Donor-specific HLA Antibodies Research Articles

NAVIGATING THE UNCHARTED WATERS: MANAGING CONCURRENT CYTOMEGALOVIRUS MYOCARDITIS AND ALLOGRAFT REJECTION IN A PATIENT WITH HEART TRANSPLANTATION

Abstract A 49–year–old female with hypertrophic cardiomyopathy underwent orthotopic heart transplantation. She was cytomegalovirus(CMV)–positive and received a heart from a CMV–negative donor, so prophylaptic therapy was not administered. Initial immunosuppression included cyclosporine and induction with Basiliximab in a steroid–free regimen. The first routine biopsy (EMB) showed mild focal cellular rejection (grade 1A) and C4d positivity with granular aspect. Mycophenolate mofetil (MMF) was added. The second EMB, 7 days later, showed mild diffuse cellular rejection (grade 1B) and diffuse C4d positivity. Anti–HLA donor–specific antibodies (DSA) had increased anti–B13 levels (MFI=21298) compared to pre–transplantation tests. Echocardiography showed a normal ejection fraction (LVEF) and mild concentric hypertrophy (LVH). Treatment included pulse methylprednisolone therapy (1 g/day for three days) followed by oral prednisone with a slow tapering regimen, and increased MMF. CMV count weekly monitored was negative. The next two EMBs performed one week apart from each other described a grade 1A mild focal pattern while the 5th EMB (1 month and 2 weeks post–transplant) showed a grade 1A rejection together with focal mild fibrosis and eosinophils, so we replaced cyclosporine with tacrolimus, increased MMF and prednisone. At this time, the patient had no overt clinical signs of CMV infection but a CMV–PCR of 100847 IU/mL so ganciclovir 5 mg/Kg bid was started and CMV–PCR decreased to 42308 IU/mL after 1 week. Subsequently, she developed sinus tachycardia, mildly reduced LVEF, moderate LVH and diastolic dysfunction. Troponin was elevated. A new EMB revealed interstitial edema, diffuse ischemic injury, and eosinophilic inclusions suggesting CMV myocarditis. Treatment involved down–titrating MMF and Prednisone, administering standard intravenous immunoglobulin (IVIG), and high–dose Ganciclovir (7.5 mg/Kg bid). Leukopenia subsequently led to Ganciclovir down–titration. In the next days, the patient improved, CMV cleared, and leukocyte count normalized. She was discharged on Tacrolimus, MMF, Prednisone and Valganciclovir. Subsequent tests showed no cellular or antibody–mediated rejection (AMR), no CMV inclusions, recovered cardiac function, and markedly reduced anti–B13 DSA (MFI 3000). This case raises some concerns: in this context, the use of IVIG in the treatment of CMV infection and AMR appears reasonable, in support of antiviral and immunosuppressive therapy.

Very Low Frequency of Pathological Findings in One-year Protocol Biopsies of Uneventful Standard Risk Kidney Transplant Recipients: Results From the Nordic Protocol Biopsy Study.

The clinical significance of kidney transplant protocol biopsies has been debated. We studied the frequency of borderline changes and T cell-mediated rejection (TCMR) in 1-y protocol biopsies in standard risk kidney transplant recipients. Consecutive non-HLA-sensitized recipients of kidney transplants between 2006 and 2017, who underwent a protocol biopsy at 1 y in 2 national transplant centers were studied retrospectively (N = 1546). Donor-specific HLA antibodies (DSAs), graft function (plasma creatinine), and proteinuria were measured at the time of 1-y protocol biopsy. The occurrence of subclinical acute TCMR (i2t2v0 or higher) or borderline changes suspicious of TCMR (i1t1v0 or higher) in the protocol biopsy was studied, together with frequency of findings causing changes in the composite score iBox. Subclinical acute TCMR was detected in 30 of 1546 (1.9%) of the protocol biopsies, and borderline or TCMR in 179 of 1546 (12%). Among patients with no history of acute rejection, and no proteinuria or DSA, TCMR was detected in only 1 of 974 (0.1%) and borderline or TCMR in only 48 of 974 (4.9%) patients at 1 y. In the absence of proteinuria (<30 mg/g, or equivalent as measured with a negative dipstick proteinuria) or DSA, or history of acute rejection, only 50 of 974 (5.1%) biopsies showed any lesions significant for the iBox score. The likelihood of pathological findings in 1-y protocol biopsies in non-HLA-sensitized patients without previous immunological events is low. Clinical usefulness of protocol biopsies seems limited in these patients.

