Combining Blood Gene Expression and Cellfree DNA to Diagnose Subclinical Rejection in Kidney Transplant Recipients

Background and objectivesSubclinical acute rejection is associated with poor outcomes in kidney transplant recipients. As an alternative to surveillance biopsies, noninvasive screening has been established with a blood gene expression profile. Donor-derived cellfree DNA (cfDNA) has been used to detect rejection in patients with allograft dysfunction but not tested extensively in stable patients. We hypothesized that we could complement noninvasive diagnostic performance for subclinical rejection by combining a donor-derived cfDNA and a gene expression profile assay.Design, setting, participants, & measurementsWe performed a post hoc analysis of simultaneous blood gene expression profile and donor-derived cfDNA assays in 428 samples paired with surveillance biopsies from 208 subjects enrolled in an observational clinical trial (Clinical Trials in Organ Transplantation-08). Assay results were analyzed as binary variables, and then, their continuous scores were combined using logistic regression. The performance of each assay alone and in combination was compared.ResultsFor diagnosing subclinical rejection, the gene expression profile demonstrated a negative predictive value of 82%, a positive predictive value of 47%, a balanced accuracy of 64%, and an area under the receiver operating curve of 0.75. The donor-derived cfDNA assay showed similar negative predictive value (84%), positive predictive value (56%), balanced accuracy (68%), and area under the receiver operating curve (0.72). When both assays were negative, negative predictive value increased to 88%. When both assays were positive, positive predictive value increased to 81%. Combining assays using multivariable logistic regression, area under the receiver operating curve was 0.81, significantly higher than the gene expression profile (P<0.001) or donor-derived cfDNA alone (P=0.006). Notably, when cases were separated on the basis of rejection type, the gene expression profile was significantly better at detecting cellular rejection (area under the receiver operating curve, 0.80 versus 0.62; P=0.001), whereas the donor-derived cfDNA was significantly better at detecting antibody-mediated rejection (area under the receiver operating curve, 0.84 versus 0.71; P=0.003).ConclusionsA combination of blood-based biomarkers can improve detection and provide less invasive monitoring for subclinical rejection. In this study, the gene expression profile detected more cellular rejection, whereas donor-derived cfDNA detected more antibody-mediated rejection.

Subclinical Rejection and Immunosuppression in Pediatric Kidney Transplant Recipients : Single Centre Study

Background: By the time of histological confirmation of rejection is achieved, renal scarring may for treatment as a realistic option . This study aims to study the subclinical pathological graft data and to evaluate the histopathological impact of different immunosuppression protocols in pediatric renal transplant recipients. Methods: This is a case series that included twenty living donor renal transplant recipients. All included cases received the classic triple immunotherapy for at least one month post-transplantation [Steroids, calconurine inhibitors (CNI), and mycofenlolic mofetile (MMF)]. Based on their immunological risk stratification; included cases were divided into 2 groups: group (A) continued on CNI based triple therapy protocol; group (B) shifted to evirolimus /low dose CNI protocol. Surveillance biopsies were done for all cases at one and four month post-transplantation. Results: One and four month biopsies revealed subclinical rejection (including borderline changes) in 4 (20%) cases and 6 (30%) cases respectively. The number of patients received tacrolimus/MMF therapy significantly increased (p=0.02) while that of patients on everloimus/low dose CNI significantly decreased (p=0.014) due to drug modifications based on four month surveillance biopsy data. Conclusion: Subclinical rejection is not uncommon in pediatric renal graft recipients which makes surveillance biopsy might be of help. Early usage of evirolimus/low CNI protocol is associated with higher rejection rate than triple therapy.

