Prolonged wait times among highly sensitized HLA-B46 homozygous kidney transplant candidates.

Immunized platelet transfusion refractoriness (immunized-PTR) correlates with poor outcomes in hematological malignancies (HM) patients during allogeneic hematopoietic stem-cell transplantation (allo-HSCT). The efficacy and outcomes of desensitization with rituximab (RTX) and therapeutic plasma exchange (TPE) for immunized-PTR remain controversial. The study included 83 Ab-negative non-PTR patients and 65 patients with severe immunized-PTR who underwent allo-HSCT. Based on pre-transplant desensitization, 30 and 35 patients were classified into the Ab-positive PTR-treatment and Ab-positive PTR groups. After desensitization, HLA antibodies were decreased (71 ± 19 vs. 26 ± 22, P <0.01), 14h-corrected count increment (CCI) was increased(14.64 ± 5.36 vs. 2.00 ± 1.33, P <0.001), and the cumulative incidence of platelet engraftment at day 21 and 6 months was higher (93.3% vs. 82.9% in Ab-positive PTR group, P = 0.0002 and 90.0% vs. 67.1% in Ab-positive PTR group, P = 0.0005), significantly. Compared to Ab-positive PTR groups, desensitization leads to a better prognosis in Ab-positive PTR-treatment group (3-year OS: 93.3% vs. 47.0%, P <0.0001; 3-year LFS: 83.0% vs. 48.0%,respectively, P <0.0001). Multivariable analysis revealed that desensitization was independently associated with better OS (HR, 0.068; 95%CI, 0.014-340; P = 0.001) and LFS (HR, 0.090; 95%CI, 0.023-358; P = 0.001) for PTR patients. This result indicates that immunized-PTR is associated with poor outcomes in HM patients after allo-HSCT, and desensitization with RTX and TPE achieve favorable outcomes.

Kidney transplantation remains the most effective treatment for end-stage kidney disease. Still, the development of de novo donor-specific antibodies (dnDSA) increases the risk of rejection and allograft failure. While molecular matching algorithms assess B-cell and T-cell epitope mismatches, no single method fully captures rejection risk across immune pathways. This study combines the HLA Epitope Registry (Epregistry), PIRCHE-T2, and PIRCHE-B scores to enhance risk stratification, allowing for early intervention in high-risk recipients and improving long-term outcomes. A retrospective study of 594 kidney transplant recipients in Saskatchewan (1981-2021), Canada, was conducted, tracking de novo donor-specific antibodies (dnDSA) development until January 2024. Epitope mismatch scores were calculated using Epregistry, PIRCHE-T2, and PIRCHE-B, and receiver operating characteristic (ROC) curve analysis determined the optimal cutoff values for predicting dnDSA formation. Patients were categorized into high-risk (all scores > cutoff), intermediate-risk (one algorithm > cutoff), and low-risk (all scores < cutoff) groups. Kaplan-Meier survival analysis evaluated dnDSA-free survival across risk categories. Among 594 recipients, 104 individuals (17.5%) developed de novo DSA; of these, 29 patients developed more than one, resulting in a total of 146 dnDSA events. The most frequently targeted locus was HLA-DQ (72/146, 49.3%), followed by HLA-DR (25/146, 17.1%) and HLA-A (24/146, 16.4%). The optimal cutoff values for predicting dnDSA were 22.5 (Epregistry), 30.5 (PIRCHE-T2), and 5.5 (PIRCHE-B) for Class I, and 15.5 (Epregistry), 17.5 (PIRCHE-T2), and 5.5 (PIRCHE-B) for Class II (all p < 0.05). Across all molecular mismatch load metrics, Kaplan-Meier analysis demonstrated significantly lower dnDSA-free and antibody-mediated rejection (ABMR)-free survival among high-risk recipients compared with low-risk recipients (log-rank p < 0.001). In addition, both the PIRCHE-T2 score at HLA Class I loci and the overall PIRCHE-T2 score were significantly associated with T-cell mediated rejection (TCMR) (p < 0.01). Integrating Epregistry, PIRCHE-T2, and PIRCHE-B enhances risk stratification for kidney transplant recipients. Epregistry and PIRCHE-B evaluate HLA antibody epitope mismatches, and PIRCHE-T2 focuses on T-cell mismatches. Applied in conjunction, the methods show improved predictive accuracy, making this multi-algorithm approach more effective in identifying high-risk patients. By enabling earlier interventions and personalized immunosuppressive strategies, this model has the potential to improve long-term transplant success.

