HLA Epitopes Research Articles

Quantifying HLA Mismatches at Epitope Level in Haplo-HSCT: Impact in the Outcome in Strategies Using PTCy.

Haploidentical haematopoietic stem cell transplantation (haplo-HSCT) is one of the most effective therapies for treating malignant haematological disorders. However, HLA disparities are significant barriers to the success of this process since they increase the risk of graft versus host disease (GvHD). HLA disparities quantification could help to anticipate the probability and degree of GvHD, but the best tool for such quantification remains a challenge. The aim of this study was to quantify the degree of HLA epitope incompatibilities using PIRCHE (Predicted Indirectly Recognisable HLA Epitopes) algorithm for immunogenicity prediction and their potential relationship with GvHD degree and clinical outcome. We studied 145 patients who underwent a haplo-HSCT with post-transplant cyclophosphamide (PTCy) from a related donor between 2013 and 2020 at our centre evaluating molecular HLA mismatches by PIRCHE-algorithm. Patients with PIRCHE Score (PS) I + II > 38 in GvH direction, developed acute GvHD grade II-IV earlier than their counterparts (HR: 1.71, 95% CI: 1.02-2.87, p = 0.032). In addition, PS-B > 1 in GvH direction was an independent risk factor for chronic GvHD, (HR: 2.51, 95% CI 1.17-5.36, p = 0.017). A higher incidence of relapse was found for patients with PS-II > 28 HvG (HR: 9.16, 95% CI: 2.47-33.92, p < 0.001) which also favoured a worse GvHD/relapse-free survival in patients with PS-II > 27 in HvG direction (HR: 1.88, 95% CI: 1.11-3.18, p = 0.017). These findings indicate that the alloreactivity inferred by epitope HLA disparities is associated with post-transplant outcomes. Thus, analysing PS could benefit the selection of the most suitable donors allowing patient stratification based on HLA mismatches in the context of haplo-HSCT with PTCy.

Paternal HLA-Derived Epitopes and Live Birth in SecondaryRecurrent Pregnancy Loss: New Insights FromaClinical Trial.

Recurrent pregnancy loss (RPL), defined as two or more pregnancy losses before the 24th week of gestation, affects 1%-3% of women worldwide. Approximately, 40% of RPL cases are secondary RPL (sRPL), where women have given birth before facing pregnancy losses. The underlying causes of RPL remain unclear, but immune-related factors may play a role. Previously, a randomised controlled trial using immunoglobulin (IVIG) in sRPL women with a history of four pregnancy losses performed in our RPL unit did not show significant effects of IVIG treatment overall. Yet, some evidence suggests potential benefits for a subset of sRPL patients. In the cohort used for the randomised controlled trial, we examined the role of maternal HLA class II-presented fetal HLA-derived epitopes in sRPL using the predicted indirectly recognisable HLA epitopes (PIRCHE-II) algorithm. In the placebo group, sRPL mothers with an anti-HLA antibody response had higher PIRCHE-II scores when having a live birth compared with sRPL women who experienced another pregnancy loss. This difference was not observed in the IVIG-treated group. Furthermore, as a proxy for T-cell memory, the number of overlapping peptides between the two paternal haplotypes in couples having live births without treatment displayed a larger number of overlapping peptides. This effect was primarily driven by class II-derived peptides. These results suggest that specific combinations of sRPL mothers and fathers, particularly those with an anti-HLA antibody response, may generate higher PIRCHE-II scores, which could contribute to successful live births. Understanding these immune interactions may provide insights for personalised diagnostic and therapeutic strategies in sRPL.

Foetal Microchimerism Correlates With Foetal-Maternal Histocompatibility Both During Pregnancy and Postpartum.

