CLINICAL RESEARCH www.jasn.org Value of Donor–Specific Anti–HLA Antibody Monitoring and Characterization for Risk Stratification of Kidney Allograft Loss † †‡ | Denis Viglietti,* Alexandre Loupy, Dewi Vernerey,§ Carol Bentlejewski, Clément Gosset,¶ † † †‡ Olivier Aubert, Jean-Paul Duong van Huyen,** Xavier Jouven, Christophe Legendre, † | † Denis Glotz,* Adriana Zeevi, and Carmen Lefaucheur* Departments of *Nephrology and Kidney Transplantation and ¶Pathology, Saint Louis Hospital and Departments of ‡Kidney Transplantation and **Pathology, Necker Hospital, Assistance Publique Hôpitaux de Paris, Paris, France; †Paris Translational Research Center for Organ Transplantation, Institut National de la Santé et de la Recherche Médicale, UMR-S970, Paris, France; §Methodology Unit (EA 3181) CHRU de Besançon, France; and |University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania ABSTRACT The diagnosis system for allograft loss lacks accurate individual risk stratification on the basis of donor– specific anti–HLA antibody (anti-HLA DSA) characterization. We investigated whether systematic moni- toring of DSA with extensive characterization increases performance in predicting kidney allograft loss. This prospective study included 851 kidney recipients transplanted between 2008 and 2010 who were systematically screened for DSA at transplant, 1 and 2 years post-transplant, and the time of post– transplant clinical events. We assessed DSA characteristics and performed systematic allograft biopsies at the time of post–transplant serum evaluation. At transplant, 110 (12.9%) patients had DSAs; post- transplant screening identified 186 (21.9%) DSA-positive patients. Post–transplant DSA monitoring im- proved the prediction of allograft loss when added to a model that included traditional determinants of allograft loss (increase in c statisticfrom0.67;95%confidence interval [95% CI], 0.62 to 0.73 to 0.72; 95% CI, 0.67 to 0.77). Addition of DSA IgG3 positivity or C1q binding capacity increased discrimination per- formance of the traditional model at transplant and post-transplant. Compared with DSA mean fluores- cence intensity, DSA IgG3 positivity and C1q binding capacity adequately reclassified patients at lower or higher risk for allograft loss at transplant (category–free net reclassification index, 1.30; 95% CI, 0.94 to 1.67; P,0.001 and 0.93; 95% CI, 0.49 to 1.36; P,0.001, respectively) and post-transplant (category–free net reclassification index, 1.33; 95% CI, 1.03 to 1.62; P,0.001 and 0.95; 95% CI, 0.62 to 1.28; P,0.001, respectively). Thus, pre– and post–transplant DSA monitoring and characterization may improve individual risk stratification for kidney allograft loss. J Am Soc Nephrol 28: 702–715, 2017. doi: 10.1681/ASN.2016030368 Donor–specificanti–HLA antibodies (anti-HLA Received March 29, 2016. Accepted June 29, 2016. DSAs) have been extensively reported to be D. Viglietti and A.L. are co-first authors and contributed equally strongly associated with increased risks of rejec- to this work. A.Z. and C. Lefaucheur are co-last authors and tion and allograft loss.1–6 Although their value contributed equally to this work. for accurate risk stratification of transplant out- Published online ahead of print. Publication date available at comes has not been determined in the current www.jasn.org. literature, the detrimental influence of anti-HLA DSAs on transplant outcomes has placed anti- Correspondence: Dr. Carmen Lefaucheur, Service de Néphrologie et Transplantation, Hôpital Saint-Louis, 1 Avenue Claude Vellefaux, HLA antibodies at the center of national and 75010 Paris, France. Email: [email protected] local allocation policies in the United States – and Europe.7 10 Copyright © 2017 by the American Society of Nephrology 702 ISSN : 1046-6673/2802-702 J Am Soc Nephrol 28: 702–715, 2017 www.jasn.org CLINICAL RESEARCH Today, anti-HLA DSAs are considered to be among the most subclasses) to improve individual risk stratification for important biomarkers for predicting allograft injury and loss. allograft loss. However, there is no consensus for defining their pathogenicity and no standard for their evaluation to guide clinical decision making.11,12 The current conventional approach to pre– and RESULTS post–transplant immunologic risk evaluation is on the basis of the assessment by sensitive techniques of anti–HLA antibody Patient Characteristics specificity and strength, most frequently expressed by the This prospective study enrolled 851 patients among 906 mean fluorescence intensity (MFI) provided by single– consecutive recipients undergoing kidney transplantation antigen flow bead techniques.8–10,13 between January 1, 2008 and December 31, 2010. The study Recently, significant advances have occurred in our ability to flow chart