Acute Cellular Rejection Grade Research Articles

Challenging endomyocardial biopsies' gold standard status in rejection screening: a prospective blinded study on cardiovascular magnetic resonance in heart transplant patients

Abstract Background The detection of rejection is crucial for graft survival after heart transplantation. Endomyocardial biopsies (EMBs) serve as the gold standard for rejection surveillance despite its limitations, while cardiac magnetic resonance (CMR), in conjunction with donor-derived cell-free DNA (dd-cfDNA) analysis, holds promise as a non-invasive approach. Purpose In this prospective blinded study, our aim was to evaluate feasibility of CMR for rejection screening, validating it against EMB, dd-cfDNA, and clinical status. Methods We blindly analyzed 250 studies involving CMR, EMB, dd-cfDNA, and clinical data in 58 heart transplant recipients (17 children and 41 adults). Of these 250 studies, 239 (96%) were assessed within 1 to 28 months after transplantation, and 11 studies (4%) 3-14 years post-transplant. In CMR T1- and T2-mapping analyses, acute rejection was defined by T1 or T2 times exceeding the specified thresholds (T1 ≥ 1060 ms, T2 ≥ 55 ms) in at least 2/16 heart segments. EMB findings were graded according to the ISHLT criteria (2005 and 2013), and acute cellular rejection (ACR) grade ≥ 2 or antibody-mediated rejection (AMR) grade ≥ 1 were considered significant. The cutoff for abnormal dd-cfDNA was set at 0.2%. Solely clinical rejection included patients with absent or normal EMB (ACR grade 0-1 and AMR grade 0), but with symptoms or ECG/echocardiographic findings indicative of acute rejection. Results Our study revealed 22 acute rejections, with 15 confirmed through EMB and 7 diagnosed solely based on clinical indicators. Among the EMB-confirmed cases, 3/15 exhibited AMR grade 1, 10/15 demonstrated ACR grade 2 and 2/15 ACR grade 3. CMR identified rejection in 77% (17/22) of cases. Specifically, it successfully identified rejection in 67% (10/15) of EMB-confirmed cases and 100% (7/7) of solely clinically diagnosed cases. Examining the concordance between CMR findings and dd-cfDNA levels, 35% (6/17) of the CMR-identified rejection cases exhibited abnormal dd-cfDNA, while 30% (5/17) showed normal levels. Notably, in one case, despite normal CMR results, both EMB and dd-cfDNA indicated rejection. Conversely, in four instances where EMB indicated acute rejection, both CMR and dd-cfDNA levels were normal. These findings highlight a potential discrepancy in the diagnostic accuracy of EMB, suggesting its tendency to both over- and underdiagnose acute heart transplant rejection. The integration of CMR and dd-cfDNA in the diagnostic process may provide a more comprehensive and reliable assessment of rejection episodes, offering valuable insights for clinical decision-making in heart transplant recipients. Conclusions EMB presents challenges in both under- and overdiagnosing rejection in heart transplant surveillance. A strategy based on CMR, combined with dd-cfDNA analyses, shows promise as a new gold standard for rejection screening.Flowchart

Incidence of acute cellular and antibody mediated rejection in heart transplantation from a tertiary care center

