Amino Acid Mismatches Research Articles

Novel Scoring System for Ranking Hematopoietic Stem Cell Transplantation.

When human leukocyte antigen (HLA)-matched donors are not available for hematopoietic stem cell transplants (HSCT), there are no well-accepted guidelines for ranking 7/8 HLA-matched unrelated donors to achieve optimal transplant outcomes. A novel scoring system for ranking HLA mismatches for these donors was investigated. High-resolution HLA types were used to determine amino acid mismatches located in the HLA antigen-recognition domain. The location and physicochemical properties of mismatched amino acids were used to assign scores for peptide binding, T-cell receptor docking, and HLA structure/function. The scores were tested using a cohort of 2319 patients with leukemia or myelodysplastic syndrome who received their first unrelated donor transplant using conventional graft-versus-host disease (GVHD) prophylaxis between 2000 and 2014. Donors were 7/8 HLA-matched with a single HLA Class I mismatch. Primary outcomes were overall survival and acute GVHD. The scores did not significantly (p<0.01) associate with transplant outcomes, although a Peptide Score = 0 (i.e., no differences in peptide binding; N=146, 6.3%) appears to have lower transplant-related mortality (TRM) compared to higher scores (p=0.019). HLA mismatches with Peptide Score=0 were predominately HLA-C*03:03/03:04 (62%), previously reported to be a permissive mismatch, and a group of 28 other HLA mismatches (38%) that showed similar associations with TRM. This study suggests that HLA mismatches that do not alter peptide binding or orientation (Peptide Score=0) could expand the number of permissive HLA mismatches. Further investigation is needed to confirm this observation and to explore alternative scoring systems for ranking HLA mismatched donors.

Risk of kidney graft failure (GF) associated with HLA class II amino acid mismatches (AAMM) is highest during the first year after transplant for sensitized patients

Risk of kidney graft failure (GF) associated with HLA class II amino acid mismatches (AAMM) is highest during the first year after transplant for sensitized patients

An extended machine learning approach for binning donor-recipient HLA amino acid mismatches to predict kidney graft failure risk

An extended machine learning approach for binning donor-recipient HLA amino acid mismatches to predict kidney graft failure risk

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.

Five amino acid mismatches in the zinc-finger domains of Cellulose Synthase 5 and Cellulose Synthase 6 cooperatively modulate their functional properties by controlling homodimerization in Arabidopsis.

Cellulose synthase 5 (CESA5) and CESA6 are known to share substantial functional overlap. In the zinc-finger domain (ZN) of CESA5, there are five amino acid (AA) mismatches when compared to CESA6. These mismatches in CESA5 were replaced with their CESA6 counterparts one by one until all were replaced, generating nine engineered CESA5s. Each N-terminal enhanced yellow fluorescent protein-tagged engineered CESA5 was introduced to prc1-1, a cesa6 null mutant, and resulting mutants were subjected to phenotypic analyses. We found that five single AA-replaced CESA5 proteins partially rescue the prc1-1 mutant phenotypes to different extents. Multi-AA replaced CESA5s further rescued the mutant phenotypes in an additive manner, culminating in full recovery by CESA5G43R + S49T+S54P+S80A+Y88F. Investigations in cellulose content, cellulose synthase complex (CSC) motility, and cellulose microfibril organization in the same mutants support the results of the phenotypic analyses. Bimolecular fluorescence complementation assays demonstrated that the level of homodimerization in every engineered CESA5 is substantially higher than CESA5. The mean fluorescence intensity of CSCs carrying each engineered CESA5 fluctuates with the degree to which the prc1-1 mutant phenotypes are rescued by introducing a corresponding engineered CESA5. Taken together, these five AA mismatches in the ZNs of CESA5 and CESA6 cooperatively modulate the functional properties of these CESAs by controlling their homodimerization capacity, which in turn imposes proportional changes on the incorporation of these CESAs into CSCs.

Dual template (epitope) imprinted electrode for sensing bacterial protein with high selectivity.

