Not All Antibodies Are Created Equal: Factors That Influence Antibody Mediated Rejection

Consistent with Dr. Paul Terasaki’s “humoral theory of rejection” numerous studies have shown that HLA antibodies can cause acute and chronic antibody mediated rejection (AMR) and decreased graft survival. New evidence also supports a role for antibodies to non-HLA antigens in AMR and allograft injury. Despite the remarkable efforts by leaders in the field who pioneered single antigen bead technology for detection of donor specific antibodies, a considerable amount of work is still needed to better define the antibody attributes that are associated with AMR pathology. This review highlights what is currently known about the clinical context of pre and posttransplant antibodies, antibody characteristics that influence AMR, and the paths after donor specific antibody production (no rejection, subclinical rejection, and clinical dysfunction with AMR).

Download Full-text

Intensity of de novo DSA detected by Immucor Lifecodes assay and C3d fixing antibodies are not predictive of subclinical ABMR after Kidney Transplantation

PLoS ONE ◽

10.1371/journal.pone.0249934 ◽

2021 ◽

Vol 16 (4) ◽

pp. e0249934

Author(s):

Dominique Bertrand ◽

Rangolie Kaveri ◽

Charlotte Laurent ◽

Philippe Gatault ◽

Maïté Jauréguy ◽

...

Keyword(s):

Kidney Transplantation ◽

Prognostic Value ◽

De Novo ◽

Added Value ◽

Multicentric Study ◽

Specific Antibodies ◽

Antibody Mediated Rejection ◽

Single Antigen ◽

Donor Specific Antibodies ◽

Allograft Loss

De novo donor-specific antibodies (dnDSA) are associated with antibody-mediated rejection (ABMR) and allograft loss. We tested Immucor* (IM) Luminex Single-antigen beads (LSAB) assay and C3d-fixing antibodies in the setting of dnDSA and subclinical (s) ABMR. This retrospective multicentric study included 123 patients biopsied because of the presence of subclinical de novo DSA detected by One Lamda* Labscreen (MFI > 1000). In 112 patients, sera of the day of the biopsy were available and tested in a central lab with IM Lifecodes LSAB and C3d fixing antibodies assays. In 16 patients (14.3%), no DSA was detected using Immucor test. In 96 patients, at least one DSA was determined with IM. Systematic biopsies showed active sABMR in 30 patients (31.2%), chronic active sABMR in 17 patients (17.7%) and no lesions of sABMR in 49 KT recipients (51%). Intensitity criteria (BCM, BCR and AD-BCR) of DSA were not statistically different between these 3 histological groups. The proportion of patients with C3d-fixing DSA was not statistically different between the 3 groups and did not offer any prognostic value regarding graft survival. Performing biopsy for dnDSA could not be guided by the intensity criteria of IM LSAB assay. C3d-fixing DSA do not offer added value.

Download Full-text

CRISPR/Cas9-Engineered HLA-Deleted Glomerular Endothelial Cells as a Tool to Predict Pathogenic Non-HLA Antibodies in Kidney Transplant Recipients

Journal of the American Society of Nephrology ◽

10.1681/asn.2021050689 ◽

2021 ◽

Vol 32 (12) ◽

pp. 3231-3251

Author(s):

Baptiste Lamarthée ◽

Carole Burger ◽

Charlotte Leclaire ◽

Emilie Lebraud ◽

Aniela Zablocki ◽

...

