Leukopenia in autosomal dominant polycystic kidney disease: a single-center cohort of kidney transplant candidates with post-transplantation follow-up.

Background: Autosomal dominant polycystic kidney disease (ADPKD) has occasionally been associated with lower peripheral white blood cell (WBC) counts. This study aimed to investigate the peripheral blood cell counts in a large cohort of kidney transplant recipients before and after kidney transplantation and its potential impact on post-transplant outcomes. Methods: This was a retrospective study with long-term follow-up data of 2090 patients who underwent a first kidney transplantation in the Leuven University Hospitals, of whom 392 had ADPKD. Results: In total, 2090 patients who underwent a first kidney transplantation in the Leuven University Hospitals were included, of whom 392 had ADPKD. Both pre- and post-transplantation, ADPKD patients had significantly lower total WBC counts, and more specifically lower neutrophil, lymphocyte and eosinophil counts compared with the non-ADPKD patients. This observation was independent of potential confounders such as level of inflammation, smoking habit, vitamins and pre-transplant medication. Overall survival and kidney transplant survival were significantly better in ADPKD vs non-ADPKD transplant recipients and a longer time to first infection was observed. However, no association between blood cell counts and outcome differences was found. Conclusions: In conclusion, this large single-center study reports a strong and independent association between ADPKD and lower peripheral WBC counts both before and after kidney transplantation. Considering the role of inflammation in disease progression, further investigation into the role of WBC in ADPKD is needed.

Automated Urinary Chemokine Assays for Noninvasive Detection of Kidney Transplant Rejection: A Prospective Cohort Study.

RATIONALE & OBJECTIVE: Prior studies have demonstrated the diagnostic potential of urinary chemokines C-X-C motif ligand 9 (CXCL9) and CXCL10 for kidney transplant rejection. However, their benefit in addition to clinical information has not been demonstrated. We evaluated the diagnostic performance for detecting acute rejection of urinary CXCL9 and CXCL10 when integrated with clinical information. STUDY DESIGN: Single-center prospective cohort study. SETTING & PARTICIPANTS: We analyzed 1,559 biopsy-paired urinary samples from 622 kidney transplants performed between April 2013 and July 2019 at a single transplant center in Belgium. External validation was performed in 986 biopsy-paired urinary samples. TESTS COMPARED: We quantified urinary CXCL9 (uCXCL9) and CXCL10 (uCXCL10) using an automated immunoassay platform and normalized the values to urinary creatinine. Urinary chemokines were incorporated into a multivariable model with routine clinical markers (estimated glomerular filtration rate, donor-specific antibodies, and polyoma viremia) (integrated model). This model was then compared with the tissue diagnosis according to the Banff classification for acute rejection. OUTCOME: Acute rejection detected on kidney biopsy using the Banff classification. RESULTS: Chemokines integrated with routine clinical markers had high diagnostic value for detection of acute rejection (n=150) (receiver operating characteristic area under the curve 81.3% [95% CI, 77.6-85.0]). The integrated model would help avoid 59 protocol biopsies per 100 patients when the risk for rejection is predicted to be below 10%. The performance of the integrated model was similar in the external validation cohort. LIMITATIONS: The cross-sectional nature obviates investigating the evolution over time and prediction of future rejection. CONCLUSIONS: The use of an integrated model of urinary chemokines and clinical markers for noninvasive monitoring of rejection could enable a reduction in the number of biopsies. Urinary chemokines may be useful noninvasive biomarkers whose use should be further studied in prospective randomized trials to clarify their role in guiding clinical care and the use of biopsies to detect rejection after kidney transplantation. PLAIN-LANGUAGE SUMMARY: Urinary chemokines CXCL9 and CXCL10 have been suggested to be good noninvasive biomarkers of kidney transplant rejection. However, defining a context of use and integration with clinical information is necessary before clinical implementation can begin. In this study, we demonstrated that urinary chemokines CXCL9 and CXCL10, together with clinical information, have substantial diagnostic accuracy for the detection of acute kidney transplant rejection. Application of urinary chemokines together with clinical information may guide biopsy practices following kidney transplantation and potentially reduce the need for kidney transplant biopsies.