Epitope imprinting has shown better prospects to synthesize synthetic receptors for proteins. Here, dual epitope imprinted polymer electrode (DEIP) matrix was fabricated on gold surface of electrochemical quartz crystal microbalance (EQCM) for recognition of target epitope sequence in blood samples of patients suffering from brain fever. Epitope sequences from outer membrane protein Por B of Neisseria meningitidis (MC58) bacteria predicted through immunoinformatic tools were chosen for imprinting. Self-assembled monolayers (SAM) of cysteine appended epitope sequences on gold nanoparticles were subjected to polymerization prior to electrodeposition on gold coated EQCM electrode. The polymeric matrix was woven around the cysteine appended epitope SAMs through multiple monomers (3-sulfo propyl methacrylate potassium salt (3-SPMAP), benzyl methacrylate (BMA)) and crosslinker (N, N'-methylene-bis-acrylamide). On extraction of the peptide sequences, imprinted cavities were able to selectively and specifically bind targeted epitope sequences in laboratory samples as well as 'real' samples of patients. Selectivity of sensor was examined through mismatched peptide sequences and certain plasma proteins also. The sensor was able to show specific binding towards the blood samples of infected patients, even in the presence of 'matrix' and other plasma proteins such as albumin and globulin. Even other peptide sequences, similar to epitope sequences only with one or two amino acid mismatches were also unable to show any binding. The analytical performance of DEIP-EQCM sensor was tested through selectivity, specificity, matrix effect, detection limit (0.68-1.01 nM), quantification limit (2.05-3.05 nM) and reproducibility (RSD ~ 5%). Hence, a diagnostic tool for bacterium causing meningitis is successfully fabricated in a facile manner which will broaden the clinical access and make efficient population screening feasible.

HLA molecular mismatches and induced donor-specific tolerance in combined living donor kidney and hematopoietic stem cell transplantation.

We investigated the potential role of HLA molecular mismatches (MM) in achieving stable chimerism, allowing for donor-specific tolerance in patients undergoing combined living donor kidney and hematopoietic stem cell transplantation (HSCT). All patients with available DNA samples (N=32) who participated in a phase 2 clinical trial (NCT00498160) where they received an HLA mismatched co-transplantation of living donor kidney and facilitating cell-enriched HSCT were included in this study. High-resolution HLA genotyping data were used to calculate HLA amino acid mismatches (AAMM), Eplet MM, three-dimensional electrostatic mismatch scores (EMS-3D), PIRCHE scores, HLA-DPB1 T-cell epitope group MM, HLA-B leader sequence MM, and KIR ligands MM between the donor and recipient in both directions. HLA MM were analyzed to test for correlation with the development of chimerism, graft vs. host disease (GvHD), de novo DSA, and graft rejection. Follow-up time of this cohort was 6-13.5 years. Of the 32 patients, 26 developed high-level donor or mixed stable chimerism, followed by complete withdrawal of immunosuppression (IS) in 25 patients. The remaining six of the 32 patients had transient chimerism or no engraftment and were maintained on IS (On-IS). In host versus graft direction, a trend toward higher median number of HLA-DRB1 MM scores was seen in patients On-IS compared to patients with high-level donor/mixed chimerism, using any of the HLA MM modalities; however, initial statistical significance was observed only for the EMS-3D score (0.45 [IQR, 0.30-0.61] vs. 0.24 [IQR, 0.18-0.36], respectively; p=0.036), which was lost when applying the Bonferroni correction. No statistically significant differences between the two groups were observed for AAMM, EMS-3D, Eplet MM, and PIRCHE-II scores calculated in graft versus host direction. No associations were found between development of chimerism and GvHD and non-permissive HLA-DPB1 T-cell epitope group MM, HLA-B leader sequence, and KIR ligands MM. Our results suggest an association between HLA-DRB1 molecular mismatches and achieving stable chimerism, particularly when electrostatic quality of the mismatch is considered. The non-permissive HLA-DPB1 T-cell epitope group, HLA-B leader sequence, and KIR ligands MM do not predict chimerism and GvHD in this combined kidney/HSCT transplant patient cohort. Further work is needed to validate our findings. https://clinicaltrials.gov/study/NCT00498160, identifier NCT00498160.

