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Narcolepsy is hypothesized to be an autoimmune disease targeting the hypocretin/orexin producing neurons within the hypothalamus and is categorised into subsets based on the symptoms presented. Epidemiological studies have proposed that there is an additional requirement for an environmental trigger combined with potential genetic predisposition to trigger the onset of this spectrum of disorders. Despite epidemiological evidence which suggests the condition arises due to a breach of immunological tolerance, the absence of a defined autoantigen remains an element of debate in the aetiology of the disorder. Currently there are no definitive diagnostically useful genetic variants observed in narcolepsy suffers which could be exploited at disease onset to support its diagnosis or clinical managementThe most impactful genetic risk factor is the presence of Human Leukocyte Antigen (HLA) allele, DQB1*06:02, which increases the risk of Narcolepsy over 20-fold. The discovery of the strong association of HLA-DQB1*06:02 and disease onset was the original trigger to shift the focus from its classification as a neurological disease to that of an immune-mediated condition. However, the presence of HLA-DQB1*06:02 alone cannot be the sole genetic risk factor to explain disease onset. Primarily, not all narcolepsy patients express DQB1*06:02 (>95%) and more importantly not all people who express DQB1*06:02 suffer from narcolepsy – only 16% of the global population who express DQB1*06:02 suffer from Narcolepsy. This has resulted in low usage of this definition test in the clinical setting due to its lack of specificity and because it is of most clinical utility if the result is negative, reducing the likelihood of a diagnosis of narcolepsy.The diagnosis of narcolepsy is often suspected from the clinical history of the patient, supported by the results of a multiple sleep latency test (MSLT). Combined with the determination of hypocretin/orexin in the cerebral spinal fluid, these two tests are still considered the gold standard tests for diagnosis. However, there are also documented issues surrounding both tests in terms of level of sensitivity, specificity and potential complications which limit their reliability as a diagnostic resource. This review focuses on emerging evidence of genetic and environmental risk factors linked with narcolepsy onset, their link to its pathophysiology and we speculate how genetic traits could be better utilised to effectively aid a diagnosis.

The action of hematopoietic cell transplantation in controlling leukemia is principally mediated by donor T cells directed against residual recipient malignant cells. However, its utility is limited by graft-versus-host disease (GVHD), where alloreactivity is extended beyond leukemic and marrow cells. In a human/murine chimeric model, we previously showed that the preferential infiltration of cord blood (CB) CD8+ T cells eradicates an Epstein-Barr virus–driven lymphoblastoid tumor without causing xenogeneic GVHD. In the clinic, however, cord blood CD8+ T-cell reconstitution is significantly delayed, and the observation of such a robust antileukemia effect mediated by cord blood CD8+ T cells has not been reported. We describe an observation of very early T-cell expansion in 4 high-risk pediatric leukemia patients receiving third-party, pooled granulocytes after T cell–replete CB transplantation (CBT). The T-cell expansion was transient but robust, including expansion of CD8+ T cells, in contrast to the delayed CD8+ T-cell expansion ordinarily observed after T cell–replete CBT. The CD8+ T cells were polyclonal, rapidly switched to memory phenotype, and had the ability to mediate cytotoxicity. This phenomenon is reproducible, and each patient remains in long-term remission without GVHD. The results suggest that fetal-derived CB CD8+ T cells can be exploited to generate robust antileukemia effects without GVHD.

Susceptibility to viral infection, development of immunity, response to treatment and patient clinical outcomes are all under the control of heritable factors in the host. In the context of the current SARS‐Cov‐2 pandemic, this review considers existing immunogenetic knowledge of virus‐immune system interactions. A major focus is to highlight areas in which work is required in order to improve understanding of antiviral immune responses and to move towards improved patient management.

Human leukocyte antigens (HLA) are present on the surface of all nucleated cells, with the level of expression dependent on the particular HLA locus, the cell type and cellular activation state. Human umbilical vein endothelial cells (HUVECs) are easily isolated from umbilical cords and may aid our understanding of HLA expression on the vascular endothelium in the setting of transplantation. Endothelial cells on the donor-recipient interface form the barrier between transplanted organs and the host immune system. Increased knowledge of the variation in levels of individual HLA specificities may inform the assessment of transplant risk. HUVECs from 48 full term babies born consecutively following planned caesarean section were isolated, HLA typed and grown on gelatin coated culture wells. Once confluent, cells were stimulated with optimal concentrations of the cytokines TNF-α and IFN-γ for 24 hours and HLA-C expression on both unstimulated and stimulated cells was quantified by flow cytometry using the fluorescent labelled monoclonal antibody DT-9 PE. Unstimulated HLA-C expression varied by over 60% between allotypes (ANOVA, P = .004). Following stimulation, HLA-C levels increased over 15-fold and showed the same variation of expression between allotypes (P

Umbilical Cord blood is an intuitively attractive stem cell source, but its use has declined since it is associated with an increased procedure-related morbidity and transplant related mortality. Some of this reflects that cord blood transplants are more often HLA-mismatched compared to other unrelated donor transplants. The ability to transplant in such a setting, indeed without high rates of chronic Graft versus Host Disease (GVHD), constitutes an advantage compared to other unrelated donor cell sources and there are other advantages specifically associated with cord blood as a donor cell source. These advantages must be weighed against its disadvantage, and we have utilised cord blood preferentially as a donor cell source in certain clinical situations in paediatric medicine. In non-malignant diseases, outcomes in metabolic disease are critically dependent on age at transplant and the enzyme delivered by that transplant, and in cord blood transplantation then the time to transplant can be minimised and the engrafted recipients have higher chimerism that delivers higher enzyme levels. In malignant diseases, studies have described reduced relapse rate and better GVHD-free survival, and so we have prioritised cord as a donor cell source where the risk of relapse is highest, and the effects of higher transplant related mortality is most clearly offset by the reduced relapse rates.

