Reduction In Donor-specific Antibodies Research Articles

Reduction of HLA donor specific antibodies in heart transplant patients treated with proteasome inhibitors for antibody mediated rejection.

In this project, we describe proteasome inhibitor (PI) treatment of antibody-mediated rejection (AMR) in heart transplantation (HTX). From January 2018 to September 2021, 10 patients were treated with PI for AMR: carfilzomib (CFZ) n=8; bortezomib (BTZ) n=2. Patients received 1-3 cycles of PI. All patients had ≥1 strong donor-specific antibody (DSA) (mean fluorescence intensity [MFI]>8000) in undiluted serum. Most DSAs (20/21) had HLA class II specificity. The MFI of strong DSAs had a median reduction of 56% (IQR=13%-89%) in undiluted serum and 92% (IQR=53%-95%) at 1:16 dilution. Seventeen DSAs in seven patients were reduced>50% at 1:16 dilution after treatment. Four DSAs from three patients did not respond. DSA with MFI>8000 at 1:16 dilution was less responsive totreatment. 60% (6/10) patients presented with graft dysfunction; 4/6 recovered ejection fraction>40% after treatment. Pathologic AMR was resolved in 5/7 (71.4%) of patients within 1year after treatment. 9/10 (90%) patients survived to 1year after AMR diagnosis. Using PI in AMR resulted in significant DSA reduction with some resolution of graft dysfunction. Larger studies are needed to evaluate PI for AMR.

Research Highlights.

Research Highlights.

Photopheresis in Antibody Mediated Rejection: A Single Center Experience

<h3>Purpose</h3> Antibody mediated rejection (AMR) remains a major source of morbidity and mortality for heart transplant (HT) recipients with diverse treatment strategies across transplant centers. The role of extracorporeal photopheresis (ECP), a cellular immunotherapy, as a potential adjunctive therapy for AMR in HT remains controversial. <h3>Methods</h3> A retrospective analysis was performed of all HT patients at our institution who received at least 3 months ECP for AMR between 09/2011-09/2020. Clinical response to ECP and outcomes within 1 year of therapy are described. Endomyocardial biopsy (EMB) grading of AMR was defined per 2013 ISHLT nomenclature. Donor specific antibodies (DSA) were considered positive if identified de-novo, at mean fluorescence intensity (MFI) greater than 4000. <h3>Results</h3> Results are reported in Table 1. The analysis included 19 HT recipients treated with ECP for AMR at a median of 723 days (IQR 112.5, 1749) post HT. At diagnosis, 7/19 (36.8%) had pAMR1I+, 11/19 (57.9%) had decline in left ventricular ejection fraction (LVEF) by at least 10%, and 13/19 (68.4%) had presence of DSA. Following initial immunosuppressive therapy for AMR, 6/11 (54.5%) had at least a 10% improvement in LVEF, 4/7 (57.1 %) had resolution of AMR on EMB, and 6/13 (46.1%) had reduction in DSA below threshold MFI. In patients without improvement in LVEF, EMB, or DSA following initial treatment; 3 patients (75%) had improvement in LVEF by at least 10% and 2 patients (33.3%) had reduction in DSA after 6 months of ECP. All 3 patients (100%) with persistent pAMR1I+ had resolution of biopsy-confirmed AMR after 3 months of ECP. In patients with clinical improvement following initial therapy, 3 (60%) patients maintained or improved LVEF and 4 (80%) maintained reduction in DSA after 6 months of ECP. <h3>Conclusion</h3> In this retrospective cohort, ECP was well tolerated with improvement and maintenance in LVEF, decrease in DSA positivity, and resolution of AMR on EMB. This modality may be a reasonable adjunctive therapy following initial treatment for AMR after HT.

