Correction of a Factor VIII genomic inversion with designer-recombinases

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 a 140 kb genomic inversion of the F8 gene, which is frequently found in patients diagnosed with severe Hemophilia A. Employing substrate-linked directed molecular evolution, we developed a fused 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, resulted in prominent correction of the inversion and restored Factor VIII mRNA expression. Our data suggests that designer-recombinases may represent efficient and specific means towards treatment of monogenic diseases caused by large gene inversions.