Donor-specific Alloantibodies Research Articles

Post-transplant CFHR5 mutation-related atypical HUS: The need for pre-transplant diagnosis

Atypical hemolytic uremic syndrome (aHUS) is a rare form of thrombotic microangiopathy (TMA) affecting multiple organs and can be sporadic or familial. It is most commonly caused by dysregulation of the alternative complement pathway. aHUS can occur at any age with a high rate of progression to end-stage kidney disease. We describe the case of a 24-year-old man with chronic kidney disease, severe hypertension, and antineutrophilic antibody by IF positive. On biopsy, diffuse global glomerulosclerosis with TMA and IF findings of full house pattern suggestive of lupus nephritis were present. Considering a lupus nephritis case, after 2 years of hemodialysis underwent live-related renal transplant (Father Donor). Immediate post-transplant period developed severe cortical necrosis and TMA. An etiological workup was done to ascertain the cause of post-transplant TMA. After excluding common causes of antibody-mediated rejection (C4d and donor-specific alloantibody neg), calcineurin inhibitor toxicity, and infection, we detected an abnormal complement CFHR5 mutation with an autosomal dominant pattern of inheritance. Pre-transplant diagnosis could have prevented taking the kidney from the father for transplant and further prevented recurrence. Systemic lupus erythematosus and TMA both can have alternate complement pathway dysregulation leading to full house IF pattern and misdiagnosis.

Immunotherapy targeting B cells and long-lived plasma cells effectively eliminates pre-existing donor-specific allo-antibodies

Pre-existing anti-human leukocyte antigen (HLA) allo-antibodies constitute a major barrier to transplantation. Current desensitization approaches fail due to ineffective depletion of allo-specific memory B cells (Bmems) and long-lived plasma cells (LLPCs). We evaluate the efficacy of chimeric antigen receptor (CAR) Tcells targeting CD19 and B cell maturation antigen (BCMA) to eliminate allo-antibodies in a skin pre-sensitized murine model of isletallo-transplantation. We find that treatment of allo-sensitized hosts with CAR Tcells targeting Bmems and LLPCs eliminates donor-specific allo-antibodies (DSAs) and mitigates hyperacute rejection of subsequent isletallografts. We then assess the clinical efficacy of the CAR T therapy for desensitization in patients with multiple myeloma (MM) with pre-existing HLA allo-antibodies who were treated with the combination of CART-BCMA and CART-19 (ClinicalTrials.gov: NCT03549442) and observe clinically meaningful allo-antibody reduction. These findings provide logical rationale for clinical evaluation of CAR T-based immunotherapy in highly sensitized candidates to promote successful transplantation.

Preformed Donor-Specific Antibodies in Kidney Transplant: A Center Experience

The presence of preformed donor-specific alloantibodies (DSAs) at the time of transplantation has been associated with an increased risk of rejection, dysfunction, and shorter survival. Detection and identification of these antibodies have improved with more sensitive assays, but their clinical relevance and influence on long-term outcomes are unclear. We investigate the impact of pretransplant DSAs on the outcomes of kidney transplantation. A retrospective analysis was conducted on all patients who received a deceased donor kidney transplant in our center between January 2017 and December 2021. The study population consisted of 75 kidney transplantations, and DSAs were detected in 15 patients (20%) before transplantation. There was no significant difference in delayed graft function, serum creatinine level at discharge and the first year after transplant, acute rejection rates, or graft survival in patients with and without preformed DSAs. Detecting pretransplant DSAs by highly sensitive assays may not necessarily impact long-term graft outcomes, and the mismatch should be individually evaluated.