An observational cohort study of histological screening for BK polyomavirus nephropathy following viral replication in plasma.

Systematic screening for BKPyV-DNAemia has been advocated to aid prevention and treatment of polyomavirus associated nephropathy (PyVAN), an important cause of kidney graft failure. The added value of performing a biopsy at time of BKPyV-DNAemia, to distinguish presumptive PyVAN (negative SV40 immunohistochemistry) and proven PyVAN (positive SV40) has not been established. Therefore, we studied an unselected cohort of 950 transplantations, performed between 2008-2017. BKPyV-DNAemia was detected in 250 (26.3%) transplant recipients, and positive SV40 in 91 cases (9.6%). Among 209 patients with a concurrent biopsy at time of first BKPyV-DNAemia, 60 (28.7%) biopsies were SV40 positive. Plasma viral load showed high diagnostic value for concurrent SV40 positivity (ROC-AUC 0.950, 95% confidence interval 0.916-0.978) and the semiquantitatively scored percentage of tubules with evidence of polyomavirus replication (pvl score) (0.979, 0.968-0.988). SV40 positivity was highly unlikely when plasma viral load is below 4 log10 copies/ml (negative predictive value 0.989, 0.979-0.994). In SV40 positive patients, higher plasma BKPyV-DNA load and higher pvl scores were associated with slower viral clearance from the blood (hazard ratio 0.712, 95% confidence interval 0.604-0.839, and 0.327, 0.161-0.668, respectively), whereas the dichotomy positivity/negativity of SV40 immunohistochemistry did not predict viral clearance. Although the pvl score offers some prognostic value for viral clearance on top of plasma viral load, the latter provided good guidance for when a biopsy was unnecessary to exclude PyVAN. Thus, the distinction between presumptive and proven PyVAN, based on SV40 immunohistochemistry, has limited clinical value. Hence, management of BKPyV-DNAemia and immunosuppression reduction should be weighed against the risk of occurrence of rejection, or exacerbation of rejection observed concomitantly.

Association of Predicted HLA T-Cell Epitope Targets and T-Cell-Mediated Rejection After Kidney Transplantation.

RATIONALE & OBJECTIVE: The relationship between human leukocyte antigen (HLA) molecular mismatches and T-cell-mediated rejection (TCMR) is unknown. We investigated the associations between the different donor HLA-derived T-cell targets and the occurrence of TCMR and borderline histologic changes suggestive of TCMR after kidney transplantation. STUDY DESIGN: Retrospective cohort study. SETTING & PARTICIPANTS: All kidney transplant recipients at a single center between 2004 and 2013 with available biopsy data and a DNA sample for high-resolution HLA donor/recipient typing (N = 893). EXPOSURE: Scores calculated by the HLA matching algorithm PIRCHE-II and HLA eplet mismatches. OUTCOME: TCMR, borderline changes suggestive of TCMR, and allograft failure. ANALYTICAL APPROACH: Multivariable cause-specific hazards models were fit to characterize the association between HLA epitopes targets and study outcomes. RESULTS: We found 277 patients developed TCMR, and 134 developed only borderline changes suggestive of TCMR on at least 1 biopsy. In multivariable analyses, only the PIRCHE-II scores for HLA-DRB1 and HLA-DQB1 were independently associated with the occurrence of TCMR and with allograft failure; this was not the case for HLA class I molecules. If restricted to rejection episodes within the first 3 months after transplantation, only the T-cell epitope targets originating from the donor's HLA-DRB1 and HLA-DQB1, but not class I molecules, were associated with the early acute TCMR. Also, the median PIRCHE-II score for HLA class II was statistically different between the patients with TCMR compared to the patients without TCMR (129 [IQR, 60-240] vs 201 [IQR, 96-298], respectively; P < 0.0001). These differences were not observed for class I PIRCHE-II scores. LIMITATIONS: Observational clinical data and residual confounding. CONCLUSIONS: In the absence of HLA-DSA, HLA class II but not class I mismatches are associated with early episodes of acute TCMR and allograft failure. This suggests that current immunosuppressive therapies are largely able to abort the most deleterious HLA class I-directed alloimmune processes; however, alloresponses against HLA-DRB1 and HLA-DQB1 molecular mismatches remain insufficiently suppressed. PLAIN-LANGUAGE SUMMARY: Genetic differences in the human leukocyte antigen (HLA) complex between kidney transplant donors and recipients play a central role in T-cell-mediated rejection (TCMR), which can lead to failure of the transplanted kidney. Evaluating this genetic disparity (mismatch) in the HLA complex at the molecular (epitope) level could contribute to better prediction of the immune response to the donor organ posttransplantation. We investigated the associations of the different donor HLA-derived T-cell epitope targets and scores obtained from virtual crossmatch algorithms with the occurrence of TCMR, borderline TCMR, and graft failure after kidney transplantation after taking into account the influence of donor-specific anti-HLA antibodies. This study illustrates the greater importance of the molecular mismatches in class II molecules compared to class I HLA molecules.