Significant Reduction of Donor Specific Antibodies in Heart Transplant Recipients Treated with Proteasome Inhibitors for Antibody Mediated Rejection

<h3>Purpose</h3> Antibody-mediated rejection (AMR) results in deleterious outcomes, including coronary artery vasculopathy (CAV), graft failure, and death. Proteasome inhibitors (PI) were used in pre-transplant desensitization to reduce antibodies to Human Leucocytes Antigen (HLA). We report our experience treating heart transplant recipients with AMR using PI, plasmapheresis (PLEX), and intravenous immunoglobulin (IVIG). <h3>Methods</h3> We retrospectively analyzed all patients treated with PI, either bortezomib (BTZ) or carfilzomib (CFZ), for AMR at our institution from 2018-present. CFZ (20mg/m2) or BTZ (1.3mg/m2) was given in combination with PLEX on days 1,2, 8, 9, 15 and 16, followed with IVIG 2g/kg. Mean fluorescence intensity (MFI) for donor-specific antibody (DSA) was evaluated by Luminex single antigen beads (One Lambda). Graft dysfunction (GD) was defined by left ventricular ejection fraction (LVEF) < 40%. <h3>Results</h3> Eleven patients were treated with PI for AMR (10 with CFZ, 1 with BTZ). Patients received 1-3 cycles of PI (1 cycle n=4; 2 cycles n=4; 3 cycles n=3), based upon the response in DSA. Before treatment, all patients except one had one or more strong DSAs (MFI> 8000) at neat serum. Most of the 22 strong DSAs observed were specific for HLA class II (DQ, n=14, 64%; DR, n=5, 23%). The treatment reduced MFI by 47+/- 46% at neat serum and by 70+/- 31% at 1:16 dilution (Figure 1). Four DSAs from 3 patients did not respond to treatment at 1:16 dilutions: one patient had high-titer DSA before treatment (MFI=14027 at 1:1024 dilution), another had rebound DSA from pre-transplantation. 5/11 patients presented with GD at onset (median EF reduction by 67%); 3/5 recovered LVEF to >40% after treatment. 9/11 (81.8%) patients survived to 1 year after AMR diagnosis. <h3>Conclusion</h3> Using PI in AMR treatment resulted in significant DSA reduction in 8/11 patients. Large studies are needed to further evaluate PI in AMR treatment.

Efficacy of Plasmapheresis and Intravenous Immunoglobulin on the Reduction of Donor Specific Antibodies in Heart Transplant Recipients

<h3>Purpose</h3> Class II human leukocyte antigen (HLA) donor specific antibodies (DSA) have been associated with a high risk of graft loss and resistance to antibody removal therapy. We aimed to evaluate class II DSA reduction after plasmapheresis (PLEX) and intravenous immunoglobulin (IVIG) in heart transplant recipients (HTR). <h3>Methods</h3> All HTR who received IVIG and PLEX for class II DSAs from 1/1/2017-6/1/2021 were included. Multiorgan, retransplant, and transplant at other centers were excluded. Baseline demographics and mean immunodominant fluorescent intensity (MFI) of DSA were retrieved. The immunodominant DSA was defined as the DSA with highest MFI in any given serum sample. Median MFI reduction rates after PLEX and after IVIG +/- rituximab were calculated. <h3>Results</h3> Thirty encounters involving 21 HTR were identified. At baseline, the immunodominant class II DSA MFI was 9705 (1712,14337.3). Sixteen (76.2%) and five (23.8%) patients had DQ and DR immunodominant DSA, respectively. There was a median reduction of 5325.5 (1539.5,10003.3) in class II DSA MFI following treatment with PLEX from baseline and a subsequent increase of 1235.5 (0,5786) from post PLEX to post IVIG. Overall, there was a reduction from baseline to post PLEX and IVIG of 1864 (1034,3996.8). Four patients (19%) achieved DSA clearance after PLEX, with a median time to clearance of 34 days (26-60) from development. The patients who cleared were all non-African American, had DSAs to only one antigen, were not desensitized pre-transplant, did not have C1q positivity, and had immunodominant MFI <2000. Significantly more patients who failed to clear DSA were taking prednisone (0% vs 76.5%, p=0.005). <h3>Conclusion</h3> PLEX and IVIG can reduce the MFI of class II DSAs in some HTR. PLEX therapy is more efficacious to reduce class II DSA MFI than originally hypothesized. Results from this analysis suggest that DSA to DQ antigens may be less responsive to therapy with PLEX/IVIG at MFI >15,000 or when C1q positive.