A new slow-releasing formulation of butyrate prolongs allograft survival

Abstract The microbiota consists of diverse microbes producing metabolites that can influence the immune system locally as well as systemically. These bioactive metabolites (postbiotics) have therapeutic potential and may prove useful to facilitate graft acceptance. Short-chain fatty acids (SCFA) promote intestinal homeostasis and have been linked to health and disease. The Hubbell-Nagler laboratories at the University of Chicago have developed a micelle-based platform that can deliver the SCFA butyrate systemically, with long-term retention and induction of tolerogenic responses. We hypothesized that the administration of this butyrate formulation will improve graft survival by suppressing the alloimmune response. Our preliminary data showed that only oral delivery of the butyrate formulation prolonged survival of a C57Bl/6 male to female minor-mismatched skin graft significantly, whereas sub-cutaneous and intra-peritoneal administration were ineffective. Improved graft survival was associated with diminished production of TNFα by splenic and skin-draining lymph node myeloid cells upon LPS stimulation, suggesting that butyrate modulates the myeloid compartment to reduce levels of pro-inflammatory cytokines. The postbiotic also enhanced survival of a BALB/c to C57Bl/6 major-mismatched skin graft, without the use of immunosuppressive drugs, and resulted in lower levels of donor-specific alloantibodies. Prolonging the administration of butyrate post-transplantation ameliorated graft survival even further, confirming the micelle-based platform has therapeutic potential. Current efforts focus on determining if the butyrate formulation drives myeloid cell polarization toward an anti-inflammatory phenotype. This project was supported by grant NIH/NIAID 2R01AI115716

Autoantibodies against DNA topoisomerase I promote renal allograft rejection by increasing alloreactive T cell responses

Antibodies reactive to self-antigens are an important component of posttransplant immune responses. The generation requirements and functions of autoantibodies, as well as the mechanisms of their influence on alloimmune responses, still remain to be determined. Our study investigated the contribution of autoimmunity during rejection of renal allografts. We have previously characterized a mouse model in which the acute rejection of a life-supporting kidney allograft is mediated by antibodies. At rejection, recipient sera screening against >4000 potential autoantigens revealed DNA topoisomerase I peptide 205-219 (TI-I205-219) as the most prominent epitope. Subsequent analysis showed TI-I205-219-reactive autoantibodies are induced in nonsensitized recipients of major histocompatibility complex–mismatched kidney allografts in a T cell–dependent manner. Immunization with TI-I205-219 broke self-tolerance, elicited TI-I205-219 immunoglobin G autoantibodies, and resulted in acute rejection of allogeneic but not syngeneic renal transplants. The graft loss was associated with increased priming of donor-reactive T cells but not with donor-specific alloantibodies elevation. Similarly, passive transfer of anti-TI-I205-219 sera following transplantation increased donor-reactive T cell activation with minimal effects on donor-specific alloantibody levels. The results identify DNA topoisomerase I as a novel self-antigen in transplant settings and demonstrate that autoantibodies enhance activation of donor-reactive T cells following renal transplantation.

Outcomes of ABO Incompatible Kidney Transplantation:A Single Center Experience

Background: Kidney transplantation (KT) is the treatment of choice for patients with end-stage renal disease (ESRD). General organ donor shortage remains an important limitation on KT, given the increasing number of patients waiting for transplantation. This has led to the adoption of different strategies to increase the pool of donors, such as ABO incompatible (ABOi) living KT. In this single center study, we retrospectively evaluated all donor/recipient pairs proposed for ABOi KT since the beginning of this transplantation program in our hospital in 2014. Methods: Between November 2014 and March 2022, 609 deceased donor and 223 living donor KT were performed in our center. Seventy-one donor/recipient pairs were proposed for ABOi KT and were evaluated for ABO/Rh group, Coombs tests, human leukocyte antigen (HLA) type I (A, B, C) and II (DR) high resolution genotyping and receptor to donor isoagglutinin titers and HLA antibodies. Recipients with immunoglobulins G (IgG) and/or M (IgM) isoagglutinin titers > 1:512 and/or with HLA donor specific alloantibodies (DSAs) were excluded from the ABOi transplantation program. Isoagglutinins removal was performed recurring to therapeutic plasma exchange (TPE) and/or immunoadsorption techniques. Transplanted patients were evaluated for demographic data, diagnosis, relationship with donor, sessions of antibodies removal, pre- and post-KT isoagglutinin titers, graft function, transfusion support, significant complications and overall patient survival. Results: Eighteen patients (14 males and 4 females) were transplanted with an ABOi graft out of 71 ABOi studied pairs. Median baseline IgG and IgM isoagglutinin titers were 1:32 (min 1:2, max 1:256) and 1:8 (min 1:2, max 1:32), respectively. Fifteen patients (83.3%) had pre-KT TPE and/or immunoadsorption sessions for isoagglutinin removal (mean 2.4 ± 1.8 session per patient). Seven patients (36.8%) were submitted to post-KT TPE (mean 1.9 ± 3.2 session per patient). No acute antibody mediated rejection was observed and overall graft survival was 100% on a follow-up period between 3 and 92 months (47.6 ± 25.2 months). All patients were dialysis-free with serum creatinine steady levels (median 1.4 mg/dL) at 47.6 ± 25.2 months of follow up. Conclusion: These results confirm that ABOi KT is a viable treatment for patients with ESRD, thus expanding living donor pool and reducing access time to transplantation.

