Proteomics in Acute Heart Transplant Rejection, On Behalf of the GRAfT Investigators.

Background: Proteomic phenotyping can provide insights into rejection pathophysiology, novel biomarkers, and therapeutic targets.

Methods: Within the prospective, multicenter Genomic Research Alliance for Transplantation study, 181 proteins were evaluated from blood drawn at the time of endomyocardial biopsy; protein fold change, logistic regression, and pathway analyses were conducted, with protein discovery adjusted for a 5% false discovery rate.

Results: Among 104 adult heart transplant patients (31% female sex, 53% Black race, median age 52 y), 74 had no rejection, 18 developed acute cellular rejection (ACR), and 12 developed antibody-mediated rejection (AMR).

Pathologist interrater reliability and clinical implications of elevated donor-derived cell-free DNA beyond heart transplant rejection, on behalf of the GRAfT investigators.

Background: There is significant variability among pathologists in the histopathological interpretation of the endomyocardial biopsy (EMB) for acute cellular rejection (ACR), and assessment of variability in the interpretation of antibody-mediated rejection (AMR) has not been reported. In contemporary practice, the strategy of allograft surveillance with donor-derived cell-free DNA (dd-cfDNA) compared to EMB has not been compared with a focus on long-term clinical outcomes beyond acute rejection (AR).

Methods: The Genomic Research Alliance for Transplantation is a multicenter, prospective cohort study that enrolled patients from 2015 to 2020.

European Society for Organ Transplantation (ESOT) Consensus Statement on the Use of Non-invasive Biomarkers for Cardiothoracic Transplant Rejection Surveillance.

While allograft rejection (AR) continues to threaten the success of cardiothoracic transplantation, lack of accurate and repeatable surveillance tools to diagnose AR is a major unmet need in the clinical management of cardiothoracic transplant recipients. Endomyocardial biopsy (EMB) and transbronchial biopsy (TBBx) have been the cornerstone of rejection monitoring since the field's incipience, but both suffer from significant limitations, including poor concordance of biopsy interpretation among pathologists. In recent years, novel molecular tools for AR monitoring have emerged and their performance characteristics have been evaluated in multiple studies.

Baseline Lung Allograft Dysfunction After Bilateral Lung Transplantation Is Associated With an Increased Risk of Death: Results From a Multicenter Cohort Study.

Background: A prior single-center, retrospective cohort study identified baseline lung allograft dysfunction (BLAD) as a risk factor for death in bilateral lung transplant recipients. In this multicenter prospective cohort study, we test the association of BLAD with death in bilateral lung transplant recipients, identify clinical risk factors for BLAD, and assess its association with allograft injury on the molecular level.

Methods: This multicenter, prospective cohort study included 173 bilateral lung transplant recipients that underwent serial pulmonary function testing and plasma collection for donor-derived cell-free DNA at prespecified time points.

Extreme elevations of donor-derived cell-free DNA increases the risk of chronic lung allograft dysfunction and death, even without clinical manifestations of disease.

Background: Lung transplant recipients are traditionally monitored with pulmonary function testing (PFT) and lung biopsy to detect post-transplant complications and guide treatment. Plasma donor-derived cell free DNA (dd-cfDNA) is a novel molecular approach of assessing allograft injury, including subclinical allograft dysfunction. The aim of this study was to determine if episodes of extreme molecular injury (EMI) in lung transplant recipients increases the risk of chronic lung allograft dysfunction (CLAD) or death.

Sex-specific patterns of donor-derived cell-free DNA in heart transplant rejection: An analysis from the Genomic Research Alliance for Transplantation (GRAfT).

Background: Noninvasive methods for surveillance of acute rejection are increasingly used in heart transplantation (HT), including donor-derived cell-free DNA (dd-cfDNA). As other cardiac biomarkers differ by sex, we hypothesized that there may be sex-specific differences in the performance of dd-cfDNA for the detection of acute rejection. The purpose of the current study was to examine patterns of dd-cfDNA seen in quiescence and acute rejection in male and female transplant recipients.

Racial Differences in Donor-Derived Cell-Free DNA and Mitochondrial DNA After Heart Transplantation, on Behalf of the GRAfT Investigators.

Background: Black heart transplant patients are at higher risk of acute rejection (AR) and death than White patients. We hypothesized that this risk may be associated with higher levels of donor-derived cell-free DNA (dd-cfDNA) and cell-free mitochondrial DNA.

Methods: The Genomic Research Alliance for Transplantation is a multicenter, prospective, longitudinal cohort study.

Emergency Myelopoiesis Distinguishes Multisystem Inflammatory Syndrome in Children From Pediatric Severe Coronavirus Disease 2019.

Background: Multisystem inflammatory syndrome in children (MIS-C) is a hyperinflammatory condition caused by recent infection with severe acute respiratory syndrome coronavirus 2, but the underlying immunological mechanisms driving this distinct syndrome are unknown.

Methods: We utilized high-dimensional flow cytometry, cell-free (cf) DNA, and cytokine and chemokine profiling to identify mechanisms of critical illness distinguishing MIS-C from severe acute coronavirus disease 2019 (SAC).

Results: Compared to SAC, MIS-C patients demonstrated profound innate immune cell death and features of emergency myelopoiesis (EM), an understudied phenomenon observed in severe inflammation.

Cell-Free DNA Maps Tissue Injury and Correlates with Disease Severity in Lung Transplant Candidates.