Selective Loss of Donor Specific HLA Antibodies in a Highly Sensitized Patient After Lung Transplantation

<h3>Introduction</h3> Lung transplantation in the presence of donor specific antibodies (DSA) is safe with peri-operative desensitization, although persistence of DSA is common. Herein, we present a unique case of a highly sensitized lung transplant recipient with selective loss of DSA post-transplantation. <h3>Case Report</h3> A 65-year-old female with a calculated PRA (cPRA) value of of 17% (class I) and 100% (class II), underwent bilateral lung transplantation for chronic obstructive pulmonary disease. Virtual crossmatch (VCM) was positive, involving DPB1*04:01, DR17, and DR52. The subsequent flow cytometry B-cell crossmatch was strongly positive (median channel shift 296). She received peri-operative desensitization protocol for VCM positive patients, which includes intra-operative and post-operative plasmapheresis, intravenous immune globulin, and anti-thymocyte globulin. Maintenance immunosuppression included cyclosporine (target trough 250-350 umol/L), mycophenolic acid 900mg BID and prednisone 20 mg OD. An increase in DSA reactivity was observed one month after transplant (DPB1*04:01 (MFI=17639), DR17 (MFI=17812), and DR52 (MFI=19414). Repeat HLA testing at 3, 4, and 6 months post-transplant showed sequential reduction of DSAs to below the laboratory cut-off for positivity point despite persistence of non-DSA alloantibodies. The patient continues to maintain good lung function with an FEV1 of 2L (86% of predicted) at 6 months post-transplant. Surveillance transbronchial biopsies at 3 and 6 months were free of cellular rejection. <h3>Summary</h3> Despite initial rebound of pre-formed DSA following peri-operative desensitization, sustained selective clearance of the DSA subsequently occurred in this case, which likely benefited graft function. Similar cases have been reported after combined kidney-liver transplant. Robust laboratory research and prospective studies are required to understand the mechanism of selective DSA elimination in organ transplantation.

Tocilizumab in a patient affected by chronic active antibody mediated rejection; histological improvement, reduction of proteinuria and renal function stabilization

Introduction: Chronic active antibody-mediated rejection (cAMR) is a significant and rapid destructive form of allograft rejection and it is related to donor specific antibodies (DSA). Interleukin 6 (IL-6) plays an important role in mediating the allograft rejection by promoting CD4+ T cells differentiation to Th17 phenotype while inhibiting Treg. Tocilizumab is a humanized monoclonal antibody directed to IL-6 receptor (IL6-R). The aim of the study is to demonstrate the efficacy of tocilizumab as rescue therapy for cAMR. Case Presentation: A 50-year-old man with Alport syndrome and with positive DSA against B7 e B55 underwent a second kidney transplant (HLA 2 mismatch). He received thymoglobulin and three plasma exchanges as induction therapy. Proteinuria (1-1.3 g/24 h) and decline in kidney function (serum creatinine; 1.5 mg/dL) appeared at 9 months. Kidney biopsy showed endocapillary proliferation, mononuclear cells infiltration, glomerular basal membrane duplication and tubulitis suggestive of cAMR. The patient has been treated with tocilizumab (6 mg/kg/mon) for six months. Reduction of proteinuria (0.6 g/24 h) and mild improvement of kidney function (serum creatinine; 1.3 mg/dL) were observed after tocilizumab treatment. A second biopsy revealed a significant decrease of glomerulitis and peritubular capillaritis. A significant reduction in DSA was detected. Conclusion: Inhibition of the IL-6 receptor by tocilizumab may represent a novel and cheering approach to treat cAMR.

Antibody-Mediated Rejection Management Following Lung Transplantation.