Abstract Background Heart transplant greatly increases the survival in patients with end stage heart failure. However, cardiologists and cardiothoracic surgeons face difficulty in detecting rejection at times. Post-cardiac transplant surveillance biopsy remains the gold standard for diagnosing rejection and therapeutic intervention. Purpose The objective is to report our experience of monitoring rejection by endomyocardial biopsy (EMB). Methods The index study is an ambispective one where we evaluated all the patients who underwent heart transplant at our center from 1994 to 2021. Results Heart transplant were carried out in 72 patients within a period of 28 years, with majority being done after 2016. The patients mean age was 31.6 years (10-59 years). The commonest indication was dilated cardiomyopathy (67%) followed by ischemic cardiomyopathy (2.5%), congenital heart disease (7%) and other miscellaneous causes (5%). The mean age of donors was 31.8 years (14-55 years). Eight of the 72 patients died without any post cardiac transplant endomyocardial biopsy. The remaining 64 patients underwent cumulatively a total of 271 endomyocardial biopsy procedures. Majority of endomyocaridal biopsies contained three cores (range 1-7). First EMB procedure was performed in 17 (26.6%) patients within first seven days, 35(53.1%) within the second week and 12(20.3%) after 14 days. Histologically rejection was identified in 42 (66%) patients. Thirty-nine (61%) had only acute cellular rejection (ACR), one (1.6%) had only antibody mediated rejection (AMR) and two (3.1%) had mixed ACR and AMR. These 42 patients with rejection had a total of 90 (33.2%) episodes of ACR detected in 271 endomyocardial biopsies. Out of 90 ACR, 44 (48.9%) occurred in less than six months; seventeen (18.9% of 90 ACR) episodes occurred between six and 12 months and 29 (32.2%) after one year. The mean of presentation of first ACR episode was 36 weeks (range 5 days to 3.6 years). There were 172 endomyocardial procedures within one year of transplantation of which 61 (35%) showed rejection. Ninety-nine endomyocardial procedures were performed one year of transplantation of which 29 (29%) showed rejection. Most common grade in both the time period was ACR grade 1R (International Society for Heart and Lung Transplantation (ISHLT) 2004) (72.1% in ≤1 year and 65.5% in >1 year) followed by 2R and 3R. Thirty-eight (90.4%) of 42 patients expired in less than five years since transplantation. Two (4.8%) patients survived between five to ten years post transplantation and two (4.8%) survived more than ten years. The longest cardiac transplant recipient had completed 23 years since cardiac transplant and is alive till date. Conclusions EMB being the gold standard for cardiac allograft rejection, the cardiac pathologists need to be aware of the various grades of rejection including mixed rejection.Acute cellular rejection (ISHLT 2004)

The role of C4d and donor specific antibodies in face and hand transplantation-a systematic review.

To date, little is known about the mechanisms of rejection in vascularized composite allotransplantation, particularly for antibody mediated rejection. Additionally, no clear guidelines exist for the diagnosis and management of antibody-mediated rejection in vascularized composite allotransplantation. A systematic review of electronic databases (Embase and PubMed) was conducted to evaluate the relationship of donor specific antibodies and C4d deposition in correlation with cellular rejection following hand and face transplantation reported by centers between 1998 and July 2023. We extracted data on serum donor specific antibodies at the time of biopsy proven rejection according to Banff classification and C4d staining of target tissues. Mann-Whitney U tests were performed to compare rejection grade between groups divided by status of C4d deposition and serum donor specific antibodies, and Fisher's Exact test was used to assess association between the two markers. This review adhered to PRISMA guidelines. A total of 26 patients (5 face, 21 hand) were identified and data on 90 acute rejection episodes with information on Banff grade, donor specific antibody status, and C4d deposition were available. Donor specific antibodies were found to be associated with higher rejection grade (p = 0.005). C4d was not found to be associated with higher rejection grade (p = 0.33). Finally, no significant association was found between concurrent status of the two markers (p = 0.23). These findings suggest that the presence of donor specifc antibodies may be associated with higher grades of acute cellular rejection following hand and face transplantation. More consistent reporting on rejection episodes is needed in order to better understand antibody-mediated rejection in vascularized composite allotransplantation.

Clinical utility of plasma percent donor-derived cell-free DNA for lung allograft surveillance: A real-world single-center experience

BackgroundPlasma percent donor-derived cell-free DNA (%dd-cfDNA) has been investigated as a biomarker of allograft injury after lung transplantation (LT). We sought to determine the clinical utility of plasma percent donor-derived cell-free DNA (%dd-cfDNA) as a screening tool for acute cellular rejection (ACR) and respiratory infections (RIs) among LT recipients (LTRs). MethodsWe retrospectively analyzed %dd-cfDNA results from 95 plasma samples collected from 81 bilateral LTRs at our center with a paired transbronchial biopsy within 24 hours after sample collection. We calculated sensitivity, specificity, negative predictive value (NPV), and positive predictive value (PPV) of %dd-cfDNA to detect ACR and RIs and used a generalized estimating equation model to compare %dd-cfDNA between groups. ResultsA dd-cfDNA threshold of 0.5% had low sensitivity to detect ACR among LTRs >45 days after LT (41.67%), as did a 70% increase in %dd-cfDNA (50.00%). The NPV was high (88.89% and 87.50%, respectively) but driven by the low prevalence of ACR (12/95 [12.6%]). The area under the receiver operating characteristic curve (AUC) was 0.499 (95%CI [0.326-0.672]) and 0.360 (95%CI [0.132-0.588]) for the detection of ACR and ACR grade ≥A2, respectively. The adjusted mean %dd-cfDNA trended higher in LTRs with a definite or possible RI (1.218, 95%CI [0.671-2.212]) than in LTRs without microbial isolation (0.731, 95%CI [0.525-1.017], p=0.059), but was not significantly different from those with microbial colonization (0.873, 95%CI [0.538-1.415], p=0.390). There was no significant difference in the adjusted mean %dd-cfDNA between patients with microbial colonization and those without microbial isolation (p=0.464). %dd-cfDNA testing had an AUC of 0.573 (95%CI [0.431-0.716]) for the detection of lung allograft dysfunction due to ACR and/or infection. Conclusions%dd-cfDNA may have limited utility as a screening tool to detect ACR among LTRs.

