Pathologic Antibody-mediated Rejection 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.

Validation of donor fraction cell-free DNA with biopsy-proven cardiac allograft rejection in children and adults

ObjectivesDonor-specific cell-free DNA shows promise as a noninvasive marker for allograft rejection, but as yet has not been validated in both adult and pediatric recipients. The study objective was to validate donor fraction cell-free DNA as a noninvasive test to assess for risk of acute cellular rejection and antibody-mediated rejection after heart transplantation in pediatric and adult recipients. MethodsPediatric and adult heart transplant recipients were enrolled from 7 participating sites and followed for 12 months or more with plasma samples collected immediately before all endomyocardial biopsies. Donor fraction cell-free DNA was extracted, and quantitative genotyping was performed. Blinded donor fraction cell-free DNA and clinical data were analyzed and compared with a previously determined threshold of 0.14%. Sensitivity, specificity, negative predictive value, positive predictive value, and receiver operating characteristic curves were calculated. ResultsA total of 987 samples from 144 subjects were collected. After applying predefined clinical and technical exclusions, 745 samples from 130 subjects produced 54 rejection samples associated with the composite outcome of acute cellular rejection grade 2R or greater and pathologic antibody-mediated rejection 2 or greater and 323 healthy samples. For all participants, donor fraction cell-free DNA at a threshold of 0.14% had a sensitivity of 67%, a specificity of 79%, a positive predictive value of 34%, and a negative predictive value of 94% with an area under the curve of 0.78 for detecting rejection. When analyzed independently, these results held true for both pediatric and adult cohorts at the same threshold of 0.14% (negative predictive value 92% and 95%, respectively). ConclusionsDonor fraction cell-free DNA at a threshold of 0.14% can be used to assess for risk of rejection after heart transplantation in both pediatric and adult patients with excellent negative predictive value.

Antibody Mediated Rejection in Pediatric Heart Transplant Recipients

<h3>Purpose</h3> Pediatric heart transplant recipients who experience antibody mediated rejection (AMR) are at increased risk for poor graft outcomes. We report the incidence, treatment, and outcomes of AMR at our center. <h3>Methods</h3> Transplant recipients treated for AMR between 2010 and 2021 were identified. We reviewed AMR episodes, therapies, and graft outcomes. <h3>Results</h3> Of 183 transplants performed, 25 patients experienced 37 episodes of AMR, representing an incidence of 14%. 89% of patients were transplanted for congenital heart disease, 40% were sensitized pre-transplant, and 28% had a crossmatch positive transplant. In 33 of 37 episodes (89%), there was both strong DSA (>4,000 mfi) and findings of pathologic AMR (pAMR) on endomyocardial biopsy. Two patients had DSA <4,000 mfi with pAMR findings, one had strong DSA but died prior to biopsy, and one had strong DSA with negative pAMR but with graft dysfunction without other explanation. The first rejection episode occurred at a median of 50 days (IQR 17-635 days) from transplant. 19% of episodes were with hemodynamic compromise and 36% of patients had recurrence of AMR. Biopsy findings showed pAMR 0 in 3%, pAMR 1 (H+) in 47%, pAMR 1 (I+) in 17%, and pAMR 2 in 33%. 75% had mixed ACR/pAMR. Therapy included anti-thymocyte globulin in 13% (predominately for rejection with hemodynamic compromise), methylprednisolone in 92%, IVIG in 97%, bortezomib in 78%, rituximab in 73%, plasmapheresis in 35%, and eculizumab in 22%. A combination of 2 or greater antibody directed therapies was used in 89% of episodes and 3 or greater therapies in 70%. IVIG/bortezomib/rituximab was the most common therapy combination, used in 32% of episodes. pAMR findings resolved in 64% at 3 months and 79% at 1 year. During a median follow up of 4 years (IQR 1-6 years) from first AMR, there were 8 graft losses (2 retransplants and 6 deaths). The median time from AMR to graft loss was 7 months (IQR 2 months - 4.5 years). 20% of patients developed cardiac allograft vasculopathy at a median duration of 4.8 years (IQR 4-7) from AMR and 8 years (IQR 5-9) from transplant. Retransplant free survival from first AMR was 83% at 1 year, 76% at 3 years, and 63% at 5 years. <h3>Conclusion</h3> AMR occurred early post-transplant and recurrence was common. Treatment strategies varied, however combination therapy was nearly always employed. Despite relatively early graft loss after first AMR, AMR can be treated to prolong graft survival.