Introduction: Kidney dysfunction is a highly significant disease, both in the United Kingdom and globally. Many previous studies have reported associations between human leukocyte antigens (HLA) and renal function; this systematic review attempts to identify, summarize and appraise all published studies of these associations. Methods: A literature search was performed using Medline, Embase and Cochrane Central Register of Controlled Trials to identify papers whose keywords included each of the following concepts: HLA, renal failure and genetic association. A total of 245 papers were identified and assessed for eligibility; 35 of these were included in the final study. Results: A total of 95 HLA types and 14 three-locus haplotypes were reported to be associated with either increased or decreased renal function. A number of these findings were replicated by independent studies that reported 16 types were protective against renal dysfunction and 15 types were associated with reduced renal function. A total of 20 HLA types were associated with both increased risk of renal disease and decreased risk by independent studies. Discussion: There is very little consensus on which HLA types have a protective or deleterious effect on renal function. Ethnicity may play a role, with HLA types possibly having different effects among different populations, and it is possible that the different primary diseases that lead to ESRD may have different HLA associations. Some of the studies may contain type I and type II errors caused by insufficient sample sizes, cohort selection and statistical methods. Although we have compiled a comprehensive list of published associations between renal function and HLA, in many cases, it is unclear which associations are reliable. Further studies are required to confirm or refute these findings.

Umbilical cord blood (UCB) is the preferred donor cell source for children with Hurler syndrome undergoing transplant, and its use has been associated with improved “engrafted survival” rates. However, as in other pediatric recipients of UCB transplants for nonmalignant disease, immune-mediated cytopenia (IMC) is a significant complication. This article describes 8 episodes of IMC in 36 patients with Hurler syndrome undergoing UCB transplant. The incidence of IMC was increased in those with a higher preconditioning absolute lymphocyte count and in those conditioned with fludarabine-containing regimens rather than cyclophosphamide, and it included red cell alloantibodies directed at cord blood group antigens that are novel to the recipient. In several cases, IMC was a precursor to immune-mediated complete graft rejection. We describe IMC as part of a spectrum of graft rejection by a residual competent host immune system and a forme fruste of complete graft rejection.

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.

Human leukocyte antigens (HLA) have been associated with renal function, but previous studies report contradictory findings with little consensus on the exact nature or impact of this observation. This study included 401,307 white British subjects aged 39–73 when they were recruited by UK Biobank. Subjects’ HLA types were imputed using HLA*IMP:02 software. Regression analysis was used to compare 362 imputed HLA types with estimated glomerular filtration rate (eGFR) as a primary outcome and clinical indications as secondary outcome measures. 22 imputed HLA types were associated with increased eGFR (and therefore increased renal function). Decreased eGFR (decreased renal function) was associated with 11 imputed HLA types, seven of which were also associated with increased risk of end-stage renal disease and/or chronic kidney disease. Many of these HLA types are commonly inherited together in established haplotypes, for example: HLA-A*01:01, B*08:01, C*07:01, DRB1*03:01, DQB1*02:01. This haplotype has a population frequency of 9.5% in England and each allele was associated with decreased renal function. 33 imputed HLA types were associated with kidney function in white British subjects. Linkage disequilibrium in HLA heritance suggests that this is not random and particularly affects carriers of established haplotypes. This could have important applications for the diagnosis and treatment of renal disease and global population health.

Human leukocyte antigens (HLA) have been associated with renal function, but previous studies report contradictory findings. There has been a lack of research into how HLA affects renal function in Black, Asian and Minority Ethnic (BAME) people in the UK, despite BAME people being disproportionately affected by renal dysfunction. This study included >27 000 UK Biobank subjects of six ethnicities (>12 100 Irish, >5400 Indian, >4000 Black Caribbean, >3000 Black African, >1600 Pakistani, and > 1400 Chinese) aged 39-73. Subjects' high-resolution HLA genotypes were imputed using HLA*IMP:02 software. Regression analysis was used to compare 108 imputed HLA alleles with two measures of estimated glomerular filtration rate (eGFR): one based on serum creatinine; one based on serum cystatin. Secondary analysis compared CKD stage 2 subjects to healthy controls. Nine imputed HLA alleles were associated with eGFR (adjusted P < 0.05). Six associations were based on creatinine in Black African subjects: HLA-B*53:01 (beta = -2.628, adjusted P = 4.69x10-4 ); C*04:01 (beta = -1.667, adjusted P = 0.0269); DPA1*02:01 (beta = -1.569, adjusted P = 0.0182); and DPA1*02:02 (beta = -1.716, adjusted P = 0.0251) were linked to decreased renal function, while DRB1*03:01 (beta = 3.200, adjusted P = 3.99x10-3 ) and DPA1*01:03 (beta = 2.276, adjusted P = 2.31x10-5 ) were linked to increased renal function. Two of these (HLA-B*53:01 and C*04:01) are commonly inherited together. In Irish subjects, HLA-DRB1*04:01 (beta = 1.075, adjusted P = 0.0138) was linked to increased eGFR (based on cystatin); in Indian subjects, HLA-DRB1*03:01 (beta = -1.72, adjusted P = 4.78x10-3 ) and DQB1*02:01 (beta = -1.755, adjusted P = 2.26x10-3 )were associated with decreased eGFR (based on cystatin). No associations were found in the other three ethnic groups. Nine HLA alleles appear to be associated with kidney function in BAME people in the UK. This could have applications for the diagnosis and treatment of renal disease and could help reduce health inequalities in the UK. This article is protected by copyright. All rights reserved.