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

BACKGROUND AND OBJECTIVES: 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. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: 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. RESULTS: 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. CONCLUSIONS: 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.

Circulating Donor-Specific Anti-HLA Antibodies Associate With Immune Activation Independent of Kidney Transplant Histopathological Findings.

Despite the critical role of cytokines in allograft rejection, the relation of peripheral blood cytokine profiles to clinical kidney transplant rejection has not been fully elucidated. We assessed 28 cytokines through multiplex assay in 293 blood samples from kidney transplant recipients at time of graft dysfunction. Unsupervised hierarchical clustering identified a subset of patients with increased pro-inflammatory cytokine levels. This patient subset was hallmarked by a high prevalence (75%) of donor-specific anti-human leukocyte antigen antibodies (HLA-DSA) and histological rejection (70%) and had worse graft survival compared to the group with low cytokine levels (HLA-DSA in 1.7% and rejection in 33.7%). Thirty percent of patients with high pro-inflammatory cytokine levels and HLA-DSA did not have histological rejection. Exploring the cellular origin of these cytokines, we found a corresponding expression in endothelial cells, monocytes, and natural killer cells in single-cell RNASeq data from kidney transplant biopsies. Finally, we confirmed secretion of these cytokines in HLA-DSA-mediated cross talk between endothelial cells, NK cells, and monocytes. In conclusion, blood pro-inflammatory cytokines are increased in kidney transplant patients with HLA-DSA, even in the absence of histology of rejection. These observations challenge the concept that histology is the gold standard for identification of ongoing allo-immune activation after transplantation.

Forecasting of Patient-Specific Kidney Transplant Function With a Sequence-to-Sequence Deep Learning Model.