(789) - Outcomes in Patients with Acute Cellular Rejection Grade 1R: Is the Debate Over?

(789) - Outcomes in Patients with Acute Cellular Rejection Grade 1R: Is the Debate Over?

Treatment Responses in Histologic Versus Molecular Diagnoses of Lung Rejection.

Histologic evaluation of allograft biopsies after lung transplantation has several limitations, suggesting that molecular assessment using tissue transcriptomics could improve biopsy interpretation. This single-center, retrospective cohort study evaluated discrepancies between the histology of transbronchial biopsies (TBBs) with no rejection (NR) and T-cell mediated rejection (TCMR) by molecular diagnosis. The accuracy of diagnosis was assessed based on response to treatment. 54 TBBs from Prague Lung Transplant Program obtained between December 2015 and January 2020 were included. Patients with acute cellular rejection (ACR) grade ≥ 1 by histology received anti-rejection treatment. Response to therapy was defined as an increase in FEV1 of ≥ 10% 4weeks post-biopsy compared to the pre-biopsy value. Among the 54 analyzed TBBs, 25 (46%) were concordant with histology, while 29 (54%) showed discrepancies. ACR grade 0 was found in 12 TBBs (22%) and grade A1 ≥ 1 in 42 TBBs (78%). Treatment response was present in 14% in the NR group and in 50% in the TCMR group (p = 0.024). Our findings suggest that low-grade acute cellular rejection is less likely to be associated with molecular TCMR, which might better identify lung transplant recipients who benefit from therapy.

Donor-derived cell-free DNA kinetics in the first year post-transplant in patients without acute cellular rejection

Abstract Background Donor-derived cell-free DNA (dd-cfDNA) appears to be an excellent biomarker for acute cellular rejection (ACR) in heart transplant (HT) recipients. However, a significant rise in the immediate post-transplant phase probably related to direct myocardial damage during harvesting-implantation process has been described, and could potentially affect the biomarker’s performance in the first post-HT month. We sought to describe the biomarker’s kinetics in the first year post-HT in patients without ACR. Methods Prospective, multicentre study performed between 2019 and 2022. All patients underwent surveillance endomyocardial biopsy (EMB) at 7 pre-specified time points in the first post-HT year (0.5, 1, 2, 3, 4, 6 and 12 months). Dd-cfDNA levels were determined prior to each EMB, using Next Generation Sequencing technology. Results A total of 1006 pairs of EMB-dd-cfDNA from 206 patients were available for analysis (mean age 54 ±12 years, 74% male). Of these, 44 (4.3%) had clinically relevant rejection, defined as ACR grade ≥2, and 962 (95.7%) did not (562 0R and 400 1R). 15 samples that had pAMR≥1 were excluded from this analysis. We divided the 947 samples with ACR 0-1 into three subgroups, according to the time of blood sampling: at 15 days (time point 1), at 30 days (time point 2) and 2 to 12 months post-HT (time point 3). Samples with 0R that were drawn at 15 and 30 days showed significantly higher levels of dd-cfDNA than those drawn between the second month and first year post-transplant (p <0.001) (Figure 1A). When we included patients with 1R, a similar trend was observed (p <0.001) (Figure 1B). Median dd-cfDNA levels for 0R and for 0-1R at the three time points are shown in Figure 2. Conclusions Dd-cfDNA levels in patients without clinically relevant rejection are significantly higher in the first month post-transplant. Further studies are needed in order to determine if a different cut-off point for ACR during the first post-HT month is useful for ACR diagnosis.Donor-derived cell-free DNA kineticsDd-cfDNA values at three time points

