Ann-Margaret Little

HLA-A alleles and infectious mononucleosis suggest a critical role for cytotoxic T-cell response in EBV-related Hodgkin lymphoma

A proportion of classical Hodgkin lymphoma (HL) is believed to be causally related to infection with the ubiquitous lymphotropic EBV. The determining factors for development of EBV-related HL remain poorly understood, but likely involve immunological control of the viral infection. Accordingly, markers of the HLA class I region have been associated with risk of EBV-related HL. To study the host genetic component of EBV-related HL further, we investigated the lymphomas association with HLA-A*01 and HLA-A*02 simultaneously in the setting of infectious mononucleosis (IM), a risk factor for EBV-related HL, in a case-series analysis including 278 EBV-related and 656 EBV-unrelated cases of HL. By logistic regression, HLA-A*01 alleles [odds ratio (OR) per allele, 2.15; 95% CI, 1.60–2.88] were associated with increased and HLA-A*02 alleles (OR per allele, 0.70; 95% CI, 0.51–0.97) with decreased risk of EBV-related HL. These allele-specific associations corresponded to nearly 10-fold variation in risk of EBV-related HL between HLA-A*01 and HLA-A*02 homozygotes. History of IM was also associated with risk of EBV-related HL (OR, 3.40; 95% CI, 1.74–6.66). The association between history of IM and EBV-related HL was not seen in the presence of HLA-A*02 because this allele appeared to neutralize the effect of IM on EBV-related HL risk. Our findings suggest that HLA class I-restricted EBV-specific cytotoxic T-cell responses and events in the early immune response to EBV infection in IM play critical roles in the pathogenesis of EBV-related HL.

Major histocompatibility complex class I-related chain A allele mismatching, antibodies, and rejection in renal transplantation

Even when kidney allografts are well matched for human leukocyte antigen (HLA) and anti-HLA antibodies are not detected, graft rejection can still occur. There is evidence that some patients who lose their graft have antibodies specific for major histocompatibility complex (MHC) class I-related chain A (MICA) antigens. We investigated whether mismatching MICA alleles associates with MICA antibody production and graft rejection or dysfunction. MICA and HLA antibody screening in 442 recipients was performed, and specificities were confirmed in a subgroup of 227 recipients using single-antigen multiplex technology. For assignment of MICA antibody specificity, we used three independent assays. In addition, MICA alleles of 227 recipients and donors were determined by DNA sequencing. In all, 17 patients (7.5%) had MICA antibodies, and 13 patients (6%) developed MICA donor-specific antibodies (DSA). Multivariate analysis revealed MICA mismatching, as an independent significant factor associated with the presence of MICA antibodies (p = 0.009), and 14 mismatched MICA residues significantly correlated with MICA antibody production. MICA and HLA antibodies significantly associated with acute rejection (AR) and MICA DSA and HLA DSA correlated with decreased graft function by univariate and multivariate analysis. We conclude that mismatching for MICA epitopes in renal transplantation is a mechanism leading to production of MICA antibodies that associate with AR and graft dysfunction.

Strategies to work with HLA data in human populations for histocompatibility, clinical transplantation, epidemiology and population genetics: HLA‐NET methodological recommendations

HLA‐NET (a European COST Action) aims at networking researchers working in bone marrow transplantation, epidemiology and population genetics to improve the molecular characterization of the HLA genetic diversity of human populations, with an expected strong impact on both public health and fundamental research. Such improvements involve finding consensual strategies to characterize human populations and samples and report HLA molecular typings and ambiguities; proposing user‐friendly access to databases and computer tools and defining minimal requirements related to ethical aspects. The overall outcome is the provision of population genetic characterizations and comparisons in a standard way by all interested laboratories. This article reports the recommendations of four working groups (WG1‐4) of the HLA‐NET network at the mid‐term of its activities. WG1 (Population definitions and sampling strategies for population genetics’ analyses) recommends avoiding outdated racial classifications and population names (e.g. ‘Caucasian’) and using instead geographic and/or cultural (e.g. linguistic) criteria to describe human populations (e.g. ‘pan‐European’). A standard ‘HLA‐NET POPULATION DATA QUESTIONNAIRE’ has been finalized and is available for the whole HLA community. WG2 (HLA typing standards for population genetics analyses) recommends retaining maximal information when reporting HLA typing results. Rather than using the National Marrow Donor Program coding system, all ambiguities should be provided by listing all allele pairs required to explain each genotype, according to the formats proposed in ‘HLA‐NET GUIDELINES FOR REPORTING HLA TYPINGS’. The group also suggests taking into account a preliminary list of alleles defined by polymorphisms outside the peptide‐binding sites that may affect population genetic statistics because of significant frequencies. WG3 (Bioinformatic strategies for HLA population data storage and analysis) recommends the use of programs capable of dealing with ambiguous data, such as the ‘gene[rate]’ computer tools to estimate frequencies, test for Hardy–Weinberg equilibrium and selective neutrality on data containing any number and kind of ambiguities. WG4 (Ethical issues) proposes to adopt thorough general principles for any HLA population study to ensure that it conforms to (inter)national legislation or recommendations/guidelines. All HLA‐NET guidelines and tools are available through its website http://hla‐net.eu.