Although antibody-mediated rejection (AMR) is described in other solid organ transplant populations, the literature describing the management following lung transplantation is limited. The purpose of this study is to evaluate the management strategies of AMR in lung transplant recipients. This single-center, retrospective study described the management of AMR in adult lung transplant recipients who received treatment with rabbit antithymocyte globulin, bortezomib, rituximab, intravenous immune globulin (IVIG), and/or plasmapheresis between September 2015 and June 2019. A total of 270 medication orders for 55 patient admissions were included in the primary outcome analysis. The most commonly used regimen consisted of IVIG, plasmapheresis, and rituximab (49.1%; n = 27), followed by IVIG and plasmapheresis alone (27.3%, n = 15). A total of 51 patients (93%) received plasmapheresis as part of their AMR treatment, with a median of 4 [3, 5] sessions per encounter; 86% of patients with positive donor-specific antibodies (DSAs) had a reduction in DSAs following AMR treatment. Overall, 23.5% of patients had noted allograft failure or need for retransplantation. A total of 10 patients died during the AMR treatment hospital admission, and an additional 11 patients died within 1 year of the initial encounter. This represents the largest report describing management strategies of AMR in lung transplant recipients. Although practice varied, the most commonly used regimen consisted of plasmapheresis, IVIG, and rituximab.

DNA-PKcs Inhibition Extends Allogeneic Skin Graft Survival.

Organ transplantation is life-saving and continued investigations into immunologic mechanisms that drive organ rejection are needed to improve immunosuppression therapies and prevent graft failure. DNA-dependent protein kinase catalytic subunit, DNA dependent-protein kinase catalytic subunit (DNA-PKcs), is a critical component of both the cellular and humoral immune responses. In this study, we investigate the contribution of DNA-PKcs to allogeneic skin graft rejection to potentially highlight a novel strategy for inhibiting transplant rejection. Fully MHC mismatched murine allogeneic skin graft studies were performed by transplanting skin from BalbC mice to C57bl6 mice and treating with either vehicle or the DNA-PKcs inhibitor NU7441. Graft rejection, cytokine production, immune cell infiltration, and donor-specific antibody formation were analyzed. DNA-PKcs inhibition significantly reduced necrosis and extended graft survival compared with controls (mean survival 14 d versus 9 d, respectively). Inhibition reduced the production of the cytokines interleukin (IL)-2, IL-4, IL-6, IL-10, TNF-α, and IFN-γ and the infiltration of CD3+ lymphocytes into grafts. Furthermore, DNA-PKcs inhibition reduced the number of CD19+ B cells and CD19+ CD138+ plasma cells coinciding with a significant reduction in donor-specific antibodies. At a molecular level, we determined that the immunosuppressive effects of DNA-PKcs inhibition were mediated, in part, via inhibition of nuclear factor kappa-light-chain-enhancer of activated B cells signaling through reduced expression of the p65 subunit. Our data confirm that DNA-PKcs contributes to allogeneic graft rejection and highlight a novel immunologic function for DNA-PKcs in the regulation of nuclear factor kappa-light-chain-enhancer of activated B cells and concomitant cytokine production.

Early effects of first-line treatment with anti-interleukin-6 receptor antibody tocilizumab for chronic active antibody-mediated rejection in kidney transplantation.

Chronic active antibody-mediated rejection (cAMR) is a major determinant of late allograft failure. Rituximab/immunoglobulins (IVIg) + plasma exchange (PLEX) showed controversial results in cAMR treatment. Tocilizumab (TCZ), a humanized anti-interleukin 6 receptor antibody, has been recently used as rescue therapy in patients non-responsive to rituximab/IVIg/PLEX with favorable outcomes. Whether TCZ acts "per se" or requires a priming effect from previous treatments is currently unknown. Fifteen patients with cAMR were treated with TCZ as a first-line therapy and followed for a median time of 20.7months. Despite the majority of patients experiencing advanced transplant glomerulopathy (TG) at diagnosis (60% with cg3), glomerular filtration rate and proteinuria stabilized during the follow-up, with a significant reduction in donor-specific antibodies. Protocol biopsies after 6months demonstrated significant amelioration of microvascular inflammation and no TG, C4d deposition, or IF/TA progression. Gene-expression and immunofluorescence analysis showed upregulation of three genes (TJP-1, AKR1C3, and CASK) involved in podocyte, mesangial, and tubular restoration. Tocilizumab adopted as a first-line approach in cAMR was associated with early serological and histological improvements and functional stabilization even in advanced TG, suggesting a role for the use of TCZ alone with the avoidance of unnecessary previous immunosuppressants.