Foetal cells are detectable in women decades postpartum, a state termed foetal microchimerism. The interplay between these semi-allogeneic foetal cells and the mother could be affected by genetic mismatches in the HLA loci. Here, we relate HLA allele and molecular mismatch values to the presence and quantity of foetal microchimerism in the maternal circulation during pregnancy and postpartum. A total of 76 pregnant women were included, of which 59 were followed up 1-8 years postpartum. Maternal and foetal DNA was genotyped for HLA class I and II loci. Foetal cells in maternal buffy coat were detected by qPCR, targeting inherited paternal alleles. Antibody-verified eplet mismatch and Predicted Indirectly Recognisable HLA Epitopes (PIRCHE) scores were calculated to quantify foetal-maternal histocompatibility from the mother's perspective. Circulating foetal cells were detected in 50.0% (38/76) of women during pregnancy, and 25.4% (15/59) postpartum. During pregnancy, HLA class II antibody-verified eplet mismatch load and PIRCHE scores correlated negatively with the presence and quantity of foetal cells in the maternal circulation. Postpartum, HLA class I allele mismatches correlated negatively with foetal microchimerism presence, while HLA class II allele mismatches, HLA class I and II antibody-verified eplet mismatch load, and PIRCHE-I and PIRCHE-II scores correlated negatively with both microchimerism presence and quantity. The correlation between mismatch parameters aimed at evaluating the risk of humoral and T cell-mediated allorecognition and foetal microchimerism was more evident postpartum than during pregnancy. The observed predictive effect of foetal-maternal histocompatibility on foetal microchimerism suggests that circulating foetal cells are subject to clearance by the maternal immune system. We propose that allorecognition of foetal cells in the maternal circulation and tissues influences any long-term effect that foetal microchimerism may have on maternal health.

Allogeneic HLA Humoral Immunogenicity and the Prediction of Donor-Specific HLA Antibody Development.

The development of de novo donor-specific HLA antibodies (dnDSAs) following solid organ transplantation is considered a major risk factor for poor long-term allograft outcomes. The prediction of dnDSA development is a boon to transplant recipients, yet the assessment of allo-HLA immunogenicity remains imprecise. Despite the recent technological advances, a comprehensive evaluation of allo-HLA immunogenicity, which includes both B and T cell allorecognition, is still warranted. Recent studies have proposed using mismatched HLA epitopes (antibody and T cell) as a prognostic biomarker for humoral alloimmunity. However, the identification of immunogenic HLA mismatches has not progressed despite significant improvements in the identification of permissible mismatches. Certainly, the prediction of dnDSA development may benefit permissible HLA mismatched organ transplantations, personalized immunosuppression, and clinical trial design. However, characteristics that go beyond the listing of mismatched HLA antibody epitopes and T cell epitopes, such as the generation of HLA T cell epitope repertoires, recipient's HLA class II phenotype, and immunosuppressive regiments, are required for the precise assessment of allo-HLA immunogenicity.

Permissible HLA mismatches in 9/10 unrelated donor pediatric stem cell transplants using HLA-EMMA: an EBMT Inborn Errors Working Party study

AbstractAllogeneic hematopoietic stem cell transplantation (HSCT) with mismatched unrelated donors (MMUD) is associated with inferior outcome compared with matched unrelated donors (MUDs). We aimed to identify permissible mismatches using HLA epitope mismatch algorithm, which determines permissibility by analyzing amino acid sequences, in a single-center cohort of 70 pediatric 9/10 MMUD HSCTs and 157 10/10 MUDs for comparison. Amino acid matching was evaluated for the whole HLA protein, the α-helices, and the β-sheets, in both host vs graft (HvG) and graft vs host (GvH) direction. Superior event-free survival (EFS) was found in 13 patients permissibly mismatched in the HvG direction (totalHvG, 92% vs 58% at 1 year; P = .009) and in 21 patients matched on the α-helices (αHvG, 90% vs 53%; P = .002). These rates were similar to EFS rates in patients with 10/10 MUDs (90% vs 80%; P = .60). EFS was not related to β-sheet amino acid matching, nor to matching in the GvH direction. Overall survival (OS) rates trended similarly to those of EFS for amino acid mismatches (totalHvG, 92% vs 74%; P = .075; αHvG, 90% vs 71%; P = .072). These findings were reproduced in an EBMT Registry inborn errors cohort of 271 pediatric 9/10 MMUD HSCTs and 929 10/10 MUD HSCTs, showing a significant effect of αHvG matching on both OS and EFS and similar OS and EFS between αHvG matched MMUDs and 10/10 MUDs. In summary, HvG amino acid matching on the α-helices identifies 9/10 MMUDs with permissible mismatches, which are correlated with favorable transplant outcomes similar to those of matched donors.

