Low-level Donor-specific Antibodies Research Articles

Utilizing proficiency testing survey data to create advanced educational content: the virtual crossmatch challenge model.

Proficiency testing (PT) surveys include data from laboratories across the world and are ideal for creating advanced educational content, beyond just consensus grading. Educational challenges provide a unique opportunity to probe common laboratory practices and risk assessment, especially in cases where there is no "analyte" tested. Human leukocyte antigen (HLA) compatibility evaluation between donor and recipient pairs has been traditionally assessed using T-cell and B-cell physical crossmatches. However, advancements in our ability to identify and characterize HLA antibodies using solid phase assays, in combination with changing deceased donor allocation schemes and improved HLA typing, have shifted the paradigm from performing physical crossmatches to the use of the virtual crossmatch (VXM). VXM is a compatibility assessment relying on the interpretation of pre-transplant HLA laboratory data and as such, it is not an "analyte". However, VXM results are used in clinical decision-making. The VXM assessment depends on patient characteristics as well as laboratory and transplant center practices but must ensure safe transplantation outcomes while maintaining equity in access to transplantation. In this manuscript, we describe the American Society for Histocompatibility and Immunogenetics (ASHI) PT Educational VXM Challenge, as a model for creating educational content using PT survey data. We discuss the different components of the VXM Challenge and highlight major findings and learning points acquired from ASHI VXM Challenges performed between 2018-2022, such as the lack of correlation between the VXM and the physical crossmatch in the presence of low level donor-specific antibodies (DSA), or when the DSA were aimed against donor alleles that are not present on the antibody panel, and in the presence of an antibody to a shared eplet. Finally, we show that the VXM Educational Challenge serves as a valuable tool to highlight the strengths and pitfalls of the VXM assessment and reveals differences in testing and result interpretation among participating HLA laboratories.

Impact of Low-Level Donor-Specific Anti-HLA Antibody on Posttransplant Clinical Outcomes in Kidney Transplant Recipients.

The clinical significance of low-level donor-specific anti-HLA antibody (low-DSA) remains controversial. We investigated the impact of low-DSA on posttransplant clinical outcomes in kidney transplant (KT) recipients. We retrospectively reviewed 1,027 KT recipients, namely, 629 living donor KT (LDKT) recipients and 398 deceased donor KT (DDKT) recipients, in Seoul St. Mary's Hospital (Seoul, Korea) between 2010 and 2018. Low-DSA was defined as a positive anti-HLA-DSA result in the Luminex single antigen assay (LABScreen single antigen HLA class I - combi and class II - group 1 kits; One Lambda, Canoga Park, CA, USA) but a negative result in a crossmatch test. We compared the incidence of biopsy-proven allograft rejection (BPAR), changes in allograft function, allograft survival, patient survival, and posttransplant infections between subgroups according to pretransplant low-DSA. The incidence of overall BPAR and T cell-mediated rejection did not differ between the subgroups. However, antibody-mediated rejection (ABMR) developed more frequently in patients with low-DSA than in those without low-DSA in the total cohort and the LDKT and DDKT subgroups. In multivariate analysis, low-DSA was identified as a risk factor for ABMR development. Its impact was more pronounced in DDKT (odds ratio [OR]: 9.60, 95% confidence interval [CI]: 1.79-51.56) than in LDKT (OR: 3.76, 95% CI: 0.99-14.26) recipients. There were no significant differences in other outcomes according to pretransplant low-DSA. Pretransplant low-DSA has a significant impact on the development of ABMR, and more so in DDKT recipients than in LDKT recipients, but not on long-term outcomes.

Impact of low-level donor-specific anti-HLA antibody on posttransplant clinical outcomes of kidney transplant recipients: analysis from Korean Organ Transplantation Registry data

Haeun Lee, Jung Hwan Park, Jong-Won Park, Tae Hyun Ban, Jaeseok Yang, Myoung Soo Kim, Chul Woo Yang, Byung Ha Chung, Korean Organ Transplantation Registry. Korean J Transplant 2022;36:47. https://doi.org/10.4285/ATW2022.F-1621