Background/Objectives: While standard Luminex single antigen bead (SAB) detects total IgG antibodies, qualitative differences among IgG subclasses may influence their immunologic risk. In particular, complement fixing ability, assessed via C1q binding, is linked to poor transplant outcomes. This study aimed to evaluate the relationship between IgG subclasses and C1q-binding activity in HLA antibodies and to define clinically relevant subclass-specific mean fluorescence intensity (MFI) thresholds for predicting complement binding. Methods: We analyzed 4189 HLA IgG bead reactions from sera of 37 kidney transplant recipients using SAB assays for total IgG, IgG1-4 subclasses, and C1q-binding. IgG subclasses were assessed using a modified SAB assay with subclass-specific monoclonal secondary antibodies. Results: IgG reactivity (MFI ≥ 1000) was observed in 15.3% of beads (639/4189), with 31.0% (198/639) also positive for C1q binding. IgG+C1q+ beads exhibited significantly higher MFIs compared with IgG+C1q− beads. IgG1 showed positive correlations with both total IgG (rs = 0.5439, p < 0.0001) and C1q MFIs (rs = 0.4042, p < 0.0001), with the strongest correlations at HLA-DQ. Among subclass-positive beads, IgG1 predominated and was strongly associated with C1q binding, whereas isolated IgG2 or IgG4 positivity was rarely C1q-binding. ROC analysis identified an IgG1 MFI threshold of >837 to predict C1q positivity with 73.2% sensitivity and 92.3% specificity, while the cutoff for total IgG MFI was >7881 with 85.4% sensitivity and 88.9% specificity. At the patient level, IgG1-positive immunodominant DSAs were more frequent in antibody-mediated rejection than in non-rejection biopsies Conclusions: IgG1 predominates among complement-fixing antibodies and correlates strongly with total IgG and C1q binding. Quantitative IgG subclass assessment, especially IgG1, may serve as a useful predictor of complement activation.

Pregnancy requires local immune tolerization. Oocyte donation (OD) pregnancies, with extensive fetal-maternal human leukocyte antigen (HLA) mismatching, are at higher risk of pre-eclampsia. We hypothesize that immune adaptations are needed at the fetal-maternal interface to maintain healthy pregnancy despite high HLA dissimilarity. By multispectral imaging, myeloid cells constituted 65% of the decidual immune cells and encompassed 12 distinct clusters. Fully-allogeneic healthy OD pregnancies displayed a higher frequency of CD163+HLA-DR+ myeloid cells and FoxP3+CD4+ Tregs near CD4+ T cells compared to semi-allogeneic healthy pregnancies and pre-eclampsia, together with a Treg-reinforcing gene signature. Contrastingly, pre-eclampsia was characterized by enhanced inflammatory chemokine expression and oxidative-stress-gene imbalance. Pregnancy outcomes were unaffected by decidual pathology, maternal HLA antibodies, or fetal HLA-C/maternal KIR haplotypes. This study highlights the frequency, phenotypic diversity, and T cell proximity of decidual myeloid cells in OD pregnancies and suggests their immune regulatory effects to compensate for higher fetal-maternal HLA mismatch loads.

Practice Variability in the Evaluation and Management of HLA Alloimmunization in Platelet Refractoriness: A Multi-Institutional Survey in the U.S.

Sensitization in Organ Transplantation: Assessment of Risk (STAR) 2025 Meeting Group Report.