Importance: Like other clinical biomarkers, trajectories of estimated glomerular filtration rate (eGFR) after kidney transplant are characterized by intra-individual variability. These fluctuations hamper the distinction between alarming graft functional deterioration or harmless fluctuation within the patient-specific expected reference range of eGFR. Objective: To determine whether a deep learning model could accurately predict the patient-specific expected reference range of eGFR after kidney transplant. Design, Setting, and Participants: A multicenter diagnostic study consisted of a derivation cohort of 933 patients who received a kidney transplant between 2004 and 2013 with 100 867 eGFR measurements from University Hospitals Leuven, Belgium, and 2 independent test cohorts: with 39 999 eGFR measurements from 1 170 patients, 1 from University Hospitals Leuven, Belgium, receiving transplants between 2013 and 2018 and 1 from Hannover Medical School, Germany, receiving transplants between 2003 and 2007. Patients receiving a single kidney transplant, with consecutive eGFR measurements were included. Data were analyzed from February 2019 to April 2021. Exposures: In the derivation cohort 100 867 eGFR measurements were available for analysis and 39 999 eGFR measurements from the independent test cohorts. Main Outcomes and Measures: A sequence-to-sequence model was developed for prediction of a patient-specific expected range of eGFR, based on previous eGFR values. The primary outcome was the performance of the deep learning sequence-to-sequence model in the 2 independent cohorts. Results: In this diagnostic study, a total of 933 patients in the training sets (mean [SD] age, 53.5 [13.3] years; 570 male [61.1%]) and 1170 patients in the independent test sets (cohort 1 [n = 621]: mean [SD] age, 58.5 [12.1] years; 400 male [64.4%]; cohort 2 [n = 549]: mean [SD] age, 50.1 [13.0] years; 316 male [57.6%]) who received a single kidney transplant most frequently from deceased donors, the sequence-to-sequence models accurately predicted future patient-specific eGFR trajectories within the first 3 months after transplant, based on the previous graft eGFR values (root mean square error, 6.4-8.9 mL/min/1.73 m2). The sequence-to-sequence model predictions outperformed the more conventional autoregressive integrated moving average prediction model, at all input/output number of eGFR values. Conclusions and Relevance: In this diagnostic study, a sequence-to-sequence deep learning model was developed and validated for individual forecasting of kidney transplant function. The patient-specific sequence predictions could be used in clinical practice to guide physicians on deviations from the expected intra-individual variability, rather than relating the individual results to the reference range of the healthy population.

The evolution of histological changes suggestive of antibody-mediated injury, in the presence and absence of donor-specific anti-HLA antibodies.

The interplay between donor-specific anti-HLA antibodies (HLA-DSA), histology of active antibody-mediated rejection (aABMRh ), transplant glomerulopathy (cg), and graft failure in kidney transplantation remains insufficiently understood. We performed a single-center cohort study (n = 1000) including 2761 protocol and 833 indication biopsies. Patients with pretransplant HLA-DSA were more prone to develop aABMRh (OR 22.7, 95% CI, 11.8-43.7, P < 0.001), cg (OR 5.76, 95% CI, 1.67-19.8, P = 0.006), and aABMRh/cg (OR 19.5, 95% CI, 10.6-35.9, P < 0.001). The negative impact of pre-transplant HLA-DSA on graft survival (HR 2.12, 95% CI, 1.41-3.20, P < 0.001) was partially mediated through aABMRh and cg occurrence. When adjusted for time-dependent HLA-DSA (HR 4.03, 95% CI, 2.21-7.15, P = 0.002), graft failure was only affected by aABMRh when cg was evident. In HLA-DSA negative patients, aABMRh was associated with impaired graft outcome only when evolving to cg (HR 1.32, 95% CI, 1.07-1.61, P = 0.008). We conclude that the kinetics of HLA-DSA are important to estimate the rate of graft failure, and that histological follow-up is necessary to discover, often subclinical, ABMR and cg. In the absence of HLA-DSA, patients experience similar histological lesions and the evolution to transplant glomerulopathy associates with impaired graft outcome.

Missing Self-Induced Microvascular Rejection of Kidney Allografts: A Population-Based Study.

BACKGROUND: Circulating anti-HLA donor-specific antibodies (HLA-DSA) are often absent in kidney transplant recipients with microvascular inflammation (MVI). Missing self, the inability of donor endothelial cells to provide HLA I-mediated signals to inhibitory killer cell Ig-like receptors (KIRs) on recipient natural killer cells, can cause endothelial damage in vitro, and has been associated with HLA-DSA-negative MVI. However, missing self's clinical importance as a nonhumoral trigger of allograft rejection remains unclear. METHODS: In a population-based study of 924 consecutive kidney transplantations between March 2004 and February 2013, we performed high-resolution donor and recipient HLA typing and recipient KIR genotyping. Missing self was defined as the absence of A3/A11, Bw4, C1, or C2 donor genotype, with the presence of the corresponding educated recipient inhibitory KIR gene. RESULTS: We identified missing self in 399 of 924 transplantations. Co-occurrence of missing self types had an additive effect in increasing MVI risk, with a threshold at two concurrent types (hazard ratio [HR], 1.78; 95% confidence interval [95% CI], 1.26 to 2.53), independent of HLA-DSA (HR, 5.65; 95% CI, 4.01 to 7.96). Missing self and lesions of cellular rejection were not associated. No HLA-DSAs were detectable in 146 of 222 recipients with MVI; 28 of the 146 had at least two missing self types. Missing self associated with transplant glomerulopathy after MVI (HR, 2.51; 95% CI, 1.12 to 5.62), although allograft survival was better than with HLA-DSA-associated MVI. CONCLUSION: Missing self specifically and cumulatively increases MVI risk after kidney transplantation, independent of HLA-DSA. Systematic evaluation of missing self improves understanding of HLA-DSA-negative MVI and might be relevant for improved diagnostic classification and patient risk stratification.