AMR After Transplant in a Child with Complex CHD and Ruptured LV Aneurysm

<h3>Introduction</h3> Antibody mediated rejection (AMR) is a mechanism of allograft dysfunction and a risk factor for transplant outcomes. Left ventricular (LV) aneurysms are rare in children and may be congenital or acquired. This is a unique case of both AMR and acquired LV aneurysm. <h3>Case Report</h3> We present a 1.9 kg Caucasian female born at 35 6/7 weeks gestation with a prenatal diagnosis of D-transposition of the great arteries and a large ventricular septal defect encompassing most of the trabecular ventricular septum. After pulmonary artery banding and balloon atrial septostomy, a Damus-Kaye-Stansel amalgamation (DKS) was performed with de-banding of the pulmonary artery and a hemi-Fontan procedure. After the DKS, she developed ventricular fibrillatory arrest requiring extracorporeal membrane oxygenation (ECMO) for 12 days. During this time her DKS was taken down. She was found to have an intramural course of her right coronary artery causing ischemia and arrhythmia. She developed biventricular dysfunction. After discharge the patient was lost to follow up for 4 years. Upon her return, a computed tomography scan showed interval development of a left ventricular aneurysm with rupture into the right ventricle [Fig 1]. Heart transplantation was pursued. Despite a weakly positive cross match for B flow, she received a heart transplant. One week post-transplant pathological AMR class two and donor specific antibody (DSA) production were identified. Her immunosuppression regimen was increased and she received plasmapheresis, but on post-operative day 16 she suffered a cardiac arrest and required ECMO. Additional steroids, anti-thymocyte globulin, Rituximab and Bortezomib were administered with recovery of function and decreased DSAs. She was decannulated after 8 days and subsequent biopsies were free of rejection. She was discharged home neurologically intact. <h3>Summary</h3> This case illustrates accurate diagnosis and treatment of AMR in a child with complex congenital heart disease and ruptured LV aneurysm status post heart transplant.

Correlation Between Microvascular Inflammation in Endomyocardial Biopsies and Rejection Transcripts, Donor-specific Antibodies, and Graft Dysfunction in Antibody-mediated Rejection.

The pathology-based diagnosis of cardiac antibody-mediated rejection (AMR) relies on the 2013 International Society for Heart and Lung Transplantation Working Formulation, in which microvascular inflammation (MVI) is considered as present or absent regardless of its extent. This work assessed the biological and clinical value of a semiquantitative evaluation of the extent of MVI in endomyocardial biopsies (EMBs). We retrospectively graded the extent of MVI in 291 EMB from 291 patients according to a 4-point scale in which MVI scores of 0, 1, 2, and 3 represented 0%, 1%-10%, 11%-50%, and >50% of the myocardial area, respectively. We analyzed the association between the MVI score and tissue rejection molecular activity assessed by microarrays or reverse transcriptase multiplex ligation-dependent probe amplification, current pathology classification (pathologic AMR [pAMR]), anti-HLA donor-specific antibodies, and graft dysfunction. Overall, 172 (59.1%), 33 (11.4%), 42 (14.4%), and 44 (15.1%) EMB were given MVI scores of 0, 1, 2, and 3, respectively. pAMR1(H+) and pAMR2/3 categories were found to be heterogeneous in terms of MVI score. Acute cellular rejection grades did not influence the MVI score. In both molecular approaches, we observed a stepwise increase in the expression of AMR-related transcripts with increasing MVI scores, independent of the C4d or CD68 status (P < 0.001). Both the frequency and mean fluorescence intensity of donor-specific antibodies gradually increased with the MVI score (P < 0.001). Acute graft dysfunction was more frequent in MVI score 3 (P < 0.001). The intensity of MVI in EMB, based on a semiquantitative evaluation of its extent, has biological and clinical importance.