IntroductionXenotransplantation using genetically modified pigs is a promising solution to organ shortages, particularly for highly sensitized patients with broad anti-HLA sensitization who lack compatible allografts. However, preformed human anti-HLA antibodies may cross-react with porcine SLA, posing a barrier for clinical application. This study aims to characterize the extent and specificity of cross-reactive antibody responses against genetically engineered pig PBMCs, particularly from quadruple knockout (QKO) pigs.MethodsWe evaluated antibody binding and cytotoxicity of 68 human sera stratified by HLA class I and II antibody profiles using flow cytometric crossmatch (FCXM), complement-dependent cytotoxicity (CDC) assays (CDC-NIH and CDC-AHG), antibody elution, and single antigen bead assays. Porcine PBMCs from wild-type and gene-edited pigs (GTKO, TKO, QKO) were used. High-resolution SLA epitope mapping was performed with antibody eluted from select sera followed by in silico sequence and structural analyses.ResultsHuman sera showed strong IgG and IgM binding to wild-type pig PBMCs, which was significantly reduced by RBC adsorption, whereas binding to QKO pig PBMCs lacking key glycan xenoantigens was minimal and unaffected by RBC adsorption. Sensitized human sera with both HLA class I and II antibodies demonstrated significantly elevated IgG binding to QKO pig PBMC T and B cells compared to antibody-negative sera (p < 0.05). CDC-AHG assays revealed increased cytotoxicity titers (≥1:8) in HLA antibody-positive sera versus negatives (p < 0.01). Antibody elution from five crossmatch-positive sera identified predominant class I eplets (62EE, 162GLS, 163LG, 163LS/G, 166ES, 199V) and class II DR epitopes (13SE, 37F, 47F, 70D, 70DA) that target SLA 4.5/6.7 haplotypes. In contrast, anti-HLA-DQ and -DP reactivity was limited post-elution. Structural modeling confirmed that these epitopes are conserved and surface-exposed in SLA alleles.ConclusionCross-reactive anti-SLA antibodies are common in highly sensitized human sera, driven by antibody specificity and epitope conservation. Despite glycan xenoantigen deletion, sensitized sera maintain IgG-mediated cross-reactivity and cytotoxicity against gene-edited pig cells. These findings highlight the need for detailed epitope-level analysis to refine immunologic risk assessment and recipient selection to reduce antibody-mediated rejection in clinical xenotransplantation.

BackgroundAn increased risk of severe infectious disease and acute antibody-mediated rejection (AMR) has been described after ABOi renal transplantation. We performed a prospective renal transplant study up to 5 years posttransplant to detect long-term immunological effects of rituximab administration.MethodsMononuclear cell subsets in peripheral blood, regional lymph nodes and protocol biopsies, in-vitro T and B cell responses, serum sCD30 and neopterin were assessed in 85 renal transplant recipients (living donation: n=25 ABOi, n=30 ABO compatible (ABOc); deceased donation (DD): n=30, all ABO compatible), and IgG anti-HLA antibodies by single antigen assay in ABOc and ABOi patients.ResultsAn increased frequency of severe infectious diseases in ABOi recipients (doubled versus ABOc within 2 years, P = 0.042) coincided with profoundly downregulated peripheral blood B cell subsets for at least 2 years, impaired in-vitro B cell responses (T-dependent: 2 years, versus ABOc: P = 0.004) and significantly lower CD4+ and CD8+ T cell counts (versus ABOc; 6 months, P = 0.046 and 3 months, P = 0.011, respectively). In regional lymph nodes, we found a significant downregulation of naive B cells (P = 0.031) and short lived plasma cells (P<0.0005) at the time of transplantation. In protocol graft biopsies, rituximab induced B cell depletion at 3 months (P<0.001), but counter-regulatory enhanced counts of T cells (P = 0.041), macrophages (P = 0.021) and plasma cells (P = 0.033) at 1 year. This immune activation was associated with a temporary rise in neopterin levels (P ≤ 0.024 versus ABOc, day 14 until 1 year), CD4 helper activity (P = 0.019 versus ABOc at 2 years) and NK cell counts (P = 0.034 versus ABOc, 4 years; P ≤ 0.040 versus DD, 3–5 years), a missing impact of rituximab on sCD30 levels and HLA antibody formation, and an increased frequency of biopsy-proven acute rejection (3–12 months, P = 0.003) and AMR (P = 0.008 within 5 years).ConclusionsAn increased frequency of severe infectious diseases in ABOi renal transplant recipients may be explained by rituximab-induced long-term immunological effects on CD4+ and CD8+ T cell counts and the prolonged depletion of B cell subsets together with compromised B cell responses. In protocol graft biopsies, rituximab induced early B cell depletion but counter-regulatory proinflammatory effects coinciding with an increased acute rejection frequency.Registration DetailsClinicalTrials.gov, identifier NCT01136395; EudraCT, identifier No.: 2009-012198-36.