The importance of avoiding time-dependent bias when testing the prognostic value of an intervening event — Two acute cellular rejection examples in intestinal transplantation

In testing the prognostic value of the occurrence of an intervening event (clinical event that occurs posttransplant), 3 proper statistical methodologies for testing its prognostic value exist (time-dependent covariate, landmark, and semi-Markov modeling methods). However, time-dependent bias has appeared in many clinical reports, whereby the intervening event is statistically treated as a baseline variable (as if it occurred at transplant). Using a single-center cohort of 445 intestinal transplant cases to test the prognostic value of first acute cellular rejection (ACR) and severe (grade of) ACR on the hazard rate of developing graft loss, we demonstrate how the inclusion of such time-dependent bias can lead to severe underestimation of the true hazard ratio (HR). The (statistically more powerful) time-dependent covariate method in Cox’s multivariable model yielded significantly unfavorable effects of first ACR (P < .0001; HR = 2.492) and severe ACR (P < .0001; HR = 4.531). In contrast, when using the time-dependent biased approach, multivariable analysis yielded an incorrect conclusion for the prognostic value of first ACR (P = .31, HR = 0.877, 35.2% of 2.492) and a much smaller estimated effect of severe ACR (P = .0008; HR = 1.589; 35.1% of 4.531). In conclusion, this study demonstrates the importance of avoiding time-dependent bias when testing the prognostic value of an intervening event.

The Role of Left Atrial Longitudinal Strain in the Diagnosis of Acute Cellular Rejection in Heart Transplant Recipients.

Our aim was to investigate the role of left atrial longitudinal strain (LALS) in the non-invasive diagnosis of acute cellular rejection (ACR) episodes in heart transplant (HTx) recipients. Methods: We performed successive echocardiographic exams in 18 consecutive adult HTx recipients in their first year after HTx within 3 h of the routine surveillance endomyocardial biopsies (EMB) in a single center. LALS parameters were analyzed with two different software. We investigated LALS association with ACR presence, as well as inter-vendor variability in comparable LALS values. Results: A total of 147 pairs of EMB and echo exams were carried out. Lower values of LALS were significantly associated with any grade of ACR presence. Peak atrial longitudinal strain (PALS) offered the best diagnostic value for any grade of ACR, with a C statistic of 0.77 using one software (95% CI 0.68–0.84, p < 0.0005) and 0.64 with the other (95% CI 0.54–0.73, p = 0.013) (p = 0.02 for comparison between both curves). Reproducibility between comparable LALS parameters was poor (intraclass correlation coefficients were 0.60 for PALS, 95% CI 0.42–0.73, p < 0.0005; and 0.42 for PALS rate, 95% CI −0.13–0.68, p < 0.0005). Conclusions: LALS variables might be a sensitive marker of ACR in HTx recipients, principally discriminating between those studies without rejection and those with any grade of ACR. Inter-vendor variability was significant.

Retrospective Evaluation of Rabbit Antithymocyte Globulin Induction in Heart Transplant Patients.

Background.The dosing intensity of antithymocyte globulin as induction therapy in heart transplantation remains controversial. We sought to evaluate the efficacy and safety of rabbit antithymocyte globulin at a total dose of 4.5 mg/kg compared with <4.5 mg/kg.Methods.This was a retrospective study of consecutive patients who underwent heart transplantation from January 2016 to December 2018 at a single quaternary care center. Exposure was defined as full antithymocyte globulin (4.5 mg/kg total) induction compared with partial (<4.5 mg/kg) induction. The primary outcome was the incidence of The International Society for Heart and Lung Transplantation 1990 acute cellular rejection grade 2 or above at 2 y. Secondary outcomes were all-cause mortality, number of infections, and time to therapeutic tacrolimus levels. Cox proportional hazard models were used to compare rejection rates and mortality.Results.Of 201 patients, 61 received partial and 140 received full induction. There was no difference in the cumulative incidence of cellular rejection grade 2 or above (18% versus 11.4%, P = 0.209) within 2 y. The adjusted hazard ratio was 1.45 (confidence interval: 0.62-3.37, P = 0.388) for partial compared with full induction for any grade rejection. Landmark survival analysis conditional on survival to 1 mo showed no difference in mortality (P = 0.239). There was no difference in the incidence of infection within 3 mo of transplant (partial 29.5% versus full 20.0%, P = 0.140). Both groups achieved therapeutic tacrolimus levels by day 7 after initiation.Conclusions.There was no difference in overall risk for any grade cellular rejection between partial or full dose induction therapy. Additionally, there was no difference in medium-term mortality from landmark survival analysis.