Keyword(s):

Endothelial Cells ◽

Kidney Transplant ◽

Transplant Recipients ◽

Kidney Transplant Recipients ◽

Hla Antibodies ◽

Specific Antibodies ◽

Antibody Mediated Rejection ◽

Biopsy Specimens ◽

Glomerular Endothelial Cells ◽

Donor Specific Antibodies

BackgroundAfter kidney transplantation, donor-specific antibodies against human leukocyte antigen donor-specific antibodies (HLA-DSAs) drive antibody-mediated rejection (ABMR) and are associated with poor transplant outcomes. However, ABMR histology (ABMRh) is increasingly reported in kidney transplant recipients (KTRs) without HLA-DSAs, highlighting the emerging role of non-HLA antibodies (Abs).MethodsW e designed a non-HLA Ab detection immunoassay (NHADIA) using HLA class I and II–deficient glomerular endothelial cells (CiGEnCΔHLA) that had been previously generated through CRISPR/Cas9-induced B2M and CIITA gene disruption. Flow cytometry assessed the reactivity to non-HLA antigens of pretransplantation serum samples from 389 consecutive KTRs. The intensity of the signal observed with the NHADIA was associated with post-transplant graft histology assessed in 951 adequate biopsy specimens.ResultsW e sequentially applied CRISPR/Cas9 to delete the B2M and CIITA genes to obtain a CiGEnCΔHLA clone. CiGEnCΔHLA cells remained indistinguishable from the parental cell line, CiGEnC, in terms of morphology and phenotype. Previous transplantation was the main determinant of the pretransplantation NHADIA result (P<0.001). Stratification of 3-month allograft biopsy specimens (n=298) according to pretransplantation NHADIA tertiles demonstrated that higher levels of non-HLA Abs positively correlated with increased glomerulitis (P=0.002), microvascular inflammation (P=0.003), and ABMRh (P=0.03). A pretransplantation NHADIA threshold of 1.87 strongly discriminated the KTRs with the highest risk of ABMRh (P=0.005, log-rank test). A multivariate Cox model confirmed that NHADIA status and HLA-DSAs were independent, yet synergistic, predictors of ABMRh.ConclusionThe NHADIA identifies non-HLA Abs and strongly predicts graft endothelial injury independent of HLA-DSAs.

Download Full-text

Autoantibodies Against Ro/SS-A, CENP-B, and La/SS-B are Increased in Patients with Kidney Allograft Antibody-Mediated Rejection

10.1101/2020.12.02.408922 ◽

2020 ◽

Author(s):

Sergi Clotet-Freixas ◽

Max Kotlyar ◽

Caitriona M. McEvoy ◽

Chiara Pastrello ◽

Sonia Rodríguez-Ramírez ◽

...

Keyword(s):

Kidney Transplant ◽

Hla Antigens ◽

Kidney Graft ◽

Hla Antibodies ◽

Antibody Mediated Rejection ◽

Transplant Patients ◽

Kidney Transplant Patients ◽

Donor Specific Antibodies ◽

Cellular Rejection ◽

Independent Cohort

ABSTRACTAntibody-mediated rejection (AMR) causes >50% of late kidney graft losses. Although donor-specific antibodies (DSA) against HLA cause AMR, antibodies against non-HLA antigens are also linked to rejection. Identifying key non-HLA antibodies will improve our understanding of antibody-mediated injury.We analyzed non-HLA antibodies using protein microarrays in sera from 91 kidney transplant patients with AMR, mixed rejection, acute cellular rejection (ACR), or acute tubular necrosis (ATN). IgM and IgG antibodies against 134 non-HLA antigens were measured pre-transplant, at the time of biopsy-proven diagnosis, and post-diagnosis. Findings were validated in 60 kidney transplant patients from an independent cohort.Seventeen non-HLA antibodies were significantly increased (p<0.05) in AMR and mixed rejection compared to ACR or ATN pre-transplant, nine at diagnosis and six post-diagnosis. AMR and mixed cases showed significantly increased pre-transplant levels of IgG anti-Ro/SS-A and anti-CENP-B, compared to ACR. Together with IgM anti-CENP-B and anti-La/SS-B, these antibodies were also significantly increased in AMR/mixed rejection at diagnosis. Increased IgG anti-Ro/SS-A and anti-CENP-B pre-transplant and at diagnosis, and IgM anti-La/SS-B at diagnosis, were associated with the presence of microvascular lesions, but not with tubulitis or interstitial/total inflammation. All three antibodies were associated with the presence of class-II DSA (p<0.05). Significantly increased IgG anti-Ro/SS-A in AMR/mixed compared to ACR (p=0.01), and numerically increased IgM anti-CENP-B (p=0.05) and anti-La/SS-B (p=0.06), were validated in the independent cohort.This is the first study that implicates autoantibodies against Ro/SS-A and CENP-B in AMR. These non-HLA antibodies may participate in the crosstalk between autoimmunity and alloimmunity in kidney AMR.SIGNIFICANCE STATEMENTAntibody-mediated rejection (AMR) causes >50% of kidney graft losses. Although donor-specific antibodies against HLA cause AMR, antibodies against non-HLA antigens are also linked to rejection. Serum samples of 91 kidney transplant patients were analyzed using protein arrays against 134 non-HLA antigens. AMR and mixed rejection cases showed significantly increased pre-transplant levels of IgG anti-Ro/SS-A and anti-CENP-B, compared to acute cellular rejection. Together with IgM anti-CENP-B and anti-La/SS-B, these antibodies were significantly increased in AMR/mixed rejection at diagnosis and were validated in a second, independent cohort. Increased IgG anti-Ro/SS-A, IgG anti-CENP-B and IgM anti-La/SS-B were associated with the presence of microvascular lesions and anti-HLA class-II antibodies. This is the first study to implicate anti-Ro/SS-A, anti-La/SS-B and anti-CENP-B autoantibodies in AMR.

