Diminished Immune Cell Adhesion in Hypoimmune ICAM-1 Knockout Human Pluripotent Stem Cells
Gene edited human pluripotent stem cells (hPSCs) are a promising platform for developing reparative cellular therapies that evade immune rejection. Existing first-generation hypoimmune strategies have used CRISPR/Cas9 editing to modulate genes associated with adaptive immune responses, but have largely not addressed the innate immune cells, such as neutrophils, that mediate inflammation and rejection processes occurring early after graft transplantation. We identified the adhesion molecule ICAM-1 as a hypoimmune target that plays multiple critical roles in both adaptive and innate immune responses post-transplantation. In our experiments, we found that ICAM-1 blocking or knock-out (KO) in hPSC-derived cardiovascular therapies imparted significantly diminished binding of multiple immune cell types. ICAM-1 KO resulted in diminished T cell proliferation responses in vitro and in longer in vivo retention/protection of KO grafts following immune cell encounter in NeoThy humanized mice. The ICAM-1 KO edit was also introduced into existing first-generation hypoimmune hPSCs and prevented immune cell binding. This promising hypoimmune editing strategy has the potential to improve transplantation outcomes for regenerative therapies in the setting of cardiovascular pathologies and several other diseases. Overall design: The iPSCs were originally from a thymus donor, and reprogrammed using episomal methods. Wild-type and ICAM-1 KO iPSCs, as well as WT-derived cardiomyocytes (CMs), in biological replicate cell culture wells were harvested for mRNA isolation. Librarys were made and the samples were sequenced on an Illumina sequencer