Desensitization therapy for elevated donor-specific antibody levels in haploidentical transplantation in Chilean patients

Allogeneic stem cell transplantation (ALOSCT) is curative for several hematological diseases. Unrelated donors and cord blood stem cells are valid options but haploidentical donors (HAPLO) have been considered the main source of stem cells in several countries, partly due to easy access and low cost of the transplantation process. Unfortunately, some patients have antibodies against the HLA epitopes from family haploidentical donors (donor-specific antibodies [DSA]) which are associated with engraftment failure and lethality. A few strategies exist to reduce or eliminate HLA antibodies that bind to these receptors. In this study, we present our experience with DSA desensitization by retrospectively examining a cohort from our hospital program. Between 2012 and 2023, we performed 243 ALOSCTs, of which 142 were from HAPLO and 56 were from unrelated donors. Nine patients (7%) had elevated DSA levels, most of which were female patients and mostly HAPLO. The median fluorescence intensity for these patients was 22,490 (19,000-28,560). Most of these high DSA patients (80%) received a desensitization procedure that involves plasmapheresis, rituximab, and immunoglobulin. The remaining 20% had severe infections during transplantation, and received rituximab monotherapy instead. The median dose of stem cells infused was 6.5 x 10^6 CD34/kg. Graft-versus-host disease (GVHD) preventative measures for all patients involved post-transplantation cyclophosphamide. Primary graft failure was observed in 45% of DSA elevated patients. For the remaining patients, median granulocyte and platelet engraftment were 14 d (12-16) and 16 d (13-18), respectively. Mortality in patients who did not receive engraftment was 100%. The incidence of mild, chronic GVHD was 15%. In conclusion, the desensitization of DSA in patients provided a 55% rate of engraftment and survival. However, a 45% rate of primary graft failure continued to pose a challenge in patients with DSA and required the development of improved strategies to reduce elevated transplant-related mortality.

Early progression of chronic histologic lesions in kidney transplant biopsies is not associated with HLA histocompatibility.

Early progression of chronic histologic lesions in kidney allografts represents the main finding in graft attrition. The objective of this retrospective cohort study was to elucidate whether HLA histocompatibility is associated with progression of chronic histologic lesions in the first year post-transplant. Established associations of de novo donor-specific antibody (dnDSA) formation with HLA mismatch and microvascular inflammation (MVI) were calculated to allow for comparability with other study cohorts. We included 117 adult kidney transplant recipients, transplanted between 2016 and 2020 from predominantly deceased donors, who had surveillance biopsies at 3 and 12 months. Histologic lesion scores were assessed according to the Banff classification. HLA mismatch scores [i.e. eplet, predicted indirectly recognizable HLA-epitopes algorithm (PIRCHE-II), HLA epitope mismatch algorithm (HLA-EMMA), HLA whole antigen A/B/DR] were calculated for all transplant pairs. Formation of dnDSAs was quantified by single antigen beads. More than one-third of patients exhibited a progression of chronic lesion scores by at least one Banff grade in tubular atrophy (ct), interstitial fibrosis (ci), arteriolar hyalinosis (ah) and inflammation in the area of interstitial fibrosis and tubular atrophy (i-IFTA) from the 3- to the 12-month biopsy. Multivariable proportional odds logistic regression models revealed no association of HLA mismatch scores with progression of histologic lesions, except for ah and especially HLA-EMMA DRB1 [odds ratio (OR)=1.10, 95% confidence interval (CI) 1.03-1.18]. Furthermore, the established associations of dnDSA formation with HLA mismatch and MVI (OR=5.31, 95% CI 1.19-22.57) could be confirmed in our cohort. These data support the association of HLA mismatch and alloimmune response, while suggesting that other factors contribute to early progression of chronic histologic lesions.