Download Full-text

Long-term survival of the transplanted kidney and the clinical relevance of donor-specific antibodies

Molecular and experimental biology in medicine ◽

10.33602/mebm.3.1.3 ◽

2020 ◽

Vol 3 (1) ◽

pp. 17-22

Author(s):

Eva Svobodova ◽

Sarka Valhova ◽

Ondrej Viklicky ◽

Ilja Striz ◽

Jelena Skibova ◽

...

Keyword(s):

Graft Survival ◽

Solid Phase ◽

Kidney Graft ◽

Term Survival ◽

Specific Antibodies ◽

Antibody Mediated Rejection ◽

Number Of Patients ◽

Single Antigen ◽

Donor Specific Antibodies

The aim of our study was to evaluate the relevance of donor-specific antibodies (DSA) as defined by solid-phase single-antigen (SA) assays for predicting long-term graft survival after kidney transplantation. Sera from 132 kidney transplant recipients were retrospectively tested before, 3, 6 and 12 months after transplantation. The incidence of rejection and graft survival was followed up for 7 years. We found 29 episodes of acute cellular rejection (CR), 21 cases of antibody-mediated rejection (AMR) and 18 graft failures due to immunological reasons. Pre-transplant DSA and DSA three months after transplantation correlated with an increased rate of AMR and impaired graft function. After the fourth year, recipients with persistent DSA were at a higher risk of graft failure (p = 0.0317). Antibody specificity was prevailingly to HLA class I antigens (66.6% DSA, 75% non-DSA). During the first year after transplantation, the number of patients with non-DSA decreased (30.3% to 10.7%), while, due to de novo production of antibodies, the number of DSA positive patients remained constant. Conclusion: Detection of antibodies to HLA antigens using solid-phase assays, especially single-antigen bead technology before and three months after transplantation is predictive for increased incidence of antibody-mediated rejection and impaired long-term kidney graft survival.

Download Full-text

FC 128NON-HLA ANTIBODIES AND EPLET MISMATCHES IN KIDNEY TRANSPLANT RECIPIENTS WITH A HISTOLOGICAL PICTURE OF ANTIBODY-MEDIATED REJECTION WITH AND WITHOUT HLA DONOR-SPECIFIC ANTIBODIES

Nephrology Dialysis Transplantation ◽

10.1093/ndt/gfab148.003 ◽

2021 ◽

Vol 36 (Supplement_1) ◽

Author(s):

Marta Crespo ◽

Laura Llinas ◽

Dolores Redondo Pachon ◽

Carrie L Butler ◽

Javier Gimeno ◽

...