Management of Antibody Mediated Rejection (AMR) in Heart Transplant Patients: The Real World in a French Single Center Treating All Pathological AMR

<h3>Purpose</h3> There is no consensus for the treatment of antibody mediated rejection (AMR) in heart transplant patients (HTP), only expert recommendation with a grey zone in patients without graft dysfunction. The purpose of the study was to report our AMR management since we perform routine screening of donor-specific antibodies (DSA) with EMB in case of positive DSA and treat AMR irrespective of graft dysfunction, severity of AMR or presence of DSA. <h3>Methods</h3> We retrospectively reviewed HTP between 2011 and 2019 and studied patients with AMR defined only on endomyocardial biopsy (EMB) according to the latest ISHLT criteria. <h3>Results</h3> A total of 15 patients developed AMR (5 patients with pAMR 1 and 10 with pAMR2) (Figure1), followed for a mean of 4.6 years; 3 patients (20%) had a graft dysfunction (EF<40%) and 3 (20%) did not have circulating DSA. All patients received intravenous steroids, plasmapheresis (5 to7 sessions only for DSA+ patients) IVig for 2 days and rituximab. The 7 more recent patients had additional IVig therapy (high-dose regimen) for 6 weeks and 5 patients received bortezomib. At the end of the treatment, 12 patients (60%) had normal EMB, 6 patients (50%) had persistent DSA with MFI>5000, 1 patient had persistent graft dysfunction and 80% were alive (Figure1). <h3>Conclusion</h3> despite a short follow up, our experience is encouraging to treat aggressively all pathological AMR irrespective of graft dysfunction or presence of DSA to prevent graft dysfunction and poor outcomes.

Sarcolemmal Staining by Anti-C4d/C3d in Cardiac Transplant Endomyocardial Biopsies and Associated Serum HLA Antibodies and Pathologic Rejection

<h3>Purpose</h3> Immunologic evidence of complement fixation in endomyocardial biopsies (EMB) is a feature used to assess pathologic antibody-mediated rejection in cardiac transplants. Uncommonly, sarcolemmal staining (SS) of myocytes is observed. The significance of this is unclear and under-investigated. This study aims to temporally correlate SS with changes in HLA antibodies and pathologic rejection. <h3>Methods</h3> All EMBs that had SS by anti-C4d and/or C3d on immunofluorescence or immunohistochemistry (Figure 1) were identified via the laboratory information system between 10/1/2016 and 10/1/2019. The HLA antibodies and all other EMB results since time of transplant for each case were reviewed and temporally correlated to the biopsy that showed SS. <h3>Results</h3> 28 patients (mean: 31 yrs; range: 6-73 yrs) with 31 SS events were identified. Associated features are summarized in Table 1. Of note, 29/31 events of SS had HLA antibodies measured within a 2 week period before/after the time of biopsy. A temporally associated expansion of HLA Class I/II antibodies was noted in 12 (39%) cases. 21 (68%) cases had prior donor specific antibodies before EMB showing SS, with less immediate temporal association. <h3>Conclusion</h3> Sarcolemmal staining of complement split products in EMBs was associated with a concurrent expansion in strength and/or breadth of HLA antibodies in 39% of cases. This is a novel observation that may partially explain the appearance of SS by immunologic assays. Further investigation is required to fully elucidate the significance of SS in EMB.

Angiotensin II Type 1 Receptor Antibody Mediated Rejection Following Orthotopic Heart Transplant: A Single Center Experience