Reducing Donor-specific Antibody During Acute Rejection Diminishes Long-term Renal Allograft Loss: Comparison of Early and Late Rejection

Reduction in donor-specific antibody (DSA) has been associated with improved renal allograft survival after antibody-mediated rejection (AMR). These observations have not been separately analyzed for early and late AMR and mixed acute rejection (MAR). The purpose of this study was to evaluate long-term responses to proteasome inhibitor-based therapy for 4 rejection phenotypes and to determine factors that predict allograft survival. Retrospective cohort study evaluating renal transplant recipients with first AMR episodes treated with proteasome inhibitor-based therapy from January 2005 to July 2015. A total of 108 patients were included in the analysis. Immunodominant DSA reduction at 14 days differed significantly (early AMR 79.6%, early MAR 54.7%, late AMR 23.4%, late MAR 21.1%, P < 0.001). Death-censored graft survival (DCGS) differed at 3 years postrejection (early AMR 88.3% versus early MAR 77.8% versus late AMR 56.7% versus late MAR 54.9%, P = 0.02). Multivariate analysis revealed that immunodominant DSA reduction > 50% at 14 days was associated with improved DCGS (odds ratio, 0.12, 95% CI, 0.02-0.52, P = 0.01). In summary, significant differences exist across rejection phenotypes with respect to histological and DSA responses. The data suggest that DSA reduction may be associated with improved DCGS in both early and late AMR.

Daratumumab in Sensitized Kidney Transplantation: Potentials and Limitations of Experimental and Clinical Use.

Donor-specific antibodies are associated with increased risk of antibody-mediated rejection and decreased allograft survival. Therefore, reducing the risk of these antibodies remains a clinical need in transplantation. Plasma cells are a logical target of therapy given their critical role in antibody production. To target plasma cells, we treated sensitized rhesus macaques with daratumumab (anti-CD38 mAb). Before transplant, we sensitized eight macaques with two sequential skin grafts from MHC-mismatched donors; four of them were also desensitized with daratumumab and plerixafor (anti-CXCR4). We also treated two patients with daratumumab in the context of transplant. The animals treated with daratumumab had significantly reduced donor-specific antibody levels compared with untreated controls (57.9% versus 13% reduction; P<0.05) and prolonged renal graft survival (28.0 days versus 5.2 days; P<0.01). However, the reduction in donor-specific antibodies was not maintained because all recipients demonstrated rapid rebound of antibodies, with profound T cell-mediated rejection. In the two clinical patients, a combined heart and kidney transplant recipient with refractory antibody-mediated rejection and a highly sensitized heart transplant candidate, we also observed a significant decrease in class 1 and 2 donor-specific antibodies that led to clinical improvement of antibody-mediated rejection and to heart graft access. Targeting CD38 with daratumumab significantly reduced anti-HLA antibodies and anti-HLA donor-specific antibodies in a nonhuman primate model and in two transplant clinical cases before and after transplant. This supports investigation of daratumumab as a potential therapeutic strategy; however, further research is needed regarding its use for both antibody-mediated rejection and desensitization.

Therapeutic Plasma Exchange in Pediatric Renal Transplantation Experience of One Decade and 389 Sessions

