Minor histocompatibility antigens (MiHA) have long been known to be key factors in the rejection of solid organ allografts and in the development of graft-versus-host-disease (GVHD) following bone marrow transplantation (BMT) in animal models. Recent evidence suggests that MiHA in humans also provoke transplantation immunity and function as targets of GVHD in bone marrow transplants between HLA-identical donor/recipient pairs, necessitating life long pharmacologic immunosuppression of 15-35 percent of such recipients. MiHA have been shown to be peptides derived from normal cellular proteins that are presented by class I and class II MHC molecules and recognized by MHC-restricted T cells that can be isolated from peripheral blood lymphocytes of BMT recipients. However, because of the difficulty in characterizing such antigenic peptides, the nature of MiHA in both humans and mice has remained largely unknown. Little information is currently available on the total number of human mH loci, their allelic forms, population frequencies, and immunogenetic potential. The program announcement to which this application responds recognizes that this is a major problem in understanding the role that these antigens play in human GVHD. The 3 laboratories represented in this application have established a collaboration that has led to the definition of four MiHA in humans, which represents the entire database of such antigens so far elucidated for that species. This application proposes a major expansion of that work in order to dramatically increase the number of identified antigens and the genetic basis for their expression, in order to allow a systematic study of their immunodominance and involvement in GVHD. Accordingly, the specific aims of this proposal are: (1) To generate class I MHC restricted donor anti-host T cell lines and clones from HLA matched bone marrow transplant patients who exhibit GVHD, and to characterize these lines and clones for their anti-host specificity, MHC restriction, phenotype frequencies and tissue specificity; (2) To chemically identify the peptide antigens recognized by these class I MHC restricted T cell clones using peptide extraction, T cell epitope reconstitution, and tandem mass spectrometry; (3) To identify the genes encoding these peptide antigens, establish the basis for differences in antigen expression within the human population, and develop the means to molecularly type for MiHA alleles; (4) To establish whether there are immunodominant antigens among the group of peptide antigens associated with individual class I MHC molecule and whether they are significantly associated with GVHD.