We analysed data from 80 patients who tested positive for SARS‐CoV‐2 RNA who had previously been HLA typed to support transplantation. Data were combined from two adjacent centres in Manchester and Leeds to achieve a sufficient number for early analysis. HLA frequencies observed were compared against two control populations: first, against published frequencies in a UK deceased donor population (n = 10,000) representing the target population of the virus, and second, using a cohort of individuals from the combined transplant waiting lists of both centres (n = 308), representing a comparator group of unaffected individuals of the same demographic. We report a significant HLA association with HLA‐ DQB1*06 (53% vs. 36%; p

ObjectiveTo identify whether renal transplant activity varies in a reproducible manner across the year.DesignRetrospective cohort study using NHS Blood and Transplant data.SettingAll renal transplant centres in the UK.ParticipantsA total of 24 270 patients who underwent renal transplantation between 2005 and 2014.Primary outcomeMonthly transplant activity was analysed to see if transplant activity showed variation during the year.Secondary outcomeThe number of organs rejected due to healthcare capacity was analysed to see if this affected transplantation rates.ResultsAnalysis of national transplant data revealed a reproducible yearly variance in transplant activity. This activity increased in late autumn and early winter (p=0.05) and could be attributed to increased rates of living (October and November) and deceased organ donation (November and December). An increase in deceased donation was attributed to a rise in donors following cerebrovascular accidents and hypoxic brain injury. Other causes of death (infections and road traffic accidents) were more seasonal in nature peaking in the winter or summer, respectively. Only 1.4% of transplants to intended recipients were redirected due to a lack of healthcare capacity, suggesting that capacity pressures in the National Health Service did not significantly affect transplant activity.ConclusionUK renal transplant activity peaks in late autumn/winter in contrast to other countries. Currently, healthcare capacity, though under strain, does not affect transplant activity; however, this may change if transplantation activity increases in line with national strategies as the spike in transplant activity coincides with peak activity in the national healthcare system.

The detection and semiquantitative measurement of circulating human leucocyte antigen (HLA)-specific antibodies is essential for the management of patients before and after transplantation. In addition, the pretransplant cross-match to assess the reactivity of recipient HLA antibody against donor lymphocytes has long been the gold standard to prevent hyperacute rejection. Whilst both of these tests assume that recipient HLA-specific antibody is the only variable in the assessment of transplant risk, this is not the case. Transplant immunologists recognize that some HLA antigens are expressed at levels a magnitude lower than others (e.g., HLA-C, HLA-DQ), but within loci, and between different cell types there are many factors that influence HLA expression in both resting and activated cells. HLA is not usually expressed without the specific promoter proteins NLRC5, for HLA class I, and CIITA, for class II. The quantity of HLA protein production is then affected by factors including promoter region polymorphisms, alternative exon splice sites, methylation and microRNA-directed degradation. Different loci are influenced by multiple combinations of these control mechanisms making prediction of HLA regulation difficult, but an ability to measure the cellular expression of each HLA antigen, in conjunction with knowledge of circulating HLA-specific antibody, would lead to a more informed algorithm to assess transplant risk.

In vivo T-cell depletion, using alemtuzumab therapy prior to SCT, can reduce the incidence of GVHD. This treatment has a potential to delay immune reconstitution resulting in increased morbidity due to viral illnesses. We retrospectively analyzed data on all pediatric patients with non-malignant disorders who received alemtuzumab-based conditioning regimens in our center over the last 10 yr (n = 91). Our data show an OS of 91.2%. The incidence of acute (grade 2-4) GVHD was 18.7% and that of chronic GVHD 5.5%. Viremia due to adenovirus, EBV and CMV was seen in 19.8%, 64.8% and 39.6% patients, respectively, with only two deaths attributed to viral infection (adenovirus). Chimerism level at three month was predictive of graft outcome. Nine patients, who had graft failure after first SCT, were salvaged with a second SCT using RIC and same donor (if available). Based on these results, we conclude that the use of in vivo T-cell depletion is safe, achieves good chimerism and does not lead to increased morbidity and mortality due to viral infections. It is associated with a reduced incidence of chronic GVHD.

Background Human embryonic stem cells (hESCs) hold tremendous promise for cell replacement therapies for a range of degenerative diseases. In order to provide cost-effective treatments affordable by public health systems, HLA-matched allogeneic tissue banks of the highest quality clinical-grade hESCs will be required. However only a small number of existing hESC lines are suitable for clinical use; they are limited by moral and ethical concerns and none of them apply Good Manufacturing Practice (GMP) standards to the earliest and critical stages of gamete and embryo procurement. We thus aimed to derive new clinical grade hESC lines of highest quality from fresh surplus GMP grade human embryos. Methods A comprehensive screen was performed for suitable combinations of culture media with supporting feeder cells or feeder-free matrix, at different stages, to support expansion of the inner cell mass and to establish new hESC lines. Results We developed a novel two-step and sequential media system of clinical-grade hESC derivation and successfully generated seven new hESC lines of widely varying HLA type, carefully screened for genetic health, from human embryos donated under the highest ethical and moral standards under an integrated GMP system which extends from hESC banking all the way back to gamete and embryo procurement. Conclusions The present study, for the first time, reports the successful derivation of highest-quality clinical-grade hESC lines from fresh poor-quality surplus human embryos generated in a GMP-grade IVF laboratory. The availability of hESC lines of this status represents an important step towards more widespread application of regenerative medicine therapies. Electronic supplementary material The online version of this article (doi:10.1186/s13287-017-0561-y) contains supplementary material, which is available to authorized users.

