Humanized six-gene edited porcine erythrocytes in treatment of acute hemorrhagic shock in rhesus monkeys

Background Clinical shortage of blood products is a world problem. Porcine erythrocytes, with many similarities as human erythrocytes, are expected to solve this problem.

Objective To explore the possibility of xenotransfusion of humanized gene-edited porcine (GTKO/β4GalNT2KO/CMAHKO/hCD55/hCD46/hTBM) erythrocytes into rhesus monkeys.

Methods Blood agglutination of six gene-edited porcine erythrocytes (6GE-pRBC) with rhesus monkeys was detected by saline-mediated cross-matching method. The safety of xenotransfusion was verified by IgG and IgM binding assay and complement-mediated cytotoxicity assay. An equal amount of fluorescein isothiocyanate (FITC)-labeled 6GE-pRBC was transfused into rhesus monkeys after 25% of the blood volume was withdrawn from the monkeys, and the survival status in their bodies was monitored by flow cytometry, and the validity of the xenotransfusion was verified by collecting the recipient's blood samples at regular intervals. The possibility of 6GE-pRBC transfusion for the treatment of hemorrhagic shock in rhesus monkeys was observed.

Results The primary and secondary crossmatch blood tests of 6GE-pRBC with rhesus monkeys were negative. The six-gene porcine genotyping conformed to the expected results. The results of IgG and IgM binding assay of rhesus monkey serum with 6GE-pRBC showed low expression of IgG (3.35%) and IgM (4.13%). The complement-mediated cytotoxicity (CDC) of 6GE-pRBC was significantly lower than that of wild porcine erythrocytes (P < 0.000 1). The 6-GE-pRBC survived in rhesus monkeys until 60 h after blood transfusion. Erythrocyte pressure volume was 28.6% after blood loss, and then increased to 35% at 60 h. The hemoglobin content was 92 g/L after blood loss, and then increased to 116 g/L at 60 h and was maintained at the normal pre-blood loss level.

Conclusion 6-GE-pRBC infusion to rhesus monkeys is effective in correcting the symptoms of shock in rhesus monkeys within a certain period of time, and no immune rejection is found. With the continuous innovation of gene editing technology, gene edited porcine erythrocytes are expected to replace human erythrocytes for the treatment of acute hemorrhagic shock in humans.