High-Throughput Characterization of 10 New Minor Histocompatibility Antigens by Whole Genome Association Scanning

Abstract

Patients with malignant diseases can be effectively treated with allogeneic hematopoietic stem cell transplantation (allo-SCT). Polymorphic peptides presented in HLA molecules, the so-called minor histocompatibility antigens (MiHA), play a crucial role in antitumor immunity as targets for alloreactive donor T cells. Identification of multiple MiHAs is essential to understand and manipulate the development of clinical responses after allo-SCT. In this study, CD8+ T-cell clones were isolated from leukemia patients who entered complete remission after allo-SCT, and MiHA-specific T-cell clones were efficiently selected for analysis of recognition of a panel of EBV-transformed B cells positive for the HLA restriction elements of the selected T-cell clones. One million single nucleotide polymorphisms (SNP) were determined in the panel cell lines and investigated for matching with the T-cell recognition data by whole genome association scanning (WGAs). Significant association with 12 genomic regions was found, and detailed analysis of genes located within these genomic regions revealed SNP disparities encoding polymorphic peptides in 10 cases. Differential recognition of patient-type, but not donor-type, peptides validated the identification of these MiHAs. Using tetramers, distinct populations of MiHA-specific CD8 + T cells were detected, demonstrating that our WGAs strategy allows high-throughput discovery of relevant targets in antitumor immunity after allo-SCT.

Isolation of Human CD4/CD8 Double-Positive, Graft-Versus-Host Disease–Protective, Minor Histocompatibility Antigen–Specific Regulatory T Cells and of a Novel HLA-DR7–Restricted HY-Specific CD4 Clone

“…Recently, a novel strategy using whole-genome association scanning reported characterization of 10 new HLA class I-restricted minor H Ags (17). However, identification of HLA class IIrestricted minor H Ags has been more difficult.…”

Identification of minor histocompatibility antigens based on the 1000 Genomes Project

“…12,24,25 At the same time, other investigators developed similar approaches, either based on the HapMap databases or using mHag-phenotyped custom panels of EBV-LCLs, which they genotyped for up to one million SNPs (Table 1). 23,26,31 Nonetheless, even the most comprehensive approach could not facilitate the identification of all mHags as, for example, the HapMap database did not contain the SNP encoding mHag UTDP4-1. The extensive analysis of a large adjacent region of SNP rs12551834 based on a rather weak correlation revealed by the HapMap approach was in this case not sufficient to identify the true mHag-encoding SNP.…”

“…[14][15][16][17][18] After the discovery that mHags are products of polymorphic genes, these techniques were replaced by genetic strategies that made use of the correlation of mHag phenotypes with large numbers of genetic variations, derived from databases such as the HapMap Project or self-assembled panels. 12,[19][20][21][22][23][24][25][26][27] Although these genetic techniques proved to be successful and revealed the identity of more than 15 mHags, they all are still limited by the fact that the databases used do not include all existing genetic variations and variation types (i.e. indels and CNVs).…”

Minor histocompatibility Ags: identification strategies, clinical results and translational perspectives

“…For this purpose, many investigators made use of large pedigrees from the CEPH reference family collections, 14 and also self-assembled panels of individuals were used. 15 The mHag phenotypes of these individuals were usually determined by measuring the response of mHag-specific T-cell clones to Epstein-Barr virus-transformed B cells derived from those individuals. In the very first approach, the genomic locus of the mHag was identified by pairwise correlation of the mHag phenotypes with about 32 000 SNPs and microsatellite markers throughout the whole genome.…”