ABSTRACT Background Kidney transplantation is the preferred therapy for end-stage renal disease (ESRD), yet long-term graft survival remains limited. Effective preservation of transplanted kidneys is essential amid a persistent organ shortage. Post-transplant graft injury arises from both immune and nonimmune mechanisms. Advances in multi-omic technologies have increasingly unraveled pathways underlying graft rejection and tolerance. While most studies have centered on intragraft immune-cell infiltration and circulating biomarkers, the role of antibodies in acute rejection and other graft injuries is not fully studied. Research design and methods In this exploratory study, we employed a high-throughput antibody-profiling microarray to quantitatively assess IgG antibodies against Human Leukocyte Antigen (HLA) and non-HLA (nHLA) in urine samples from kidney transplant recipients experiencing graft injury, including acute rejection. Results We identified multiple HLA and nHLA antibodies that were selectively enriched in urine at the time of graft injury and acute rejection, indicating antigen-specific humoral responses detectable at the site of injury. Conclusions This first-of-its-kind urinary antibody-profiling study reveals promising antibody signatures associated with graft injury. These findings support the potential development of noninvasive, personalized biomarkers for routine monitoring and earlier detection of rejection in kidney transplant recipients.

Induction of immune education in type 1 diabetes through controlled allogeneic islet rejection at onset: a monocentric open-label pilot study

Transplant outcomes after de novo donor specific antibody (DSA) development correlate with persistence of DSA.

Humoral rejection in heart transplantation in the current era

ABSTRACTBackgroundFollowing pediatric liver transplantation (pLT), the significance and management of donor‐specific antibodies (DSA) against human leukocyte antigen (HLA) remain undefined. The aim of this single‐center study was to investigate the occurrence of DSA, their clinical impact on predictors for and protectors against DSA.Patients and MethodsWe compared anti‐HLA DSA (cutoff for mean fluorescence intensity (MFI) ≥ 1000), clinical and laboratory results and outcome in a routine (RG, n = 142, standard DSA testing) and a hepatopathy group (HG, n = 19, DSA testing following indeterminate hepatopathy) in 161 pLT patients, treated 2000–2021, retrospectively.Results40% of RG and 32% of HG patients were DSA+ (39% of all patients, of which 13% with antibody‐mediated rejection [AMR]). Most frequent DSA subtypes were HLA‐DQ3, ‐DQ1, ‐DQ2 in RG and HLA‐DQ2, ‐DR15 in HG. MFI was higher for anti‐HLA II DSA (15 257 DSA+ vs. 5500 DSA−, p = 0.003), especially with AMR (21 000 DSA+ with AMR vs. 14 584 DSA+ without AMR, p = 0.042). Predictors for DSA included age at pLT, re‐pLT, and cystic fibrosis. Living donation and cold ischemia time <8 h appeared to offer protection. Graft survival was poorer with DSA (RG 78% DSA+ vs. 97% DSA−, p = 0.018, HG 67% DSA+ vs. 100% DSA−, p = 0.0007). Patient survival was 97% for the entire cohort.ConclusionsDSA were detectable in 39% and associated with AMR in 13% of children post‐pLT in addition to worse graft survival in all patients. Patient survival of 97% was not influenced. Potential DSA and predictors and protectors were identified. Therefore, DSA diagnostics are recommended after pLT.

Understanding Human Leukocyte Antigens in Vascularized Composite Allotransplantation.

Acute kidney allograft dysfunction is associated with increased number of non-HLA antibodies in absence of donor specific HLA antibodies

Abstract Objectives Immune-mediated platelet refractoriness is a challenge in appropriate clinical management of hematopoietic stem cell transplant (HSCT) and cancer patients. Our objective was to assess the real-world clinical utility of recommendations from published guidelines that two consecutive post-transfusion one-hour corrected count increments (CCIs) be collected prior to acquiring specific class I HLA antibody information by a more specific assay. Methods We assessed three distinct aspects of post-transfusion CCIs at our institution. We examined institutional adherence to standard recommendations that two post-transfusion platelet counts (CCIs) (1) be collected in advance of anti-HLA assessment by Luminex™; (2) when done, be collected within 10-60 min of transfusion; and (3) are superior to next-day CCIs. Thrombocytopenic HSCT and cancer patients between Aug23 - Aug24 having initial evaluation of anti-HLA Abs by Luminex™ were identified in the EMR. (1) Manual chart review identified whether none, one, or two CCIs were obtained in advance of sample collection for Luminex. (2) Through analysis of EMR data, we determined the actual time interval between end of transfusion and collection of the “one-hour” post-cbc. (3) For patients with available data, 10-60 min and next-day CCI’s were compared by linear regression. Results For the first analysis, 77 patients with initial Luminex™ collections were evaluated. The percentage of Luminex™ results positive for antibodies to HLA (Panel Reactive Antibodies (PRA) &amp;gt; 0%) was: 38% (14/37) when no CCI had been collected in advance of Luminex™ testing; 36% (9/25), when one CCI had been collected in advance, and only 27% when the recommended two CCIs had been collected in advance. Based on two papers published in 1980 and 1989, current guidelines state that the post-platelet count is to be collected within 10-60 min of the end of transfusion. In the dataset examined, only 25% of CCI’s were collected within the recommended 10-60 min timeframe; the rest were obtained either before 10 min (17%) or between 1-7 h post-transfusion (58%). For outpatients undergoing daily prophylactic platelet transfusions (n = 50), true 10-60 min and next-day CCIs (n = 50) were highly correlated (r² = 0.54; p &amp;lt; 0.0001). Conclusions Published literature/guidelines indicate that collecting two consecutive 10-60 min post-platelet-transfusion counts is clinically useful in identifying patients likely to be immune refractory due to anti-HLA antibodies, and therefore likely to benefit from transfusion of HLA-selected (antigen-negative) platelet units. Our clinical practice data show that it is a challenge to have one-hour post CCIs consistently collected, that most such CCIs are collected outside the 10-60 min recommended window, and that they may not be superior to next day platelet counts. These findings call into question the utility of 10-60 min CCIs as screening assays prior to more specific testing by Luminex™.