Impact of Donor Characteristics on AlloSure Scores

<h3>Purpose</h3> AlloSure dd-cfDNA (AS) has been demonstrated to detect allograft injury after heart transplantation in a large and diverse patient cohort in the United States, however, little is known about the association between donor and recipient characteristics and AS results in the early post-transplant period. <h3>Methods</h3> A total of 2029 patients from the SHORE registry were analyzed. To evaluate the impact of donor/recipient variables on early post-transplant AS values, we compared clinically stable patients with and without persistent AS elevation during the first 6 months, specifically excluding those patients with clinical events (rejection, death, dnDSA, CAV and graft dysfunction, defined as LVEF < 40% or LVEF decline >= 25% between consecutive visits) during the 2-year study follow-up. Not all donor/recipient variables were available for every analysis performed. <h3>Results</h3> Among 767 patients meeting the inclusion criteria, we identified 82 patients with median scores >=0.15% during the first 6 months after transplant; the remaining 685 had median 6m AS scores <0.15% and were used as the comparator group. No differences were observed with regards to donor age, gender, cause of death, CMV status, or recipient variables including cPRA, induction therapy, HLA mismatches (A, B, C, DR, DQ loci), or need for pre-transplant mechanical support [Table 1]. <h3>Conclusion</h3> Our investigation did not identify significant associations between donor/recipient variables and sustained elevations in AS in the early post-transplant period. Found in only a small minority of cohort patients (<5% of the SHORE registry), these elevations, suggestive of ongoing myocardial injury, may reflect the impact of immunologic variables not included in our analysis, such as the effect of and adherence to immunosuppressive therapy or the presence of low-level donor-specific antibodies. Further prospective studies are needed to better understand the etiologies and significance of these elevations.

Use of Human Leukocyte Antigen (HLA)-Incompatible Platelet Units in HLA Platelet-Refractory Patients With Limited Number of or Low-Level HLA Donor-Specific Antibodies Results in Permissive Transfusions.

In human leukocyte antigen (HLA)-mediated alloimmune platelet refractoriness, HLA-incompatible platelets may produce adequate posttransfusion corrected count increment ("permissive transfusion") and increase the donor pool. To determine if a lower number of or low-level anti-HLA donor-specific antibodies (DSAs) predict permissive transfusion and could be used to prioritize platelet selection. We categorized platelets administered from 2016 to 2018 as HLA-compatible or HLA-incompatible based on presence of DSAs against the donor unit. We further divided HLA-incompatible units based on the number of DSAs and the level of DSAs (measured by mean fluorescence intensity [MFI]), where cumulative MFI ≥6000 defines high-level DSA. We compared posttransfusion corrected count increments (CCIs) and transfusion reactions among these transfusions. Of 279 HLA-selected units transfused into 26 platelet-refractory patients, we resorted to using 39 HLA-incompatible units (14%). Posttransfusion CCI and transfusion reaction frequency were similar among units targeted by 1 or low-level DSAs and HLA-compatible units. Units targeted by ≥2 distinct or high-level DSAs produced lower CCIs. Regardless of ABO compatibility, similarly HLA-categorized units yielded comparable CCIs and comparable frequency of transfusion reactions. HLA-incompatible platelets transfused across 1 or low-level DSAs were commonly permissive, whereas those transfused across ≥2 DSAs or high levels of DSA (MFI ≥6000) were nonpermissive. The use of such donor units offers transfusion services alternative platelet units for support of platelet-refractory patients.

Belatacept in Kidney Transplant Recipients With Failed Allografts for the Prevention of Humoral Sensitization: A Pilot Randomized Controlled Trial.

Belatacept in Kidney Transplant Recipients With Failed Allografts for the Prevention of Humoral Sensitization: A Pilot Randomized Controlled Trial.

Lung transplantation outcomes after crossing low-level donor specific antibodies without planned augmented immunosuppression.