Following haematopoietic stem cell transplantation, monitoring the proportion of donor and recipient haematopoiesis in the patient (chimerism) is an influential tool in directing further treatment choices. Short tandem repeat (STR) analysis is a method of chimerism monitoring using DNA isolated from peripheral blood, bone marrow or specific isolated cell lineages such as CD3+ T cells. For lineage-specific STR analysis on cell populations isolated from peripheral blood, a qualitative estimation of the purity of each isolated population is essential for the correct interpretation of the test data. We describe a rapid, inexpensive method for the determination of purity using a simple flow cytometry method. The method described for assessing the purity of sorted CD3+ cells can be applied to any cell population isolated using the same technology. Data obtained were comparable to results from a commercial polymerase chain reaction (PCR)-based method for the assessment of purity (Non-T Genomic Detection Kit, Accumol, Calgary, AB, Canada) (P = 0.59). Of the 303 samples tested by flow cytometry, 290 (95.7%) exceeded 90% purity, and 215 (70.95%) were over 99% pure. There were some outlying samples, showing diversity between samples and the unpredictability of purity of isolated cell populations. This flow cytometry method can be easily assimilated into routine testing protocols, allowing purity assessment in multiple-sorted cell populations for lineage-specific chimerism monitoring using a single secondary antibody and giving results comparable to a PCR-based method. As purity of isolated cell lineages is affected by time after venepuncture and storage temperature, assessment of each sample is recommended to give a reliable indication of sample quality and confidence in the interpretation of the results.

Summary Accurate human leucocyte antigen (HLA) typing results are essential in determining the degree of compatibility between donor and recipient in both solid organ (SO) and hematopoietic stem cell (HSC) transplantation. Current HLA typing methodologies can generate ambiguous results which may need resolving. This group-specific sequencing approach allowed investigation into the presence of the low expressor HLA-A*24:02:01:02L allele and the rare HLA-A*02:64 allele in a SO transplant recipient and a HSC transplant recipient, respectively. Locus-specific amplification of HLA-A was performed. Exons 2 and 3 were sequenced in both directions followed by group-specific sequencing to resolve ambiguities. Hemizygous sequence data of intron 2 generated from the HLA-A*24 allele indicated the presence of the HLA-A*24:02:01:01 allele. HLA-A*02:64 was identified by sequencing the allele in isolation over exons 2 and 3 and allowed confirmation of this allele sequence with the IMGT/HLA database (Accession number AY297166). This approach is cost efficient and can be modified to sequence alleles at other HLA loci. It has also been adapted to characterize the novel HLA-DQB1*06:48 allele (Accession number HE647646) as well as the non-HLA gene, UGT2B17, making it a useful tool to augment existing typing methodologies.

Antibodies to the phospholipase A2 receptor 1 (PLA2R1) have been reported in 70% of cases of idiopathic membranous nephropathy (IMN). The genetic susceptibility of IMN has been accounted for by HLA DQA1 and PLA2R1 genes. Here we retrospectively quantified PLA2R antibodies by ELISA, and genotyped DQ alleles and PLA2R1 single-nucleotide polymorphisms for association with clinical criteria for disease activity at the time of first sample and with outcome over a median total follow-up of 90 months. In 90 prevalent patients with biopsy-proven IMN, anti-PLA2R antibodies were present in 75% of patients with IMN with active disease and were significantly higher than in patients in partial or complete remission at the time of antibody measurement. There was a differential IgG subclass response (4>2>3>1) at an early stage, i.e., within 6 months of biopsy. Levels of PLA2R antibodies were significantly linked to DQA1*05:01 and DQB1*02:01. Survival analysis of patients with IMN showed that PLA2R antibodies are significantly linked with outcome. Thus, high levels of PLA2R antibodies are linked with active disease and a higher risk of declining renal function during follow-up. Future therapeutic trials in IMN should monitor anti-PLA2R, as patients with a high antibody burden may benefit from earlier therapeutic intervention.

Hematopoietic stem cell transplantation (HSCT) is the standard of care in children with Hurler syndrome (HS) as it is the only therapy that can arrest disease progression. We examined the incidence, patterns and outcomes of graft failure in all HS children undergoing first HSCT at the Royal Manchester Children's Hospital or the University of Minnesota Children's Hospital from 1983 to 2016. Implementation of busulfan pharmacokinetic monitoring started in 2004 in both institutions. Two hundred and forty HS children were included in this analysis (historical era (pre-2004), n=131; current era (post 2004), n=109). The proportion of patients with graft failure was significantly lower in the current era compared with the historical era (37.2% vs 10.1%, respectively). Of 49 patients with graft failure in the historical era, 1 had aplasia and 48 had autologous reconstitution. All the 11 graft failures of the current era occurred in recipients of cord blood transplants (7 aplasia and 4 autologous reconstitution). The outcomes of second transplant in these patients has improved, with 89% of such patients alive and engrafted in the current era compared with 58% in the historical era. The pattern of graft failure has changed from autologous reconstitution, likely secondary to inadequate myelosuppression in the historical era, to aplasia in the current era, likely due to imperfect immunosuppression.