Antibody mediated rejection (AMR) following heart transplant (OTHx) causes significant morbidity and mortality. In particular, diagnosis and management of non-HLA AMR remains challenging. Antibodies to the angiotensin II type 1 receptor (AT1R) causing rejection after kidney transplant have been described, but there is limited data on AT1R-Ab mediated rejection following OHTx. Here, we present our single center experience. We retrospectively reviewed all OHTx cases from the past 2 years with clinically suspected AMR without evidence of canonical donor-specific AMR who had a positive AT1R-Ab titer. Clinical and pathological features were documented. We identified 7 patients, the majority were female (71%), African American (71%), and had prior LVAD support (71%). All patients had NICM pretransplant. Patients presented both early (∼2 weeks) and late (∼7 years) with heart failure; 3 patients presented within 1 year post OHTx. Echocardiography showed progressive graft dysfunction, primarily decreased EF, ventricular wall thickening, and diastolic dysfunction. The average AT1R-Ab titer was 22U/mL at 1:100 dilution (range: 3 to >40 U/mL). Biopsies showed reactive endothelium and increased interstitial macrophages, without pathologic AMR features (negative C4d and <10% intravascular macrophages by CD68). Five patients were empirically treated with plasmapheresis, IVIG and rituximab. One patient did not tolerate plasmapheresis, and treatment was deferred on one patient given low titers and clinical improvement with standard heart failure treatment. All patients were started on an angiotensin receptor blocker (ARB). There were no deaths, and the majority of patients had clinically significant improvement in symptoms and graft function following treatment with an ARB at an average 6 months of follow-up. AT1R-Ab mediated rejection is an underdiagnosed and emerging entity following OHTx that remains ill-defined; it requires a high index of suspicion and should be considered for all OHTx recipients with evidence of graft dysfunction or decompensation. Empiric AMR treatment followed by oral ARB therapy anecdotally improves patient outcomes; however, future studies are needed to better define the pathophysiology, long-term clinical sequelae, and optimal treatment modalities of this disease.

Sarcolemmal Staining by Anti-C4d/C3d in Cardiac Transplant Endomyocardial Biopsies and Associated Serum HLA Antibodies and Pathologic Rejection

Immunologic evidence of complement fixation in endomyocardial biopsies (EMB) is a feature used to assess pathologic antibody-mediated rejection in cardiac transplants. Uncommonly, sarcolemmal staining (SS) of myocytes is observed. The significance of this is unclear and under-investigated. This study aims to temporally correlate SS with changes in HLA antibodies and pathologic rejection. All EMBs that had SS by anti-C4d and/or C3d on immunofluorescence or immunohistochemistry (Figure 1) were identified via the laboratory information system between 10/1/2016 and 10/1/2019. The HLA antibodies and all other EMB results since time of transplant for each case were reviewed and temporally correlated to the biopsy that showed SS. 28 patients (mean: 31 yrs; range: 6-73 yrs) with 31 SS events were identified. Associated features are summarized in Table 1. Of note, 29/31 events of SS had HLA antibodies measured within a 2 week period before/after the time of biopsy. A temporally associated expansion of HLA Class I/II antibodies was noted in 12 (39%) cases. 21 (68%) cases had prior donor specific antibodies before EMB showing SS, with less immediate temporal association. Sarcolemmal staining of complement split products in EMBs was associated with a concurrent expansion in strength and/or breadth of HLA antibodies in 39% of cases. This is a novel observation that may partially explain the appearance of SS by immunologic assays. Further investigation is required to fully elucidate the significance of SS in EMB.

Upregulation of Endothelial HLA Class II is a Marker of Antibody-Mediated Rejection in Heart Allograft Biopsies