It is unknown whether some donor specific antibodies (DSA) can be crossed at the time of lung transplant without desensitization or augmented induction immunosuppression. This study assessed whether crossing low-level pre-transplant DSA (defined as mean fluorescence intensity [MFI] 1000-6000) without augmented immunosuppression is associated with worse retransplant-free or chronic lung allograft dysfunction (CLAD)-free survival. Of the 458 included recipients, low-level pre-transplant DSA was crossed in 39 (8.6%) patients. The median follow-up time was 2.2years. There were 15 (38.5%) patients with Class I DSA and 24 (61.5%) with Class II DSA. There was no difference in adjusted overall retransplant-free survival between recipients where pre-transplant DSA was and was not crossed (HR: .98 [95% CI=.49-1.99], P=.96). There was also no difference in CLAD-free survival (HR: .71 [95% CI=.38-1.33], P=.28). There was no difference in Grade 3 PGD at 72h (OR: 1.13 [95% CI=.52-2.48], P=.75) or definite or probable AMR (HR: 2.22 [95% CI=.64-7.61], P=.21). Lung transplantation in the presence of low-level DSA without planned augmented immunosuppression is not associated with worse overall or CLAD-free survival among recipients with intermediate-term follow-up.

Impact of low-level pretransplant donor-specific antibodies on outcomes after kidney transplantation.

BackgroundThe effect of low‐level pretransplant donor‐specific antibody (DSA) on kidney transplant outcomes is not well described. The goal of this study was to compare outcomes among patients of varying immunologic risk, based on the level of pretransplant DSA.MethodsWe retrospectively reviewed all adult kidney transplant recipients who had undergone a transplant at our center between January 2013 and May 2017. Patients were grouped as negative DSA (mean fluorescence intensity, [MFISUM < 100]), low‐level DSA (MFISUM 100–1000), and positive DSA (MFISUM > 1000). Rejection, infection, graft, and patient survival were outcomes measured.ResultsOf 952 patients, 82.1% had negative DSA, 10.7% had low‐level DSA, and 7.1% had positive DSA. The positive DSA group had the highest rate of antibody‐mediated rejection (10.3%), followed by low‐level DSA (7.8%) and the negative DSA group (4.5%) (p = .034). The rate of BK viremia was highest in the positive DSA group (39.7%), followed by the low‐level group (30.4%) and the negative DSA group (25.6%), (p = .025). None of the other outcomes, including graft or patient survival, were different between the groups.ConclusionWhile low‐level DSA should not prevent proceeding with kidney transplantation, it should not be ignored. Future studies are needed to investigate the long‐term effects of varying levels of pre‐transplant DSA on outcomes.

Lung Transplantation Outcomes after Crossing Low Level Donor Specific Antibodies without Augmented Immunosuppression

<h3>Purpose</h3> Lung transplant recipients with pre-transplant donor specific antibodies (DSA) are often treated with augmented immunosuppression. It is unknown whether some DSA can be safely crossed without additional therapies. We implemented a protocol allowing transplant when crossing select low level DSA (defined as mean fluorescence intensity (MFI) 1000-5000) without planned augmented immunosuppression. <h3>Methods</h3> This was a single center retrospective cohort study between 4/1/2015-8/31/2020. All recipients received solumedrol and basiliximab induction without desensitization. Presence of low level pre-transplant DSA was recorded. All post-transplant DSA was monitored within 14 days of transplant and then at routine intervals. The primary study outcomes were overall survival, definite CLAD≥1-free survival, and definite antibody mediated rejection (AMR), all defined according to ISHLT consensus guidelines. <h3>Results</h3> Of the 453 recipients, 36 (7.9%) had a low-level pre-transplant DSA crossed at transplant (Table 1). 13 had Class I antibodies and 26 had Class II. The median historical DSA MFI was 1800 (IQR 1300-2400) and the median most recent MFI was 1200 (IQR 950-2000). Among recipients where pre-transplant DSA was crossed, 17 (47.2%) had persistent post-transplant DSA with a median peak MFI of 7400. Class II antibodies were more likely to be detected post-transplant than Class I (57.7% vs 15.4%, p=0.02). There was no statistical difference in definite AMR in recipients where pre-transplant DSA was and was not crossed (8.3% vs 3.1%, p=0.11). With a median follow-up time of 2.4 years, there were no differences in adjusted overall survival (HR=1.14, 95% CI=0.57-2.32, p=0.71) or CLAD≥1-free survival (HR=0.82, 95% CI=0.44-1.54, p=0.54). <h3>Conclusion</h3> Lung transplantation in the presence of low level DSA without planned augmented immunosuppression was not associated with worse overall or CLAD-free intermediate-term survival but may be associated with increased AMR.