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.

HLA-A, -B, -C, -DQB1, and -DRB1,3,4,5 allele and haplotype frequencies in the Costa Rica Central Valley Population and its relationship to worldwide populations.

The human leukocyte antigen (HLA) system is the most polymorphic in humans. Its allele, genotype, and haplotype frequencies vary significantly among different populations. Molecular typing data on HLA are necessary for the development of stem cell donor registries, cord blood banks, HLA-disease association studies, and anthropology studies. The Costa Rica Central Valley Population (CCVP) is the major population in this country. No previous study has characterized HLA frequencies in this population. Allele group and haplotype frequencies of HLA genes in the CCVP were determined by means of molecular typing in a sample of 130 unrelated blood donors from one of the countrys major hospitals. A comparison between these frequencies and those of 126 populations worldwide was also carried out. A minimum variance dendrogram based on squared Euclidean distances was constructed to assess the relationship between the CCVP sample and populations from all over the world. Allele group and haplotype frequencies observed in this study are consistent with a profile of a dynamic and diverse population, with a hybrid ethnic origin, predominantly Caucasian-Amerindian. Results showed that populations genetically closest to the CCVP are a Mestizo urban population from Venezuela, and another one from Guadalajara, Mexico.

Modeling HLA associations with EBV-positive and -negative Hodgkin lymphoma suggests distinct mechanisms in disease pathogenesis

HLA genotyping and genome wide association studies provide strong evidence for associations between Human Leukocyte Antigen (HLA) alleles and classical Hodgkin lymphoma (cHL). Analysis of these associations is complicated by the extensive linkage disequilibrium within the major histocompatibility region and recent data suggesting that associations with EBV‐positive and EBV‐negative cHL are largely distinct. To distinguish independent and therefore potentially causal associations from associations confounded by linkage disequilibrium, we applied a variable selection regression modeling procedure to directly typed HLA class I and II genes and selected SNPs from EBV‐stratified patient subgroups. In final models, HLA‐A*01:01 and B*37:01 were associated with an increased risk of EBV‐positive cHL whereas DRB1*15:01 and DPB1*01:01 were associated with decreased risk. Effects were independent of a prior history of infectious mononucleosis. For EBV‐negative cHL the class II SNP rs6903608 remained the strongest predictor of disease risk after adjusting for the effects of common HLA alleles. Associations with “all cHL” and differences by case EBV status reflected the subgroup analysis. In conclusion, this study extends previous findings by identifying novel HLA associations with EBV‐stratified subgroups of cHL, highlighting those alleles likely to be biologically relevant and strengthening evidence implicating genetic variation associated with the SNP rs6903608.

Identification of Molecular Markers of Delayed Graft Function Based on the Regulation of Biological Ageing

Introduction Delayed graft function is a prevalent clinical problem in renal transplantation for which there is no objective system to predict occurrence in advance. It can result in a significant increase in the necessity for hospitalisation post-transplant and is a significant risk factor for other post-transplant complications. Methodology The importance of microRNAs (miRNAs), a specific subclass of small RNA, have been clearly demonstrated to influence many pathways in health and disease. To investigate the influence of miRNAs on renal allograft performance post-transplant, the expression of a panel of miRNAs in pre-transplant renal biopsies was measured using qPCR. Expression was then related to clinical parameters and outcomes in two independent renal transplant cohorts. Results Here we demonstrate, in two independent cohorts of pre-implantation human renal allograft biopsies, that a novel pre-transplant renal performance scoring system (GRPSS), can determine the occurrence of DGF with a high sensitivity (>90%) and specificity (>60%) for donor allografts pre-transplant, using just three senescence associated microRNAs combined with donor age and type of organ donation. Conclusion These results demonstrate a relationship between pre-transplant microRNA expression levels, cellular biological ageing pathways and clinical outcomes for renal transplantation. They provide for a simple, rapid quantitative molecular pre-transplant assay to determine post-transplant allograft function and scope for future intervention. Furthermore, these results demonstrate the involvement of senescence pathways in ischaemic injury during the organ transplantation process and an indication of accelerated bio-ageing as a consequence of both warm and cold ischaemia.