The Histological Spectrum and Clinical Significance of T Cell-mediated Rejection of Kidney Allografts.

T cell-mediated rejection (TCMR) remains a significant cause of long-term kidney allograft loss, either indirectly through induction of donor-specific anti-HLA alloantibodies or directly through chronic active TCMR. Whether found by indication or protocol biopsy, Banff defined acute TCMR should be treated with antirejection therapy and maximized maintenance immunosuppression. Neither isolated interstitial inflammation in the absence of tubulitis nor isolated tubulitis in the absence of interstitial inflammation results in adverse outcomes, and neither requires antirejection treatment. RNA gene expression analysis of biopsy material may supplement conventional histology, especially in ambiguous cases. Lesser degrees of tubular and interstitial inflammation (Banff borderline) may portend adverse outcomes and should be treated when found on an indication biopsy. Borderline lesions on protocol biopsies may resolve spontaneously but require close follow-up if untreated. Following antirejection therapy of acute TCMR, surveillance protocol biopsies should be considered. Minimally invasive blood-borne assays (donor-derived cell-free DNA and gene expression profiling) are being increasingly studied as a means of following stable patients in lieu of biopsy. The clinical benefit and cost-effectiveness require confirmation in randomized controlled trials. Treatment of acute TCMR is not standardized but involves bolus corticosteroids with lymphocyte depleting antibodies for severe, refractory, or relapsing cases. Arteritis may be found with acute TCMR, active antibody-mediated rejection, or mixed rejections and should be treated accordingly. The optimal treatment ofchronic active TCMR is uncertain. Randomized controlled trials are necessary to optimally define therapy.

Donor Specific Allo-Antibody is Significantly Associated with Variability in Donor-Derived Cell-Free DNA Scores in Adult Heart Transplant Recipients

<h3>Purpose</h3> In this single-center retrospective pilot study heart transplant (HTx) patients were non-invasively monitored by donor-derived cell-free DNA (dd-cfDNA). We hypothesized that longitudinal dd-cfDNA variability scores correlate with preformed and de novo (dn) donor specific HLA allo-antibody (DSA). <h3>Methods</h3> 72 adult HTx patients with at least 3 dd-cfDNA results were included (AlloSure; CareDx Inc., Brisbane, CA). We determined dd-cfDNA variability (ASV) by standard deviation (SD) of longitudinal dd-cfDNA results, scaled x10 (ASV*10), and DSA by single antigen class I/II and MFI >1000 for HLA-A, B, DR, DQB/A and >2000 for HLA-C and DPB/A. <h3>Results</h3> Age at HTx was 49.1 years (SD 14.3). Study enrollment occurred at 270 days +/-SD 420.2 post-HTx. Biopsy-proven rejection (ACR ≥ 2R and/or pAMR ≥ 1) was detected in 16 (22 %), preformed DSA in 9 (12.5%), dnDSA in 13 (18%) and preformed + dn in 2 (2.7%) patients while 48 (66.7%) were DSA negative. Median time from HTx to dnDSA was 329d (range: 39-827). Including all values, mean ASV*10 was highest in patients with dnDSA (3.03 ± 4.74) compared to those with no DSA (0.65 ± 2.04) or preformed DSA (1.36 ± 3.40, p=0.001; <b>Table 1</b>). Patients who developed dnDSA showed a significant increase of ASV*10 after dnDSA detection (1.50 ± 2.06, p = 0.014), but not before dnDSA detection (1.24 ± 1.49, p=0.380) in comparison to patients with no DSA (0.65 ± 2.04, <b>Table 1</b>). <h3>Conclusion</h3> These findings highlight the value of longitudinal assessment of dd-cfDNA and DSA in HTx recipients. Our results show that increased dd-cfDNA variability is associated with the presence of preformed and dnDSA. Further, ASV*10 is significantly increased after identification of dnDSA, further supporting the mechanistic role of dnDSA in mediating poor outcomes after HTx. Confirmation in larger data sets may further advance the utility of longitudinal non-invasive surveillance using dd-cfDNA and dnDSA in transplant recipients.

Characteristics of pre-sensitization-related acute antibody-mediated rejection in a rat model of orthotopic liver transplantation.

