Considerable effort has focused on identification of gene expression patterns that may assist in diagnosis of acute cellular rejection. The NanoString platform offers a novel approach to directly quantitate hundreds of mRNAs in a single assay. As an initial exploration of NanoString in diagnosing rejection, we analyzed the expression of 795 genes in peripheral blood and urine sediment RNA prepared from 17 renal transplant subjects enrolled in the CTOT16 study. Samples were obtained from 7 subjects at the time of biopsy-proven acute Banff grade I or II rejection and twenty-three samples from 10 subjects with stable graft function and no evidence of rejection. All of the peripheral blood RNA samples were of high quality whereas only 6 of the 7 urine RNA samples were deemed of sufficient quality for analysis. We compared gene expression levels in each individual acute rejection sample to that of the average expression of each gene in all of the control samples to identify genes that changed at least 2-fold in every rejecting subject. In peripheral blood RNA, a range of 63-239 genes was upregulated >2-fold in the 7 samples from individuals experiencing rejection and a range of 7-298 genes was downregulated. However, there were no common genes that increased or decreased >2-fold in all of the 7 rejection peripheral blood samples. In individual urine sediment RNA samples from patients experiencing rejection, expression of a range of 250-720 genes was increased >2-fold, 86-590 genes increased >5-fold, and 30-370 genes increased >10 fold when compared to expression of the genes in urine sediment from controls. Importantly, expression of 164 of these genes was increased >2-fold and expression of 18 was increased >5-fold in all of the rejection patient urine samples. The common genes in both sets included those involved in apoptosis, T cell activation and inflammatory pathways. No genes were observed to be down-regulated in urine RNA samples at the time of acute rejection vs. controls. The results of this initial study indicate the power of the NanoString platform to measure gene expression changes in the urine that are associated with ongoing acute rejection in kidney transplants. These changes are not as substantially evident when measured in peripheral blood. This approach has the potential to develop a multi-gene signature for non-invasive diagnosis of rejection.

CITATION INFORMATION: Keslar K, Zmijewska A, Andriga J, Newell K, Mannon R, Heeger P, Fairchild R. NanoString Detection of Gene Signatures in Urine During Acute Cellular Rejection of Kidney Grafts. Am J Transplant. 2016;16 (suppl 3).

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