The Natural History of Pre-Existing Donor Specific Antibody and Amnestic Responses after Heart Transplantation

Sensitized patients have prolonged wait times for heart transplant and often low-level donor specific antibodies (DSA) will be crossed when accepting a donor heart. The purpose of this study was to determine if positive low-level DSA at the time of transplant increase in strength post transplant and increase the risk of antibody mediated rejection (AMR). We reviewed 72 heart transplant patients between 2010 and 2018 in whom low-level DSA were crossed. DSA strength, as measured by mean fluorescence intensity (MFI) and AMR on biopsy was assessed at 1, 3, 6, 12 months post transplant. Other outcomes assessed: freedom from first-year rejection (acute cellular rejection (ACR), any treated rejection (ATR)), freedom from cardiac allograft vasculopathy (CAV, as defined by stenosis ≥30% by angiography), and freedom from graft dysfunction (LVEF ≤40%). Patients were compared to a contemporaneous control group conditional to 1-year survival without DSA at transplant. Approximately one-third of patients with pre-existing DSA at transplant demonstrated an increase in DSA strength. Patients with pre-existing DSA had decreased freedom from ATR and AMR but no differences in freedom from CAV, graft dysfunction, or ACR compared to the control group. Crossing low-level DSA may increase the risk of AMR, but there is no difference in clinically important sequelae such as CAV or graft dysfunction. Thus, accepting low-level DSA at the time of transplant appears feasible and could broaden the donor pool.

Conversion From Calcineurin Inhibitors to Belatacept in HLA-sensitized Kidney Transplant Recipients With Low-level Donor-specific Antibodies.

Belatacept could be the treatment of choice in renal-transplant recipients with renal dysfunction attributed to calcineurin inhibitor (CNI) nephrotoxicity. Few studies have described its use in patients with donor-specific antibody (DSA). We retrospectively evaluated conversion from CNIs to belatacept in 29 human leukocyte antigen-immunized renal-transplant recipients. Data about acute rejection, DSA, and renal function were collected. These patients were compared with 42 nonimmunized patients treated with belatacept. Patients were converted from CNIs to belatacept a median of 444 days (interquartile range, 85-1200) after transplantation and were followed up after belatacept conversion, for a median of 308 days (interquartile range, 125-511). At conversion, 16 patients had DSA. Nineteen DSA were observed in these 16 patients, of which 11/19 were <1000 mean fluorescence intensity (MFI), 7/19 were between 1000 and 3000 MFI, and one was >3000 MFI. At last follow-up, preexisting DSA had decreased or stabilized. Seven patients still had DSA with a mean MFI of 1298 ± 930 at the last follow-up. No patient developed a de novo DSA in the DSA-positive group. In the nonimmunized group, one patient developed de novo DSA (A24-MFI 970; biopsy for cause did not show biopsy-proven acute rejection or microinflammation score). After belatacept conversion, one antibody-mediated rejection was diagnosed. The mean estimated glomerular filtration rate improved from 31.7 ± 14.2 mL/min/1.73 m to 40.7 ± 12.3 mL/min/1.73 m (P < 0.0001) at 12 months after conversion. We did not find any significant difference between groups in terms of renal function, proteinuria, or biopsy-proven acute rejection. We report on a safe conversion to belatacept in human leukocyte antigen-immunized patients with low DSA levels.

Crossing low-level donor-specific antibodies in heart transplantation.

Donor-specific antibodies (DSA) detected by solid-phase single-antigen bead (SAB) immunoassays have been associated with antibody-mediated rejection (AMR), cardiac allograft vasculopathy (CAV) and decreased survival after heart transplantation. The clinical relevance of low-level DSA is equivocal. This review examines the techniques used to define low-level DSA, the limitations of these techniques and recent clinical experience crossing low-level DSA. Solid-phase multiplex bead immunoassays were introduced to solid-organ transplantation over 15 years ago. These technologies have a much greater sensitivity and specificity than older cell-based immunoassays. It was hoped that this increased resolution would lead to better outcomes by avoiding donors with antigens that transplant candidates produced antibodies against. Although some transplant patients with DSA show increased risk of AMR and decreased survival, a subset of patients with DSA at the time of transplant have outcomes comparable with patients with no DSA. Recent studies have demonstrated that DSA delineated according to titration studies and C1q assays better define low-level DSA that are well tolerated to cross. Early experience with crossing low-level DSA shows promise in kidney and heart transplantation. Preliminary findings from heart and kidney transplant patients show acceptable outcomes after crossing low-level DSA. The policy of crossing low-level DSA increases the donor pool for sensitized heart transplant candidates.

