Peripheral Blood RNA Sequencing Unravels a Differential Signature of Coding and Noncoding Genes by Types of Kidney Allograft Rejection

Introduction Peripheral blood (PB) molecular patterns characterizing the different effector immune pathways driving distinct kidney rejection types remain to be fully elucidated. We hypothesized that transcriptome analysis using RNA sequencing (RNAseq) in samples of kidney transplant patients would enable the identification of unique protein-coding and non-coding genes that may be able to segregate different rejection phenotypes. Methods We evaluated 37 biopsy-paired PB samples from the discovery cohort, with stable [STA], antibody-mediated rejection [AMR] and T-cell mediated rejection [TCMR], by RNAseq. Advanced machine learning tools were used to perform three-way differential gene expression analysis to identify gene signatures associated with rejection. Then, we performed functional in silico analysis and validation by Fluidigm in 62 samples from two independent kidney transplant cohorts. Results We found 102 genes (63 coding and 39 non-coding genes) associated with AMR (54 up-regulated), TCMR (23 up-regulated) and STA (25 up-regulated) perfectly clustered with each rejection phenotype and highly correlated with main histological lesions (rho = 0.91). For the genes associated with AMR, we found enrichment in regulation of endoplasmic reticulum stress, adaptive immunity, and immunoglobulin class-switching. In the validation, we found that, the SIGLEC17P pseudogene and 9 SIGLEC17P-related coding genes, were highly expressed among AMR but not in TCMR and STA samples. Conclusion This analysis identifies a critical gene signature in peripheral blood in kidney transplant patients undergoing AMR, sufficient to differentiate them from patients with TCMR and immunologically quiescent kidney allografts. Our findings provide the basis for new studies dissecting the role of non-coding genes in the pathophysiology of kidney allograft rejection and their potential value as non-invasive biomarkers of the rejection process.