Risk factors, histopathological features, and graft outcome of transplant glomerulopathy in the absence of donor-specific HLA antibodies.

Transplant glomerulopathy is established as a hallmark of chronic antibody-mediated rejection in kidney transplant patients with donor-specific HLA antibodies (HLA-DSA). The clinical importance of transplant glomerulopathy in the absence of HLA-DSA is not well established. To help define this, 954 patients (encompassing 3744 biopsies) who underwent kidney transplantation 2004-2013 were studied with retrospective high-resolution HLA genotyping of both donors and recipients. The risk factors, histopathological appearance and prognosis of cases with transplant glomerulopathy in the absence of HLA-DSA were compared to those cases with HLA-DSA, and the impact of the PIRCHE-II score and eplet mismatches on development of transplant glomerulopathy evaluated. In this cohort, 10.3% developed transplant glomerulopathy, on average 3.2 years post-transplant. At the time of glomerulopathy, 23.5% had persistent pre-transplant or de novo HLA-DSA, while 76.5% were HLA-DSA negative. Only HLA-DSA was identified as a risk factor for glomerulopathy development as eplet mismatches and the PIRCHE-II score did not associate. HLA-DSA negative biopsies with glomerulopathy had less interstitial inflammation, less glomerulitis, and less C4d deposition in the peritubular capillaries compared to the HLA-DSA positive biopsies with glomerulopathy. While graft function was comparable between the two groups, HLA-DSA positive glomerulopathy was associated with a significantly higher risk of graft failure compared to HLA-DSA negative glomerulopathy (Hazard Ratio 3.84; 95% confidence interval 1.94-7.59). Landmark analysis three-years post-transplant showed that HLA-DSA negative patients with glomerulopathy still had a significant increased risk of graft failure compared to patients negative for glomerulopathy (2.62; 1.46-4.72). Thus, transplant glomerulopathy often occurs in the absence of HLA-DSA, independent of HLA molecular mismatches, and represents a different phenotype with less concomitant inflammation and better graft survival compared to that developed in the presence of HLA-DSA.

Revisiting the changes in the Banff classification for antibody-mediated rejection after kidney transplantation.

The Banff classification for antibody-mediated rejection (ABMR) has undergone important changes, mainly by inclusion of C4d-negative ABMR in Banff'13 and elimination of suspicious ABMR (sABMR) with the use of C4d as surrogate for HLA-DSA in Banff'17. We aimed to evaluate the numerical and prognostic repercussions of these changes in a single-center cohort study of 949 single kidney transplantations, comprising 3662 biopsies that were classified according to the different versions of the Banff classification. Overall, the number of ABMR and sABMR cases increased from Banff'01 to Banff'13. In Banff'17, 248 of 292 sABMR biopsies were reclassified to No ABMR, and 44 of 292 to ABMR. However, reclassified sABMR biopsies had worse and better outcome than No ABMR and ABMR, which was mainly driven by the presence of microvascular inflammation and absence of HLA-DSA, respectively. Consequently, the discriminative performance for allograft failure was lowest in Banff'17, and highest in Banff'13. Our data suggest that the clinical and histological heterogeneity of ABMR is inadequately represented in a binary classification system. This study provides a framework to evaluate the updates of the Banff classification and assess the impact of proposed changes on the number of cases and risk stratification. Two alternative classifications introducing an intermediate category are explored.