To establish an animal model of pre-sensitization following liver transplantation either with or without immunosuppressors. To study whether accelerated liver rejection or acute antibody-mediated rejection (AMR) occurred and study the characteristics and potential mechanism in the animal model. Lewis (LEW) rats were subjected to liver [liver graft of Brown Norway (BN) rat] transplantation 2 weeks after lymphocyte injection (lymphocytes of BN rat; pre-sensitization). At 2 weeks after transplantation, serum samples of recipients were collected for antibody analysis to identify donor-specific alloantibody (DSA) level. The recipients were treated with or without a low dose of immunosuppressor (2 mg/kg). The liver grafts of each group were analyzed by hematoxylin and eosin (HE) stain, Masson stain, CK19, C4d, and CD20 immunohistochemical (IHC) stain, CD3, CD68, and CD86 immunofluorescence and transmission electron microscope (TEM) to study the characteristics of liver rejection. Moreover, cytotoxin-associated genes, M1 macrophages conversion-related proteins, and interleukin-6 (IL-6) signaling pathway proteins were detected by western blotting. High level of DSA and accelerated liver rejection occurred in the pre-sensitized rat models following liver transplant. Accelerated liver graft rejection occurred in the pre-sensitized, post liver transplant rats regardless of whether a low dose immunosuppressor had been applied. Severe injury of the interlobular bile ducts and accelerated fibrosis could be observed. Moreover, evidence of endothelial injury, such as capillary inflammation, was found in the pre-sensitized, post-transplant rats. In addition, C4d deposition and M1 macrophages recruitment were also found in this sensitized followed transplant model, indicating that complement activation might occur in this model. The levels of IL-6, JAK1, STAT3, SHP2, and ERK1-2 were increased in the pre-sensitized, post-transplant rats. Pre-sensitized post liver transplant rats might be potential AMR models for further study.

The impact of donor-specific alloantibodies on chronic antibody-mediated rejection after pediatric living donor liver transplantation

Seiichi Shimizu, Seisuke Sakamoto, Akinari Fukuda, Yusuke Yanagi, Hajime Uchida, Kotaro Mimori, Toshimasa Nakao, Masaki Yamada, Takako Yoshioka, Mureo Kasahara. Korean J Transplant 2021;35:42. https://doi.org/10.4285/ATW2021.OR-1094

Measuring the Impact of Targeting FcRn-Mediated IgG Recycling on Donor-Specific Alloantibodies in a Sensitized NHP Model.

BackgroundIn transplantation, plasmapheresis and IVIg provide the mainstay of treatment directed at reducing or removing circulating donor-specific antibody (DSA), yet both have limitations. We sought to test the efficacy of targeting the IgG recycling mechanism of the neonatal Fc receptor (FcRn) using anti-FcRn mAb therapy in a sensitized non-human primate (NHP) model, as a pharmacological means of lowering DSA.MethodsSix (6) rhesus macaque monkeys, previously sensitized by skin transplantation, received a single dose of 30mg/kg anti-RhFcRn IV, and effects on total IgG, as well as DSA IgG, were measured, in addition to IgM and protective immunity. Subsequently, 60mg/kg IV was given in the setting of kidney transplantation from skin graft donors. Kidney transplant recipients received RhATG, and tacrolimus, MMF, and steroid for maintenance immunosuppression.ResultsCirculating total IgG was reduced from a baseline 100% on D0 to 32.0% (mean, SD ± 10.6) on d4 post infusion (p<0.05), while using a DSA assay. T-cell flow cross match (TFXM) was reduced to 40.6±12.5% of baseline, and B-cell FXCM to 52.2±19.3%. Circulating total IgM and DSA IgM were unaffected by treatment. Pathogen-specific antibodies (anti-gB and anti-tetanus toxin IgG) were significantly reduced for 14d post infusion. Post-transplant, circulating IgG responded to anti-FcRn mAb treatment, but DSA increased rapidly.ConclusionTargeting the FcRn-mediated recycling of IgG is an effective means of lowering circulating donor-specific IgG in the sensitized recipient, although in the setting of organ transplantation mechanisms of rapid antibody rise post-transplant remains unaffected.

Treatment of antibody-mediated rejection with double-filtration plasmapheresis, low dose IVIg plus rituximab after kidney transplantation.