In 2013, the International Society of Heart and Lung Transplant (ISHLT) introduced the working classification for pathologic changes associated with antibody-mediated rejection (AMR) of the heart allograft, known as pathologic AMR (pAMR). With 2 components associated with AMR, histopathologic changes) and immunopathologic markers, the proposed classification also suggests the use of class II HLA as a marker of endothelial integrity. It is known that during allograft rejection, endothelial cells are activated, therefore, we hypothesized that endothelial class II HLA rather than a marker of mere endothelial presence, is a marker of endothelial activation and becomes upregulated in AMR.Eight hundred thirty-eight heart allograft biopsies, collected from January 2016 to September 2018 at a single institution from patients with a current or recent diagnosis of AMR, were evaluated for both histopathologic and immunopathologic changes of AMR. Biopsies were labeled with immunofluorescence with antibodies against C4d and for immunohistochemistry with antibodies against C3d, CD68, and class II HLA. ISHLT criteria were used to classify the biopsies, and for class II HLA, both the percentage and the stain intensity were evaluated.Biopsies (74.8%) from our cohort showed either histopathologic pAMR-1, immunopathologic pAMR-1, or combined histopathologic and immunopathologic pAMR-2 evidence of AMR. Expression of endothelial HLA class II was significantly correlated with the diagnosis of AMR by percentage area (P < .0001) and intensity of staining (P < .0001). The diagnosis of AMR significantly correlated with moderate (+2) and strong (+3) staining intensity for class II HLA as follows: histopathologic and immunopathologic pAMR-2 with odds ratio (OR) = 28.3 (P < .0001);histopathologic pAMR-1 alone with OR = 22.7 (P < .0001); and immunopathologic pAMR-1 alone with OR = 32.6 (P < .0001). Interestingly, our study also suggested that the inclusion of C4d focally positive cases also significantly correlates with the diagnosis of AMR (P < .003).We confirmed our hypothesis that in heart allograft biopsies, there is a spectrum of both percentage and intensity of HLA class II expression due to endothelial activation, and that class II HLA by immunohistochemistry is a marker significantly correlated with the diagnosis of AMR. In addition, the group of focally positive C4d biopsies (10%-50%) should be considered positive for the immunopathologic component of the 2013 ISHLT classification, as this group of biopsies also correlated with the diagnosis of AMR.

CD68/CD31 immunohistochemistry double stain demonstrates increased accuracy in diagnosing pathologic antibody-mediated rejection in cardiac transplant patients.

Pathologic antibody-mediated rejection (pAMR) occurs in 10% of cardiac transplant patients and is associated with increased mortality. The endomyocardial biopsy remains the primary diagnostic tool to detect and define pAMR. However, certain challenges arise for the pathologist. Accurate identification of >10% of intravascular macrophages along with endothelial swelling, which remains a critical component of diagnosing pAMR, is one such challenge. We used double labeling with an endothelial and histiocytic marker to improve diagnostic accuracy. Twenty-two cardiac transplant endomyocardial biopsies were screened using a CD68/CD31 immunohistochemical (IHC) double stain. To determine whether pAMR diagnosis would change using the double stain, intravascular macrophage staining was compared to using CD68 alone. Twenty-two cardiac pAMR cases from patients were included. Fifty-nine percent of cases previously called >10% intravascular macrophage positive by CD68 alone were called <10% positive using the CD68/CD31 double stain. Not using the double stain was associated with a significant overcall. In C4d-negative cases, using the CD68/CD31 double stain downgraded the diagnosis of pAMR2 to pAMR1 in 32% of cases. It was concluded that more than one third of patients were overdiagnosed with pAMR using CD68 by IHC alone. We demonstrate the value of using a CD68/CD31 double stain to increase accuracy.

Early invasive assessment of the coronary microcirculation predicts subsequent acute rejection after heart transplantation

BackgroundAcute allograft rejection (AAR) plays an important role in patient and graft survival; therefore, more emphasis should be placed on its prediction. This study aimed to investigate baseline clinical and diagnostic variables associated with subsequent AAR during the first year post-transplant, especially focusing on early physiologic and anatomic measures. MethodsThis study enrolled 88 heart transplant patients who underwent fractional flow reserve (FFR), coronary flow reserve (CFR), the index of microcirculatory resistance (IMR) and intravascular ultrasound (IVUS) in the left anterior descending artery at baseline (within 8 weeks post-transplant). Cardiac index (CI), pulmonary capillary wedge pressure (PCWP), mean pulmonary artery pressure (mPAP), right atrial pressure and left ventricular ejection fraction were also evaluated. AAR was defined as acute cellular rejection of grade ≥2R and/or pathological antibody-mediated rejection of grade ≥pAMR2. ResultsDuring the first year post-transplant, 25.0% of patients experienced AAR. Patients with AAR during the first year showed higher rates of recipient obesity, lower rates of recipient-donor sex mismatch and rATG and tacrolimus uses, higher PCWP, mPAP and IMR, and lower CFR at baseline, compared with those without. In the multivariate analysis, only baseline IMR ≥ 16.0 was independently associated with AAR during the first year, demonstrating high negative predictive value (96.7%). ConclusionsInvasively assessing microvascular resistance (baseline IMR ≥ 16.0) in the early post-transplant period was an independent determinant of subsequent acute allograft rejection during the first year post-transplant, suggesting that early assessment of IMR may enhance patient risk stratification and target medical therapies to improve patient outcome.