(415) - Microvascular Responses to Donor Specific HLA Antibodies in Heart Transplantation. Sidestream Dark Field Imaging Results

(415) - Microvascular Responses to Donor Specific HLA Antibodies in Heart Transplantation. Sidestream Dark Field Imaging Results

(342) - Antithymocyte Globulin as an Adjunctive Therapy for Antibody-Mediated Rejection in Heart Transplant Recipients More Consistently Decreased Donor-Specific HLA Antibodies and Histopathologic Grading Than PP/IVIG Alone

(342) - Antithymocyte Globulin as an Adjunctive Therapy for Antibody-Mediated Rejection in Heart Transplant Recipients More Consistently Decreased Donor-Specific HLA Antibodies and Histopathologic Grading Than PP/IVIG Alone

New treatment for antibody-mediated rejection: interleukin-6 inhibitors.

Following kidney transplantation, antibody-mediated rejection (AMR) occurs when the antibodies of the immune system attack the transplanted organ, leading to damage of the kidney tissue. De novo human leukocyte antigen donor-specific antibodies (HLA-DSAs) play a key role in AMR. Current therapeutic approaches include intravenous immunoglobulin, anti-CD20 antibodies, and plasmapheresis. In cases resistant to treatment, proteasome inhibitors and C5 inhibitors may be employed. Nevertheless, a pressing need exists for new medications to improve transplant survival and reduce complications. In the context of AMR, interleukin (IL)-6 is instrumental in the development and maturation of B cells into plasma cells, which then produce HLA-DSAs targeting the allograft. IL-6 inhibitors are currently under investigation and show promise due to the essential role of IL-6 in the immune response; however, additional research is necessary.

PIRCHE application major versions 3 and 4 lead to equivalent T cell epitope mismatch scores in solid organ and stem cell transplantation modules

PIRCHE scores in organ and stem cell transplantation have been shown to correlate with increased risk of donor-specific HLA antibodies and graft-versus-host disease, respectively. With advancements of the PIRCHE application server, it is critical to compare the predicted scores with previous versions. This manuscript compares the newly introduced PIRCHE version 4.2 with its predecessor version 3.3, which was widely used in retrospective studies, using a virtual cohort of 10,000 transplant pairs. In the stem cell transplantation module, both versions yield identical results in 100% of the test population. In the solid organ module, 97% of the test population has identical PIRCHE scores. The deviating cases (3%) were attributed to refinements in the PIRCHE algorithm’s specification. Furthermore, the magnitude of the difference is likely to be below the detection limit for clinical effects, confirming the equivalence in PIRCHE scores between versions 3.3 and 4.2.

Role of HLA matching and donor specific antibody development in long-term survival, acute rejection and cardiac allograft vasculopathy

Although there are different data supporting benefits of HLA matching in kidney transplantation, its role in heart transplantation is still unclear. HLA mismatch (MM) between donor and recipient can lead to the development of donor-specific antibodies (DSA) which produces negative events on the outcome of heart transplantation. Moreover, DSAs are involved in the development of antibody-mediated rejection (AMR) and are associated with an increase in cardiac allograft vasculopathy (CAV). In this study it is analyzed retrospectively the influence of HLA matching and anti-HLA antibodies on overall survival, AMR and CAV in heart transplantation. For this retrospective study are recruited heart transplanted patients at the Cardiac Transplantation Centre of Naples between 2000 and 2019. Among the 155 heart transplant patients, the mean number of HLA-A, B, -DR MM (0 to 6) between donor and recipient was 4.5 ± 1.1. The results show a negative association between MM HLA-DR and survival (p = 0.01). Comparison of patients with 0–1 MM at each locus to all others with 2 MM, for both HLA class I and class II, has not showed significant differences in the development of CAV. Our analysis detected DSA in 38.1% of patients. The production of de novo DSA reveals that there is not an influence on survival (p = 0.72) and/or AMR (p = 0.39). Instead, there is an association between the production of DSA class II and the probability of CAV development (p = 0.03). Mean fluorescence intensity (MFI) values were significantly higher in CAV-positive patients that CAV-negative patients (p = 0.02). Prospective studies are needed to evaluate HLA class II matching as an additional parameter for heart allocation, especially considering the increment of waiting list time.

The impact of ageing on the distribution of preformed anti-HLA and anti-MICA antibody specificities in recipients from eastern China prior to initial HSCT

