BSHI guideline: HLA matching and donor selection for haematopoietic progenitor cell transplantation.

A review of the British Society for Histocompatibility and Immunogenetics (BSHI) Guideline 'HLA matching and donor selection for haematopoietic progenitor cell transplantation' published in 2016 was undertaken by a BSHI appointed writing committee. Literature searches were performed and the data extracted were presented as recommendations according to the GRADE nomenclature.

The novel HLA-DPB1*571:01 allele characterized by SMRT DNA sequencing in an African Caribbean individual.

A single nucleotide substitution in exon 3 of HLA-DPB1*105:01 results in a new allele, HLA-DPB1*571:01.

A case report-Two manufacturers SAB testing kits can reveal different HLA antibody profiles-Identifying prozone and denatured antigen.

Recent reports have identified that the presence of non-native conformation HLA to which antibody can bind upon Luminex HLA Class I single antigen beads, can vary in levels between different manufacturers kits and that the prozone effect may also be specific to particular products. We present a case in which both prozone and non-native HLA reactive antibodies were observed, which raises important questions on how SAB assays are utilised, especially in the post-transplant monitoring setting. A 56-year old, highly sensitised female patient awaiting a regraft received a HLAi renal transplant. Post-transplant monitoring showed discordant results between two SAB manufacturers assays, with one assay identifying a potential de novo HLA DSA. HLA Class I antibody reactivity was observed which was directed towards the Bw6 public epitope, which is present upon HLA molecules encoded for by numerous HLA-B alleles. However in the day 19 post-transplant sample reactivity spread beyond the Bw6 epitope. To investigate the possibility of a prozone type effect influencing the testing kit the day 19 post-transplant sample was diluted 1:10 with PBS and reanalysed. After dilution the Bw6 reactivity was observed again, however the suspect de novo DSA still persisted. An analysis of the mismatched epitopes identified one manufacturer's assay as being confounded by the presence of denatured reactivity as well as prozone.

Extending the sequences of HLA class I alleles without full-length genomic coverage using single molecule real-time DNA sequencing.

The assignment of an HLA allele name to a sequence requires a comparison between the generated target sequence and a reference sequence on the IPD-IMGT/HLA database. Absence of a full-length reference sequence can result in the inability of HLA typing software to accurately compare and assign the sequence. We sequenced the most frequently seen HLA class I alleles on the Anthony Nolan register present in the database with only a partial genomic sequence, with the aim of increasing the number of complete reference sequences. We successfully extended 95 full-length HLA class I sequences and identified 13 novel variants. Increasing the number of full-length HLA class I reference sequences in the database has aided accuracy of HLA analysis tools for all histocompatibility and immunogenetics laboratories.

Review article: the genetics of the human leucocyte antigen region in inflammatory bowel disease.

BACKGROUND: The human leucocyte antigen (HLA) complex, located at chromosome 6p21.3 is a highly polymorphic region containing the classical class I and II HLA genes. The region is highly associated with inflammatory bowel disease (IBD), largely through genome-wide association studies (GWAS). AIMS: To review the role of HLA in immune function, summarise data on risk/protective HLA genotypes for IBD, discuss the role of HLA in IBD pathogenesis, treatment and examine limitations that might be addressed by future research. METHODS: An organised search strategy was used to collate articles describing HLA genes in IBD, including Crohn's disease and ulcerative colitis. RESULTS: All classical HLA genes with variation (including HLA-A, B, C, DRB1, DQA1, DQB1, DPA1 and DPB1) harbour IBD-associated genotypes. The most implicated gene is HLA-DRB1, with HLA-DRB1*03:01 the most associated risk allele in both Crohn's disease and ulcerative colitis. Elucidating precise disease associations is challenging due to high linkage disequilibrium between HLA genotypes. The mechanisms by which risk alleles cause disease are multifactorial, with the best evidence indicating structural and electrostatic alteration impacting antigen binding and downstream signalling. Adverse medication events have been associated with HLA genotypes including with thiopurines (pancreatitis) and anti-TNF agents (antibody formation). CONCLUSIONS: The HLA complex is associated with multiple risk/protective alleles for IBD. Future research utilising long-read technology, ascertainment of zygosity and integration in disease modelling will improve the functional understanding and clinical translation of genetic findings.

Recipients Receiving Better HLA-Matched Hematopoietic Cell Transplantation Grafts, Uncovered by a Novel HLA Typing Method, Have Superior Survival: A Retrospective Study.

HLA matching at an allelic-level resolution for volunteer unrelated donor (VUD) hematopoietic cell transplantation (HCT) results in improved survival and fewer post-transplant complications. Limitations in typing technologies used for the hyperpolymorphic HLA genes have meant that variations outside of the antigen recognition domain (ARD) have not been previously characterized in HCT. Our aim was to explore the extent of diversity outside of the ARD and determine the impact of this diversity on transplant outcome. Eight hundred ninety-one VUD-HCT donors and their recipients transplanted for a hematologic malignancy in the United Kingdom were retrospectively HLA typed at an ultra-high resolution (UHR) for HLA-A, -B, -C, -DRB1, -DQB1, and -DPB1 using next-generation sequencing technology. Matching was determined at full gene level for HLA class I and at a coding DNA sequence level for HLA class II genes. The HLA matching status changed in 29.1% of pairs after UHR HLA typing. The 12/12 UHR HLA matched patients had significantly improved 5-year overall survival when compared with those believed to be 12/12 HLA matches based on their original HLA typing but were found to be mismatched after UHR HLA typing (54.8% versus 30.1%, P = .022). Survival was also significantly better in 12/12 UHR HLA-matched patients when compared with those with any degree of mismatch at this level of resolution (55.1% versus 40.1%, P = .005). This study shows that better HLA matching, found when typing is done at UHR that includes exons outside of the ARD, introns, and untranslated regions, can significantly improve outcomes for recipients of a VUD-HCT for a hematologic malignancy and should be prospectively performed at donor selection.

HLA Typing for the Next Generation.

Allele-level resolution data at primary HLA typing is the ideal for most histocompatibility testing laboratories. Many high-throughput molecular HLA typing approaches are unable to determine the phase of observed DNA sequence polymorphisms, leading to ambiguous results. The use of higher resolution methods is often restricted due to cost and time limitations. Here we report on the feasibility of using Pacific Biosciences' Single Molecule Real-Time (SMRT) DNA sequencing technology for high-resolution and high-throughput HLA typing. Seven DNA samples were typed for HLA-A, -B and -C. The results showed that SMRT DNA sequencing technology was able to generate sequences that spanned entire HLA Class I genes that allowed for accurate allele calling. Eight novel genomic HLA class I sequences were identified, four were novel alleles, three were confirmed as genomic sequence extensions and one corrected an existing genomic reference sequence. This method has the potential to revolutionize the field of HLA typing. The clinical impact of achieving this level of resolution HLA typing data is likely to considerable, particularly in applications such as organ and blood stem cell transplantation where matching donors and recipients for their HLA is of utmost importance.

An overview of HLA typing for hematopoietic stem cell transplantation.

The selection of a related or an unrelated hematopoietic stem cell donor for a patient requires accurate matching of human leukocyte antigen (HLA) genes in order to maximize the beneficial effects of the transplant. There are various different factors a laboratory must consider in order to achieve an HLA type including the number of samples being processed, level of resolution to be achieved, cost of providing the various tests, and turnaround time required. Each method has its advantages and disadvantages, and in most laboratories, a combination of methods may be used.