Apixaban in patients on haemodialysis: a single-dose pharmacokinetics study.

BACKGROUND: Apixaban, a direct oral anticoagulant inhibiting factor Xa, has been proven to reduce the risk of atrial fibrillation-related stroke and thromboembolism in patients with mild to moderate renal insufficiency. Patients on renal replacement therapy, however, were excluded from randomized controlled trials. Therefore, uncertainty remains concerning benefits, dosing and timing of intake in haemodialysis population. METHODS: We conducted a Phase II pharmacokinetics study in which 24 patients on maintenance haemodialysis were given a single dose (2.5 mg or 5 mg) of apixaban, either 30 min before or immediately after dialysis on the mid-week dialysis day. RESULTS: Apixaban 5 mg resulted in higher area under the curve (AUC0-48) in comparison with 2.5 mg, although significance could only be reached for dosing pre-dialysis (2.5 mg versus 5 mg, P = 0.008). In line, peak concentrations (Cmax) after dosing pre-dialysis were significantly higher in the 5 mg than in the 2.5 mg groups (P = 0.02). In addition, dialysis resulted in significant reduction of drug exposure. AUC0-48 pre-dialysis were on average 48% (2.5 mg) and 26% (5 mg) lower than the AUC0-48 post-dialysis, in line with Cmax. As a result, a dose of 2.5 mg post-dialysis and a dose of 5 mg pre-dialysis resulted in similar AUC0-48. In contrast, significant differences were found between the 5 mg group post-dialysis and the 2.5 mg group pre-dialysis (P = 0.02). CONCLUSIONS: Our data suggest that exposure to apixaban in patients on maintenance haemodialysis is dependent not only on drug dose but also on timing of intake relative to the haemodialysis procedure.

The Histological Picture of Indication Biopsies in the First 2 Weeks after Kidney Transplantation.

BACKGROUND AND OBJECTIVES: In preclinical studies, ischemia-reperfusion injury and older donor age are associated with graft inflammation in the early phase after transplantation. In human kidney transplantation, impaired allograft function in the first days after transplantation is often adjudicated to donor- and procedure-related characteristics, such as donor age, donor type, and ischemia times. DESIGN: , setting, participants, & measurementsIn a cohort of 984 kidney recipients, 329 indication biopsies were performed within the first 14 days after transplantation. The histologic picture of these biopsies and its relationship with alloimmune risk factors and donor- and procedure-related characteristics were studied, as well as the association with graft failure. Multivariable Cox models were applied to quantify the cause-specific hazard ratios for early rejection and early inflammatory scores, adjusted for potential confounders. For quantification of hazard ratios of early events for death-censored graft failure, landmark analyses starting from day 15 were used. RESULTS: Early indication biopsy specimens displayed microvascular inflammation score ≥2 in 30% and tubulointerstitial inflammation score ≥2 in 49%. Rejection was diagnosed in 186 of 329 (57%) biopsies and associated with the presence of pretransplant donor-specific HLA antibodies and the number of HLA mismatches, but not nonimmune risk factors in multivariable Cox proportional hazards analysis. In multivariable Cox proportional hazards analysis, delayed graft function, the graft dysfunction that prompted an early indication biopsy, HLA mismatches, and pretransplant donor-specific HLA antibodies were significantly associated with a higher risk for death-censored graft failure, whereas early acute rejection was not. CONCLUSIONS: Indication biopsies performed early after kidney transplantation display inflammatory changes related to alloimmune risk factors. Nonimmune risk factors for ischemia-reperfusion injury, such as cold and warm ischemia time, older donor age, and donor type, were not identified as strong risk factors for early inflammation after human kidney transplantation.

Eplet Mismatch Load and De Novo Occurrence of Donor-Specific Anti-HLA Antibodies, Rejection, and Graft Failure after Kidney Transplantation: An Observational Cohort Study.

