Correction of a Factor VIII genomic inversion with designer-recombinases
Felix Lansing, L. R. Mukhametzyanova, Teresa Rojo Romanos, Kentaro Iwasawa, Masaki Kimura, Maciej Paszkowski‐Rogacz, Janet Karpinski, Tobias Graß, Jan Sonntag, Martin Schneider, Ceren Güneş, Jenna Hoersten, Lukas Theo Schmitt, Natalia Rodríguez‐Muela, Ralf Knöfler, Takanori Takebe, Frank Buchholz
Abstract
Despite advances in nuclease-based genome editing technologies, correcting human disease-causing genomic inversions remains a challenge. Here, we describe the potential use of a recombinase-based system to correct the 140 kb inversion of the F8 gene frequently found in patients diagnosed with severe Hemophilia A. Employing substrate-linked directed molecular evolution, we develop a coupled heterodimeric recombinase system (RecF8) achieving 30% inversion of the target sequence in human tissue culture cells. Transient RecF8 treatment of endothelial cells, differentiated from patient-derived induced pluripotent stem cells (iPSCs) of a hemophilic donor, results in 12% correction of the inversion and restores Factor VIII mRNA expression. In this work, we present designer-recombinases as an efficient and specific means towards treatment of monogenic diseases caused by large gene inversions.