BackgroundWith the development of Hematopoietic Stem Cell Transplantation (HSCT) technology, increasing numbers of elderly patients were undergoing allogeneic HSCT and elderly patients with hematologic malignancies could benefit most from it. Preformed donor-specific human leukocyte antigen (HLA) antibodies (DSA) were associated with graft failure in HLA-mismatched allogeneic HSCT and the absence of DSA was the main criterion of selecting the donor. Except for sensitization events such as transfusion, pregnancy or previous transplantation, ageing affects the humoral immune response both quantitatively and qualitatively. To evaluate the prevalence and distribution of anti-HLA and antibodies of MHC class I chain related antigens A (MICA) specificities in different age groups before initial HSCT would provide HLA and MICA specific antibody profiles under the impact of ageing, which could provide meaningful information in the process of selecting suitable HLA-mismatched donors by avoiding preformed DSA.ResultsThere were no significant differences in the distribution of anti-HLA class I, class II and anti-MICA antibodies among the three age groups in this study except that a significant lower negative ratio of anti-HLA class I, class II antibodies and higher positive rate of MICA antibodies with maximum mean fluorescent intensity (MFI) > 5000 in the elderly than in young age group. The distribution of antibody specificities against HLA -A, -B, -C, -DR, -DQ, -DP and MICA antigens in the three age groups were generally consistent. The anti-HLA class I antibody specificities with higher frequencies were A80,A68;B76,B45;Cw17, which were unlikely to become DSA in Chinese. Anti-HLA class II antibody specificities were more likely to become potential DSA than class I.DR7, DR9, DQ7, DQ8 and DQ9 were most likely to become potential DSA.ConclusionsThe prevalence of anti-HLA and anti-MICA antibodies increased slightly as age increased. While ageing had a small impact on the distribution of antibody specificity frequencies against HLA-A, -B, -C, -DR,-DQ, -DP and MICA antigens in recipients awaiting initial HSCT from East China. The risk of developing preformed DSA was basically consistent in the three age groups and the elderly group might be more favorable in HLA-mismatched HSCT due to higher positive rate of anti-MICA antibody.

Assessment of IgM DSAs in Transplant Recipients: Relationship to De Novo IgG DSAs and Risk for Antibody Rejection.

The presence of anti-HLA donor-specific antibodies (DSAs) is associated with antibody-mediated rejection (AMR) and inferior graft survival. However, recent data suggest that ~50% of AMR episodes are IgG DSA negative and possibly related to non-HLA DSAs. After the initial activation of B cells to alloantigen, IgM is the first immunoglobulin produced. In addition, both IgM and IgG isotopes can activate the classic complement pathway and induce complement-dependent cytotoxicity to allograft targets. Current practices focus on the assessment of IgG DSAs with little concern for the assessment of IgM DSAs. Here, we examined anti-HLA IgM in a cohort of 22 patients who developed de novo IgG DSAs by a modified single-antigen bead-based test. We found IgM HLA DSAs developed before IgG DSAs. The median time from the detection of IgM DSAs to the appearance of de novo IgG DSAs was 461 d. Most patients had IgM DSAs against the same HLA-DQ antigens, for which IgG de novo DSAs were also later detected. IgM DSAs were detected in patients with biopsies suspected of AMR. The detection of IgM DSAs could be an early indicator of alloimmune responses to allografts before IgG de novo DSAs appear.

A Single-Center Analysis of How HLA Mismatch and Donor-Specific Antibodies Affect Short-Term Outcome After Lung Transplantation: A Pilot Study Before a Country-Wide Histocompatibility Study in Japan

Analyzing HLA polymorphism in lung transplantation (LTx) is important, given its impact on LTx recipient survival and graft function. Accordingly, we conducted a retrospective study to examine the influence of HLA mismatch and donor-specific antibodies (DSA) on short-term outcomes and early-phase post-LTx complications. HLA antigen or eplet mismatch in LTx patients at Tohoku University Hospital from 2018 to 2023 was determined, and DSA was measured on admission for surgery to identify preformed DSA and at weeks 4 to 12 post-LTx for de novo DSA, respectively. The participants were 45 LTx recipients, HLA-A/B/DR antigen mismatch (5-6 of 6) being identified in 57%, HLA-A/B/Cw/DR/DQ mismatch (8-10 of 10) in 57%, and HLA eplet mismatch (>61) in 46%. The prevalence of preformed DSA was 24%, and persistence (uncleared) was 16%. The incidence of de novo DSA was 16% after LTx. During the study,16 recipients experienced grade 3 primary graft dysfunction (PGD), 8 developed acute rejection, and 5 died. No HLA-related variables were significantly associated with post-LTx mortality and were not risk factors for high-grade PGD or acute rejection. Despite limitations in sample size, resulting in tentative findings, the study serves as a crucial pilot study for an ongoing multicenter prospective trial in Japan.

Determination of the clinical relevance of donor epitope-specific HLA-antibodies in kidney transplantation.

