Friday October 11, 2019 from 17:50 to 18:30
Human CTLA4-Ig therapy is associated with false-positive IgG antibody binding to pig cells
Hidetaka Hara1, Qi Li1, Hayato Iwase1, Takayuki Yamamoto1, David Ayares2, David K.C. Cooper1.
1Surgery, University of Alabama at Birmingham, Birmingham, AL, United States; 2Revivicor Inc., Blacksburg, VA, United States
Introduction: Measurement of serum anti-pig antibodies is an important parameter in immune monitoring after pig-to-nonhuman primate xenotransplantation. Pig aortic endothelial cells (pAECs) are commonly used for this purpose. However, human (h) CTLA4-Ig (abatacept/belatacept) can bind to pCD80/86 on the cells, and a secondary antibody (i.e., anti-human IgG) may recognize hCTLA4-Ig (in the absence of recipient serum anti-pig IgG antibody binding to pAECs), potentially leading to misinterpretation of the results. The aim of the present study was to determine whether hCTLA4-Ig binding to pAECs was associated with false-positive results.
Methods: Sera were obtained from (i) naïve baboons (n=3) and (ii) baboons that had undergone pig artery patch transplantation with/without hCTLA4Ig therapy (n=2). Serum IgM and IgG antibody binding to (i) AECs, (ii) red blood cells (RBCs), and (iii) CD3+T cells in peripheral blood mononuclear cells (PBMCs) from an α1,3-galactosyltransferase gene-knockout pig expressing human CD46 (GTKO/hCD46) was measured by flow cytometry in the presence or absence of hCTLA-4Ig. Complement-dependent cytotoxicity (CDC) of wild-type (WT) pAECs by hCTLA4Ig was also measured by flow cytometry.
Results: Sera containing hCTLA4-Ig demonstrated significantly increased IgG (but not IgM) binding to pAECs (relative geometric mean [RGM] = 1.3) compared to sera without hCTLA-4Ig (RGM =1.8) (p<0.01). In contrast, there was no increased binding to pRBCs or CD3+T cells. hCTLA4-Ig binding did in primates result in cytotoxicity of WT pAECs.
Conclusions: pAECs might not be an optimal cell to investigate the development of anti-pig IgG antibodies when hCTLA4-Ig is administered to the recipient, as a false-positive result may be obtained from hCTLA4-Ig binding to the pAECs. CD3+T cells (from PBMCs) would be preferable targets (compared to pRBCs) because they express both carbohydrate and MHC class I/II antigens.
NIAID U19 grant AI090959 .