Comparison of human leukocyte antigen immunologic risk stratification methods in lung transplantation

Outcomes after lung transplantation (LTx) remain poor, despite advances in sequencing technology and development of algorithms defining immunologic compatibility. Presently, there is no consensus regarding the best approach to define human leukocyte antigen (HLA) compatibility in LTx. In this study, we compared 5 different HLA compatibility tools in a high-resolution HLA-typed, clinically characterized cohort, to determine which approach predicts outcomes after LTx. In this retrospective single-center study, 277 donor-recipient transplant pairs were HLA-typed using next generation sequencing. HLA compatibility was defined using HLAMatchmaker, HLA epitope mismatch algorithm (HLA-EMMA), predicted indirectly recognizable HLA epitopes (PIRCHE), electrostatic mismatch score (EMS), and amino acid mismatches (AAMMs). Associations with HLA mismatching and survival, chronic lung allograft dysfunction (CLAD), and anti-HLA donor-specific antibody (DSA) were calculated using adjusted Cox proportional modeling. Lower HLA class II mismatching was associated with improved survival as defined by HLAMatchmaker (P < .01), HLA-EMMA (P < .05), PIRCHE (P < .05), EMS (P < .001), and AAMM (P < .01). All approaches demonstrated that HLA-DRB1345 matching was associated with freedom from restrictive allograft syndrome and HLA-DQ matching with reduced DSA development. Reducing the level of HLA mismatching, in T cell or B cell epitopes, electrostatic differences, or amino acid, can improve outcomes after LTx and potentially guide immunosuppression strategies.

Requirements for anti-aquaporin 5 autoantibody production in a mouse model.

Several oral bacteria, including Prevotella melaninogenica (Pm), have aquaporin (AQP) proteins homologous to human AQP5, a major water channel protein targeted in Sjogren's syndrome. This study aimed to understand the antigenic characteristics that induce autoantibodies against an AQP5 "E" epitope (AQP5E) in a mouse model using C57BL/6 mice. Immunization with a PmE-L peptide derived from Pm AQP, which contains amino acid mismatches both at the B- and T-cell epitopes, efficiently induced anti-AQP5E autoantibodies accompanied by increased germinal center (GC) B and follicular helper T cells in the draining lymph nodes. However, PmE, a peptide lacking a T-cell epitope, and AQP5E-L, an AQP5-derived self-peptide, hardly induced either anti-AQP5E autoantibodies or GC responses. Surprisingly, OTII-AQP5E, a peptide that replaced the self T-cell epitope of AQP5E-L with an ovalbumin-derived foreign T-cell epitope, was not any better than AQP5E-L in the induction of anti-AQP5E autoantibodies and GC response, despite the substantial expansion of CD4+ T cells and production of anti-OTII-AQP5E antibodies. The complex of biotinylated PmE-L peptide and highly immunogenic streptavidin (SA) induced a strong extrafollicular B-cell response skewed toward the expansion of SA-specific B cells. However, the expansion of AQP5E-specific GC B cells was limited, resulting in the inefficient induction of anti-AQP5E autoantibodies. Collectively, our results have demonstrated that anti-AQP5E autoantibody production is only allowed when foreign B- and T-cell epitopes drive a strong GC response of AQP5E-specific B cells for affinity maturation. This study helps explain why cross-reactive anti-AQP5 autoantibodies are not produced during the immune response to Pm in most healthy people.

O7 Combining allele-specific solvent-accessible surface-protruding amino acid mismatches and predicted indirect T cell epitopes improves HLA matching in predicting kidney graft survival

O7 Combining allele-specific solvent-accessible surface-protruding amino acid mismatches and predicted indirect T cell epitopes improves HLA matching in predicting kidney graft survival

HLA amino acid Mismatch-Based risk stratification of kidney allograft failure using a novel Machine learning algorithm