P121 Correlation of HLA epitope mismatches with de novo donor-specific antibody formation in renal transplant recipients – A comparison of snowflake to HLAMatchmaker and PIRCHE scores

P121 Correlation of HLA epitope mismatches with de novo donor-specific antibody formation in renal transplant recipients – A comparison of snowflake to HLAMatchmaker and PIRCHE scores

P507 Frequency of immunizing HLA epitopes and immungenicity: A new approach to immunological risk assessment?

P507 Frequency of immunizing HLA epitopes and immungenicity: A new approach to immunological risk assessment?

P477 PIRCHE (PREDICTED INDIRECTLY RECOGNIZABLE HLA EPITOPES) ALGORITHM AS A POSSIBLE PREDICTOR OF AMR (ANTIBODY MEDIATED REJECTION) FOLLOWING HEART TRANSPLANT, FROM LAB EXPERIENCE TO CLINICAL PRACTICE: A CASE REPORT

Abstract PIRCHE–II is an algorithm used to estimate the risk for developing alloreactivity towards HLA mismatches. A 35–year–old man with idiopathic dilated cardiomyopathy with several previous hospitalizations for heart failure is admitted to our unit for a new episode in August 2021. After evaluation, the patient was placed on the waiting list for transplant. The following month the patient underwent heart transplant. The donor had a compatible CDC crossmatch and was negative for anti–HLA antibodies. After 46 days, the patient was discharged at home in stable conditions after 2 negative endomyocardial biopsies (EMB): ISHLT’04 0. In January 2021, the patient was newly admitted for dyspnea and oliguria. Imaging revealed bilateral severe pleural effusion, biventricular cardiac disfunction; EMB showed acute rejection (ISHLT’04 3R). Panel–reactive antibody (PRA) screening found 93 % of class I PRA, 3 % of class II PRA with IgM and IgG donor–specific antibodies (DSA), C1q–positive. After 2 months, the patient was placed again on the transplant waitlist for a persistent severe biventricular disfunction, massive tricuspid regurgitation and diffuse myocardial fibrosis; class I PRA was decreasing (41 % in March 2022, 31 % in April 2022). Unfortunately, the patient died of infection before re–transplantation. After the event, which was recognized as AMR caused by class I DSA, a form rarely described in literature, we employed the PIRCHE (Predicted Indirectly Recognizable HLA Epitopes) algorithm to predict indirect donor–recipient alloreactivity: PIRCHE–II was 54. Is a high PIRCHE–II score a reliable predictor of AMR following heart transplant? More data and deeper understanding is surely needed. A correct after–transplant serologic monitoring is definitely needed to search for de–novo DSA.

Epitope Mismatch at HLA-DRB1 Associates with Reduced Relapse Risk in Cord Blood Transplantation for Standard-Risk Hematologic Malignancy

Cord blood transplantation (CBT) is an attractive therapeutic option for patients with hematological malignancies. CBT tolerates HLA mismatches between donors and recipients, but which HLA mismatches generate graft-versus-tumor (GVT) effects is unknown. Given that HLA molecules contain epitopes comprising polymorphic amino acids that determine their immunogenicity, we investigated associations between epitope-level HLA mismatches and relapse following single-unit CBT. A total of 492 patients with hematological malignancies who received single unit, T cell replete CBT were included in this multi-center retrospective study. HLA epitope mismatches (EM) were quantified using HLA matchmaker software from donor and patient HLA-A, B, C and DRB1 allele data. Patients were dichotomized by median EM value and divided into groups transplanted in complete/partial remission (standard stage: 62.4%) and others (advanced stage: 37.6%). Median EM numbers in the graft-versus-host direction (GVH-EM) at HLA-class I and HLA-DRB1 were 3 (range, 0-16) and 1 (range, 0-7), respectively. Higher HLA-class I GVH-EM increased non-relapse mortality (NRM) in the advanced stage group (adjusted hazard ratio [HR], 2.12; p=0.021), with no significant advantage for relapse in either stage. On the other hand, higher HLA-DRB1 GVH-EM was associated with better disease-free survival in the standard stage group (adjusted HR, 0.63; p=0.020), which was attributed to lower relapse risk (adjusted HR, 0.46; p=0.014). These associations were also observed even within HLA-DRB1 allele-mismatched transplants in the standard stage group, indicating that EM might have impacts on relapse risk independently of allele mismatch. High HLA-DRB1 GVH-EM did not increase NRM in either stage. High HLA-DRB1 GVH-EM may lead to potent GVT effects and favorable prognosis following CBT especially in the patients transplanted in the standard stage. This approach may facilitate appropriate unit selection and improve the overall prognosis of patients with hematological malignancies who receive CBT.

