Enzymatic Removal of A-Antigen in a Mouse Model of ABO-Incompatible (ABOi) Transplantation.

The ABO histo-blood group system presents a major barrier in organ transplantation that limits organ availability. A reduction in donor A/B antigens may represent a novel approach to allow safe ABOi transplantation. The utility of FpGalNAc deacetylase and FpGalactosaminidase (Azymes) to convert blood group A-antigen to H-antigen in an ex vivo human lung perfusion model was recently demonstrated. However, as A-glycosyltransferases are constitutively expressed, the duration of A-antigen removal remains unclear. A-transgenic (Tg) mice developed by our group constitutively express A-antigen on vascular endothelium and erythrocytes and have been used to model ABOi Tx. Using this model, we assessed A-antigen removal and re-expression following Azyme administration in vitro and in vivo. Azyme function was studied using A-Tg BALB/c (male n=5, female n=3) and A-Tg C57BL/6 (male n=2, female n=3) mice (10-44 wk). In vitro : Red blood cells (RBC) were assessed for A- and H-antigen expression by hemagglutination at various times (0.5-4 hr) post-Azyme treatment (2-50 µg/mL) of whole blood. In vivo : A-Tg mice were injected iv with Azyme (0.4-0.8 mg/kg); blood was sampled at various times and RBC were tested for A-antigen by flow cytometry and for A- and H-antigens by hemagglutination. Heart and lung tissue were harvested at various times (1-96 hr) and assessed for expression of A- and H- antigens by immunohistochemistry. In vitro , Azyme cleaved A-antigen from A-Tg mouse RBC, resulting in H-antigen. In vivo , Azyme treatment resulted in the absence of A-antigen and the appearance of H-antigen on RBC up to 2 hr post-injection, with detection of A-antigen reappearing after 4 hr. Heart and lung tissue showed a reduction in A-antigen and the appearance of H-antigen up to 8-hr post-injection. No significant differences between males and females were observed. Our preliminary data confirm that Azymes effectively remove A-antigen from RBC and heart and lung tissues in a mouse model. Future studies will assess the efficacy of higher doses of Azymes and the resulting timing of A-antigen re-expression. Clinical application of Azyme technology has the potential to expand ABOi organ transplantation, allowing life-saving transplants in individuals for whom compatible organs cannot be found, and use of donated organs otherwise discarded due to lack of compatible recipients. [ABSTRACT FROM AUTHOR]