A review of the British Society for Histocompatibility and Immunogenetics (BSHI) "Guideline for selection and HLA matching of related, adult unrelated donors and umbilical cord units for haematopoietic progenitor cell transplantation" 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.

Background: Hurler's syndrome is the most severe phenotype of Mucopolysaccharidosis 1 (MPS1H) with severe deficiency of the lysosomal enzyme alpha-L-iduronidase causing heparan and dermatan sulfate accumulation, CNS and somatic manifestations and premature death. Haematopoeitic Stem Cell Transplant (HSCT) alters the disease phenotype and prevents early death, and is standard of care in this condition. The use of an established 'donor hierarchy' recommending Unrelated Cord Blood Transplantation (UCBT) in the absence of a HLA-matched, non-carrier sibling, has resulted in improved engrafted and surviving rates and better post-enzyme levels with improved somatic and CNS clinical outcomes. However, UCBT for malignant and non-malignant conditions has led to high rates of post-transplant autoimmune disease, most frequently Autoimmune Cytopenia (AIC). We report a large cohort of MPS1H patients receiving busulfan-based UCBT and show that AIC is a frequently encountered complication of treatment with significant associated morbidity and we identify important risk factors for its occurrence. Methods: This was a retrospective observational study of patients undergoing first UCBT for Hurler's Syndrome at the Royal Manchester Children's Hospital between 30th September 2004 and 15th March 2018. An episode of AIC was defined as an episode of single or multi-lineage cytopenia occurring post-engraftment, confirmed by the presence of antibodies or diagnosed on the basis of clinical, biochemical and pathologic features. All patients received pK targeted busulfan to achieve myeloablative levels between 80-90 mg/L x hr. Conditioning between 2004 and May 2010 was with Busulfan and Cyclophosphamide (BuCy), and Fludarabine and Busulfan (FluBu) thereafter based on the reported equivocal efficacy and favourable toxicity profile of FluBu conditioning. All patients received serotherapy with Anti-Thymocyte Globulin (ATG) 10mg/kg for In Vivo T-Cell depletion. Timing of ATG was either distal (day -4 to -1) or proximal (day - 9 to -6) in accordance with international consensus at the time. Variables assessed in statistical analysis were conditioning drugs, ATG timing, Absolute Lymphocyte Count pre-conditioning, age-at-transplant, Graft-versus-Host Disease (GVHD) and Total Nucleated Cell dose. Results and Discussion: Thirty-six patients underwent first UCBT for Hurler's Syndrome. There were 8 episodes of AIC (22%), with median onset post-UCBT of 66 days (range 22-236 days). The median number of therapies required was 4 (range 0-8), with duration of illness ranging from 10 to 215 days. There were no deaths, though there were 2 episodes of life-threatening bleeding. Absolute Lymphocyte Count pre-conditioning (p=0.004), FluBu conditioning (p=0.03) and use of proximal-ATG (p=0.016) were significant risk factors for AIC in univariate analysis(see figure 1). Absolute Lymphocyte Count pre-conditioning was the most significant predictor in multivariable analysis (AOR 5.796, CI 0.921-36.474). This confirms that AIC is a frequent, serious but survivable complication of UCBT for Hurler's Syndrome. This data indicates that the balance between recipient immunity and graft immunity impacts AIC risk and severity, mandating a larger international review of registry data to define the frequency of AIC in this population, and an analysis of the risk factors that predispose to it. By better defining these risk factors, we can identify patients in whom conditioning might be altered to reduce risk. Figure 1. Univariate analysis of variables predicting an episode of Autoimmune Cytopenia. ALC (p=0.004), ATG timing (0.016) and Conditioning drugs (p=0.032) were significant risk factors. GVHD= Graft-versus-Host Disease ATG= Anti-Thymocyte Globulin Disclosures Wynn: Orchard SAB: Membership on an entity's Board of Directors or advisory committees; Orchard Therapeutics: Equity Ownership; Chimerix: Research Funding; Genzyme: Honoraria; Bluebird Bio: Consultancy; Orchard Therapeutics: Consultancy; Chimerix: Consultancy.

Aim: This study was undertaken to investigate whether a single G deletion within a series of seven G residues (codon 196) at the exon 9–intron 9 boundary of the enamelin gene ENAM and a tri-nucleotide deletion at codon 180 in exon 7 (GGA vs deletion) of ameloblastin gene AMBN could have a role in autosomal amelogenesis imperfecta among affected Syrian families. Methods: A new technique – size-dependent, deletion screening – was developed to detect nucleotide deletion in ENAM and AMBN genes. Twelve Syrian families with autosomal-dominant or -recessive amelogenesis imperfecta were included. Results: A homozygous/heterozygous mutation in the ENAM gene (152/152, 152/153) was identified in affected members of three families with autosomal-dominant amelogenesis imperfecta and one family with autosomal-recessive amelogenesis imperfecta. A heterozygous mutation (222/225) in the AMBN gene was identified. However, no disease causing mutations was found. The present findings provide useful information for the implication of ENAM gene polymorphism in autosomal-dominant/-recessive amelogenesis imperfecta. Conclusion: Further investigations are required to identify other genes responsible for the various clinical phenotypes.