ObjectiveWhile associations between HLA antigen-level mismatches (Ag-MM) and kidney allograft failure are well established, HLA amino acid-level mismatches (AA-MM) have been less explored. Ag-MM fails to consider the substantial variability in the number of MMs at polymorphic amino acid (AA) sites within any given Ag-MM category, which may conceal variable impact on allorecognition. In this study we aim to develop a novel Feature Inclusion Bin Evolver for Risk Stratification (FIBERS) and apply it to automatically discover bins of HLA amino acid mismatches that stratify donor-recipient pairs into low versus high graft survival risk groups. MethodsUsing data from the Scientific Registry of Transplant Recipients, we applied FIBERS on a multiethnic population of 166,574 kidney transplants between 2000 and 2017. FIBERS was applied (1) across all HLA-A, B, C, DRB1, and DQB1 locus AA-MMs with comparison to 0-ABDR Ag-MM risk stratification, (2) on AA-MMs within each HLA locus individually, and (3) using cross validation to evaluate FIBERS generalizability. The predictive power of graft failure risk stratification was evaluated while adjusting for donor/recipient characteristics and HLA-A, B, C, DRB1, and DQB1 Ag-MMs as covariates. ResultsFIBERS’s best-performing bin (on AA-MMs across all loci) added significant predictive power (hazard ratio = 1.10, Bonferroni adj. p < 0.001) in stratifying graft failure risk (where low-risk is defined as zero AA-MMs and high-risk is one or more AA-MMs) even after adjusting for Ag-MMs and donor/recipient covariates. The best bin also categorized more than twice as many patients to the low-risk category, compared to traditional 0-ABDR Ag mismatching (∼24.4% vs ∼ 9.1%). When HLA loci were binned individually, the bin for DRB1 exhibited the strongest risk stratification; relative to zero AA-MM, one or more MMs in the bin yielded HR = 1.11, p < 0.005 in a fully adjusted Cox model. AA-MMs at HLA-DRB1 peptide contact sites contributed most to incremental risk of graft failure. Additionally, FIBERS points to possible risk associated with HLA-DQB1 AA-MMs at positions that determine specificity of peptide anchor residues and HLA-DQ heterodimer stability. ConclusionFIBERS’s performance suggests potential for discovery of HLA immunogenetics-based risk stratification of kidney graft failure that outperforms traditional assessment.

HLA diversity in ethnic populations can affect detection of donor-specific antibodies by single antigen beads.

In solid-organ transplantation, human leukocyte antigen (HLA) donor-specific antibodies (DSA) are strongly associated with graft rejection, graft loss, and patient death. The predominant tests used for detecting HLA DSA before and after solid-organ transplantation are HLA single antigen bead (SAB) assays. However, SAB assays may not detect antibodies directed against HLA epitopes that are not represented in the SAB. The prevalence and potential impact of unrepresented HLA epitopes are expected to vary by ethnicity, but have not been thoroughly investigated. To address this knowledge gap, HLA allele frequencies from seven ethnic populations were compared with HLA proteins present in SAB products from two manufacturers to determine unrepresented HLA proteins. Allele frequencies were obtained from the Common, Intermediate, and Well Documented HLA catalog v3.0, and frequencies of unrepresented HLA types were calculated. Next-generation sequencing was used to determine HLA types of 60 deceased solid-organ donors, and results were used to determine if their HLA-A, -B, -C, and -DRB1 proteins were not present in SAB reagents from two vendors. Unrepresented HLA proteins were compared with the most similar protein in SAB assays from either vendor and then visualized using modeling software to assess potential HLA epitopes. For the seven ethnic populations, 0.5% to 11.8% of each population had HLA proteins not included in SAB assays from one vendor. Non-European populations had greater numbers of unrepresented alleles. Among the deceased donors, 26.7% (16/60) had at least one unrepresented HLA-A, -B, -C, or -DRB1 protein. Structural modeling demonstrated that a subset of these had potential HLA epitopes that are solvent accessible amino acid mismatches and are likely to be accessible to B cell receptors. In conclusion, SAB assays cannot completely rule out the presence of HLA DSA. HLA epitopes not represented in those assays vary by ethnicity and should not be overlooked, especially in non-European populations. Allele-level HLA typing can help determine the potential for HLA antibodies that could evade detection.

Snowflake epitope matching correlates with child-specific antibodies during pregnancy and donor-specific antibodies after kidney transplantation.