Rationale:Platelet transfusions are the key treatment for patients with severely low platelet counts and significant bleeding symptoms. Platelet transfusion refractorines (PTR) is defined as unsatisfactory transfusion of platelets with at least 2 ABO blood groups matched and stored for <72 hours.Patient concerns:A 37-year-old woman presented for medical evaluation due to the cessation of fetal heart activity after 50 days of gestation.Diagnoses:AML-M2, PTR.Interventions:To explore potential immune-mediated causes of platelet refractoriness, the following diagnostic evaluations were conducted: Platelet antibody screening using the PAK-Lx assay (Immucor) to detect anti-HLA class I and HPA antibodies. Anti-HLA class I specificity determination via LIFECODES LSA Single Antigen bead assays (Luminex). Molecular typing of HLA class I and HPA alleles using PCR-sequence based typing (PCR-SBT), as well as CD36 phenotyping via flow cytometry. Confirmation testing for anti-HPA-5b antibodies using chloroquine-treated platelet solid-phase assays.Outcomes:The patient had high-fluorescence-intensity HLA antibodies along with HPA antibodies, resulting in severe refractoriness to platelet transfusion.Lessons:Various methods were used to comprehensively analyze the specificity of anti-platelet antibodies in the samples of patients with ineffective platelet transfusion, thus providing a reference for the clinical diagnosis and treatment of PTR.

Human leukocyte antigen sensitization from red cell transfusions: Rate and risk factors in renal and nonrenal patients requiring transfusion.

Fetal/neonatal alloimmune thrombocytopenia (FNAIT) and neonatal alloimmune neutropenia (NAIN) result from maternal alloantibodies targeting fetal human platelet antigen (HPA) and human neutrophil antigen (HNA) antigens, respectively. However, increasing evidence supports the pathogenic role of HLA class I alloantibodies in these conditions. Since the simultaneous occurrence of FNAIT and NAIN has not been systematically investigated, this study aimed to determine its prevalence, characterize the specificity and strength of associated alloantibodies, and correlate findings with neonatal cell counts. In this cross-sectional study, 10,000 umbilical cord blood samples were analyzed for platelet and neutrophil counts. Neonates with thrombocytopenia and neutropenia were selected. Genotyping for HPA, HNA, and HLA class I was performed in mother-infant pairs to assess incompatibilities. Maternal sera were tested for anti-HPA, anti-HNA, and anti-HLA antibodies. Ten cases (0.1%) of concurrent cytopenias were identified. Alloantibodies were detected in four cases: one with combined anti-HPA-5b, HNA-2, and HLA-A2 antibodies; and three with isolated high-mean fluorescence intensity (MFI) HLA antibodies (anti-HLA-A2, HLA-A3, HLA-B7). Anti-HLA-A2 was linked to the lowest neutrophil counts, and anti-HLA-B7 to severe thrombocytopenia. The estimated prevalence of simultaneous FNAIT and NAIN was 0.04% (1 in 2500 neonates). This is the first large-scale study to document the co-occurrence of FNAIT and NAIN. Our findings explore the serological and molecular features of these immune syndromes and underscore the potential pathogenic role of maternal anti-HLA class I antibodies, even in the absence of detectable anti-HPA or anti-HNA, and support including HLA testing in the diagnostic workup of neonatal cytopenias.