For certain drugs, pharmacogenetic tests can reduce adverse drug reactions and improve treatment efficacy. However, the adoption of pharmacogenetics into clinical practice has been relatively slow. One potential barrier is the capacity of laboratories to meet the demands of a clinical pharmacogenetic service. We aimed to establish the range, capacity to deliver, and demand for germline pharmacogenetic testing in the United Kingdom and Ireland, through an e-survey of 34 molecular genetics and 28 histocompatibility and immunogenetics (HI) laboratories.Thirty-five percent of molecular genetics laboratories and 54% of HI laboratories responded to the survey. The majority of HI laboratories (93%) offered pharmacogenetic testing, whereas only one molecular genetics laboratory provided a pharmacogenetic service. HLA-B*5701 was most commonly tested to identify those at risk of abacavir hypersensitivity among patients with HIV. A number of barriers to testing were identified, including lack of clinician knowledge and a lack of scientific evidence. All molecular genetics laboratories believed that national coordination of clinical pharmacogenetic services was required, whereas only 50% of HI laboratories supported this view.In the United Kingdom, pharmacogenetic testing is currently being predominantly provided through HI laboratories for a limited number of indications. The number of laboratories offering pharmacogenetic tests is increasing and is likely to continue to increase over the coming years.

CD31 gene polymorphisms are implicated in the pathogenesis of graft-versus-host disease (GvHD) following haematopoietic stem cell transplantation (HST). We investigated the influence of CD31 genotype on the incidence of GvHD following HST from an human leukocyte antigen (HLA)-identical sibling donor. Donor and recipient CD31 codons 125, 563 and 670 DNA polymorphisms were determined in 85 cases of HLA identical sibling HST from two transplant centres. A correlation between CD31 genotype and acute GvHD was considered significant if observed in patients from both transplant centres independently. A strong correlation was identified between donor CD31 codon 125 genotype and the incidence of acute GvHD. Acute GvHD grades II-IV occurred in 27 of 46 (59%) recipients with a CD31 codon 125 leucine / valine heterozygous donor compared to nine of 39 (23%) recipients with a CD31 codon 125 homozygous donor (P=0.0019, relative-risk 2.45, 95% confidence interval 1.3-4.5). This correlation was significant in patients from both transplant centres (P=0.015 and P=0.019). We suggest that CD31 genotype may influence the function of donor-derived leukocytes and may be informative when there is a choice of comparable donors.

This study was undertaken to investigate whether a single G deletion within a series of seven G residues (codon 196) at the exon 9-intron 9 boundary of the enamelin gene ENAM and a tri-nucleotide deletion at codon 180 in exon 7 (GGA vs deletion) of ameloblastin gene AMBN could have a role in autosomal amelogenesis imperfecta among affected Syrian families. A new technique - size-dependent, deletion screening - was developed to detect nucleotide deletion in ENAM and AMBN genes. Twelve Syrian families with autosomal-dominant or -recessive amelogenesis imperfecta were included. A homozygous/heterozygous mutation in the ENAM gene (152/152, 152/153) was identified in affected members of three families with autosomal-dominant amelogenesis imperfecta and one family with autosomal-recessive amelogenesis imperfecta. A heterozygous mutation (222/225) in the AMBN gene was identified. However, no disease causing mutations was found. The present findings provide useful information for the implication of ENAM gene polymorphism in autosomal-dominant/-recessive amelogenesis imperfecta. Further investigations are required to identify other genes responsible for the various clinical phenotypes.

Glomerulonephritis is a significant cause of chronic kidney disease requiring renal replacement therapy. For patients receiving a transplant, it is known that specific primary pathologies such as membranous nephropathy, IgA nephropathy and FSGS have a high risk of recurrence in the transplant but the reasons for this are unknown and the ability to predict recurrence is poor. The recent discovery that primary MN is an autoimmune disease characterised by an autoantibody to phospholipase A2 receptor 1 and the identification of two genes, PLA2R1 and DQA1 which account for the genetic susceptibility to the disease, open up the potential to understand the mechanism of recurrent MN and therefore to design and manage therapy to prevent recurrence. Transplantation offers a unique ethical experimental context in which to explore the genetic contribution to recurrent autoimmune membranous nephropathy. By analysing the genetic changes in the kidney transplant in the context of anti-PLA2R status post transplant, it may be possible to link genetic markers, anti-PLAR regulation with recurrence and non-recurrence of disease. If successful, similar strategies may help unravel mechanisms of recurrent IgA nephropathy and FSGS.

Objectives. To investigate the influence of HLA-DRB1 alleles encoding the QK/RRAA shared epitope (SE) on radiological outcome in rheumatoid arthritis (RA), and to determine whether it is modulated by alleles carrying the putative rheumatoid arthritis-protective (RAP) sequence DERAA. Patients and methods. The association between erosive damage and HLA-DRB1 status was examined in 315 RA patients with a disease duration of 5‐30 yr. Radiological outcome was measured by scoring X-rays of the hands and feet using the standard radiographs of Larsen (Larsen score). HLA-DRB1 typing was carried out using polymerase chain reaction methodology. Results. Patients with two alleles encoding the QK/RRAA SE had significantly higher Larsen scores than SE-negative patients (96.9 vs 83.3; P= 0.04, after correction for multiple testing), with DRB1*0401/*0401 homozygotes demonstrating the greatest radiological damage (99.9). The lowest Larsen score (65.6) was observed in patients carrying the DERAA motif without an accompanying SE allele (RAP+/SE’ ). This was significantly lower than in patients with RAP+/SE+ (105.6; P= 0.04), RAP’/SE’ (88.2; P= 0.05) and RAP’/SE+ (95.8; P= 0.009), after correction for multiple testing. There was no evidence that the RAP sequence was modulating the eVect of the SE since radiological outcome in RAP+/SE+ patients was not significantly diVerent to that in RAP’/SE+ individuals. Conclusions. Our data support a possible role for DRB1 alleles encoding the DERAA motif in protection against severe erosive damage in patients lacking the QK/RRAA SE, but not in patients heterozygous for the SE. This suggests that DRB1 alleles encoding the SE have a dominant influence over ‘protective alleles’ and are not merely ‘non-protective’.

