Developing Universal Blood Type Donor Lungs Using Ex Vivo ABO Enzymatic Treatment

Purpose We explored the feasibility and safety of using recently developed enzymes (FpGalNAc deacetylase and FpGalNase) to remove histo-blood type antigens from donor lungs during ex vivo lung perfusion (EVLP). The enzymes have been reported to efficiently convert Type A whole blood to O (Withers, Nature Microbiology, 2019). Methods Human type A red blood cells (RBCs, n=5/group) and arteries (aorta and pulmonary arteries, n=3/group) were treated with enzyme-containing EVLP solution and lung preservation solution. Human donor lungs (type A) declined for transplantation were treated with the enzymes (1µg/mL) during normothermic EVLP for 1 and 3 h. Lung physiological parameters were monitored throughout treatment. Cell and tissue samples were taken before and after treatment. The distribution and expression levels of blood type antigens were analyzed by flow cytometry and immunohistochemistry. Results The enzymes achieved over 99% antigen removal from RBC within 1h at a dose of 4 µg/mL (Fig1A) in EVLP solution and lung preservation solution. The histo-blood type antigens in human arteries were observed primarily on endothelial cells - blood type A (BTA) antigens co-localized with endothelial marker CD31 (Fig1B, Pre-treatment). In vitro static treatment of arteries demonstrated 92±1% of BTA antigen removal with 1 µg/mL enzyme (Fig1B). Blood type antigen in human lungs were observed on both endothelial and epithelial cells. Within 1h of perfusion, human lungs treated with enzyme had remarkable clearance of intravascular BTA antigens (Fig1C). Similar results were observed after 3h treatment. No acute physiological side effects were observed from the enzymatic treatment. Conclusion Ex vivo enzymatic treatment can efficiently remove blood type antigen in donor lungs without acute side effects. This treatment has potential to be developed as an adjuvant therapy in ABO-incompatible organ transplantation, minimizing the need for recipient antibody removal procedures or augmented immunosuppression.