BACKGROUND: In kidney transplantation, evaluating mismatches of HLA eplets-small patches of surface-exposed amino acids of the HLA molecule-instead of antigen mismatches might offer a better approach to assessing donor-recipient HLA incompatibility and improve risk assessment and prediction of transplant outcomes. METHODS: To evaluate the effect of number of eplet mismatches (mismatch load) on de novo formation of donor-specific HLA antibodies (DSAs) and transplant outcomes, we conducted a cohort study that included consecutive adult kidney recipients transplanted at a single center from March 2004 to February 2013. We performed retrospective high-resolution genotyping of HLA loci of 926 transplant pairs and used the HLAMatchmaker computer algorithm to count HLA eplet mismatches. RESULTS: De novo DSAs occurred in 43 (4.6%) patients. Multivariable analysis showed a significant independent association between antibody-verified eplet mismatch load and de novo DSA occurrence and graft failure, mainly explained by DQ antibody-verified eplet effects. The association with DQ antibody-verified eplet mismatches was linear, without a safe threshold at which de novo DSA did not occur. Odds for T cell- or antibody-mediated rejection increased by 5% and 12%, respectively, per antibody-verified DQ eplet mismatch. CONCLUSIONS: Eplet mismatches in HLA-DQ confer substantial risk for de novo DSA formation, graft rejection, and graft failure after kidney transplantation. Mismatches in other loci seem to have less effect. The results suggest that antibody-verified HLA-DQ eplet mismatch load could be used to guide personalized post-transplant immunosuppression. Adoption of molecular matching for DQA1 and DQB1 alleles could also help to minimize de novo DSA formation and potentially improve transplant outcomes.

Clinical importance of extended second field high-resolution HLA genotyping for kidney transplantation.

The need for extended second field high-resolution (2F-HR) HLA genotyping in kidney transplantation is debated. In a cohort of 1000 kidney transplants, we evaluated the impact of different HLA genotyping levels on the assignment of donor-specific anti-HLA antibodies (DSA) and investigated whether inference of 2F-HR genotypes from low-resolution (LR) genotypes could be used to correctly assign DSA. Based on LR genotypes, 224 pretransplant DSAs were present in 140 patients and absent in 860 patients (DSAneg group). With extended 2F-HR HLA genotyping, we confirmed 173 DSA (77.2%) in 108 (77.1%) patients (2F-HRpos LRpos DSA group) and excluded DSA in 32 patients (22.9%) (2F-HRneg LRpos DSA group). Kaplan-Meier curves showed that 10-year graft survival rates were similar between the DSAneg and 2F-HRneg LRpos DSA groups (82.4% vs 93.8%; P = .27) and confirmed that DSA determined using LR typing but not confirmed using 2F-HR typing were indeed misclassified. By inferring 2F-HR genotypes using HaploStats, DSA still could not be correctly assigned in 23.3% of cases. We conclude that extended 2F-HR typing of the donor-recipient pairs is relevant for the correct assessment of DSA. Although inference of 2F-HR genotypes may improve the assessment of DSA in some cases, significant misclassification occurs, and warrants caution in using inferred HLA results for clinical and research purposes.

Assessing the Complex Causes of Kidney Allograft Loss.

BACKGROUND: Although graft loss is a primary endpoint in many studies in kidney transplantation and a broad spectrum of risk factors has been identified, the eventual causes of graft failure in individual cases remain ill studied. METHODS: We performed a single-center cohort study in 1000 renal allograft recipients, transplanted between March 2004 and February 2013. RESULTS: In total, 365 graft losses (36.5%) were identified, of which 211 (57.8%) were due to recipient death with a functioning graft and 154 (42.2%) to graft failure defined as return to dialysis or retransplantation. The main causes of recipient death were malignancy, infections, and cardiovascular disease. The main causes of graft failure were distinct for early failures, where structural issues and primary nonfunction prevailed, compared to later failures with a shift towards chronic injury. In contrast to the main focus of current research efforts, pure alloimmune causes accounted for only 17.5% of graft failures and only 7.4% of overall graft losses, although 72.7% of cases with chronic injury as presumed reason for graft failure had prior rejection episodes, potentially suggesting that alloimmune phenomena contributed to the chronic injury. CONCLUSIONS: In conclusion, this study provides better insight in the eventual causes of graft failure, and their relative contribution, highlighting the weight of nonimmune causes. Future efforts aimed to improve outcome after kidney transplantation should align with the relative weight and expected impact of targeting these causes.

