HLA diversity in the Russian population assessed by next generation sequencing

Next generation sequencing is used to determine full-length sequences of HLA genes at the 4-field (allelic) resolution. The study was aimed at determining frequency and diversity of HLA alleles in a cohort of blood donors from the Registry of the National Research Center for Hematology who design ated themselves as Russians (including some not routinely typed variations in HLA gene regions). The studied population consisted of 1510 donors. HLA typing was performed by next generation sequencing. Libraries were performed with AllType NGS Amplification Kits (One Lambda, USA) and sequenced using MiSeq (Illumina, USA). Data analysis used the TypeStream Visual Software V2.0.0.68 (One Lambda, USA) and IPD-IMGT/HLA database 3.40.0.1. Arlequin 3.5 software was used for estimation of allele and haplotype frequencies, deviation from Hardy-Weinberg equilibrium. 82 HLA-A, 156 HLA-В and 85 HLA-С alleles were identified with four-field resolution. 45 HLA-DRB1 and 18 HLA-DQB1 alleles were identified with 2-3-field resolution. Considerable HLA diversity was found among the donors self-designated as Russians: the population had large numbers of distinct alleles at each HLA gene, high percentage of alleles (25-32% of HLA class I) were revealed only once. Sufficient numbers of new alleles were registered which are absent in the IPD-IMGT/HLA database. Considerable allelic diversity in Russian population is due to low-incidence alleles. Despite this diversity, the majority of HLA alleles detected at each locus were common. Significant HLA diversity of the donors was connected with a large number of alleles with rare occurrence. The novel alleles identified in our study differed from the known alleles by single nucleotide substitutions. The most common alleles at the four-field level were as follows: A*02:01:01:01 (27.1%), C*07:02:01:03 (13.1%), A*03:01:01:01 (13.0%), B*07:02:01:01 (13.0%), A*01:01:01:01 (11.6%) and C*07:01:01:01/16 (10.4%). The HLA alleles, which are common for Russian populations, are not always common or well-documented alleles in present catalogues. The data obtained in this study may be used as a reference sample for estimation of HLA allele frequencies in Russian population, for proper frequency evaluation of specific alleles when searching donors for allogeneic hematopoietic stem cell transplantation, as well as for association studies between HLA alleles and different diseases, and for research in population genetics

CLINICALLY RELEVANT MINOR HISTOCOMPATIBILITY ANTIGENS FOR RUSSIAN PATIENTS UNDERGOING HEMATOPOIETIC STEM CELL TRANSPLANTATION

Hematopoietic stem cell transplantation (HSCT) from healthy donors is used for blood cancer treatment. Alloreactive graft-versus-host disease (GvHD) is one of the post-transplant detrimental side effects, and the main reason for GVHD after HSCT fully matched for human leukocyte peptide antigens (HLA) presented by HLA molecules on cell surface. These polymorphic peptides, minor histocompatibility antigens (MiHA), arise from any genes, including those expressed at hematopoietic tissues. The latter may lead to the s.c. graft-versus-leukemia effect (GvL), thus preventing relapse of a malignancy. A*02:01 is one of the most frequent HLA alleles for European part of Russia. We assessed frequencies for 20 MiHA-encoded genetic polymorphisms, presented via A*02:01 allele, for plausible bone marrow donors, or hematopoietic stem cells (HSC) from the Donor Registry at Russian National Research Center for Hematology, we have also determined a number of immunogenic mismatches for these 20 MiHA in real donor – recipient pairs. A total of 608 potential donors, 90 donors and 92 recipients were genotyped. Using public data, we have shown that frequencies for MiHA coding genes are most close to appropriate frequencies among the European population. We have calculated probability of MiHA-specific alloimmune response after HSCT: there are chances of 33 and 75% for three or more immunogenic mismatches (IM) for related and unrelated HSCTs, respectively. Real frequencies for immune mismatch in 20 related and 20 unrelated donor – recipient pairs are in accordance with estimated theoretical probabilities. As based on the calculated frequencies, we suggest the LB-NDC80- 1P/A, LB-CCL4- 1T, and HA-1 MiHA to be the most promising minor antigens for targeted cell therapies of hematopoietic tissue malignancies. The data obtained could be used for planning allo-HSCTs in Russian patients

IMPACT OF HLA-DPB1 INCOMPATIBILITY ON THE RESULTS OF ALLOGENEIC HEMATOPOIETIC STEM CELLS TRANSPLANTATION FROM HLA-A-B-C–DRB1-DQB1-COMPATIBLE UNRELATED DONOR

Introduction. An accepted fact in allogeneic hematopoietic stem cells transplantation (allo-HSCT) from unrelated donors is that matching for HLA genes is critical to ensure the best outcomes for patients. The current gold standard is an unrelated donor matched for 10/10 HLA alleles (HLA-A, -B, -C, -DRB1, -DQB1). In the HLA-mismatched setting graft-versus-host disease (GVHD) is increased, and overall survival becomes significantly worse. The importance of HLA-DPB1matching is more controversial.The aim of our study was to evaluate the impact of HLA-DPB1 mismatches on outcome of patients who underwent 10/10 HLA matched unrelated HSCT.Materials and methods. 49 patients treated with allo-HSCT in transplant center of National Research Center for Hematology from 10/10 HLA-matched unrelated donors were included in the study. High-resolution typing for HLA-DPB1 was done by PCR with sequence specific primers (SSP) using Olerup typing kits. HLA-DPB1 permissive/nonpermissive mismatches were examined according to TCE groups. Overall survival, event-free survival and survival without acute graft-versus-host disease were calculated by Kaplan–Meier method, and compared with log-rank test. Proportional hazard Cox models were used for multivariate analysis.Results. The 3-year overall survival after allo-HSCT was 68 %, event-free survival – 51 %, acute GVHD-free survival – 62 %. No significant impacts of HLA-DPB1 disparities on overall, event-free survival and probability of acute graft-versus-host disease were observed. Patients with nonpermissive HLA-DPB1-incompatible donors had a tendency to increased event-free survival. The factors significantly increasing risk of acute GVHD were peripheral blood grafts, advanced-stage disease and male gender of recipients.Conclusion. The factors associated with acute GVHD are peripheral blood grafts, advanced-stage disease and male sex of recipients. For more precise evaluation of impact of HLA-DPB1-incompatibility on results of allo-HSCT further investigations are needed