Genome Editing With TALEN, CRISPR-Cas9 and CRISPR-Cas12a in Combination With AAV6 Homology Donor Restores T Cell Function for XLP

• DOI: 10.3389/fgeed.2022.828489
• Publication Date: 2022-05-23T04:57:32Z
• Authors: Benjamin C. Houghton; Neelam Panchal; Simone A. Haas; Kay O. Chmielewski; Markus Hildenbeutel; Thomas Whittaker; Claudio Mussolino; Toni Cathomen; Adrian J Thrasher; Claire Booth
• Keywords: Homology directed repair

X-linked lymphoproliferative disease is a rare inherited immune disorder, caused by mutations or deletions in the SH2D1A gene that encodes an intracellular adapter protein SAP (Slam-associated protein). essential for mediating several key processes and system - T cells particular are dysregulated its absence. Patients present with spectrum of clinical manifestations, including haemophagocytic lymphohistiocytosis (HLH), dysgammaglobulinemia, lymphoma autoimmunity. Treatment options limited, patients rarely survive to adulthood without allogeneic haematopoietic stem cell transplant (HSCT). However, this procedure can have poor outcomes mismatched donor setting presence active HLH, leaving unmet need. Autologous haematopoeitic therapy may offer alternative treatment options, removing need find suitable HSCT any risk alloreactivity. has tightly controlled expression profile conventional lentiviral delivery platform not be able fully replicate. A editing approach could preserve more endogenous regulatory elements govern expression, potentially providing optimum therapy. Here, we assessed ability TALEN, CRISPR-Cas9 CRISPR-Cas12a nucleases drive targeted insertion cDNA at first exon locus using adeno-associated virus serotype 6 (AAV6)-based vector containing template. All nuclease platforms were capable high efficiency editing, which was optimised serum-free AAV6 transduction protocol. We show from XLP corrected tools restored physiological levels restore SAP-dependent functions, indicating new therapeutic opportunity patients.