HLA-A, -B, -DR allele group frequencies in 7007 kidney transplant list patients in 27 UK centres

This observational study aims to determine the HLA specificity frequencies of patients on the UK renal transplant list, which can be used as a resource for those laboratories that support the UK renal transplant programme. Whilst the HLA specificity frequencies may differ from that of the general population, it is the individuals on the transplant list who are in need of a new kidney, which has to be provided from the general population. Any differences in protein allele frequencies between this patient population and the general population are likely to be minimal because of the very large number of patients included. The HLA‐A, ‐B and ‐DR allele group frequencies from 7007 patients on the UK kidney transplant list (August, 2009) were analysed. HLA types had been submitted to NHSBT to register patients on the UK deceased donor kidney waiting list. The data were submitted from 27 different registering centres throughout the UK. Within this data set, 25 different HLA‐A, 50 HLA‐B and 18 HLA‐DR allele groups were present. The most common allele groups at each locus were ‐A2 (phenotype frequency 42.6%), ‐B44 (phenotype frequency 23.3%) and –DR4 (phenotype frequency 29.8%). The least common allele groups at each locus were ‐A19, ‐ A43, ‐B16, ‐B21, ‐B22, ‐B83 and ‐DR5. Reports of HLA frequency (protein allotype) data from populations as large as this are not readily available adding value to this observational study.

Prioritizing renal transplantation based on clinical need: the role of an 'urgent' kidney waiting list.

Dear Editors, In the United Kingdom, deceased donor kidneys are allocated for transplantation according to the National Health Service – Blood and Transplant (NHS-BT) Deceased Donor Organ Allocation Policy [1]. The complex matching algorithm attempts to provide equity of access by prioritizing based on factors such as waiting time, HLA match, blood group and age difference. However, unlike other solid organ transplantation (e.g. liver and heart), there is no opportunity to prioritize for renal transplantation based on clinical need [2,3]. The median waiting time for a deceased donor kidney transplant is 3 years [4]. Most patients with endstage renal disease can be maintained on dialysis until an organ becomes available; however, there is a small subset of patients with precarious vascular access for whom transplantation becomes a priority, without which they may die through vascular access loss [5]. Locally, in the West of Scotland, we recognized a small cohort of patients with precarious vascular access whose lives were threatened by potential access loss. It was acknowledged that these patients might benefit from priority allocation of high Kidney Donor Risk Index (KDRI) organs, which might not be suitable for other recipients. Within the context of national allocation, it remained possible to locally allocate donation after circulatory death (DCD) kidneys from donors ≥50 years old or with other adverse prognostic features or tier E donation after brainstem death (DBD) kidneys preferentially to patients with ‘end-stage’ vascular access (ESVA) (bilateral central venous occlusion, failed or contraindication to peritoneal dialysis and survival deemed by the multidisciplinary team to be <1 year on haemodialysis as a result of predicted access failure). We describe our early experience with this approach. Over a 4-year period, 22 patients with ESVA were identified. Eighteen were transplanted during the study period (nine via the ‘priority’ list, six via the national allocation policy, three live donors). Half of those who were transplanted had cRF >95%. Median donor age was 66 years (range: 7–71 years). All but one were DCD organs. Mean KDRI was 2.5. Median CIT was 10 h (range: 6–18 h). Recipient age and wait time were comparable to the overall transplant cohort. One-year patient and graft survival was 88.9%. Mean eGFR at 1 year was comparable to the general transplant cohort (62.0 13.4 vs. 58.4 20.9 ml/min/1.73 m; P = 0.71). Transplantation in this cohort of patients reduced both the number and length of hospital admission in the year following transplantation (Table 1). In all but one case, the patient, who would have been allocated the kidney according to the national algorithm, was transplanted within the subsequent year. The two patients with ESVA who remain on the ‘priority’ list and have not yet been transplanted both have a cRF of 100% and match score of one. Within the Eurotransplant Zone, there is the option to prioritize a ‘medically urgent’ recipient for a deceased donor kidney [6]. Latterly, the Kidney Advisory Group in the UK has discussed the issue of failing access and a national appeals panel has been created. Individual patients with failing vascular access can be discussed for national prioritization on a case-by-case basis [7]. Some clinicians have criticized this approach, suggesting it promotes or rewards bad vascular access practice. Conversely, our strategy, which permits local flexibility in the allocation of high KDRI kidneys within the confines of the national allocation scheme, encourages uniform vascular access practices within the individual transplant centre without disadvantaging those patients who find themselves in a precarious position. It also permits a tailored approach to providing the right organ for the right patient in this unique patient cohort.