Antibody-mediated rejection (ABMR) at early or late post-transplantation remains challenging. We performed a single-center single-arm study where four cases of acute ABMR and nine cases of chronic active ABMR (defined by Banff classification) were treated with double-filtration plasmapheresis (two cycles of three consecutive daily sessions with a 4-day gap between). At the end of the third and sixth DFPP sessions, the patients received rituximab 375 mg/m2 . After a median follow-up of 1078 (61-1676) days, kidney-allograft survival was 50%. Before DFPP/rituximab therapy, the median donor-specific alloantibody (DSA) mean fluorescence intensity (MFI) was 9160 (4000-15 400); 45 days (D45) later it had significantly decreased to 7375 (215-18 100) (P=.018). In addition, at one-year (Y1) post-therapy, MFI had decreased further, that is, 4060 (400-7850) (P=.001). In two patients, DSA MFIs decreased and remained below 2000. The slope of estimated glomerular-filtration rate within the 6 months preceding intervention was -1.18 mL/min/month and remained unchanged at -1.29 mL/min/month within the year after intervention. Proteinuria remained unchanged. Baseline Banff scores on repeat allograft biopsies (post-therapy D45, Y1) did not show any improvement. Side-effects were mild to moderate. We conclude that the combined DFPP/rituximab significantly decreased DSAs in ABMR kidney-transplant recipients but did not improve renal function or renal histology at 1-year follow-up.

De Novo Donor Specific Antibody and Long-Term Outcome After Liver Transplantation: A Systematic Review and Meta-Analysis.

BackgroundThe impact of de novo anti-HLA donor-specific alloantibodies (DSA) which develop after long-term liver transplantation (LT) remains controversial and unclear. The aim of this study was to investigate the role of de novo DSAs on the outcome in LT.MethodsWe did a systematic review and meta-analysis of observational studies published until Dec 31, 2019, that reported de novo DSA outcome data (≥1 year of follow-up) after liver transplant. A literature search in the MEDLINE/PubMed, EMBASE, Cochrane Library, Scopus and Web of Science Core Collection databases was performed.ResultsOf 5,325 studies identified, 15 fulfilled our inclusion criteria. The studies which reported 2016 liver transplant recipients with de novo DSAs showed an increased complication risk, i.e. graft loss and chronic rejection (OR 3.61; 95% CI 1.94–6.71, P < 0.001; I2 58.19%), and allograft rejection alone (OR 6.43; 95% CI: 3.17–13.04; P < 0.001; I2 49.77%); they were compared to patients without de novo DSAs. The association between de novo DSAs and overall outcome failure was consistent across all subgroups and sensitivity analysis.ConclusionsOur study suggested that de novo DSAs had a significant deleterious impact on the liver transplant risk of rejection. The routine detection of de novo DSAs may be beneficial as noninvasive biomarker-guided risk stratification.

Long-term outcome after hand and forearm transplantation - a retrospective study.

SummaryBetween 2000 and 2014, five patients received bilateral hand (n = 3), bilateral forearm (n = 1), and unilateral hand (n = 1) transplants at the Innsbruck Medical University Hospital. We provide a comprehensive report of the long‐term results at 20 years. During the 6–20 years follow‐up, 43 rejection episodes were recorded in total. Of these, 27.9% were antibody‐related with serum donor‐specific alloantibodies (DSA) and skin‐infiltrating B‐cells. The cell phenotype in rejecting skin biopsies changed and C4d‐staining increased with time post‐transplantation. In the long‐term, a change in hand appearance was observed. The functional outcome was highly depending on the level of amputation. The number and severity of rejections did not correlate with hand function, but negatively impacted on the patients´ well‐being and quality of life. Patient satisfaction significantly correlated with upper limb function. One hand allograft eventually developed severe allograft vasculopathy and was amputated at 7 years. The patient later died due to progressive gastric cancer. The other four patients are currently rejection‐free with moderate levels of immunosuppression. Hand transplantation remains a therapeutic option for carefully selected patients. A stable immunologic situation with optimized and individually adopted immunosuppression favors good compliance and patient satisfaction and may prevent development of DSA.

De Novo Membranous Nephropathy and Donor-Specific Alloantibodies: A Path to the Pathophysiology?

De Novo Membranous Nephropathy and Donor-Specific Alloantibodies: A Path to the Pathophysiology?