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.

Prevalence of Clinical Events Following High Donor Derived Cell Free DNA in the Absence of Rejection in Heart Transplant Recipients

Purpose Donor-derived cell free DNA (dd-cfDNA) is a noninvasive biomarker used to assess graft injury following heart transplant (HTx). While the relationship between increased levels of dd-cfDNA and acute cellular rejection (ACR) and antibody mediated rejection (AMR) has been well studied, less is known about the frequency of other processes that damage the allograft following an elevated dd-cfDNA measurement. The goal of this analysis was to assess the prevalence of clinical events following a high dd-cfDNA measurement in the absence of rejection. Methods This retrospective study included 223 HTx recipients from the Donor-Derived Cell-Free DNA-Outcomes AlloMap Registry (D-OAR) with elevated dd-cfDNA (AlloSure®), defined as a level ≥0.20% that did not occur within 14 days of biopsy proven ACR (grade ≥2R) or AMR (pAMR≥1). Clinical events included the presence of donor specific antibodies (DSA), cardiac allograft vasculopathy (CAV; ISHLT grade ≥1), graft dysfunction [left ventricular ejection fraction (LVEF) ≤45% or a proportional decrease in LVEF ≥25% from the first D-OAR study visit], ACR, AMR and death. Clinical data were collected for 180 days following the first elevated dd-cfDNA. Allosensitization was defined as a pre-transplant panel reactive antibody ≥10%. Results The study consisted of 68% men and 71% Caucasians (mean ± SD age at HTx 52 ± 14 years), with non-ischemic cardiomyopathy (46%) representing the primary reason for HTx. The median time post-HTx to the first non-rejection elevated dd-cfDNA was 241 days (IQR 154-362). Fifty-seven of the 223 patients (26%), of whom 6 were allosensitized, experienced 70 clinical events in the 180 days post-elevated dd-cfDNA. The presence of at least one DSA (n=36) was the most common clinical event, of which 28/36 (78%) were de novo DSA. Graft dysfunction (n=14), CAV (n=9), AMR (n=5), death (n=4), and ACR (n=2) were the next most common. Median time (IQR) from elevated dd-cfDNA to these events was as follows: DSA, 37 days (15-104); graft dysfunction, 22 days (0-77); AMR, 70 days (42-91); and death, 100 days (75-124). Conclusion Elevated dd-cfDNA levels in the absence of allograft rejection appeared to precede the development of clinically relevant events within the next 6 months. Further studies are needed to determine whether elevated dd-cfDNA should trigger a change in clinical management. Donor-derived cell free DNA (dd-cfDNA) is a noninvasive biomarker used to assess graft injury following heart transplant (HTx). While the relationship between increased levels of dd-cfDNA and acute cellular rejection (ACR) and antibody mediated rejection (AMR) has been well studied, less is known about the frequency of other processes that damage the allograft following an elevated dd-cfDNA measurement. The goal of this analysis was to assess the prevalence of clinical events following a high dd-cfDNA measurement in the absence of rejection. This retrospective study included 223 HTx recipients from the Donor-Derived Cell-Free DNA-Outcomes AlloMap Registry (D-OAR) with elevated dd-cfDNA (AlloSure®), defined as a level ≥0.20% that did not occur within 14 days of biopsy proven ACR (grade ≥2R) or AMR (pAMR≥1). Clinical events included the presence of donor specific antibodies (DSA), cardiac allograft vasculopathy (CAV; ISHLT grade ≥1), graft dysfunction [left ventricular ejection fraction (LVEF) ≤45% or a proportional decrease in LVEF ≥25% from the first D-OAR study visit], ACR, AMR and death. Clinical data were collected for 180 days following the first elevated dd-cfDNA. Allosensitization was defined as a pre-transplant panel reactive antibody ≥10%. The study consisted of 68% men and 71% Caucasians (mean ± SD age at HTx 52 ± 14 years), with non-ischemic cardiomyopathy (46%) representing the primary reason for HTx. The median time post-HTx to the first non-rejection elevated dd-cfDNA was 241 days (IQR 154-362). Fifty-seven of the 223 patients (26%), of whom 6 were allosensitized, experienced 70 clinical events in the 180 days post-elevated dd-cfDNA. The presence of at least one DSA (n=36) was the most common clinical event, of which 28/36 (78%) were de novo DSA. Graft dysfunction (n=14), CAV (n=9), AMR (n=5), death (n=4), and ACR (n=2) were the next most common. Median time (IQR) from elevated dd-cfDNA to these events was as follows: DSA, 37 days (15-104); graft dysfunction, 22 days (0-77); AMR, 70 days (42-91); and death, 100 days (75-124). Elevated dd-cfDNA levels in the absence of allograft rejection appeared to precede the development of clinically relevant events within the next 6 months. Further studies are needed to determine whether elevated dd-cfDNA should trigger a change in clinical management.