is provided in Figure 1. diagnose patients with antibody-mediated rejection (ABMR) and The characteristics of the study population at the time of link anti–HLA antibody characteristics to transplant outcomes. transplantation are summarized in Table 1. The median follow- Theseadvancesincludetheassessmentofthecapacityofanti- up after transplantation was 5.3 years (interquartile range, HLA antibodies to bind complement, particularly C1q binding, 4.6–6.2). and the characterization of their IgG subclass composition. Con- verging evidence has supported that the capacity of anti-HLA Anti–HLA DSA Characteristics According to Time of DSA to bind complement is associated with an increased risk Detection of antibody-mediated injury and poor allograft survival in not Anti–HLA DSA Characteristics at the Time of Transplantation only kidney transplant14–20 but also, heart,21,22 liver,23 and lung24 Among the 110 (12.9%) patients with circulating anti–HLA transplant. Furthermore, emerging data have emphasized the DSA at the time of transplantation, the DSA with the highest clinical relevance of the IgG subclass composition of anti-HLA MFI level, the immunodominant donor–specific antibody DSAs and their relationships with allograft injury phenotype25 (iDSA), was HLA class 1 in 52 (47.3%) patients and HLA class and survival in kidney25–27 and liver23,28 transplantation. 2 in 58 (52.7%) patients, with a mean MFI of 5952.164213.4 Considering that one of the most pressing unmet needs in and C1q binding capacity in 35 (31.8%) patients. IgG1 was transplant medicine involves delineating the characteristics of positive for 82 (74.6%) iDSAs, IgG2 was positive for 48 circulating anti–HLA antibodies that confer pathogenesis and (43.6%) iDSAs, IgG3 was positive for 31 (28.2%) iDSAs, influence transplant outcomes, the Transplantation Society An- and IgG4 was positive for 30 (27.3%) iDSAs. tibody Consensus Group issued a call to action in 2013 and encouraged the transplant community to focus future efforts Post–Transplant Anti–HLA DSA Characteristics on clinical trials that include serial anti–HLA DSA monitoring Among the186 (21.9%)patients identified with anti-HLA DSA with the assessment of anti–HLA DSA characteristics, including after transplantation, 86 (46.2%) patients were positive for their complement binding capacity and IgG subclass composi- anti-HLA DSA at the time of a clinical event, 55 (29.6%) tion.9 After decades of studies emphasizing the associations be- patients were identified at 1 year after transplantation, and 45 tween anti-HLA antibodies and kidney transplant outcomes,29 (24.2%) patients were identified at 2 years after transplanta- the key issue today is to evaluate whether systematic anti–HLA tion. In total, the iDSA was HLA class 1 in 76 (40.9%) patients DSA monitoring integrating the assessment of antibody char- and HLA class 2 in 110 (59.1%) patients, with a mean MFI of acteristics might improve risk stratification for allograft loss.30 5746.764627.7 and C1q binding capacity in 57 (30.7%) pa- Stratifying patients by their immunologic risk has the potential tients. IgG1 was positive for 137 (73.7%) iDSAs, IgG2 was to resolve the puzzle of alloimmune conditions determining positive for 80 (43.0%) iDSAs, IgG3 was positive for 42 allograft outcomes and increase long–term allograft and patient (22.6%) iDSAs, and IgG4 was positive for 46 (24.7%) iDSAs. survival by improving the efficacy of allocation policies and The characteristics of post–transplant anti–HLA DSA at the therapeutic strategies.31 time of detection are shown in Table 2. Our hypothesis was that systematic monitoring and precise A comparison of post–transplant anti–HLA iDSA charac- characterization of anti-HLA DSAs, including their comple- teristics according to their preformed/de novo status is shown ment binding capacity and IgG subclass composition, might in Supplemental Table 1. add to the predictive value for allograft loss of the conventional approach on the basis of their detection and strength assessed Clinical and Histologic Characteristics at the Time of by MFI level. To test this hypothesis, we specifically designed Post–Transplant Anti–HLA DSA Detection a prospective study performed in a large and unselected All of the patients with post–transplant anti–HLA DSA (n=186) population of kidney transplant recipients who underwent underwent kidney allograft biopsy at the time of anti–HLA DSA standardized monitoring of anti-HLA DSAs together with detection. Among them, 65 (34.9%) patients had clinical ABMR, systematic allograft biopsies. Weassessed the performance of and 67 (36.0%) patients had subclinical ABMR. The clinical and prospective systematic monitoring and characterization of histologic characteristics, according to the time of post–transplant anti-HLA