In kidney transplantation, survival rates are still partly impaired due to the deleterious effects of donor specific HLA antibodies (DSA). However, not all luminex-defined DSA appear to be clinically relevant. Further analysis of DSA recognizing polymorphic amino acid configurations, called eplets or functional epitopes, might improve the discrimination between clinically relevant vs. irrelevant HLA antibodies. To evaluate which donor epitope-specific HLA antibodies (DESAs) are clinically important in kidney graft survival, relevant and irrelevant DESAs were discerned in a Dutch cohort of 4690 patients using Kaplan-Meier analysis and tested in a cox proportional hazard (CPH) model including nonimmunological variables. Pre-transplant DESAs were detected in 439 patients (9.4%). The presence of certain clinically relevant DESAs was significantly associated with increased risk on graft loss in deceased donor transplantations (p < 0.0001). The antibodies recognized six epitopes of HLA Class I, 3 of HLA-DR, and 1 of HLA-DQ, and most antibodies were directed to HLA-B (47%). Fifty-three patients (69.7%) had DESA against one donor epitope (range 1-5). Long-term graft survival rate in patients with clinically relevant DESA was 32%, rendering DESA a superior parameter to classical DSA (60%). In the CPH model, the hazard ratio (95% CI) of clinically relevant DESAs was 2.45 (1.84-3.25) in deceased donation, and 2.22 (1.25-3.95) in living donation. In conclusion, the developed model shows the deleterious effect of clinically relevant DESAs on graft outcome which outperformed traditional DSA-based risk analysis on antigen level.

Antibody verification of HLA class I and class II eplets by human monoclonal HLA antibodies.

In solid organ transplantation, formation of de novo donor-specific HLA antibodies is induced by mismatched eplets on donor HLA molecules. While several studies have shown a strong correlation between the number of eplet mismatches and inferior outcomes, not every eplet mismatch is immunogenic. Eplets are theoretically defined entities, necessitating formal proof that they can be recognised and bound by antibodies. This antibody verification is pivotal to ensure that clinically relevant eplets are considered in studies on molecular matching. Recombinant human HLA-specific monoclonal antibodies (mAbs) were generated from HLA-reactive B cell clones isolated from HLA immunised individuals using recombinant HLA molecules. Subsequently, the reactivity patterns of the mAbs obtained from single antigen bead assay were analysed using HLA-EMMA software to identify single or configurations of solvent accessible amino acids uniquely present on the reactive HLA alleles and were mapped to eplets. Two HLA class I and seven HLA class II-specific human mAbs were generated from four individuals. Extensive mAb reactivity analysis, led to antibody verification of three HLA-DR-specific eplets, and conversion of five eplets (one HLA-A, one HLA-B, two HLA-DR, and one HLA-DP), from provisionally verified to truly antibody-verified. Finally, one HLA-DQ-specific eplet was upgraded from level A2 to level A1 verification evidence. The generation of recombinant human HLA-specific mAbs with different specificities contributes significantly to the antibody verification of eplets and therefore is instrumental for implementation of eplet matching in the clinical setting.

Fixed low dose versus concentration-controlled initial tacrolimus dosing with reduced target levels in the course after kidney transplantation: results from a prospective randomized controlled non-inferiority trial (Slow & Low study)

Optimal initial tacrolimus dosing and early exposure of tacrolimus after renal transplantation is not well studied. In this open-label, 6 months, multicenter, randomized controlled, non-inferiority study, we randomly assigned 432 renal allograft recipients to receive basiliximab induction, mycophenolate and steroids and either standard prolonged-release tacrolimus (trough levels: 7-9ng/ml; Standard Care arm), or an initial 7-day fixed 5mg/day dose of prolonged-release tacrolimus followed by lower tacrolimus predose levels (trough levels: 5-7ng/ml; Slow & Low arm). The primary end point was the combined incidence rate of biopsy-proven acute rejections (BPAR; including borderline), graft failure, or death at 6 months with a non-inferiority margin of 12.5%. (EudraCT-Nr: 2013-001770-19. The combined primary endpoint in the Slow & Low arm was non-inferior compared to the Standard Care arm (22.1% versus 20.7%; difference: 1.4%, 90% CI-5.5% to 8.3%). The overall rate of BPAR including borderlines was similar (Slow & Low 17.4% versus Standard Care 16.6%). Safety parameters such as delayed graft function, kidney function, donor specific HLA-antibodies, infections, or post-transplantation diabetes mellitus did not differ. This is the first study to show that an initial fixed dose of 5mg per day followed by lower tacrolimus exposure is non-inferior compared to standard tacrolimus therapy and equally efficient and safe within 6 months after renal transplantation. These data suggest that therapeutic drug monitoring for prolonged release tacrolimus can be abandoned until start of the second week after transplantation. Investigator-initiated trial, financial support by Astellas Pharma GmbH.