Development of donor-specific human leukocyte antigen (HLA) antibodies (DSA) remains a major risk factor for graft loss following organ transplantation, where DSA are directed towards patches on the three-dimensional structure of the respective organ donor's HLA proteins. Matching donors and recipients based on HLA epitopes appears beneficial for the avoidance of DSA. Defining surface epitopes however remains challenging and the concepts underlying their characterization are not fully understood. Based on our recently implemented computational deep learning pipeline to define HLA Class I protein-specific surface residues, we hypothesized a correlation between the number of HLA protein-specific solvent-accessible interlocus amino acid mismatches (arbitrarily called Snowflake) and the incidence of DSA. To validate our hypothesis, we considered two cohorts simultaneously. The kidney transplant cohort (KTC) considers 305 kidney-transplanted patients without DSA prior to transplantation. During the follow-up, HLA antibody screening was performed regularly to identify DSA. The pregnancy cohort (PC) considers 231 women without major sensitization events prior to pregnancy who gave live birth. Post-delivery serum was screened for HLA antibodies directed against the child's inherited paternal haplotype (CSA). Based on the involved individuals' HLA typings, the numbers of interlocus-mismatched antibody-verified eplets (AbvEPS), the T cell epitope PIRCHE-II model and Snowflake were calculated locus-specific (HLA-A, -B and -C), normalized and pooled. In both cohorts, Snowflake numbers were significantly elevated in recipients/mothers that developed DSA/CSA. Univariable regression revealed significant positive correlation between DSA/CSA and AbvEPS, PIRCHE-II and Snowflake. Snowflake numbers showed stronger correlation with numbers of AbvEPS compared to Snowflake numbers with PIRCHE-II. Our data shows correlation between Snowflake scores and the incidence of DSA after allo-immunization. Given both AbvEPS and Snowflake are B cell epitope models, their stronger correlation compared to PIRCHE-II and Snowflake appears plausible. Our data confirms that exploring solvent accessibility is a valuable approach for refining B cell epitope definitions.

De novo assisted AFB1-Specific monoclonal antibody sequence assembly and comprehensive molecular characterization

Despite their widely used and access as biological reagents in analytical methods, the detailed structural features for most of the antibodies were rarely known. Here, a new antibody for AFB1 with high specificity in constructing ELISA was studied in detail. The molecular structure and modification were elucidated mainly by nano-electrospray ionization mass spectrometry. The mass experiments, including MALDI-TOF MS, revealed complete and specific fragments, including antibody molecular weight, peptides, glycopeptide, and N-glycoform. By proteolytic treatment of pepsin and trypsin and high-resolution tandem-MS, the primary structure of the newly developed anti-AFB1 antibody was assembled by several rounds of Database search process assisted with the de novo results. The antibody CDR annotation and constraint-based multiple alignment tool were used to differentiate and align the sequences. The method uses only two proteases to generate numerous peptides for de novo sequencing. This artificial assembled AFB1-specific monoclonal antibody sequence was validated by comparison with the sequencing results of the immunoglobulin gene. The results showed that this method achieves full sequence coverage of anti-AFB1 monoclonal antibody, with an accuracy of 100% in the CDR regions of light chain and four amino acid mismatch in heavy chain. This simple and low-cost method was confirmed by treating a public dataset. The secondary structure information of intact antibody was also elucidated from the results of circular dichroism spectrum.

340.3: Comparison of HLA Compatibility Algorithms to Predict Longterm Survival and CLAD Following Lung Transplantation