Does Antibody-Mediated Rejection in the First-Month after Heart Transplant Lead to Poor Longer-Term Outcome?

Purpose Antibody-mediated rejection (AMR) has been recently well described in the 2011 ISHLT AMR biopsy-grading scale. AMR has been found to correlate with poor survival and greater development of cardiac allograft vasculopathy (CAV). There is concern that early AMR occurring in the first month after heart transplant (HTx) sets the stage for early activation of the humoral system resulting in poor outcome. Therefore, we chose to assess the timing of AMR within the first-month after HTx and subsequent outcomes. Methods Between 2000 and 2013, we assessed 20 heart transplant recipients who developed biopsy-proven AMR≥1 in the first-month post-HTx (n=9) or 2-12 months post-HTx (n=11). Endpoints included subsequent 5-year survival, and subsequent 5-year freedom from CAV (as defined by stenosis ≥ 30% by angiography) and non-fatal major adverse cardiac events (NF-MACE: myocardial infarction , new congestive heart failure , percutaneous coronary intervention, implantable cardioverter defibrillator/pacemaker implant, stroke). Of interest was whether early AMR in the first month post-transplant led to increased recurrence of pathology AMR in the rest of the first-year. Results 45% of AMR in the first-year after HTx occurred in the first-month post-HTx (early AMR) and 55% occurred between 2-12 months post-HTx (late AMR). None of the patients with early or late AMR had recurrence of antibody-mediated rejection in the first-year. In the long-term, late AMR was associated with significantly less freedom from NF-MACE. However, there was no difference in 5-year survival or 5-year freedom from CAV in patients with early or late AMR. (see Table) Conclusion AMR in the first-month post-HTx does not appear to lead to recurrent AMR or poor 5-year outcome. Late AMR (between 2-12 months post-HTx) may increase the risk for NF-MACE. More aggressive anti-humoral immunosuppression strategies may be warranted in this patient group.

DOES PATHOLOGY ANTIBODY-MEDIATED REJECTION LEAD TO INTIMAL THICKENING IN THE FIRST-YEAR AFTER HEART TRANSPLANTATION?

Cardiac allograft vasculopathy (CAV) may be antibody-mediated. The 2011 ISHLT grading scale includes antibody-mediated rejection (AMR). It is believed that pathology AMR includes not only AMR 2 but also AMR 1H via intragraft mRNA transcript findings. It appears that the molecular signature, meaning

Pathological antibody-mediated rejection in pediatric heart transplant recipients: Immunologic risk factors, hemodynamic significance, and outcomes.

Biopsy-diagnosed pAMR has been observed in over half of pediatric HT recipients within 6years of transplantation. We report the incidence and outcomes of pAMR at our center. All endomyocardial biopsies for all HT recipients transplanted between 2010 and 2015 were reviewed and classified using contemporary ISHLT guidelines. Graft dysfunction was defined as a qualitative decrement in systolic function by echocardiogram or an increase of ≥3mmHg in atrial filling pressure by direct measurement. Among 96 patients, pAMR2 occurred in 7 (7%) over a median follow-up period of 3.1years, while no cases of pAMR3 occurred. A history of CHD, DSA at transplant, and elevated filling pressures were associated with pAMR2. Five-sixths (83%) of patients developed new C1q+ DSA at the time of pAMR diagnosis. There was a trend toward reduced survival, with 43% of patients dying within 2.3years of pAMR diagnosis.

Pathology Antibody-Mediated Rejection Plus Circulating Donor-Specific Antibodies Increase Development of Cardiac Allograft Vasculopathy After Heart Transplantation

Pathology Antibody-Mediated Rejection Plus Circulating Donor-Specific Antibodies Increase Development of Cardiac Allograft Vasculopathy After Heart Transplantation

Gene Expression Profiling for the Identification and Classification of Antibody-Mediated Heart Rejection.