Development and validation of a peripheral blood mRNA assay for the assessment of antibody-mediated kidney allograft rejection: A multicentre, prospective study.

BACKGROUND: Antibody-mediated rejection, a leading cause of renal allograft graft failure, is diagnosed by histological assessment of invasive allograft biopsies. Accurate non-invasive biomarkers are not available. METHODS: In the multicentre, prospective BIOMARGIN study, blood samples were prospectively collected at time of renal allograft biopsies between June 2011 and August 2016 and analyzed in three phases. The discovery and derivation phases of the study (N = 117 and N = 183 respectively) followed a case-control design and included whole genome transcriptomics and targeted mRNA expression analysis to construct and lock a multigene model. The primary end point was the diagnostic accuracy of the locked multigene assay for antibody-mediated rejection in a third validation cohort of serially collected blood samples (N = 387). This trial is registered with ClinicalTrials.gov, number NCT02832661. FINDINGS: We identified and locked an 8-gene assay (CXCL10, FCGR1A, FCGR1B, GBP1, GBP4, IL15, KLRC1, TIMP1) in blood samples from the discovery and derivation phases for discrimination between cases with (N = 49) and without (N = 134) antibody-mediated rejection. In the validation cohort, this 8-gene assay discriminated between cases with (N = 41) and without antibody-mediated rejection (N = 346) with good diagnostic accuracy (ROC AUC 79·9%; 95% CI 72·6 to 87·2, p < 0·0001). The diagnostic accuracy of the 8-gene assay was retained both at time of stable graft function and of graft dysfunction, within the first year and also later after transplantation. The 8-gene assay is correlated with microvascular inflammation and transplant glomerulopathy, but not with the histological lesions of T-cell mediated rejection. INTERPRETATION: We identified and validated a novel 8-gene expression assay that can be used for non-invasive diagnosis of antibody-mediated rejection. FUNDING: The Seventh Framework Programme (FP7) of the European Commission.

Specificity, strength, and evolution of pretransplant donor-specific HLA antibodies determine outcome after kidney transplantation.

In this cohort study (N = 924), we investigated the evolution and clinical significance of pretransplant donor-specific HLA antibodies (preDSA), detected in the single-antigen beads assay but complement-dependent cytotoxicity crossmatch-negative. Donor specificity of the preDSA (N = 107) was determined by high-resolution genotyping of donor-recipient pairs. We found that in 52% of the patients with preDSA, preDSA spontaneously resolved within the first 3 months posttransplant. PreDSA that persisted posttransplant had higher pretransplant median fluorescence intensity values and more specificity against DQ. Patients with both resolved and persistent DSA had a high incidence of histological picture of antibody-mediated rejection (ABMRh ; 54% and 59% respectively). Patients with preDSA that persisted posttransplant had worse 10-year graft survival compared to resolved DSA and preDSA-negative patients. Compared to cases without preDSA, Cox modeling revealed an increased risk of graft failure only in the patients with persistent DSA, in the presence (hazard ratio [HR] = 8.3) but also in the absence (HR = 4.3) of ABMRh . In contrast, no increased risk of graft failure was seen in patients with resolved DSA. We conclude that persistence of preDSA posttransplant has a negative impact on graft survival, beyond ABMRh . Even in the absence of antibody-targeting therapy, low median fluorescence intensity DSA and non-DQ preDSA often disappear early posttransplantation and are not deleterious for graft outcome.