Immuno-bioinformatic approach for designing of multi-epitope merozoite surface antigen of Babesia bigemina and evaluation of its immunogenicity in inoculated calves.

Babesiosis is a parasitic disease caused by intraerythrocytic parasites of the genus Babesia, which infect both wild and domestic animals. Merozoite surface antigens (MSAs) have been identified as efficient immunogens in Babesia-infected animals. MSAs play a key role in the invasion process and have been proposed as potential targets for vaccine development. Epitope-based vaccines offer several advantages over whole protein vaccines as the immunogenic proteins are small and can induce both Th1 and Th2 immune responses, which are desirable for protection. However, the MSA, particularly gp45, is polymorphic in Babesia bigemina, posing a challenge to vaccine development. The purpose of this study was to develop a recombinant gpME (gp45-multi-epitope) for a vaccine against Babesia bigemina. B-cell, T-cell, and HLA epitope predictions were used to synthesize the gpME sequence from the consensus sequence of gp45. The gpME sequence was synthesized and cloned in the pET28α vector through the commercial biotechnology company to get pET28-gpME. The plasmid cloned with the gpME sequence comprising 1068 bp was expressed in a bacterial expression system. A band of 39 kDa of rec-gpME was obtained via SDS-PAGE and Western blotting. Rec-gpME @200ng was injected in calves 3 times at 2 weeks interval. The humoral response was evaluated through the indirect ELISA method. The ELISA with rec-gp45 protein showed a significant value of optical density. The recombinant protein containing multiple epitopes from the MSA gp45 may represent a promising candidate for a vaccine against Babesia bigemina.

Molecular disparity of HLA-DPB1 is associated with the development of subsequent solid cancer after allogeneic hematopoietic stem cell transplantation.

An increased incidence of subsequent solid cancers (SSCs) has been reported in long-term survivors of allogeneic hematopoietic stem cell transplantation (allo-HSCT), and SSC is associated with inferior mortality and morbidity. Previous studies showed that the incidence of SSC is significantly higher in those who underwent allo-HSCT from HLA-mismatched donors, suggesting that persistent alloimmunity may predispose patients to SSCs. It was recently reported that, in a cohort of patients who received allo-HSCT from an unrelated donor matched at HLA-A, -B, -C, -DRB1/3/4/5, and -DQB1 loci, HLA-DPB1 alloimmunity determined by high mismatched eplets (MEs) and Predicted Indirectly Recognizable HLA Epitopes (PIRCHE) score (PS), was associated with relapse protection and increased risk of acute graft-versus-host disease (GVHD). In the present study, the impact of HLA-DPB1 alloimmunity assessed by molecular mismatch algorithms on the development of SSCs in a cohort of 1514 patients who underwent allo-HSCT for hematologic malignancies was further investigated. ME load at the HLA-DPB1 locus was measured using the HLAMatchmaker module incorporated in HLA Fusion software, and the PS for mismatched HLA-DPB1 was calculated using the HSCT module from the PIRCHE online matching service. In multivariable analysis after adjusting for baseline risk factors, higher ME, PS-I, and PS-II in the GVH direction, but not in the HVG direction, were associated with an increased risk of SSCs (ME: subdistribution hazard ratio [SHR] 1.58, p= .01; PS-I: SHR 1.59, p= .009; PS-II: SHR 1.71, p= .003). In contrast, nonpermissive HLA-DPB1 mismatches defined by the conventional T-cell epitope algorithm were not predictive of the risk of SSCs. Moreover, posttransplant cyclophosphamide-based GVHD prophylaxis was associated with a reduced risk of subsequent solid cancer (SHR 0.34, p= .021). These results indicate for the first time that increased GVH alloreactivity could contribute to the development of SSCs in allo-HSCT survivors.