Positive Crossmatch Kidney Transplantation: State of the Art and Future Perspectives

The organ shortage for kidney transplantation remains a challenging issue worldwide. Incompatibility between donor-recipient pairs, commonly occurring among transplant candidates who were sensitized from prior antigen exposure, serves as a significant barrier to kidney transplantation. In efforts to overcome this obstacle, living and deceased donor kidney transplantation across human leukocyte antigen barriers following desensitization has been pursued via positive crossmatch transplantation. The goal of desensitization therapy is to remove or denigrate donor-specific alloantibodies prior to transplantation in order to permit transplantation across the human leukocyte antigen barrier and prevent rejection. Various desensitization regimens have been utilized, including the use of plasmapheresis, intravenous immunoglobulin, and or immunoadsorption. Although long-term allograft outcomes for positive crossmatch kidney transplantation following desensitization therapy have been shown to be inferior to compatible transplantation, particularly with increasing strength of the crossmatch, there is an established survival benefit for positive crossmatch transplant recipients compared with remaining on the transplant waitlist. However, positive crossmatch transplantation may confer higher risks of infection and malignancy. Despite the fact that some of these heightened risks, positive crossmatch transplantation has also been demonstrated to have cost-savings compared with remaining on dialysis and may, therefore, be a cost-effective treatment option for sensitized patients who would face long waiting times and may never be able to achieve compatible transplantation. This review highlights both the risks and benefits of positive crossmatch transplantation and its role in the broader field of kidney transplantation.

EARLY POST-TRANSPLANT INFUSION OF EX VIVO-EXPANDED AUTOLOGOUS POLYCLONAL REGULATORY T CELLS (TREG) PROLONGS KIDNEY ALLOGRAFT SURVIVAL IN NONLYMPHODEPLETED, CTLA4IG-TREATED RHESUS MONKEYS

Introduction: Rodent studies have demonstrated the utility of regulatory T cell (Treg) infusion for the promotion of tolerance to organ allografts. Here, we evaluated the therapeutic potential of ex vivo-expanded autologous polyclonal Treg in rhesus monkeys with life-supporting kidney allografts. Methods: One week before transplantation (Tx), CD4+CD25+Treg were isolated from peripheral blood (using MACS), followed by polyclonal expansion using artificial APCs (L-cells) and IL-2. Recipients and donors were MHC-mismatched. Immunosuppression in the form of co-stimulation blockade (CTLA4Ig; 20mg/kg, with tapering until d180) and tacrolimus (trough levels 10-15ng/ml, with tapering until d60) was given to recipients without (n=3) or with (n=4) Treg infusion. Purity and phenotype of the expanded Treg were validated by flow cytometry. Treg infusions (total: 8 per graft recipient) were administered on d6 & d10 (10x106/kg per infusion), d13 & d16 (30x106/kg), and d20, d23, 27 & d30 (50x106/kg). Graft survival was determined by serum creatinine level, weight loss and urinary protein/creatinine ratio. Results: With no Treg infusion, graft survival times were 15, 35, and 40 days (median 35 days), compared to 38, 58, 86, and 234 days (median 72 days) with Treg infusion (p<0.05). In peripheral blood, no significant differences in memory CD4+ and CD8+T cell subsets were observed between recipients with or without Treg infusion. However, the percentages of CD4+CD28-CD95+ T cells in peripheral blood were significantly lower in recipients with Treg infusion (p<0.05). In both groups, no increase in donor-specific allo-antibodies was observed. With or without Treg infusion, the incidences of CD4+CD25hiFoxp3hi Treg in peripheral blood were markedly reduced after Tx. On the day of Tx (d0), the incidences of CD4+CD25hiFoxp3hi Treg in mesenteric lymph nodes (LN) were similar in both groups. However, on d35, mesenteric LN from Treg-infused monkeys exhibited significantly higher percentages of CD4+CD25hiFoxp3hi Treg than monkeys without Treg infusion (p<0.05). Conclusions: Multiple infusions of ex vivo-expanded polyclonal autologous Treg are safe and delay kidney allograft rejection in non-lymphodepleted monkeys treated with CTLA4Ig and tacrolimus. Furthermore, Treg were enriched in secondary lymphoid organs of recipients given Treg infusions. NIH grant U19 AI131453.

Natural soluble human leukocyte antigen class I in donor serum neutralizes donor-specific HLA alloantibodies in recipient serum.