ObjectiveThere are no specific recommendations for therapeutic plasma exchange (TPE) in children after renal transplantation. The purpose of this study was to report the experience with TPE in a pediatric transplant setting. Materials and Methods59 patients (mean age 12.5 ± 4.5 years) undergoing renal transplantation. Indications for TPE included the recurrence of nephrotic syndrome (NS; n = 30) and atypical hemolytic uremic syndrome (n = 6), chronic antibody-mediated rejection (cAMR; n = 20), sensitization (n = 2), and immune thrombocytopenia (n = 1). The single-filtration TPE was performed in all cases. In 74.7% of patients, fresh frozen plasma was used as a replacement fluid. In 25.3% of patients, 4% albumin solution was used as a replacement fluid. Criteria for TPE efficacy included a decrease of proteinuria and normalization of renal function in NS; a normalization of platelet count, C3, and hemoglobin concentration in aHUS; improvement in renal function; and reduction of donor-specific antibodies in cAMR; and removal of antiplatelet antibodies in immune thrombocytopenia. ResultsEfficacy results for patients with NS: 59.3% achieved remission, 25.9% achieved partial remission, and 14.8% achieved no remission, respectively. For patients with atypical hemolytic uremic syndrome there was remission in 66.6% and no remission in 33.4%. For patients with cAMR there was remission in 75% and no remission in 25%. Antiplatelet antibodies disappeared after TPE in 1 patient. In 9% of TPE procedures, minor complications were noted. All patients were on posttransplant maintenance immunosuppression and several children received additional treatment (intravenous immunoglobulin therapy or rituximab) during TPE therapy. ConclusionTPE therapy (combined with immunosuppression) was an effective tool in most pediatric cases after renal transplantation with low incidence of minor adverse events.

Early and Sustained Reduction in Donor-Specific Antibodies in Desensitized Living Donor Kidney Transplant Recipients: A 3-Year Prospective Study.

BackgroundDesensitization with IVIG and rituximab allows acceptable graft survival in sensitized kidney transplant recipients with preexisting donor-specific antibodies (DSAs) and a positive crossmatch. There is little published data reporting the durability of DSA removal in kidney transplant recipients treated with IVIG and rituximab.MethodsWe conducted a 3-year prospective DSA monitoring study in living donor kidney recipients with preexisting DSA to assess the durability of DSA removal after a perioperative protocol of IVIG and rituximab. All recipients had flow crossmatch titers less than 1:32. Data were analyzed using linear mixed effects models and Kaplan-Meier survival methods.ResultsThe longitudinal database comprised 210 mean fluorescence intensity (MFI) determinations. Forty-two DSAs were identified in 29 patients. Pretreatment MFI averaged 4715 ± 3962 (range, 947-20 129). At 1 month posttransplant, 18 patients (62%) had a complete response (MFI < 1000) and an additional 9 patients (31%) had a partial response (MFI reduced but >1000). There was a 46% reduction (P < 0.001) in DSA MFI at 1 month posttransplant that was sustained throughout the 3-year follow-up period and was observed for both class I and II DSAs regardless of pretreatment MFI levels. With a mean posttransplant follow-up of 1048 ± 574 days, 3-year patient and graft survivals were 95% and 90%. Four patients (14%) had acute rejection between days 125 and 560.ConclusionsDesensitization with IVIG and rituximab results in early and sustained DSA removal over a 3-year posttransplant period in living donor kidney transplant recipients with pretransplant DSA and a positive crossmatch, excellent patient and graft survivals and a low incidence of acute rejection.

A closer look at rituximab induction on HLA antibody rebound following HLA-incompatible kidney transplantation

Rituximab has been used to increase the efficacy of desensitization protocols for HLA incompatible kidney transplantation, however, controlled comparisons have not been reported. Here we examined 256 post-transplant HLA antibody levels in 25 recipients desensitized with or 25 without rituximab induction, to determine the impact of B cell depletion. We found significantly less HLA antibody rebound in the rituximab-treated patients (7% of donor specific antibodies (DSAs) and 33% of non-DSAs) compared to a control cohort desensitized and transplanted without rituximab (32% DSAs and 55% non-DSAs). The magnitude of the increase was significantly larger among patients who did not receive rituximab. Interestingly, in rituximab treated patients, of the 39 HLA antibodies that increased post-transplant, 34 were specific for HLA mismatches present in previous allografts or pregnancies, implying limited efficacy in memory B cell depletion. Compared to controls, rituximab-treated patients had a significantly greater mean reduction in DSA (−2505 versus −292 mean fluorescence intensity), but a similar rate of DSA persistence (52% in rituximab treated and 40% in non-treated recipients). Thus, rituximab induction in HLA incompatible recipients reduced the incidence and magnitude of HLA antibody rebound, but did not impact DSA elimination, antibody mediated rejection, or 5 year allograft survival when compared to recipients desensitized and transplanted without rituximab.

