CHARACTERIZATION OF INDUCED IgG AND IgM XENOANTIBODY RESPONSES TO PIG ENDOTHELIUM IN HUMANS

Abstract

446 The severe shortage of allogeneic organs available for human transplantation has led to the consideration of the use of pigs as organ donors. Human preformed and induced antibody responses to pig xenografts, however, present a major barrier to the successful use of pig organs for human transplantation. We have recently identified the IgVH genes responsible for encoding IgM xenoantibody responses in humans exposed to pig tissues in a bioartificial liver support device. These responses are encoded by Ig genes that are members of the VH3 family and are restricted to genes encoded by the IgHV3-11 and IGHV3-74 germline progenitors. Induced xenoantibody responses in human patients at 10 days post BAL exposure demonstrate a clonal expansion of a VH gene with a unique VDJ gene configuration. We have recently demonstrated that this clonally expanded gene encodes a functional antibody that reacts with the α-gal epitope. In this report we have extended our analysis of the host response to the Ig genes used to encode IgG xenoantibodies. We have generated IgG gene libraries established from lymphocytes of patients at days 0 and 22 following treatment with a bioartificial liver containing pig hepatocytes and endothelial cells. cDNA libraries representing the VH3 family were screened by colony filter hybridization to identify increases in the frequency of expression of specific VH genes encoding α-gal antibodies. Immunoglobulin genes derived from the IGHV3-11 germline progenitor demonstrate an increase in frequency of expression from 2.9% at day 0 to 20% at day 22. A CDR3 specific probe was used to identify an IgM to IgG isotype switch during the maturation of the antibody response. A unique, clonally expanded IGHV3-11 gene identified in IgM clones at day 10 represent the majority of IgG clones expressed at high levels at day 22. The identify of this clone was confirmed by DNA sequencing. IgVH genes encoded by the IGHV3-74 family demonstrate an increase in expression from 0.8% at day 0 to 5.5% at day 22. Genes related to V3-7 germline progenitors do not contribute to the xenoantibody response. These results demonstrate that the xenoantibody response in humans is encoded by IgVH genes restricted to IGHV3-11 and IGHV3-74 germline families. A specific xenoantibody reactive with the α-gal epitope undergoes a clonal expansion and an isotype switch during the maturation of the xenoantibody response. The restricted nature of the humoral response to pig tissues is of potential importance for the development of therapeutic strategies to prevent xenograft rejection.