Introduction: Outcomes following lung transplantation (LTx) remain poor, despite recent advances in sequencing technology and the development of multiple algorithms defining immunological compatibility between recipient and donor. No consensus has been achieved regarding the best approach to define HLA compatibility in LTx. Here, we compared numerous available HLA compatibility approaches, including T- and B-cell epitopes, electrostatic charge and amino acid mismatching, using a high resolution typed, well characterised cohort, to determine which approach best predicts outcomes following LTx. Methods: In this retrospective single-center study, 277 donor-recipient transplant pairs were retrospectively HLA typed by Next generation sequencing (NGS) for all HLA loci. (HLA-A-DPA). HLA mismatching was defined using HLAMatchmaker eplets (epMM), HLA-EMMA epitopes (EMMA), PIRCHE T-cell epitopes (PIRCHE), electrostatic differences (EMS) and amino Acid mismatches (AAMM). Associations with mismatching scores generated by the different algorithms with CLAD (BOS/RAS) and overall survival were calculated using adjusted cox proportional modelling and Kaplan-Meier survival curves to demonstrate time to outcomes (SPSS v.21). Results: Lower HLA-class II compatibility was associated with increased overall survival for all algorithms, with lower HLA-DR, -DQ and –DQA mismatches generated by PIRCHE and epMM independently associated with increased survival. Lower HLA-DR compatibility was significantly associated with increased time to RAS for all algorithms. Lower epMM and PIRCHE HLA-DQA was also associated with increased time to RAS, however the algorithms that did not separate DQA and DQB, did not reach significance. Conclusion: Lower class II mismatching, specifically for HLA-DR and DQ, for all approaches associated with improved graft and patient survival. The PIRCHE and HLAMatchmatcher algorithms that separate compatibility based on individual alleles/chains provide a higher degree of granularity, which may be useful to select the best matched recipient. Regardless, reducing the level of HLA mismatching, in either T- or B-cell epitopes, electrostatic differences or amino acid would improve outcomes following LTx and potentially guide immunosuppression strategies.

Neutralizing antibody correlates of sequence specific dengue disease in a tetravalent dengue vaccine efficacy trial in Asia

In the CYD14 trial of the CYD-TDV dengue vaccine in 2–14 year-olds, neutralizing antibody (nAb) titers to the vaccine-insert dengue strains correlated inversely with symptomatic, virologically-confirmed dengue (VCD). Also, vaccine efficacy against VCD was higher against dengue prM/E amino acid sequences closer to the vaccine inserts. We integrated the nAb and sequence data types by assessing nAb titers as a correlate of sequence-specific VCD separately in the vaccine arm and in the placebo arm. In both vaccine and placebo recipients the correlation of nAb titer with sequence-specific VCD was stronger for dengue nAb contact site sequences closer to the vaccine (p = 0.005 and p = 0.012, respectively). The risk of VCD in vaccine (placebo) recipients was 6.7- (1.80)-fold lower at the 90th vs 10th percentile of nAb for viruses perfectly matched to CYD-TDV, compared to 2.1- (0.78)-fold lower at the 90th vs 10th percentile for viruses with five amino acid mismatches. The evidence for a stronger sequence-distance dependent correlate of risk for the vaccine arm indicates departure from the Prentice criteria for a valid sequence-distance specific surrogate endpoint and suggests that the nAb marker may affect dengue risk differently depending on whether nAbs arise from infection or also by vaccination. However, when restricting to baseline-seropositive 9–14 year-olds, the correlation pattern became more similar between the vaccine and placebo arms, supporting nAb titers as an approximate surrogate endpoint in this population. No sequence-specific nAb titer correlates of VCD were seen in baseline-seronegative participants. Integrated immune response/pathogen sequence data correlates analyses could help increase knowledge of correlates of risk and surrogate endpoints for other vaccines against genetically diverse pathogens.Trial registration: EU Clinical Trials Register 2014-001708-24; registration date 2014-05-26.

316.11: HLA-DPB1 Molecular Mismatches Are Associated With Acute Rejection and Long-term Graft Function in First Kidney Transplants

