-targeted AAV-LK03 displayed a significantly improved lobular transduction profile in this model.