Keyword(s):

Kidney Transplant ◽

Graft Survival ◽

Kidney Transplant Recipients ◽

Serum Samples ◽

Hla Antibodies ◽

Aortic Endothelial Cells ◽

Specific Antibodies ◽

Antibody Mediated Rejection ◽

Before And After ◽

Donor Specific Antibodies

Abstract Background and Aims Correlation between antibody-mediated rejection (ABMR) and HLA donor-specific antibodies (DSA) is strong but imperfect in kidney transplant (KT) recipients, raising the possibility of other detrimental antibodies contributing to ABMR. The role of non-HLA antibodies on outcomes is not well known. Method We retrospectively assessed KT biopsies scored according to Banff’15 classification. Pre- and post-KT serum samples were checked for HLA and non-HLA antibodies (MICA-Ab, angiotensin II type 1 receptor (AT1R)-Ab, endothelin-1 type A receptor (ETAR)-Ab and crossmatches with primary aortic endothelial cells (EC-XM)). We also analyzed HLA epitope mismatches between donors and recipients. Results One-hundred eighteen patients with normal (n=19), ABMR histology (n=52) or IFTA (n=47) in their biopsy were studied. Graft survival was worse in ABMR patients (p=0.003). Pre-KT HLA-DSA were more frequent in ABMR cases (p=0.006). At biopsy, 73% ABMR patients had HLA-DSA (p<0.001). Pre-KT AT1R-Ab were more frequent in ABMR compared with IFTA and normal cases (p=0.003), without differences in other non-HLA antibodies. Fourteen patients with histological ABMR (27%) had no detectable HLA-DSA post-KT and only 3 had non-HLA Ab. However, these ABMR-DSA- cases showed similar biopsy changes and graft survival compared with ABMR-DSA+. Pre- or post-KT non-HLA antibodies other than AT1R-Ab were detected similarly in ABMR and in normal or IFTA cases. Both total class II and DRB1 epitope mismatches were associated with postransplant DSA and ABMR-DSA+. Multivariate analysis showed that both pre-KT HLA-DSA and AT1R-Ab (DSA: OR: 3.39 [1.20-9.59], p=0.021; AT1R-Ab: OR: 5.31 [1.75-16.10], p=0.003) were strong independent predictors of postransplant ABMR-DSA+ (Table 1). Conclusion Despite highly prevalent HLA-DSA before and after transplantation in KT with histological ABMR, 27% of cases did not show circulating HLA-DSA. Pre-KT AT1R-Ab associated with ABMR-DSA+, but not MICA-Ab, ETAR-Ab or EC-XM+. Any of them associated significantly with ABMR-DSA-. Epitope mismatch predicted both postransplant DRB-DSA and ABMR-DSA+. Detection of pre-KT HLA-DSA and/or AT1R-Ab, together with HLA epitope mismatch assessment, are valuable tools for better DSA and ABMR prediction in KT patients.

Download Full-text

Late and Chronic Antibody-Mediated Rejection: Main Barrier to Long Term Graft Survival

Clinical and Developmental Immunology ◽

10.1155/2013/859761 ◽

2013 ◽

Vol 2013 ◽

pp. 1-7 ◽

Cited By ~ 11

Author(s):

Qiquan Sun ◽

Yang Yang

Keyword(s):

Graft Survival ◽

Graft Loss ◽

Limited Effect ◽

Hla Antibodies ◽

Specific Antibodies ◽

Antibody Mediated Rejection ◽

Donor Specific Antibodies ◽

Main Barrier ◽

Lines Of Evidence

Antibody-mediated rejection (AMR) is an important cause of graft loss after organ transplantation. It is caused by anti-donor-specific antibodies especially anti-HLA antibodies. C4d had been regarded as a diagnosis marker for AMR. Although most early AMR episodes can be successfully controlled or reversed, late and chronic AMR remains the leading cause of late graft loss. The strategies which work in early AMR have limited effect on late/chronic episodes. Here, we reviewed the lines of evidence that late/chronic AMR is the leading cause of late graft loss, characteristics of late AMR, and current strategies in managing late/chronic AMR. More effort should be put on the management of late/chronic AMR to make a better long term graft survival.