Introduction: It is generally accepted that HLA-DP mismatches (MM) do not impact the survival of first transplants but that they may exert a deleterious effect in retransplants. This study aimed to investigate the impact of HLA-DPB1 allele and molecular MM (mMM) on the occurrence of acute rejection (AR) and low 5-year graft function (5-yGF) in first kidney transplants (KT). Method: HLA-DPB1 allele and mMM were determined in 130 first KT from deceased donors, performed between 2014 and 2016. The following mMM models were investigated: expression MM, with the high expression G allele in the donor, T cell epitope (TCE) MM, classified as permissive and nonpermissive MM, epitope MM (EMM), considering all six HLA-DPB1 hypervariable regions (EMM-ABCDEF HVR) or only ABEF regions (EMM-ABEF HVR), Eplet MM (EpMM), considering all or only antibody-verified (AbVer) eplets, and solvent accessible amino acid MM (SAMM). EMM, EpletMM and SAMM loads were categorized using thresholds determined by ROC analysis for each outcome. The outcomes were AR and 5-yGF (estimated by CKD-EPI). Logistic and Cox regressions were used to investigate variables associated with 5-yGF and AR, respectively. Results: 119 (91.5%) KT were performed with one or two HLA-DPB1 allele MM. The overall incidence of AR was 17.7% and the median 5-yGF was 40.3 (27.0 - 51.7) mL/min/1.73m2. No association between HLA-DPB1 allele MM and 5-yGF or rejection was observed. Considering only recipients of standard criteria donor, five logistic models for 5-yGF were built including variables that reached a p-value <0.10 in the univariable analysis (age and donor sex, cause of brain death, ABDR MM and AR) and the mMM (TCE, SAMM, all and AbVer EpMM, EMM-ABCDEF and ABEF HVR). In the model 2, variables associated with 5-yGF≤ 40 were donor age (OR=1.08; CI95%=1.02-1.14; p=0.005) and SAMM≥7 (OR=4.40; CI95%=1.56-12.36, p<0.005), while in the model 4, donor age (OR= 1.09; CI95%=1.03-1.15; p=0.003) and EpMM(AbVer≥2 (OR=3.39; CI95%=1.2-9.45, p=0.02) were associated with low GF. The AUC-ROC for predicting 5-yGF was 0.83 (0.75-0.92) for the model 2, and 0.68 (0.57-0.79) for the model 4. Lastly, for the outcome AR in all recipients, beyond the final donor creatinine, the following mMM models were statistically significant: TCE (HR=3.01; CI95%=1.33-6.84; p=0.008), SAMM≥5 (HR=2.69; CI95%=1.10-6.56; p=0.03), AbVerEpMM ≥3 (HR=3.38; 95%CI=1.32-8.70; p=0.01), and EMM-ABCDEF HVR ≥ 6 (HR=5.08; 95%CI=2.19-11.82; p<0.001). Conclusion: To the best of our knowledge, this is the first study that shows that some mMM, in contrast to classical allele HLA-DPB1 MM, are associated with acute rejection and long-term graft function in a population of exclusively first kidney transplants.

Association of HLA Mismatches and Histology Suggestive of Antibody-Mediated Injury in the Absence of Donor-Specific Anti-HLA Antibodies.

The histology of antibody-mediated rejection after kidney transplantation is observed frequently in the absence of detectable donor-specific anti-HLA antibodies. Although there is an active interest in the role of non-HLA antibodies in this phenotype, it remains unknown whether HLA mismatches play an antibody-independent role in this phenotype of microcirculation inflammation. To study this, we used the tools HLAMatchmaker, three-dimensional electrostatic mismatch score, HLA solvent accessible amino acid mismatches, and mismatched donor HLA-derived T cell epitope targets to determine the degree of HLA molecular mismatches in 893 kidney transplant recipients with available biopsy follow-up. Multivariable Cox proportional hazards models were applied to quantify the cause-specific hazard ratios of the different types of HLA mismatch scores for developing antibody-mediated rejection or histology of antibody-mediated rejection in the absence of donor-specific anti-HLA antibodies. In all survival analyses, the patients were censored at the time of the last biopsy. In total, 121 (14%) patients developed histology of antibody-mediated rejection in the absence of donor-specific anti-HLA antibodies, of which 44 (36%) patients had concomitant T cell-mediated rejection. In multivariable Cox analysis, all different calculations of the degree of HLA mismatch associated with developing histology of antibody-mediated rejection in the absence of donor-specific anti-HLA antibodies. This association was dependent neither on the presence of missing self (potentially related to natural killer cell activation) nor on the formation of de novo HLA antibodies. Also, glomerulitis and complement C4d deposition in peritubular capillaries associated with the degree of HLA mismatch in the absence of anti-HLA antibodies. The histology of antibody-mediated rejection and its defining lesions are also observed in patients without circulating anti-HLA antibodies and relate to the degree of HLA mismatch.