Continuing a project presented at the 15th International HLA and Immunogenetics Workshop (IHIWS) on the rarity of HLA alleles, we sought to expand the number of data sources and bioinformatics tools available in the Allele Frequencies Net Database website (AFND, www.allelefrequencies.net). In this 16th IHIWS Rare Alleles project, HLA alleles described in the latest IMGT/HLA Database (release 3.8.0) were queried against different sources including data from registries (stem cell) and from 74 different laboratories around the world. We demonstrated that approximately 40% of the alleles officially named in the IMGT/HLA Database have been reported only once across all different sources. To facilitate the large-scale analysis of rare alleles, we have produced an online tool called the Rare Allele Detector that simplifies the detection of alleles that are considered to be 'very rare', 'rare' or 'frequent'. Tools and associated data can be accessed via the www.allelefrequencies.net website.

Many new HLA-C locus alleles have recently been identified by DNA sequencing, and a molecular based method for their detection using PCR with sequence specific primers has been reported. However, other methods may be more appropriate for the identification of C locus alleles in larger studies. Here we describe one such system, based on PCR sequence specific oligonucleotide probes, (SSOP) for C locus typing. Advantages of SSOP typing compared to SSP are that it is easier to detect new alleles, more cost effective and less time consuming. We have developed a DNA typing method to identify the broad C locus antigens (including those not yet defined serologically) using a minimum of probes with one amplification. We use a C locus specific sense primer in exon 2 and a consensus antisense primer in exon 3, in a two-step PCR, giving a product of 710 bp. Probes were designed with similar melting temperatures (54-56 degrees C) that would identify as many alleles as possible. The method was established using DNA from B lymphoid cell lines of known C locus type, mostly 10th workshop homozygous cell lines, plus as many other sequenced cell lines as possible. The system was able to correctly identify their C locus types using only 26 probes. DNA was tested from a panel of serologically typed individuals which included many different heterozygous combinations. We found a high concordance of results, with all discrepancies being additional antigens identified by molecular typing, filling in serological blanks. We can identify all common heterozygote combinations using this method.

Summary HLA-DQB1*06:48 has single nucleotide polymorphisms within codons 70 and 62 of exon 2 (GGG>AGG and AAG>AAC) relative to HLA-DQB1*06:02:01 and HLA-DQB1*06:37. This results in amino acid differences (G>R and K>N) that will change the polarity and charge of the encoded antigen and may therefore affect its peptide repertoire.

Summary Acute graft versus host disease (aGvHD) is a major cause of early morbidity post-haematopoietic stem cell transplantation with minor histocompatibility antigens being a contributing factor. One mHA encoded by the UDP glycosyltransferase 2 family polypeptide B17 (UGT2B17) gene has been shown to be immunogenic because of differential expression in the donor and recipient. We investigated the effects of a homozygous gene deletion of UGT2B17 on the severity of acute aGvHD post-HSCT in HLA-matched related donors. 115 donor and recipient HLA and UGT2B17 genotypes were determined using PCR-SSO and PCR-SSP, respectively. aGvHD grading was determined using routine criteria and dichotomized into either nonclinically significant (0–I) or clinically significant (II–IV). For all analyses, P-values of ≤ 0.05 were considered significant. The frequency of the gene deletion within the total cohort tested was 29.1%. A significant increase in aGvHD severity (grades II-IV) was seen in UGT2B17 recipients expressing the protein when transplanted with a UGT2B17 disparate donor (P = 0.011). We observed a significant association between UGT2B17 expressing recipients and UGT2B17 deficient donors with the severity of aGvHD. This study provides additional evidence that genomic variations may predispose to more severe aGvHD, but are not a mechanism for GvHD.

Histocompatibility testing for stem cell and solid organ transplantation has become increasingly complex as newly discovered HLA alleles are described. HLA typing assignments reported by laboratories are used by physicians and donor registries for matching donors and recipients. To communicate effectively, a common language for histocompatibility terms should be established. In early 2010, representatives from Clinical, Registry, and Histocompatibility organizations joined together as the Harmonization of Histocompatibility Typing Terms Working Group to define a consensual language for laboratories, physicians, and registries to communicate histocompatibility typing information. The Working Group defined terms for HLA typing resolution, HLA matching, and a format for reporting HLA assignments. In addition, definitions of verification typing and extended typing were addressed. The original draft of the Definitions of Histocompatibility Typing Terms was disseminated to colleagues from each organization to gain feedback and create a collaborative document. Commentary gathered during this 90-day review period were discussed and implemented for preparation of this report. Histocompatibility testing continues to evolve; thus, the definitions agreed on today probably will require refinement and perhaps additional terminology in the future.