Utility of AlloMap® Scores in Younger Pediatric Heart Transplant Recipients

<h3>Purpose</h3> AlloMap® is used to predict acute cellular rejection (ACR) post heart transplant (HT) in pediatric patients > 15 years old. Data on its utility in a younger pediatric cohort are limited. We examine the correlation of AlloMap® score with hemodynamic, echocardiographic (echo), and endomyocardial biopsy (EMB) markers of rejection in HT recipients ≤ 15 years old. <h3>Methods</h3> Retrospective study of all pediatric patients ≤ 15 years old at our institution who had AlloMap® testing with routine cardiac catheterization and EMB, or when rejection was suspected based on clinical/echo evidence. <h3>Results</h3> 438 AlloMap® scores obtained from 57 children from 2010-2017. Median age at HT was 2.4 years (IQR 0.6-11.9 years) and 56% (n=32) were male. Indication for HT was cardiomyopathy (CM) in 56.1% (n=32) and congenital heart disease (CHD) in 43.9% (n=25) of patients. AlloMap® scores did not differ between CHD and CM groups (32.8 ± 3.5 vs 32.4 ± 3.7, p=.2). Scores were highest in those <2 years old. Scores differed between patients aged < 2 versus 2-10 years (p<.001) and between patients aged 2-10 and > 10 years old (p<.001). 32 rejection episodes were identified in 18 patients; 5 patients (27.8%) were < 1 year old. ACR grade 0/1R and 2R/3R were seen in 84.4% (n=27) and 15.6% (n=5), respectively. Those with ACR 0R had lower AlloMap® scores than those with ACR 1R (p=.024), but there was no difference between 1R and higher grade rejection. AlloMap® score weakly correlated with left ventricular end diastolic pressure (LVEDP) (r=.149, p=.026), but not with echo variables. <h3>Conclusion</h3> AlloMap® scores are similar in CM and CHD patients. Scores decrease with age, which may represent differing gene expression profiles over time. Scores are lower for those with ACR 0R versus 1R, but do not differ significantly between 1R and higher grade ACR. AlloMap® score correlates weakly with LVEDP, but not echo variables. Sample size is small with few rejection episodes. A larger study is needed to determine the utility of routine AlloMap® surveillance in HT recipients ≤ 15 years of age.

Acute Cellular Rejection at Long-Term Follow-Up After Heart Transplantation: A Single Center Experience in Japan

