A Simple Molecular Tool for the Assessment of Kidney Transplant Biopsies.

Background The Banff Classification for Allograft Pathology recommendations for the diagnosis of kidney transplant rejection includes molecular assessment of the transplant biopsy. However, implementation of molecular tools in clinical practice is still limited, partly due to the required expertise and financial investment. The reverse transcriptase multiplex ligation-dependent probe amplification (RT-MLPA) assay is a simple, rapid and inexpensive assay that permits simultaneous evaluation of a restricted gene panel using paraffin-embedded tissue blocks. The aim of this study was to develop and validate a RT-MLPA assay for diagnosis and classification of rejection. Methods A retrospective cohort of 220 kidney transplant biopsies from two centres, which included with 52 antibody-mediated rejection, 51 T cell-mediated rejection and 117 no-rejection controls was assessed. A 17-gene panel was identified, based on relevant pathophysiological pathways. A support vector machine classifier was developed. A subset of 109 biopsies was also assessed using the Nanostring B-HOT panel to compare the two assays. Results The support vector machine classifier train and test accuracy scores were 0.84 and 0.83, respectively. In the test cohort, the F1-score for antibody mediated rejection, T cell-mediated rejection and control were 0.88, 0.86 and 0.69, respectively. Using ROC curves, AUC for class predictions were 0.96, 0.89 and 0.91, respectively, with a weight-average at 0.94. Classifiers' performances were highest for antibody mediated rejection diagnosis with 94% correct predictions, compared to 88% correct predictions for control biopsies and 60% for T-cell mediated rejection biopsies. Gene expression levels assessed by RT-MLPA or Nanostring correlated: r = 0.68, p < 0.001, and gene expression profiles obtained with these assays were comparable in 81% of samples. Conclusions The 17-gene panel RT-MLPA assay, developed here for formalin-fixed paraffin-embedded kidney transplant biopsies, classifies kidney transplant rejection with an overall accurate prediction ratio of 0.83.