Chronic lung allograft dysfunction is associated with an increased number of non-HLA antibodies

BackgroundChronic Lung Allograft Dysfunction (CLAD) is the major cause of adverse outcomes in lung transplant recipients. Multiple factors, such as infection, alloimmunity, and autoimmunity, may lead to CLAD. Here, we aim to examine the role of non-HLA antibodies in CLAD in a large retrospective cohort. MethodsWe analyzed non-HLA antibodies in the pre-and post-transplant sera of 226 (100 CLAD, 126 stable) lung transplant recipients from 5 centers, and we used a separate cohort to confirm our findings. ResultsA panel of 18 non-HLA antibodies was selected for analysis based on their significantly higher positive rates in CLAD vs. stable groups. The panel-18 non-HLA antibodies (n>3) may be positive pre- or post-transplant; the risk for CLAD is higher in the latter. The presence of both non-HLA antibody and HLA donor-specific antibody (DSA) was associated with an augmented risk of CLAD (HR=25.09 [5.52–14.04], p <0.001), which was higher than that for single-positive patients. In the independent confirmatory cohort of 61 (20 CLAD, 41 stable) lung transplant recipients, the risk for CLAD remained elevated in double-positive patients (HR=10.67 [0.98–115.68], p=0.052). After adjusting for non-standard immunosuppression, patients with double-positive DSA/Non-HLA antibodies had an elevated risk for graft loss (HR=2.53 [1.29–4.96], p=0.007). ConclusionsCirculating non-HLA antibodies (n>3) were independently associated with a higher risk for CLAD. Furthermore, when non-HLA antibodies and DSA were detected concomitantly, the risk for CLAD and graft loss was significantly increased. These results show that humoral immunity to HLA and non-HLA antigens may contribute to CLAD development.

Non-HLA Antibodies to G Protein-coupled Receptors in Pediatric Kidney Transplant Recipients: Short- and Long-term Clinical Outcomes.

Angiotensin II type 1 receptor antibodies (AT1R-Abs) and endothelin-type A receptor antibodies (ETAR-Abs) are G protein-coupled receptor activating autoantibodies associated with antibody-mediated rejection, vascular pathology, increased cytokines, allograft dysfunction, and allograft loss in pediatric kidney transplant recipients in the first 2 y posttransplantation. The impact of AT1R-Ab and ETAR-Ab positivity on longer-term 5-y transplant outcomes is unknown. One hundred pediatric kidney transplant recipients were tested for ETAR-Ab and AT1R-Ab on serially collected blood samples in the first 2 y posttransplant. Biopsies were collected per protocol and 6, 12, and 24 mo posttransplant and at any time during the 5-y follow-up period for clinical indication. Clinical outcomes, including renal dysfunction, rejection, HLA donor-specific antibodies, and allograft loss, were assessed through 5 y posttransplantation. AT1R-Ab or ETAR-Ab were positive in 59% of patients. AT1R-Ab or ETAR-Ab positivity was associated with greater declines in estimated glomerular filtration rate, and de novo AT1R-Ab or ETAR-Ab was associated with allograft loss in the first 2 y posttransplant. There was no association between antibody positivity and rejection, antibody-mediated rejection, or allograft loss in the first 5 y posttransplant. In a model controlled for age, sex, immunosuppression, and HLA mismatch, AT1R-Ab or ETAR-Ab positivity was significantly associated with the development of HLA donor-specific antibodies at 5 y posttransplant (odds ratio 2.87, P = 0.034). Our findings suggest temporally distinct clinical complications associated with AT1R-Ab or ETAR-Ab positivity in pediatric patients; these injury patterns are of significant interest for developing effective treatment strategies.

The Significance of Major Histocompatibility Complex Class I Chain-related Molecule A in Solid Organ and Hematopoietic Stem Cell Transplantation: A Comprehensive Overview.

Improving long-term allograft survival and minimizing recipient morbidity is of key importance in all of transplantation. Improved matching of classical HLA molecules and avoiding HLA donor-specific antibody has been a major focus; however, emerging data suggest the relevance of nonclassical HLA molecules, major histocompatibility complex class I chain-related gene A (MICA) and B, in transplant outcomes. The purpose of this review is to discuss the structure, function, polymorphisms, and genetics of the MICA molecule and relates this to clinical outcomes in solid organ and hematopoietic stem cell transplantation. The tools available for genotyping and antibody detection will be reviewed combined with a discussion of their shortcomings. Although data supporting the relevance of MICA molecules have accumulated, key knowledge gaps exist and should be addressed before widespread implementation of MICA testing for recipients pre- or posttransplantation.

Complement-dependent cytotoxicity crossmatch in solid organ transplantation: The gold standard or golden history?