<h3>Purpose</h3> Acute cellular rejection (ACR) is a major cause of death within 1-3 years after heart transplantation (HTx). However, recipients are usually asymptomatic in the early stages of ACR, and it is difficult to obtain specific findings of ACR using non-invasive tests. Therefore, regular endomyocardial biopsy plays an important role in monitoring for ACR. Although mortality from ACR decreases in the long term after HTx, there is no strong evidence of screening methods such as the appropriate frequency of biopsies. We retrospectively analyzed the outcome of ACR in our institution. <h3>Methods</h3> We enrolled 136 patients who underwent HTx at our hospital from May 1999 to April 2020. 7 Recipients under the age of 18 at the time of HTx were excluded. The current myocardial biopsy schedule is 1,2,3,5,7,11 weeks, 4.5,6,9 months, twice a year from 1 to 2 years, and once a year until 10 years after HTx. After that, it is once every two years. Tacrolimus (cyclosporine before 2005), mycophenolate mofetil, and prednisolone were used as first immunosuppressive therapies. Basiliximab was administered in patients with impaired renal function, elderly patients, and anti-donor antibody-positive patients. <h3>Results</h3> 129 HTx recipients (74% male, age at transplantation 41.8 ± 12.9 years) were analyzed. Grade 2R or higher grade of ACR were observed in 19 times in 13 patients during a mean follow-up period of 7.4 ± 5.0 years. Only one case with Grade 2R (3.5 years after HTx) combined with antibody-mediated rejection developed heart failure. All recipients with ACR recovered with steroid pulse therapy, and no deaths due to ACR were observed. The frequency of ACR of Grade 2R or higher in less than 1 year, 1-3 years, 3-5 years, 5-10 years, and 10 years or more after HTx was 4.0, 2.1, 2.2, 0.7, and 0 %/patients/years, respectively. The incidence of ACR was significantly higher in 15 patients who took cyclosporine as the first calcineurin inhibitor after HTx than in 114 patients who took tacrolimus (40% vs 6.1%, p <0.05). Late recurrence of ACR after HTx was seen in a patient with poor medication adherence and in a patient after immunosuppressive drug changes. <h3>Conclusion</h3> The frequency of developing ACR decreased over the years after HTx. Careful observation is still required in cases with a high risk of recurrence, such as inadequate medication adherence even long after HTx.

Impact of Liver Biopsy Size on Histopathologic Evaluation of Liver Allograft Rejection.

Allograft liver biopsy is the gold standard in assessing transplant recipients for graft dysfunction. The impact of biopsy sample size on the diagnosis of acute cellular rejection (ACR) has not been studied. To assess the relationship of biopsy sample length with the diagnosis and determine optimal biopsy sample size in the transplant setting. We retrospectively reviewed 68 core biopsies from patients with a history of liver transplant. Each biopsy sample was read, on 5 different occasions with differing lengths, to assess for ACR per Banff criteria. Categorical agreement was calculated from rejection severity. The length of biopsy sample strongly correlated with the number of portal tracts. ACR rates increased from 73.5% to 79.4% with increase in length from 1 cm to 2 cm, and moderate rejection increased from 27.9% to 33.82%. At 1.0 and 1.5 cm, no cases of severe rejection were detected; at 2.0 cm, 1 case was detected; and at 3.0 cm, 2 cases were detected. The major error rate was reduced to less than 10% with a length of 2.0 cm, at which length the average number of complete and partial portal triads was 10 and 13, respectively. The likelihood of diagnosing ACR and rejection grade increased substantially with increase in biopsy sample length. This study suggests that a minimum length of 2 cm, 10 complete portal triads, or 13 partial/complete portal triads should be obtained for confident exclusion and grading of ACR.

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 antibody-mediated rejection (AMR) following heart transplantation relies on the 2013 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 (EMB). We retrospectively graded the extent of MVI in 291 EMB from 291 patients according to a 4-point scale where 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 micro-arrays or reverse transcriptase multiplex ligation-dependent probe amplification, current pathology classification (pAMR), circulating anti-HLA donor-specific antibodies (DSA) 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 heterogenous 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, independently of the C4d or CD68 status ( Fig. 1 , P < 0.001). Both the frequency and mean fluorescence intensity of DSA 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.

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.

Initiation of noninvasive surveillance for allograft rejection in heart transplant patients>1 year after transplant.

Gene expression profiling (GEP) and donor-derived, cell-free DNA (dd-cfDNA) measurement are alternative methods to endomyocardial biopsy (EMB) to monitor for rejection following heart transplantation. We aim to describe our use of GEP and dd-cfDNA in heart transplant recipients>1-year post-transplantation. This is a single-center, retrospective study in post-transplant recipients. For patients who were>1-year post-transplantation and deemed to be at elevated clinical risk for rejection, we collected both GEP and dd-cfDNA every 3 months. Baseline characteristics including GEP, dd-cfDNA levels, rejection episodes, and number of biopsies were obtained. Since July 2019, there were 18 patients being followed with GEP and dd-cfDNA who were>1-year post-transplantation. Nine EMBs had been performed in seven patients due to as follows; three due to elevated GEP ({greater than or equal to} 34), one due to elevated dd-cfDNA ({greater than or equal to} .20%), two due to elevations of both GEP and dd-cfDNA, two due to clinical rejection and one to follow up a post rejection episode. One of the two biopsies due to elevations of both GEP and dd-cfDNA showed acute cellular rejection grade 2R. None of the biopsies due to either an elevation in the GEP or dd-cfDNA revealed any significant rejection. In this study, the use of both GEP and dd-cfDNA led to an increased number of EMB in patients>1-year post-transplantation. Further studies are needed to validate these findings and evaluate long-term consequences of these diagnostic tests in this population.