Crossing Low Level Donor-Specific Antibody at the Time of Heart Transplantation May Save Lives

Purpose Sensitization is an increasing problem for patients (pts) awaiting heart transplant (HTx) partly due to increased requirement for bridging with mechanical support, a risk factor for sensitization. In order to widen the donor pool for sensitized pts, our program has decided to cross low-level donor-specific antibody (DSA) at HTx. We assessed the long-term outcome of these pts. Methods Between 2010-13, we reviewed 352 HTx pts in our single-center program and identified 31 pts transplanted against a low-level DSA (MFI Results The 31 pts transplanted against a low-level DSA had significantly less freedom from first-year AMR compared to control group. However, the low-level DSA group had significantly better 5-year survival. There was no significant difference in freedom from 1-year ATR or ACR and 5-year freedom from CAV compared to control group. There was a trend for better 5-year freedom from NF-MACE in the low-level DSA group compared to control group (see table). Conclusion Crossing low-level DSA at HTx appears to have acceptable outcomes despite increased risk of first-year AMR. This strategy can expand the donor pool and potentially save lives. The survival benefit in the low-level DSA group may represent eventual accommodation however further studies are needed.

Positive Luminex and negative flow cytometry in kidney transplantation: a systematic review and meta-analysis.

The presence of pre-formed donor-specific antibodies (DSAs) in kidney transplantation is associated with worse overall outcomes compared with DSA-negative transplantation. A positive complement-dependant cytotoxic crossmatch presents a high immunological risk, while a negative flow cytometry crossmatch is at the lower end of the risk spectrum. Yet, the presence of low-level DSA detected by Luminex alone, that is, positive Luminex and negative flow (PLNF) cytometry crossmatch lacks robust scientific exploration. In this systematic review and pooled analysis, we investigate the glomerular filtration rate, acute rejection (AR), graft survival and patient survival of PLNF transplants compared with DSA-negative transplants. Our analysis identified seven retrospective studies consisting of 429 PLNF transplants and 10677 DSA-negative transplants. Pooled analysis identified no significant difference in the incidence of AR at 1 year [relative risk (RR) = 1.35, 95% confidence interval (CI) 0.90-2.02, Z = 1.46, P = 0.14, I2 = 0%], graft failure at 1 year (RR = 1.66, 95% CI 0.94-2.94, Z = 1.75, P = 0.08, I2 = 23%), graft failure at 5 years (RR = 1.29, 95% CI 0.90-1.87, Z = 1.38, P = 0.17, I2 = 0%), patient mortality at 1 year (RR = 0.89, 95% CI 0.31-2.56, Z = 0.22, P = 0.82, I2 = 0%) and patient mortality at 5 years (RR = 1.76, 95% CI 0.48-6.48, Z = 0.85, P = 0.39, I2 = 61%). Pooled analysis of graft function was not possible due to insufficient data. Current evidence suggests that low-level DSA detected by Luminex alone does not pose significant risk at least in the short-medium term. Considering the shortage of kidney transplants and the ever-increasing waiting time, the avoidance of PLNF transplants may be unwarranted especially in patients who have been enlisted for a long time.

OR2 Variable expression of HLA-C impacts T versus B cell crossmatch outcomes

Flow cytometric crossmatches (FCXM) are used to detect low-level donor-specific antibodies (DSA) and determine eligibility for transplantation and desensitization regimens. We measured HLA class I and HLA-C surface expression on T and B cells from various sources and examined the impact of HLA-C expression on FCXM reactivity in the presence of HLA-C DSA. HLA class I and HLA-C expression was examined on cells isolated from 18 donors using monoclonal antibodies (clone G46.2 and mAb DT9). T and B cells were differentiated using CD3 and CD19 specific monoclonal antibodies. Cells were acquired on a BD FACSCanto II; HLA expression was determined using median channel fluorescence (MCF) values and FCXM reactivity was assessed as a ratio of test serum to negative control serum. The mean HLA-C expression was significantly higher on T (11693 MCF) than B cells (5480 MCF, p < 0.05, Figure 1) while class I expression was lower on T (10167 MCF) than B cells (23602 MCF, p < 0.001) when comparing cells isolated from the same donors. This difference in HLA-C expression was mirrored in FCXM tests using sera containing HLA-C specific DSA. Mean reactivity with T cells as targets yielded consistently higher ratios than tests using B cells as targets (p < 0.001). Seven of 10 FCXM tests yielded positive T and negative B results. HLA-C expression is known to vary among HLA-C alleles (Apps et al., Science 2013); however, this study shows variability also exists between lymphocyte lineages. HLA-C is expressed higher on T than B cells and this difference impacts crossmatch reactivity. These findings should be considered when interpreting FCXM results.Download : Download high-res image (108KB)Download : Download full-size image