Complement-dependent cytotoxicity crossmatch (CDC-XM) has been considered for many years the standard of practice for determining compatibility in solid organ transplantation (SOT). However, as this method is laborious, time intensive and lacks sensitivity and specificity, it has been replaced in many laboratories worldwide by flow cytometry crossmatch (FCXM) and/or virtual crossmatch (vXM).With this study we intend to show the relevance of performing CDC-XM in the era of virtual crossmatching. We retrospectively analyzed 1,007 consecutive T and B cell deceased donor (DD) CDC-XMs performed in parallel using non-treated and dithiothreitol (DTT) treated sera between May 2022 and January 2023 in waitlisted patients with no donor specific antibodies (DSA) against HLA-A, B and/or DR antigens. Thirty five of 1,007 (3.5%) T cell crossmatches and 132 of 1,007 (13.1%) B cell crossmatches were positive with non-treated sera. Correlation with the vXM demonstrated no DSA in any of the positive T cell crossmatches. DSA were also absent in 126/132 positive B cell crossmatches, indicating a high rate of false positive CDC-XM. Indeed, only 4/35 T cell and 13/132 B cell CDC-XM remained positive after treatment with DTT, confirming that false positive reactivity with non-treated sera is high. Class I HLA DSA against C locus antigens were present in 17/1,007 T cell crossmatches and none were detected by CDC-XM (sensitivity = 0%). Similarly, only 6/77 B cell crossmatches with DSA targeting HLA-C, DQ and/or DP antigens were CDC-XM positive (sensitivity = 7.8%). Furthermore, only 4/6 positive B cell CDC-XM were confirmed to have complement binding potential using the C1q assay, suggesting additional false positive reactivity in 2/6 of the positive CDC-XM.Our study demonstrates that CDC-XM exhibits poor sensitivity, high false positive reactivity (especially without DTT treatment) and does not meaningfully contribute to pre-transplant compatibility testing in the context of vXM based allocation. Furthermore, the use of CDC-XM can unnecessarily delay or even prevent safe and appropriate transplant allocation.

Identification of Anti-HLA-C Autoantibody As Risk Factor for Primary Platelet Transfusion Refractoriness in Patients with Hematologic Disorders

Platelet transfusion is an important supporting treatment for hematologic disorders, especially in bone marrow failure disease and post-chemotherapy myelosuppression. However, in patients experienced multiple transfusions, platelet transfusion refractoriness (PTR) occasionally happened. Up to date, many factors had been explored contribute to the PTR, including infections, splenomegaly, antibodies against human leukocyte antigen class I (HLA-I) and antibodies against human platelet antigens (HPA). Based on our clinical practice, we also discovered that many cases had experienced primary PTR (who have been experienced PTR since the first platelet transfusion). Those patients suffered from severe thrombocytopenia can only be effectively corrected by cross-matching platelet transfusion. To our knowledge, anti-HLA-I-A and B antibodies had been reported to be in reference to immune-mediated PTR, while there are no conclusions about primary PTR. Herein, we identified anti-HLA-C autoantibodies was an independent risk factor for the primary PTR in patients with hematologic disorders, even refractoriness to cross-matching platelet transfusion or HLA-selected platelet transfusion. We retrospectively reviewed the patients that had HLA high resolution genotyping test report (potential for hematopoietic stem cell transplantation), and along with a donor-specific HLA antibodies (DSA) test or PRA testing potential for cross-matching platelet transfusion in our department between January 2019 and March 2023. 114 patients were enrolled, with a median age of 40 (14-80) years old, and the composition of disorders were aplastic anemia (65.8%), leukemia (21.9%), myelodysplastic syndrome (8.8%) and other (3.5%) (Fig A). Post-transfusion platelet efficacy was evaluated by the calculation of corrected count increment (CCI). The overall transfusion effective rate was 69.23% in this cohort, while 18.8% failed response from the first transfusion. Logistic regression analysis showed that only anti-HLA-I antibody contribute to the ineffectiveness of first and overall platelet transfusion (Fig B-E). In patients with positive anti-HLA-I antibodies, there were 35, 48 and 32 patients positive for anti-HLA-A, B and C antibodies, respectively (Fig A). Meanwhile, we surprisingly found that there were also 6 patients have anti-HLA-I autoantibody, including 6 on HLA-C and 1 have both HLA-A and C. To distinguish and gain a better understanding of the value of anti-HLA autoantibody in both TR and PTR, we further analyzed each locus of HLA-I and found that either anti-HLA-A, HLA-B or HLA-C antibody contribute to the ineffective to the overall and first platelet transfusion (Fig F-G), while logistic regression analysis revealed that only anti-HLA-C antibody was the independent risk factor (Fig H-K). We performed a logistic regression analysis with factors of anti-HLA-A, B antibody and anti-HLA-C autoantibody, and results showed that the association between positive anti-HLA-C autoantibody and the occurrence of PTR was significant ( P=0.008) (Fig J-M), while a difference approached the significance was also observed in overall platelet transfusion analysis ( P=0.054) (Fig N-O). In detail, we found that the effective overall platelet transfusion in the 6 patients who had anti-HLA-C autoantibodies was 20% (18 out of 90 transfusion), while for patients only have anti-HLA-C non-autoantibodies, it was 59.76% (47 out of 246 transfusion), a significant difference was observed ( P&amp;lt;0.001) (Fig P). For 3 patients with positive anti-HLA-C autoantibodies, all experience ineffective cross-matching platelet transfusion; while for the patients whom without anti-HLA-C autoantibodies, 21 patients experience cross-matching platelet transfusion and 17 of them response quite well, whit a significantly differences ( P=0.017) (Fig Q). In summary, our study demonstrated that positive anti-HLA-C autoantibody had a significant association with the occurrence of primary and overall PTR. The effects of cross-matching platelet transfusions in these patients are also unsatisfactory. To reduce the occurrence of PTR, our results suggested that it is necessary to anticipate the occurrence of PTR in patients with anti-HLA-C autoantibody positivity. Meanwhile, mechanisms should be explored and ways should be sought to lower the levels of such autoantibodies.