Antibody-mediated rejection (AMR) contributes to heart allograft loss. However, an important knowledge gap remains in terms of the pathophysiology of AMR and how detection of immune activity, injury degree, and stage could be improved by intragraft gene expression profiling. We prospectively monitored 617 heart transplant recipients referred from 4 French transplant centers (January 1, 2006-January 1, 2011) for AMR. We compared patients with AMR (n=55) with a matched control group of 55 patients without AMR. We characterized all patients using histopathology (ISHLT [International Society for Heart and Lung Transplantation] 2013 grades), immunostaining, and circulating anti-HLA donor-specific antibodies at the time of biopsy, together with systematic gene expression assessments of the allograft tissue, using microarrays. Effector cells were evaluated with in vitro human cell cultures. We studied a validation cohort of 98 heart recipients transplanted in Edmonton, AB, Canada, including 27 cases of AMR and 71 controls. A total of 240 heart transplant endomyocardial biopsies were assessed. AMR showed a distinct pattern of injury characterized by endothelial activation with microcirculatory inflammation by monocytes/macrophages and natural killer (NK) cells. We also observed selective changes in endothelial/angiogenesis and NK cell transcripts, including CD16A signaling and interferon-γ-inducible genes. The AMR-selective gene sets accurately discriminated patients with AMR from those without and included NK transcripts (area under the curve=0.87), endothelial activation transcripts (area under the curve=0.80), macrophage transcripts (area under the curve=0.86), and interferon-γ transcripts (area under the curve=0.84; P<0.0001 for all comparisons). These 4 gene sets showed increased expression with increasing pathological AMR (pAMR) International Society for Heart and Lung Transplantation grade (P<0.001) and association with donor-specific antibody levels. The unsupervised principal components analysis demonstrated a high proportion of molecularly inactive pAMR1(I+), and there was significant molecular overlap between pAMR1(H+) and full-blown pAMR2/3 cases. Endothelial activation transcripts, interferon-γ, and NK transcripts showed association with chronic allograft vasculopathy. The molecular architecture and selective AMR transcripts, together with gene set discrimination capacity for AMR identified in the discovery set, were reproduced in the validation cohort. Tissue-based measurements of specific pathogenesis-based transcripts reflecting NK burden, endothelial activation, macrophage burden, and interferon-γ effects accurately classify AMR and correlate with degree of injury and disease activity. This study illustrates the clinical potential of a tissue-based analysis of gene transcripts to refine diagnosis of heart transplant rejection.

Donor-specific anti-HLA antibodies with antibody-mediated rejection and long-term outcomes following heart transplantation

Donor-specific anti-HLA antibodies (DSA) are common after heart transplantation and are associated with rejection, cardiac allograft vasculopathy, and mortality. A noninvasive diagnostic test for pathologic antibody-mediated rejection (pAMR) does not exist. From January 1, 2010, through August 31, 2013, 221 consecutive adult patients underwent heart transplantation and were followed through October 1, 2015. The primary objective was to determine whether the presence of DSA could detect AMR at the time of pathologic diagnosis. Secondary analyses included association of DSA (stratified by major histocompatibility complex class and de novo status) during AMR with new graft dysfunction, graft loss (mortality or retransplantation), and development of cardiac allograft vasculopathy. During the study period, 69 patients (31.2%) had DSA (24% had de novo DSA), and there were 74 episodes of pAMR in 38 patients. Sensitivity of DSA at any mean fluorescence intensity to detect concurrent pAMR was only 54.3%. The presence of any DSA during pAMR increased the odds of graft dysfunction (odds ratio = 5.37; 95% confidence interval [CI], 1.34-21.47; p = 0.018), adjusting for age, sex, and timing of AMR. Circulating class II DSA after transplantation increased risk of future pAMR (hazard ratio = 2.97; 95% CI, 1.31-6.73; p = 0.009). Patients who developed de novo class II DSA had 151% increased risk of graft loss (contingent on 30-day survival) compared with patients who did not have DSA (95% CI, 1.11-5.69; p = 0.027). DSA were inadequate to diagnose pAMR. Class II DSA provided prognostic information regarding future pAMR, graft dysfunction with pAMR, and graft loss.