BackgroundHuman leukocyte antigen (HLA) molecules are cell-bound but can be identified in a soluble form. These soluble HLA (sHLA) molecules have an immunomodulatory function. We investigated whether natural sHLA in donor serum can neutralize donor-specific HLA alloantibodies (DSAs) in recipient serum.MethodsNeutralizing effects of donor serum on DSAs in recipient serum were measured using inhibition assay principle of flow cytometric crossmatch (FCXM), performed using sera from 143 kidney transplant recipients and their donors. The adding of donor serum to recipient serum yielded lower mean fluorescence intensity (MFI) ratios (test/control) than when diluent was added [Roswell Park Memorial Institute (RPMI) or third-party serum], which was presumed to be caused by the neutralizing effects of sHLA.ResultsIn the recipient group with class I DSAs alone (N=14), donor serum addition to recipient serum resulted in lower T cell MFI ratios [2.25 (1.31‒32.51)] than those observed on RPMI addition [3.04 (1.33‒125.39), P<0.05]. In the recipient group with class II DSAs alone (N=27), donor serum addition showed no significant difference in B cell MFI ratios [5.03 (1.41‒103.53)] compared to diluent addition: RPMI [4.50 (1.34‒145.98)] or third-party serum [5.08 (1.44‒138.47)], P>0.05 for both.ConclusionUsing inhibition FCXM, we verified that natural sHLA class I in donor serum neutralizes DSAs in recipient serum. However, no neutralizing effects of sHLA class II were revealed in this study. These potentially beneficial effects of sHLA infused via blood-derived products should be considered when desensitizing highly HLA-sensitized patients.

A new model to determine Optimal Exposure to Tacrolimus and Mycophenolate Mofetil after renal transplantation.

Drug dosing for Tacrolimus (TAC) and Mycophenolate Mofetil (MMF) after kidney transplantation remains challenging. Therapeutic drug monitoring (TDM) offers a means to individualize drug dosing and improve outcomes. In this observational study, patients having mycophenolic acid (MPA) exposure assessed by limited sampling strategy (LSS) within the first 6months were included and followed for 1year. A total of 113 clinical events occurring in 110 patients were classified into 3 groups: Group 1 Stable (n=34), Group 2 Over drug exposed (n=64) having infections or drug toxicity and Group 3 Under drug exposed (n=15) developing rejection or de novo donor-specific alloantibodies. Although TAC levels, MMF dose, MPA, and MPA Glucuronide (MPAG) exposure, expressed as area under curve (AUC), individually failed to predict outcomes, a scoring model incorporating all 3 drug levels TAC TDM×(MPA AUC+MPAG/10 AUC) correctly classified outcomes. A score over 1071 had a sensitivity and specificity of 0.94 (95% CI 0.56-0.83) and 0.84 (95% CI 0.69-0.89) for over exposure. A score below 625 had a sensitivity and specificity of 0.76 (95% CI 0.53-0.93) and 0.80 (95% CI 0.41-0.70) for under exposure. This integrated model of assessing TAC and MMF exposure may facilitate individualized therapy.

Critical roles of TIGIT+ human regulatory B cells in the suppression of antibody-mediated allograft rejection

Abstract TIGIT (T cell immunoreceptor with Ig and ITIM domains) is a co-inhibitory receptor, highly expressed by Tregs, and contributes to the suppressive function of Tregs by limiting pro-inflammatory Th1 and Th17 cell responses. Herein, we report that subsets of human regulatory B cells (Bregs) express surface TIGIT and can thus play a critical role in immune regulation. Using data from human B cell surface arrays and RNA-seq, we identified two major TIGIT+ human Bregs subsets, exhibiting CD24hiCD27+CD39hiIgD+IgMhiCD1chi marginal zone (MZ)-like and CD24hiCD27+CD39+IgD−IgM+CD1c+ memory Bregs. We further demonstrated that TIGIT+ Bregs were also capable of suppressing immune responses by their ability to express not only IL-10 and PD-L1, but also granzyme B and TGFb1. In addition, TIGIT+ Bregs can also suppress immune responses by altering dendritic cell functions. In line with these observations, liver allograft recipients with donor-specific alloantibody (DSA) had significantly lower numbers of TIGIT+ Bregs than those without DSA. Moreover, the frequency of TIGIT+ Bregs correlated with that of CD4+CD25+CD127lo Tregs, while it inversely correlated with the frequency of follicular helper T cells. We thus concluded that TIGIT+ Bregs play critical roles in immune regulation in liver allograft recipients.