Impact of Non-Immune Factors on Platelet Transfusion Responses in an Allo-Immunized Cohort

Background: Patients with hematologic malignancies and/or hematopoietic stem cell transplantation (HSCT) recipients often require platelet transfusion support for extended periods. This can result in platelet transfusion refractoriness (PTR) from HLA alloimmunization in up to 30% of individuals. These medically complex patients may also have splenomegaly, fevers and other factors which contribute to PTR. Consequently, they may demonstrate variability in immediate platelet recovery (at 0-2 hours post-transfusion), and in subsequent platelet consumption (at 18-24 hours) despite receiving HLA-selected platelet products. We sought to examine patient and product related factors which impacted platelet transfusion response in a cohort of HLA-alloimmunized platelet transfusion refractory patients. Methods: From April 2021 to June 2023, HLA-alloimmunized (PRA &amp;gt;50%) subjects who required three or more HLA selected platelet units were identified at the Fred Hutchinson Cancer Center/University of Washington. Data were obtained on patient demographics, diagnoses, Panel Reactive Antibodies (PRA %) and factors shown to be associated with poor platelet responses (spleen size, disseminated intravascular coagulation (DIC), bleeding, selected medications, history of hematopoietic stem cell transplant (HSCT) and/or graft versus host disease (GVHD), ABO compatibility, and product age). Platelet corrected count increments (CCI) were calculated for each transfusion. Non-HLA factors potentially contributing to CCIs at 0-2 and 18-24 hours post-transfusion were compared among HLA-compatible (either 4/4 HLA matched without HLA antigen mismatches, or with permissive mismatches (defined as cumulative a mean fluorescent intensity (MFI) &amp;lt;10,000 on the Luminex donor specific HLA antibody testing platform)) and HLA incompatible transfusions (cumulative MFI &amp;gt;10,000 or random donor apheresis products). Results: Of the 420 patients screened, 75 had sufficient records for inclusion. These subjects received 2729 platelet transfusions, an average of 36 transfusions per patient over 5.2 (± 4.1) months. Post transfusion platelet counts were available for 1528 transfusions. All platelet products were leukoreduced, irradiated, single donor apheresis products. Patients were predominantly Caucasian (70.7%; n=53/75), female (82.7%; n=62/75), aged 57 (±14.9) years. The primary diagnosis was Acute myelogenous leukemia or myeloproliferative disorder in most cases (88%; n=66/75). 58.7% (n=44/75) of patients were HSCT recipients (3 auto-; 41 allo-HSCT). Average PRA among patients was 72.4%±22.2%. 25 patients had splenomegaly. Among HLA-compatible transfusions, average CCIs at 0-2 and 18-24 hours were 17,088 and 6,666. In HLA-mismatched/random donor transfusions, average CCIs at 0-2 and 18-24 hours were 11,489 and 2,786; HLA compatible products increased CCIs by 5,599 and 3880 at 0-2 and 18-24 hours. In both groups splenomegaly had the most consistent association with poor CCI at both time intervals (Figures A and B). Conclusions: In this cohort of individuals with HLA-alloimmunization mediated PTR, the administration of HLA-compatible products offset the impact of other detrimental factors generating adequate CCIs at 0-2 and 18-24 hours. Among patients with splenomegaly, CCIs were lower despite HLA compatible product administration. In the same individuals, HLA-incompatible/random donor products negatively impacted CCIs overall and the negative effect of other concomitant non-immune/product-related factors was additive.Our data highlights the need to account for non-HLA related conditions while evaluating CCI response following each HLA-selected product transfusion, and to further explore any interactions among such factors. Next steps include conducting a multivariate analysis and developing a predictive model for platelet transfusion response in HLA-alloimmunized recipients who have concomitant non-immune or product related risk factors.