Prospective Post-Transplant DSA Monitoring in Kidney Transplant Recipients With Pre-Transplant DSA and a Positive Crossmatch Treated With IVIG/Rituximab.

Purpose: Desensitization with IVIG/Rituximab allows acceptable graft survival in sensitized kidney transplant recipients (KTR) with pre-transplant (pre-tx) donor-specific antibody (DSA) and a positive cross-match (XM+). We report initial data from a 3-yr prospective study of DSA monitoring in DSA+/XM+ living donor (LD) KTR to assess the efficacy of antibody removal with IVIG/Ritux and possible associations between pre-tx DSA-MFI and C1q with post-transplant outcomes. Methods: Prospective DSA monitoring with Single Antigen Luminex assay was performed in DSA+/XM+ (flow titer <1:32) LD KTR. All KTR received IVIG (2 gms/kg on days -7 and +7), rituximab (375 mg/m2 day +1), alemtuzumab (30 mg day 0), and maintenance tac/MMF/Pred. C1q binding assay was performed on pre-tx sera. Data were analyzed using paired t-tests and linear regression. Results: From 11/09-10/13, 23 DSA+/XM+ LD KTR were enrolled. Patient survival is 100% and graft survival is 96%. 3 pts (13%) had acute rejection (2/3 C4d+) at 4, 12, and 19 mos. A total of 42 DSAs were identified in 23 KTR (range 1-5 DSA/pt). Mean±SD pre-IVIG MFI was 4191+4166. 8/23 pts had positive C1q assay (>500) against donor HLA in pre-IVIG serum. We found a significant reduction from pre-IVIG MFI to post-IVIG/pre-tx MFI (p=0.030) and to 1 mo post-tx MFI (p=0.005). The magnitude of IVIG MFI pre-IVIG appears related to decrease in MFI both post-IVIG/pre-tx (beta=-0.627, p<0.001) and at 1 month post tx (Figure). Durability of DSA reduction over 36 months awaits further follow-up. Due to low incidence of AR, correlation between either pre-IVIG MFI or C1q binding or magnitude of reduction post IVIG/Ritux and subsequent AR or graft loss await longer follow-up.Figure: No Caption available.Conclusion: Desensitization with IVIG/Ritux results in excellent short-term patient and graft survival with low AR in LD KTR with DSA/+XM. There is a significant reduction of DSA 1 month post-tx and the magnitude of early MFI reduction is related to pre-treatment level. Longer follow-up is needed to determine durability of DSA removal and impact on long-term AR or graft survival. DISCLOSURE:Shaffer, D.: Grant/Research Support, Novartis, Astellas.

Rapid Reduction in Donor-Specific Anti-Human Leukocyte Antigen Antibodies and Reversal of Antibody-Mediated Rejection With Bortezomib in Pediatric Heart Transplant Patients

High titer donor-specific antibodies (DSA) and positive crossmatch in cardiac transplant recipients is associated with increased mortality from antibody-mediated rejection (AMR). Although treatment to reduce anti-human leukocyte antigen antibodies using plasmapheresis, intravenous immunoglobulin, and rituximab has been reported to be beneficial, in practice these are often ineffective. Moreover, these interventions do not affect the mature antibody producing plasma cell. Bortezomib, a proteasome inhibitor active against plasma cells, has been shown to reduce DSA in renal transplant patients with AMR. We report here the first use of bortezomib for cardiac transplant recipients in four pediatric heart recipients with biopsy-proven AMR, hemodynamic compromise, positive crossmatch, and high titer class I DSA. Patients received four intravenous dose of bortezomib (1.3 mg/m(2)) over 2 weeks with plasmapheresis and rituximab. DSA specificity and strength (mean fluorescence intensity) was determined with Luminex. All had received previous treatment with plasmapheresis, intravenous immunoglobulin, and rituximab that was ineffective. AMR resolved in all patients treated with bortezomib with improvement in systolic function, conversion of biopsy to C4d negative in three patients and IgG negative in one patient, and a prompt, precipitous reduction in DSAs. In three patients who received plasmapheresis before bortezomib, plasmapheresis failed to reduce DSA. In one case, DSA increased after bortezomib but decreased after retreatment. Bortezomib reduces DSA and may be an important adjunct to treatment of AMR in cardiac transplant recipients. Bortezomib may also be useful in desensitization protocols and in prevention of AMR in sensitized patients with positive crossmatch and elevated DSA.