Abstract 11935: The Index of Microcirculatory Resistance to Predict Subsequent Acute Cellular Rejection and Cardiac Events After Heart Transplantation

Introduction: Single center data suggest that the index of microcirculatory resistance (IMR) measured early after heart transplantation predicts subsequent acute cellular rejection. Hypothesis: To validate whether IMR measured early after transplantation can predict subsequent acute cellular rejection and long-term outcome in a large multicenter cohort. Methods: From 5 international heart transplantation cohorts, 237 patients who underwent IMR measurement early after transplantation were enrolled. The primary outcome was biopsy-proven acute cellular rejection of grade 2 or 3 within 1 year after transplantation. A key secondary outcome was major adverse cardiac events (MACE, the composite of death, re-transplantation, myocardial infarction, stroke, graft dysfunction, and readmission due to a cardiac cause) at 10 years. Results: IMR was measured at a median of 7 weeks (interquartile range: 3 to 10 weeks) post-transplantation. Patients with acute cellular rejection grade 2 or 3 at the time of IMR assessment were excluded. At 1 year, the incidence of acute cellular rejection was 14.0%. IMR was associated proportionally with the risk of acute cellular rejection (per increase of 1-unit IMR; adjusted hazard ratio [aHR], 1.04; 95% confidence interval [CI], 1.01-1.06; P=0.002). The incidence of acute cellular rejection in patients with IMR ≥ 18 was 23.8%, while the incidence of acute cellular rejection in those with IMR &lt; 18 was 5.5% (aHR, 4.09; 95% CI, 1.67-10.0; P=0.002). At 10 years, MACE occurred in 86 (36.3%) patients. IMR was significantly associated with the risk of MACE (per increase of 1-unit IMR; aHR, 1.03; 95% CI, 1.01-1.05; P=0.002). Conclusions: IMR measured early after heart transplantation is associated with subsequent acute cellular rejection at 1 year and clinical events at 10 years. Early IMR measurement after transplantation identifies patients at higher risk and may guide personalized post-transplantation management.

Sarcomere ACTC1 and MYO18A serum expression levels detect acute cellular rejection in cardiac transplant patients

Abstract Introduction and purpose The development of non-invasive approaches for the early diagnosis of cardiac allograft rejection is necessary. Given the central role of sarcomeric dysfunction in cardiomyocyte biology and sarcomere alterations, described in endomyocardial biopsies of transplant patients with rejection, we hypothesized that the serum expression levels of genes encoding sarcomeric proteins were altered in acute cellular rejection (ACR). Methods Forty consecutive serum samples from transplant recipients undergoing routine endomyocardial biopsies were included in an RNA sequencing analysis. Results We identified 61 sarcomeric genes, 19 of which were differentially expressed in patients with clinically relevant rejection (ACR grade ≥2R). A receiver operating characteristic curve was done to assess their accuracy for ACR detection, and found that 8 relevant actins, myosins, and other sarcomere-related genes showed great diagnostic capacity (AUC≥0.800); ACTC1 (AUC=1.000, p&amp;lt;0.0001) and MYO18A (AUC=1.000, p&amp;lt;0.0001) showed the best results. MYO18A had a good ability to detect mild rejection (AUC=0.789, p&amp;lt;0.05). We found a relationship between MYO18A and the left ventricular end-systolic (p&amp;lt;0.01) and end-diastolic (p&amp;lt;0.05) diameters. Conclusions ACTC1 and MYO18A also showed a significant correlation with the NT-proBNP levels (p&amp;lt;0.05). Because of their precision to detect ACR, we propose sarcomere ACTC1 and MYO18A serum expression levels as potential candidates for non-invasive ACR detection. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): National Institute of Health Carlos IIIEuropean Regional Development Fund (ERDF)