Snowflake: A deep learning-based human leukocyte antigen matching algorithm considering allele-specific surface accessibility.

Histocompatibility in solid-organ transplantation has a strong impact on long-term graft survival. Although recent advances in matching of both B-cell epitopes and T-cell epitopes have improved understanding of allorecognition, the immunogenic determinants are still not fully understood. We hypothesized that HLA solvent accessibility is allele-specific, thus supporting refinement of HLA B-cell epitope prediction. We developed a computational pipeline named Snowflake to calculate solvent accessibility of HLA Class I proteins for deposited HLA crystal structures, supplemented by constructed HLA structures through the AlphaFold protein folding predictor and peptide binding predictions of the APE-Gen docking framework. This dataset trained a four-layer long short-term memory bidirectional recurrent neural network, which in turn inferred solvent accessibility of all known HLA Class I proteins. We extracted 676 HLA Class-I experimental structures from the Protein Data Bank and supplemented it by 37 Class-I alleles for which structures were predicted. For each of the predicted structures, 10 known binding peptides as reported by the Immune Epitope DataBase were rendered into the binding groove. Although HLA Class I proteins predominantly are folded similarly, we found higher variation in root mean square difference of solvent accessibility between experimental structures of different HLAs compared to structures with identical amino acid sequence, suggesting HLA’s solvent accessible surface is protein specific. Hence, residues may be surface-accessible on e.g. HLA-A*02:01, but not on HLA-A*01:01. Mapping these data to antibody-verified epitopes as defined by the HLA Epitope Registry reveals patterns of (1) consistently accessible residues, (2) only subsets of an epitope’s residues being consistently accessible and (3) varying surface accessibility of residues of epitopes. Our data suggest B-cell epitope definitions can be refined by considering allele-specific solvent-accessibility, rather than aggregating HLA protein surface maps by HLA class or locus. To support studies on epitope analyses in organ transplantation, the calculation of donor-allele-specific solvent-accessible amino acid mismatches was implemented as a cloud-based web service.

Improving Access to HLA-Matched Kidney Transplants for African American Patients.

IntroductionKidney transplants fail more often in Black than in non-Black (White, non-Black Hispanic, and Asian) recipients. We used the estimated physicochemical immunogenicity for polymorphic amino acids of donor/recipient HLAs to select weakly immunogenic kidney transplants for Black vs. White or non-Black patients.MethodsOPTN data for 65,040 donor/recipient pairs over a 20-year period were used to calculate the individual physicochemical immunogenicity by hydrophobic, electrostatic and amino acid mismatch scores (HMS, EMS, AMS) and graft-survival outcomes for Black vs. White or vs. non-Black recipients, using Kaplan-Meier survival and Cox regression analyses. Simulations for re-matching recipients with donors were based on race-adjusted HMS thresholds with clinically achievable allocations.ResultsThe retrospective median kidney graft survival was 12.0 years in Black vs. 18.6 years in White (6.6-year difference; p>0.001) and 18.4 years in non-Black (6.4-year difference; p>0.01) recipients. Only 0.7% of Blacks received transplants matched at HLA-A/B/DR/DQ (HMS=0) vs. 8.1% in Whites (p<0.001). Among fully matched Blacks (HMS=0), graft survival was 16.1-years and in well-matched Blacks (HMS ≤ 3.0) it was 14.0-years. Whites had 21.6-years survival at HMS ≤ 3.0 and 18.7-years at HMS ≤ 7.0 whereas non-Blacks had 22.0-year at HMS ≤ 3.0 and 18.7-year at HMS ≤ 7.0, confirming that higher HMS thresholds produced excellent survival. Simulation of ABO-compatible donor-recipient pairs using race-adjusted HMS thresholds identified weakly immunogenic matches at HMS=0 for 6.1% Blacks and 18.0% at HMS ≤ 3.0. Despite prioritizing Black patients, non-Black patients could be matched at the same level as in current allocation (47.0% vs 56.5%, at HMS ≤ 7.0).ConclusionsRace-adjusted HMS (EMS, AMS)-based allocation increased the number of weakly immunogenic donors for Black patients, while still providing excellent options for non-Black recipients.