Lung Transplantation Across Donor-Specific Anti-Human Leukocyte Antigen Antibodies: Utility of Bortezomib Therapy in Early Graft Dysfunction

To report the usefulness of bortezomib therapy in a sensitized lung transplant recipient experiencing antibody-mediated rejection. During a pretransplant evaluation, a 62-year-old woman with usual interstitial pneumonitis developed a diverticular bleed requiring transfusions, which elevated her panel reactive antibody to 98% for human leukocyte antigen (HLA) class I and 71% for class II. She underwent desensitization to decrease her panel reactive antibody levels. She received a double lung transplant across a weak HLA class II incompatibility but developed respiratory failure due to early graft dysfunction. On postoperative day (POD) 14 she was found to have donor-specific antibodies (DSA) to HLA class I and class II antigens. She received intravenous immunoglobulin (IVIG), plasmapheresis, and bortezomib to reduce the DSA. Repeat DSA testing on POD 80 demonstrated a 50% reduction in DSA, which became undetectable at POD 255. Antibody-mediated rejection (AMR) is difficult to diagnose and treat in lung transplantation. Since primary treatment options such as plasmapheresis and IVIG alone may not adequately eradicate DSA, the proteasome inhibitor bortezomib can be of additional value for the treatment of AMR. Bortezomib causes apoptosis of plasma cells, thus eliminating the production of allograft-specific DSA. This is the first report describing the utility of bortezomib for early graft dysfunction in a highly sensitized lung transplant recipient. Although this patient had preformed donor-specific anti-HLA antibodies, AMR was successfully treated with a combination of plasmapheresis, IVIG, and bortezomib. At time of writing, the patient continued to have excellent graft function 2 years posttransplant. Bortezomib is a potent inhibitor of plasma cell production and it appears to be useful for the treatment of antibody-mediated graft dysfunction.

255: Rapid Reduction in Donor Specific Antibodies with Bortezomib in Pediatric Heart Transplant Recipients with Positive Crossmatch and Antibody Mediated Rejection

255: Rapid Reduction in Donor Specific Antibodies with Bortezomib in Pediatric Heart Transplant Recipients with Positive Crossmatch and Antibody Mediated Rejection

Intravenous immunoglobulin preparations have no direct effect on B cell proliferation and immunoglobulin production

Intravenous immunoglobulin (IVIg) is used for treatment of a variety of immunological disorders and in transplantation. As one of its applications in transplantation is the reduction of donor specific antibodies in the circulation, we examined the direct effect of IVIg on essential parameters of human B cell responses in vitro. Purified human B cells, human B cell hybridomas and T cells were cultured in the presence of graded concentrations of IVIg to test its effect on their proliferative capacity. To address the effect of IVIg on immunoglobulin production, we designed a novel technique making use of quantitative polymerase chain reaction to assess IgM and IgG levels. IVIg failed to inhibit proliferation of human B cells and human B cell hybridomas. In contrast, when IVIg was added to T cell cultures, a dose-dependent reduction of the proliferative capacity was observed. IVIg did not affect the levels of IgM and IgG mRNA of activated B cells. Our data show that IVIg is not capable of directly inhibiting key B cell responses. Direct B cell inhibition by IVIg seems therefore unlikely, implying that alteration in humoral immunity by IVIg is due to indirect effects on T cells and/or interactions with circulating antibodies and complement factors.