Histocompatibility testing for stem cell and solid organ transplantation has become increasingly complex as newly discovered human leukocyte antigen (HLA) alleles are described. HLA typing assignments reported by laboratories are used by physicians and donor registries for matching donors and recipients. To communicate effectively, a common language for histocompatibility terms should be established. In early 2010, representatives from clinical, registry, and histocompatibility organizations joined together as the Harmonization of Histocompatibility Typing Terms Working Group to define a consensual language for laboratories, physicians and registries to communicate histocompatibility typing information. The Working Group defined terms for HLA typing resolution, HLA matching and a format for reporting HLA assignments. In addition, definitions of verification typing and extended typing were addressed. The original draft of the Definitions of Histocompatibility Typing Terms was disseminated to colleagues from each organization to gain feedback and create a collaborative document. Commentary gathered during this 90-day review period were discussed and implemented for preparation of this report. Histocompatibility testing continues to evolve thus, the definitions agreed upon today, likely will require refinement and perhaps additional terminology in the future.

Granulocytic sarcoma (GS) in the absence of bone marrow involvement at diagnosis is unusual. Treatment of primary (aleukemic) GS should follow intensive acute myeloid leukemia (AML)-style chemotherapy protocols, as patients otherwise appear destined to rapidly declare frank medullary disease. However, the role of hematopoietic stem cell transplantation (HSCT) is less clear. Here we present an unusual case of isolated primary GS involving small bowel treated with chemotherapy consolidated by HSCT from an unrelated donor. Full donor chimerism was achieved and maintained, without relapse of the original disease. However, 28 months post-transplant our patient developed a distinct and rapidly progressive medullary AML, apparently derived from transplanted cells of donor origin. Donor cell leukemia (DCL) is a fascinating and increasingly recognized phenomenon, with at least 50 cases reported to date. Our case adds to this growing literature and highlights the ethical dilemmas concerning notification of unrelated donors in such cases, since there is little evidence that they are at increased risk of developing leukemia themselves. This also represents the first case of DCL reported after transplantation for primary GS.

Human T cell lymphotropic virus type I (HTLV-I)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) is an inflammatory disease which occurs in less than 2% of HTLV-I -infected individuals. High proviral load, high HTLV-I-specific CD8+ cytotoxic T lymphocyte frequency (CTL) and host genetic factors such as HLA all appear to be associated with HTLV-I infection. Previous studies have shown that HLA-DRB1*01 increases the risk of HAM/TSP in Japanese HTLV-I infected individuals.To investigate the association between HLA class II DRB1 alleles and HLA class I alleles (HLA-Cw*08, B54, A*02 and A-30) in HTLV-I infected individuals in Mashhad.Here we determined the frequency of HLA class II DRB1, using INNO-LIPA reverse hybridization line probe assay, and HLA class I alleles (HLA-Cw*08,B54, A*02 and A-30) by PCR-SSCP method in healthy controls, HAM/TSP patients and HTLV-I infected individuals born and resident in Mashhad.The frequency of HLA-DRB1*01 alleles in this population was different from other areas of Iran. The frequency of HLA-DRB1*01 was significantly increased in HAM/TSP patients compared with carriers (p 0.028; OR=9.4). The frequency of HLA-Cw*08 was also significantly increased in HAM/TSP patients compared with controls (p=0.03; OR=13.5).Our results may suggest that possession of HLA-DRB1*01 increases the risk of HAM/TSP in HTLV-I-infected individuals and HLA-Cw*08 correlates with low CTL immune response in HAM/TSP patients.

Objective: To study whether there is an association between the frequency of functional polymorphisms in the toll-like receptor 4 (TLR4) and cluster differentiation 14 (CD14) genes and periodontitis. Methodology: Genotyping for the TLR4 single-nucleotide polymorphisms (SNPs) Asp299Gly, Thr399Ile and the CD14 SNPs −159 and −1359 was completed for subjects with periodontal disease compared with control subjects. Two disease populations were investigated: 73 subjects with aggressive periodontitis (AgP; 28 males, 45 females) and 95 males with chronic periodontitis (CP). The TLR4 and CD14 polymorphisms were determined using SNaPshot primer extension with capillary electrophoresis. Comparison of allele and genotype frequencies for each polymorphism was by Fisher's exact test or χ2 analysis. Results: The TLR4 Asp299Gly genotype was present in a significantly (p=0.026) lower proportion of AgP subjects (5.5%) compared with control subjects (16.3%). The unadjusted odds ratio for the Asp299Gly genotype to be associated with AgP was 0.30, 95% confidence interval 0.10–0.91. No differences were found in the prevalence of the TLR4 Asp299Gly genotype in men with CP (18.9%) compared with an age-matched control group with no evidence of periodontitis (17%). In addition, there was no difference in the distribution of the CD14 polymorphisms in either the AgP or CP populations studied compared with controls. Conclusion: It is concluded that in West European Caucasians, the Asp299Gly TLR4 gene polymorphism is associated with a decreased risk of AgP but not CP. Promoter polymorphisms of the CD14 gene, however, did not influence susceptibility to inflammatory periodontitis in the population cohorts studied.

Several recently reported HLA-DPB1 alleles have only been identified in a single family or individuals and are of unknown distribution world-wide. Many new DPB1 alleles appear to arise as a result of gene conversion-like events, which may localize variant DPB1 alleles to the population in which they were first identified. Using two SSOP-based typing methods in parallel, we have identified HLA-DPB1*6301 in an individual from rural Cameroon which has previously only been reported in a family of Mexican-American origin. The presence of DPB1*6301 was confirmed by sequence-based typing of exon 2.