(769) - Outcome of Patients Proceeding to Heart Transplant with Low Level Donor-Specific Antibodies: How Do They Do?

(769) - Outcome of Patients Proceeding to Heart Transplant with Low Level Donor-Specific Antibodies: How Do They Do?

(784) - Low Level Donor Specific Antibodies at Transplant Does Not Appear to Be Associated With the development of Cardiac Allograft Vasculopathy After Heart Transplantation

Donor specific antibodies (DSA) following heart transplant has been reported to be associated with increased development of cardiac allograft vasculopathy (CAV) and risk of mortality following transplantation. In the era of virtual crossmatch, we may now elect to place donor hearts in patients with low level DSAs that are not placed as avoids. The effect of these low level DSAs on the development of CAV has not yet been established.

Pre-transplant low level HLA antibody shows a composite poor outcome in long-term outcome of renal transplant recipients

To determine the significance of low-level DSA (donor specific antibody) in patients transplanted with negative cytotoxicity AHG (antihuman immunoglobulin) crossmatch, data from 279 patients who received a kidney transplant between July 1999 and March 2006 were collected. All kidney recipients received ABO-compatible donors. A poor outcome was defined as any one of the following: death, Cr > 2.0 mmol/L, occurrence of a rejection episode. Luminex Screening and Single Antigen assays from Tepnel Life Codes were used to detect human leukocyte antigen antibodies on pre-transplant sera retrospectively. Twenty-four out of 279 recipients demonstrated the presence of solid-phase DSA (MFI > 1000) present pre-transplant. In DSA+ group, the accumulated good versus poor outcome rate was 0.30 versus 0.70, respectively. These rates were 0.49 and 0.51, respectively, in the DSA− group. The difference in composite poor outcome between DSA+ versus DSA− group was significant (p = 0.030). The DSA− group had no difference in patient survival as compared to the DSA+ group (p = 0.061). There is no statistically significant difference for either mortality or outcome results between high MFI (>2000) and low MFI (≤2000) groups. Our data suggest that solid-phase antibodies which are not strong enough to elicit a positive T-AHG crossmatch may influence long-term graft outcome.

ABO Incompatible Renal Transplantation Without Antibody Removal Using Conventional Immunosuppression Alone

ABO incompatible living donor renal transplantation (ABOi) can achieve outcomes comparable to ABO compatible transplantation (ABOc). However, with the exception of blood group A2 kidneys transplanted into recipients with low titer anti-A antibody, regimens generally include antibody removal, intensified immunosuppression and splenectomy or rituximab. We now report a series of 20 successful renal transplants across a range of blood group incompatibilities using conventional immunosuppression alone in recipients with low baseline anti-blood group antibody (ABGAb) titers. Incompatibilities were A1 to O (3), A1 to B (2), A2 to O (2), AB to A (2), AB to B (1), B to A1 (9), B to O (1); titers 1:1 to 1:16 by Ortho. At 36 months, patient and graft survival are 100%. Antibody-mediated rejection (AbMR) occurred in one patient with thrombophilia and low level donor-specific anti-HLA antibody. Four patients experienced cellular rejection (two subclinical), which responded to oral prednisolone. This series demonstrates that selected patients with low titer ABGAb can undergo ABOi with standard immunosuppression alone, suggesting baseline titer as a reliable predictor of AbMR. This reduces morbidity and cost of ABOi for patients with low titer ABGAb and increases the possibility of ABOi from deceased donors.

Heart Transplantation across Low Level Donor Specific Antibodies: Is It Bad?

Heart Transplantation across Low Level Donor Specific Antibodies: Is It Bad?