A multi-institutional evaluation of antibody-mediated rejection utilizing the Pediatric Heart Transplant Study database: Incidence, therapies and outcomes

Current knowledge of antibody-mediated rejection (AMR) after heart transplantation (HT) stems largely from adult data. Using the Pediatric Heart Transplant Study (PHTS) database, we report the incidence of AMR, describe treatment, and evaluate outcomes for treated AMR in children after HT. We queried the PHTS database for patients <18 years of age undergoing primary HT between January 2010 and December 2014. An AMR episode was defined as either a biopsy consistent with pathologic AMR or a rejection event based on immunotherapy augmentation directed against antibody production. Biopsy data, treatment strategies and survival were analyzed. An episode of AMR was identified in 179 of 1,596 (11%) HT recipients and in 246 of 705 (35%) rejection episodes. AMR was diagnosed by biopsy in 182 of 246 episodes and by immunotherapy in 64 of 179 episodes. Mixed rejection was identified in 179. Freedom from AMR was 88% and 82% at 1 and 3 years, respectively. AMR therapies included intravenous immunoglobulin (IVIg) (58%), plasmapheresis (40%), rituximab (40%), bortezomib (11%) and eculizumab (0.4%). The most commonly used combination therapies included IVIg/plasmapheresis/rituximab (13%). Thirty-three patients (16%) died after developing AMR. Patient and graft survival were lower for the AMR+ group. One- and 3-year survival after initial AMR diagnosis was 88% and 77%, respectively. In his study we report the largest experience of AMR in pediatric HT recipients. AMR was common and often occurred concurrently with acute cellular rejection. There is wide variability in the treatment of AMR. Short-term patient and graft outcomes were worse for those with treated AMR.

The impact of angiotensin II type 1 receptor antibodies on post-heart transplantation outcome in Heart Mate II bridged recipients.

Antibodies targeting angiotensin II type 1 receptor (AT1R) have been associated with malignant hypertension, autoimmune diseases and acute rejection and graft loss in solid organ transplantation. The aim of our study was to assess the impact of anti-AT1R antibodies on survival and incidence of acute cellular rejection (ACR) and pathology antibody-mediated rejection (pAMR) in a population of heart transplant recipients who were bridged to transplantation with a durable mechanical assist device Heart Mate II. Sera of 69 consecutive heart transplant recipients transplanted between October 2008 and August 2014 were tested for the presence of angiotensin II type 1 receptor antibodies before Heart Mate II device implantation and at the time of transplantation. Overall survival and post-transplant rejection-free survival were compared between antibody-negative and antibody-positive recipients using Kaplan-Meier and log-rank tests. Anti-AT1R antibodies were present in 8 patients (11.6%) before Heart Mate II implantation. During the left ventricular assist device (LVAD) bridging, 44 patients (63.8%) who were initially anti-AT1R antibody-negative became positive, leaving 17 (24.6%) anti-AT1R antibody-negative patients at the time of transplantation for all comparisons. One- and 5-year survival was 88 ± 8 and 76 ± 10% for anti-AT1R antibody-negative and 87 ± 5 and 81 ± 7% for anti-AT1R antibody-positive patients, respectively (P = 0.582). Freedom from ACR at 1 year was 68 ± 12% for anti-AT1R-negative and 75 ± 6% for anti-AT1R-positive recipients (P = 0.218). None of the anti-AT1R-negative patients developed AMR 1 year post-transplantation, whereas freedom from pAMR in anti-AT1R-positive recipients was 98 ± 2% (P = 0.198). Our data showed no difference in the overall post-heart transplant survival and freedom from acute cellular and antibody-mediated rejection between anti-AT1R-negative and anti-AT1R-positive recipients. Further research is needed to assess the role of anti-AT1R antibodies in the risk stratification of LVAD-bridged recipients on the post-heart transplantation outcomes.