Gender Disparities in Allograft Access Due to HLA-Sensitization in Multiparous Women: Implications for Evaluation of Female Patients for Alternative Donor Transplantation

Introduction: The relationship between allograft patient (pt) demographics &amp; HLA-antibody (HLA-Ab) burden, &amp; the degree to which HLA-Ab burden impacts donor type received in the current era of “donors for all”, is not established. Methods: We examined associations between pt sex/ parity, ancestry, &amp; HLA-Ab burden in consecutive adult allograft recipients (excluding HLA-identical siblings) with acute leukemia, MDS, or MPN, transplanted 1/2018-12/2022. We also assessed the impact of HLA-Ab burden on donor type received. We classified HLA-Ab burden by the number &amp; intensity of class I (HLA-A, B, -C) &amp; II (HLA-DR, -DQ, -DP) HLA-Abs. A mean fluorescence intensity (MFI) &amp;gt; 1,000 defined a positive HLA-Ab. Pts were classified as broadly sensitized (more than the median number of class I/ II HLA-Abs among Ab positive pts) &amp;/or highly sensitized (MFI &amp;gt; 10,000 for ≥ 1 HLA-Ab). For pts with &amp;gt; 1 screen, the one closest to transplant was used. We hypothesized that multiparous females have the greatest HLA-Ab burden, limiting provision of haploidentical (haplo) grafts in these pts due to the rejection risk. Results: Of 672 pts [median age 60 yrs (range 29-79), 419/672 (62%) with acute leukemia, 472/672 (70%) with European ancestry], 278/672 (41%) were female of whom 54/278 (19%) were nulliparous, 52/278 (19%) uniparous &amp; 172/278 (62%) multiparous. Overall, 367/672 (55%) received 8/8 unrelated donor (URD) &amp; 305/672 (45%) HLA-disparate grafts [137 cord blood (CB), 88 haplo, &amp; 80 5-7/8 URD (mmURD)]. As expected, non-European pts received 2x the proportion of HLA-disparate grafts [165/472 (35%) vs 141/200 (70%), p &amp;lt; .001]. Pt ancestry was not associated with HLA-Ab burden. Pt sex/ parity &amp; HLA-Ab burden associations are shown in Table 1A &amp; Fig. 1. Over one third [254/672 (38%)] of pts had a positive screen for class I/ II HLA-abs (median of 8 HLA-Abs of positive pts, range 1-113). In the 254 positive pts, 3,563 Abs were detected with the MFI distribution being 1,001-4,000 in 1,340/3,563 (38%), 4,001-10,000 in 874/3,563 (25%) &amp; &amp;gt; 10,000 in 1,349/3,563 (38%). Of the total pt cohort, 140/672 (21%) pts were broadly &amp;/or highly sensitized with 90/672 (13%) being both broadly &amp; highly sensitized. Compared with males &amp; nulliparous females, uni- &amp; multi-parous female pts had over 2x the proportion of positive screens [117/448 (26%) vs 137/224 (61%), p &amp;lt; .001]. They also had more than 8x the proportion broadly &amp; highly sensitized [17/448 (4%) vs 73/224 (33%), p &amp;lt; .001]. HLA-Ab burden by donor type is shown in Table 1B. Among HLA-disparate graft recipients, compared with recipients of CB &amp; mmURDs, fewer haplo recipients had positive screens [96/217 (44%)CB/ mmURD vs 27/88 (31%) haplo, p = .029]. Less than half the proportion were broadly &amp; highly sensitized [37/217 (17%) CB/ mmURD vs 7/88 (8%) haplo, p = .041]. Also, haplo recipients included the fewest multiparous females [16/88 (18%)] compared with making up over one quarter of 8/8 URD [98/367 (27%)], CB [36/137 (26%)], or mmURD [22/80 (28%)] transplants. Conclusion: Over 10% of transplanted pts were broadly &amp; highly sensitized to HLA. The HLA-Ab burden was predominantly in parous females (33% of our pt cohort), most of whom had HLA-Abs with one-third of this group being both broadly &amp; highly sensitized. Moreover, pts with a high HLA-Ab burden were the least likely to receive haplos, &amp; thus these transplants were less common in multiparous females. These data have immediate implications for alternative donor transplant evaluations of female patients: given that HLA-Abs should be expected among parous females, in the absence of a matched adult donor, mmURD or CB may need to be considered over haplo grafts. This could compound allograft access for multiparous female pts of non-European heritage, given many will lack a fully matched adult donor &amp; non-European pts have greatly decreased URD availability (Fingrut et al., ASH 2023). These challenges are under-appreciated in the transplant field. At initial allograft evaluation, transplant centers should obtain ancestry &amp; parity histories &amp; screen for HLA-Abs. In the absence of a suitable HLA-identical sibling donor, this information, together with the URD search prognosis (ie Davis et al., TCT 2023), should then guide triage to alternative donor allografts. Finally, prospective analysis of the impact of donor-specific HLA-Abs on donor selection &amp; transplant outcomes as well as development of effective HLA de-sensitization strategies are urgently needed.