Download Full-text

Proteomics Reveals Extracellular Matrix Injury in the Glomeruli and Tubulointerstitium of Kidney Allografts with Early Antibody-Mediated Rejection

10.1101/2020.03.03.975672 ◽

2020 ◽

Author(s):

Sergi Clotet-Freixas ◽

Caitriona M. McEvoy ◽

Ihor Batruch ◽

Julie Van ◽

Chiara Pastrello ◽

...

Keyword(s):

Extracellular Matrix ◽

Hla Antigens ◽

Acute Cellular Rejection ◽

Ecm Remodeling ◽

Specific Antibodies ◽

Antibody Mediated Rejection ◽

Tubular Necrosis ◽

Ecm Proteins ◽

Donor Specific Antibodies ◽

Cellular Rejection

ABSTRACTAntibody-mediated rejection (AMR) accounts for >50% of kidney allograft losses. AMR is caused by donor-specific antibodies (DSA) against HLA and non-HLA antigens in the glomeruli and the tubulointerstitium, which together with inflammatory cytokines such as tumor necrosis factor alpha (TNFα) and interferon gamma (IFNɣ), trigger graft injury. Unfortunately, the mechanisms governing cell-specific injury in AMR remain unclear. We studied 30 for-cause kidney biopsies with early AMR, acute cellular rejection or acute tubular necrosis (‘non-AMR’). We laser-captured and microdissected glomeruli and tubulointerstitium, and subjected them to unbiased proteome analysis. 120/2026 glomerular and 180/2399 tubulointerstitial proteins were significantly differentially expressed in AMR vs. non-AMR biopsies (P<0.05). Basement membrane and extracellular matrix (ECM) proteins were significantly decreased in both AMR compartments. We verified decreased glomerular and tubulointerstitial LAMC1 expression, and decreased glomerular NPHS1 and PTPRO expression in AMR. Cathepsin-V (CTSV) was predicted to cleave ECM-proteins in the AMR glomeruli, and CTSL, CTSS and LGMN in the tubulointerstitium. We identified galectin-1, an immunomodulatory protein upregulated in the AMR glomeruli and linked to the ECM. Anti-HLA class-I antibodies significantly increased CTSV expression, and galectin-1 expression and secretion, in human glomerular endothelial cells. Glutathione S-transferase omega-1 (GSTO1), an ECM-modifying enzyme, was significantly increased in the AMR tubulointerstitium, and in TNFα-treated proximal tubular epithelial cells. IFNɣ and TNFα significantly increased CTSS and LGMN expression in these cells. Basement membranes are often remodeled in chronic AMR, and we demonstrated that this remodeling begins early in glomeruli and tubulointerstitium. Targeting ECM-remodeling in AMR may represent a new therapeutic opportunity.SIGNIFICANCE STATEMENTAntibody-mediated rejection (AMR) accounts for >50% of kidney allograft loss, and is caused by donor-specific antibodies against HLA antigens, which induce maladaptive responses in the kidney glomeruli and tubulointerstitium. This is the first unbiased proteomics analysis of laser-captured/microdissected glomeruli and tubulointerstitium from 30 indication kidney biopsies with early AMR, acute cellular rejection or acute tubular necrosis. >2,000 proteins were quantified in each compartment. We discovered that basement membrane and extracellular matrix (ECM) proteins were significantly decreased in both AMR compartments. Two ECM-modifying proteins, LGALS1 and GSTO1, were significantly increased in glomeruli and tubulointerstitium, respectively. LGALS1 and GSTO1 were upregulated by anti-HLA antibodies or AMR-related cytokines in primary kidney cells, and may represent therapeutic targets to ameliorate ECM-remodeling in AMR.

Download Full-text