A study on the design of an in silico self-amplifying mRNA vaccine against Nipah virus using immunoinformatics

The scientific community continues to be impressed with RNA-based vaccines with great efficacy, quick synthesis and speed-to-market. The traditional vaccine may require large doses or repeat injections to achieve an expression for protection against the virus; the self-amplifying mRNA vaccine addresses this limitation. Therefore, a thorough examination of the most antigenic component of the Nipah virus was carried out to design the coding sequence of an antigen, which will provoke a virus-specific immune response. After that, we predicted and evaluated epitopes from NiV G-protein. We employed 8 HTL, 2 CTL and 3 B-cell epitopes. The study of structural compatibility was done by performing docking between HLA alleles and epitopes to get insights into the immune response of epitopes. The entire peptide coding sequence of an antigen was linked using a linker to design the structure of the vaccine. Physicochemical parameters of the designed vaccine constructs were assessed using a protparam server. Later, the vaccine sequence was converted into cDNA. We inserted a gene-expressing replicase at the start of a coding sequence for self-amplification. Next, to formulate the final version of vaccine signal sequences were added. Based on these findings, this mRNA vaccine appears to be a promising option against the Nipah virus. Communicated by Ramaswamy H. Sarma

Successful Deceased Donor Kidney Transplantation in a Highly Sensitized Patient after Reclassification of Unacceptable Antigens Based on HLA Epitope Analysis

Successful Deceased Donor Kidney Transplantation in a Highly Sensitized Patient after Reclassification of Unacceptable Antigens Based on HLA Epitope Analysis

Successful Deceased Donor Kidney Transplantation in a Highly Sensitized Patient after Reclassification of Unacceptable Antigens Based on HLA Epitope Analysis

For patients with end stage renal disease, kidney transplant offers significant survival and quality-of-life advantages compared with dialysis. But for patients seeking transplant who are highly sensitized, waiting times have traditionally been long and options limited. We present the case of a 34-year-old hypersensitized female who underwent renal retransplantation. Histocompatibility tests revealed a calculated panel-reactive antibody of 99.53% with multiple antibodies against class I and II human leucocyte antigens and an eplet analysis was performed. The donor’s potential unacceptable antigens were re-defined and the calculated panel-reactive antibody decreased to 88.38%. After one month the patient received a deceased-donor kidney transplant. Complement dependent cytotoxicity crossmatch was negative; virtual crossmatch and flow cytometry crossmatch with historical serum were positive. High-dose intravenous immunoglobulin and rituximab were added to the thymoglobulin-based induction immunosuppression. Three donor-specific antibodies were detected and plasmapheresis was performed. Renal allograft biopsy revealed no manifestations of rejection. Repeated testing observed a decrease in donor-specific antibodies median fluorescence intensity values. Four months post-transplant, the patient remained with normal graft function without proteinuria. She is receiving a standard maintenance immunosuppression regime with prednisolone, mycophenolate mofetil and tacrolimus. The careful discussion among the transplantation center and histocompatibility laboratory in association with intense immunosuppression and close laboratory monitoring allowed a successful human leukocyte antigen-incompatible deceased donor kidney transplantation in the most critical phase for the occurrence of humoral rejection. It is noteworthy that the new histocompatibility and immunogenetics methodologies provide a more affirmative and comprehensive assessment of mismatch acceptability.