MO937ASSOCIATION OF RENAL ELASTOGRAPHY WITH BANFF MICROVASCULAR INFLAMMATION IN PROTOCOL BIOPSIES FOR SUBCLINICAL RENAL GRAFT REJECTION DIAGNOSIS

Background and Aims The most common cause of renal graft failure is chronic dysfunction in 24.7% and the most common etiology of this is clinical or subclinical rejection. The incidence of subclinical rejection varies from 15 to 50% (25% in protocol biopsies in the first year after transplantation and 35% after two years). Melek E et al have shown that doppler ultrasound is a non-invasive study that, through the resistance index (RI), has traditionally been used for the early diagnosis of acute graft rejection (AR); however, it is influenced by extrarenal systemic factors. Naesens et al published that in 321 kidney transplant recipients, RI wasn´t associated with histological findings of AR in protocol biopsies. Elastography is another ultrasonographic modality for the evaluation of the kidney graft, which measures the stiffness/elasticity of the tissue expressed in Kpa (kilopascals). Stock in 2011 and Kim BJ in 2018 published studies where they showed that increased stiffness was correlated with the diagnosis of kidney graft rejection. The aim of this study was to describe the association between elastography with microvascular inflammation determined by Banff for diagnosis of renal allograft subclinical rejection. Method Observational, analytical and cross-sectional study that included kidney transplant patients who underwent protocol biopsy and renal elastography at the Central Military Hospital in Mexico City between January 2018 and December 2020. The demographic and biochemical characteristics, degree elastography stiffness and Banff 2017 lesions were determined. The sample calculation, determination of correlation degree and ROC curve elaboration were performed. Results We included 146 patients. 56.8% were men; the most common causes of CKD were undetermined and chronic glomerulonephritis with 52.7% and 17.1% respectively. 47.3% were hypertensive at biopsy time and 1.4% had chronic heart failure. The most common immunosuppression schemes were FK/MPA/steroid and FK/mTOR-i/steroid with 60.3% and 13%, respectively. The mean GFR was 65.31 ml/min which shows graft good function. The mean stiffness in the elastography was 15.73 Kpa. The rest of baseline data are shown in Table 1. Had rejection 36.3% of the biopsies, the most frequent chronic AMR C4d- with 15.1% and active AMR C4d- 8.9%. When analyzing the ROC curves, the Banff 2017 lesions AUC values that correlated better with graft stiffness were: v=0.607, i=0.594, g=0.578, C4d deposit=0.519, ptc=0.498. Figure 1. Conclusion Intimal arteritis, inflammation, and glomerulitis are the Banff lesions best associated with elastography graft stiffness in protocol biopsies. Prospective studies are recommended in patients with acute graft dysfunction to find an adequate elastography cut-off value that allows another tool for fast and non-invasive diagnosis of renal graft rejection.

A Rejection Gene Expression Score in Indication and Surveillance Biopsies Is Associated with Graft Outcome

Rejection-associated gene expression has been characterized in renal allograft biopsies for cause. The aim is to evaluate rejection gene expression in subclinical rejection and in biopsies with borderline changes or interstitial fibrosis and tubular atrophy (IFTA). We included 96 biopsies. Most differentially expressed genes between normal surveillance biopsies (n = 17) and clinical rejection (n = 12) were obtained. A rejection-associated gene (RAG) score was defined as its geometric mean. The following groups were considered: (a) subclinical rejection (REJ-S, n = 6); (b) borderline changes in biopsies for cause (BL-C, n = 13); (c) borderline changes in surveillance biopsies (BL-S, n = 12); (d) IFTA in biopsies for cause (IFTA-C, n = 20); and (e) IFTA in surveillance biopsies (IFTA-S, n = 16). The outcome variable was death-censored graft loss or glomerular filtration rate decline ≥ 30 % at 2 years. A RAG score containing 109 genes derived from normal and clinical rejection (area under the curve, AUC = 1) was employed to classify the study groups. A positive RAG score was observed in 83% REJ-S, 38% BL-C, 17% BL-S, 25% IFTA-C, and 5% IFTA-S. A positive RAG score was an independent predictor of graft outcome from histological diagnosis (hazard ratio: 3.5 and 95% confidence interval: 1.1–10.9; p = 0.031). A positive RAG score predicts graft outcome in surveillance and for cause biopsies with a less severe phenotype than clinical rejection.