Aim: In xenotransplantation, gene knock-out approaches have avoided hyper acute rejection but reports still provide evidence of acute rejection. Therefore, we focused on antigens of Non-HLA antibodies associated with rejection along with endothelial cell surface proteins. If pig proteins are perceived as foreign, human recipient immune cells can attack the xenograft. Non-HLA antigens contribute to antibody generation due to mismatches and leaky tolerance. Our aim was to decipher the extent of sequence and epitope mismatch between pig and human non-HLA antigens as this mismatch can potentially contribute to rejection.
Methods: We analyzed 40 antigens of human and pig for which non-HLA antibodies are reported and sequences of 2 endothelial cell surface proteins. Human and pig protein sequences were from NCBI. Epitopes presented by HLA-A and HLA-B and B-cell epitopes were analyzed using immune epitope database https://www.iedb.org/.
Results: The dissimilarity of 40 antigens between human and pig ranged from 0 to 40%. AT1R antibody has been observed in rejection. Human and pig AT1R sequences were 3.62% dissimilar, had 10 epitope mismatches and 4 B-cell epitope mismatches. We then analyzed antigens showing high sequence dissimilarity and presence of antibody observed in our patient population. Antibody to SSB was frequently seen in our patients and SSB has 23% sequence dissimilarity, 17 epitope mismatches and 8 B-cell epitope mismatches. Antibody to IFNG was also often observed. Human and pig IFNG has 40.36% dissimilarity, 14 epitope mismatches among class I presenting epitopes and 5 B-cell epitope mismatches. Additionally, we analyzed dissimilarity in human and pig endothelial cell surface marker PECAM1 which had 28.53% dissimilarity, 56 epitope mismatches and 28 B-cell epitope mismatches. Another marker; Endoglin had 30.45% dissimilarity, 40 mismatched epitopes and 20 B-cell epitopes. These results suggest a possible immune response to pig antigens and cell surface proteins.
Conclusion: Xenograft is prone to rejection. Therefore, it is critical that diagnostic tests be designed to predict rejection and enable timely immunosuppression. We have identified potential drivers of rejection based on human and pig antigen epitope mismatches. Our results suggest that as the xenotransplantation advances, non-HLA antibody and cell surface antibody detection will be necessary to realize life-saving promise of pig organs long-term.