Plasma cell-free DNA levels correlate with disease severity in many conditions. Pretransplant cell-free DNA may risk stratify lung transplant candidates for post-transplant complications. To evaluate if pretransplant cell-free DNA levels and tissue sources identify patients at high risk of primary graft dysfunction and other pre- and post-transplant outcomes.

Chronic graft-versus-host disease is characterized by high levels and distinctive tissue-of-origin patterns of cell-free DNA.

Chronic graft-versus-host disease (cGvHD) is a devastating complication of hematopoietic stem cell transplantation (HSCT). Effective early detection may improve the outcome of cGvHD. The potential utility of circulating cell-free DNA (cfDNA), a sensitive marker for tissue injury, in HSCT and cGvHD remains to be established.

Organizing pneumonia is associated with molecular allograft injury and the development of antibody-mediated rejection.

Background: The association between organizing pneumonia (OP) after lung transplantation with the development of acute rejection (AR) remains undefined. In addition, molecular allograft injury, as measured by donor-derived cell-free DNA (dd-cfDNA), during episodes of OP and its relationship to episodes of AR, chronic lung allograft dysfunction (CLAD), or death is unknown.

Methods: This multicenter, prospective cohort study collected serial plasma samples from 188 lung transplant recipients for dd-cfDNA at the time of bronchoscopy with biopsy.

Cell-free chromatin immunoprecipitation to detect molecular pathways in heart transplantation.

Existing monitoring approaches in heart transplantation lack the sensitivity to provide deep molecular assessments to guide management, or require endomyocardial biopsy, an invasive and blind procedure that lacks the precision to reliably obtain biopsy samples from diseased sites. This study examined plasma cell-free DNA chromatin immunoprecipitation sequencing (cfChIP-seq) as a noninvasive proxy to define molecular gene sets and sources of tissue injury in heart transplant patients. In healthy controls and in heart transplant patients, cfChIP-seq reliably detected housekeeping genes.

Cell-free DNA reveals distinct pathology of multisystem inflammatory syndrome in children.

Multisystem inflammatory syndrome in children (MIS-C) is a rare but life-threatening hyperinflammatory condition induced by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that causes pediatric COVID-19 (pCOVID-19). The relationship of the systemic tissue injury to the pathophysiology of MIS-C is poorly defined. We leveraged the high sensitivity of epigenomics analyses of plasma cell-free DNA (cfDNA) and plasma cytokine measurements to identify the spectrum of tissue injury and glean mechanistic insights.

Selection and Interpretation of Molecular Diagnostics in Heart Transplantation.

The number of heart transplants performed annually in the United States and worldwide continues to increase, but there has been little change in graft longevity and patient survival over the past 2 decades. The reference standard for diagnosis of acute cellular and antibody-mediated rejection includes histologic and immunofluorescence evaluation of endomyocardial biopsy samples, despite invasiveness and high interrater variability for grading histologic rejection. Circulating biomarkers and molecular diagnostics have shown substantial predictive value in rejection monitoring, and emerging data support their use in diagnosing other posttransplant complications.

Pseudomonas-dominant microbiome elicits sustained IL-1β upregulation in alveolar macrophages from lung transplant recipients.

Background: Isolation of Pseudomonas aeruginosa (PsA) is associated with increased BAL (bronchoalveolar lavage) inflammation and lung allograft injury in lung transplant recipients (LTR). However, the effect of PsA on macrophage responses in this population is incompletely understood. We examined human alveolar macrophage (AMΦ) responses to PsA and Pseudomonas dominant microbiome in healthy LTR.

Cell-free Chromatin Immunoprecipitation to detect molecular pathways in Physiological and Disease States.

Patient monitoring is a cornerstone in clinical practice to define disease phenotypes and guide clinical management. Unfortunately, this is often reliant on invasive and/or less sensitive methods that do not provide deep phenotype assessments of disease state to guide treatment. This paper examined plasma cell-free DNA chromatin immunoprecipitation sequencing (cfChIP-seq) to define molecular gene sets in physiological and heart transplant patients taking immunosuppression medications.

Preemptive treatment of de novo donor-specific antibodies in lung transplant patients reduces subsequent risk of chronic lung allograft dysfunction or death.

The development of donor-specific antibodies after lung transplantation is associated with downstream acute cellular rejection, antibody-mediated rejection (AMR), chronic lung allograft dysfunction (CLAD), or death. It is unknown whether preemptive (early) treatment of de novo donor-specific antibodies (dnDSAs), in the absence of clinical signs and symptoms of allograft dysfunction, reduces the risk of subsequent CLAD or death. We performed a multicenter, retrospective cohort study to determine if early treatment of dnDSAs in lung transplant patients reduces the risk of the composite endpoint of CLAD or death.

Clinical features and allograft failure rates of pulmonary antibody-mediated rejection categories.

Background: Pulmonary antibody-mediated rejection (AMR) consensus criteria categorize AMR by diagnostic certainty. This study aims to define the clinical features and associated outcomes of these recently defined AMR categories.

Methods: Adjudication committees reviewed clinical data of 335 lung transplant recipients to define clinical or subclinical AMR based on the presence of allograft dysfunction, and the primary endpoints, time from transplant to allograft failure, a composite endpoint of chronic lung allograft dysfunction and/or death.

Plasma Cell-Free DNA Predicts Survival and Maps Specific Sources of Injury in Pulmonary Arterial Hypertension.

Background: Cell-free DNA (cfDNA) is a noninvasive marker of cellular injury. Its significance in pulmonary arterial hypertension (PAH) is unknown.

Methods: Plasma cfDNA was measured in 2 PAH cohorts (A, n=48; B, n=161) and controls (n=48).