An Approach to Identify HLA Class II Immunogenic Epitopes in the Greek Population through Machine Learning Algorithms

Current pre-transplantation routine matching involves serum anti-HLA antibodies quantification but cannot always preclude unfavorable graft outcomes. Epitope-based matching is proposed as a more precise approach, but to date no epitope-matching algorithm provides a satisfactory predictive tool for transplantation outcomes. In this study, anti-HLA-II loci responses from 1748 patients were analyzed with unsupervised machine learning algorithms, namely principal component analysis (PCA) and antigenic distances, projected as dendrograms. PCA for anti-HLA-DR anti-bodies revealed three main clusters of responses: anti-HLA-DR51 combined with anti-HLA-DRB1*01, anti-HLA-DR52 combined with anti-HLA-DRB1*08 and anti-HLA-DR53 combined with anti-HLA-DRB1*10. The dendrogram for anti-HLA-DR confirmed the pattern and showed further bisection of each cluster. Common epitopes present exclusively in all HLA molecules of each cluster were determined following the HLA epitope registry. Thus, we propose that 19 out of 123 HLA-DR epitopes are those that mainly lead anti-HLA-DR responses in the studied population. Likewise, we identified 22 out of 83 epitopes responsible for anti-HLA-DQ and 13 out of 62 responsible for anti-HLA-DP responses. Interpretation of these results may elucidate mechanisms of interlocus cross-reactivity, providing an alternative way of estimating the significance of each epitope in a population and thus suggesting a novel strategy towards optimal donor selection.

Eplet-Predicted Antigens: An Attempt to Introduce Eplets into Unacceptable Antigen Determination and Calculated Panel-Reactive Antibody Calculation Facilitating Kidney Allocation.

(1) Calculated panel-reactive antibody (CPRA) is a measure of sensitization based on unacceptable antigens (UAs). Determination of UAs based on single-antigen bead assays at allele or antigen levels may be inappropriate. We aimed to introduce eplets for better assessment of sensitization; (2) 900 recipients and 1427 donors were enrolled for candidate or donor pools, respectively. Eplets were from the HLA Epitope Registry. UAs were determined by anti-HLA antibodies identified using LIFECODES Single Antigen (LSA) kits. CPRA values were calculated using a simplified method of donor filtering; (3) HLA antigens containing all eplets of an HLA antigen in LSA kits (LSA antigen) were defined as eplet-predicted (EP) antigens, the reactivity of which could be predicted by that LSA antigen. High reactivity concordance was found between LSA and EP antigens. More HLA antigens were covered by EP antigens in the population than LSA antigens. CPRA values at the EP level were higher than at the allele level and lower than at the antigen level. The EP antigens facilitated UA determination for non-LSA antigens and avoided acute rejection; (4) UA determination using EP antigens can lead to more accurate assessment of sensitization, enabling a high probability of compatible organs and a low risk of adverse outcomes.

Site-directed mutagenesis of HLA molecules reveals the functional epitope of a human HLA-A1/A36-specific monoclonal antibody.

Eplet 44KM is currently listed in the HLA Epitope Registry but does not adhere to the eplet definition of an amino acid configuration within a 3.5Å radius. Eplet 44KM has been previously redefined to the antibody-verified reactivity pattern 44K/150V/158V, based on reactivity analysis of monoclonal antibody VDK1D12. Since the three residues are always simultaneously present on common HLA alleles, methods to define which residue is crucial for antibody-induction and binding are limited. In this proof-of-concept study, we performed site-directed mutagenesis to narrow down the antibody-verified reactivity pattern 44K/150V/158V to a single amino acid and defined 44K as the eplet or functional epitope of mAb VDK1D12.

Exploring vast microbial epitope spaces.

The spectrum and origin of commensal bacterial HLA epitopes is not well understood. In this issue of Immunity, Pedersen etal. provide a systematic analysis of commensal bacterial HLA II epitopes and demonstrate plasticity of corresponding CD4+ Tcell responses.