Date: Sunday, May 3, 2015
Session Time: 5:30pm-6:30pm
Presentation Time: 5:30pm-6:30pm
Location: Exhibit Hall E
Introduction:Circulating donor-derived cell-free DNA (dd-cfDNA) has been reported as a surrogate of transplant injury in the blood but requires expensive donor and recipient DNA sequencing.
Method: We have developed a rapid and economical approach for measuring urine cfDNA in renal transplant recipients without the need for DNA sequencing using a PCR based approach with customized primer design for quantification of dd-cfDNA. Biopsy matched urine and blood samples 292 urine and 40 plasma samples were assessed for dd-cfDNA. cfDNA was quantified as copy numbers of loci of Chromosome Y, and customized primer sets PS1 and PS2 by digital QPCR.
Result: Dd-cfDNA measured in terms of Chromosome Y in female recipients of male donor organs, was highly correlated to cfDNA as measured by the PS1 primer pair (r=0.96; p<0.001), confirming that the major burden of urinary dd-cfDNA can be detected by PS1 and PS2 primers that are not restricted to localization on the Y chromosome, and thus can detect dd-cfDNA irrespective of gender donor/recipient pairing. Dd-cfDNA load was elevated in urine during different triggers of acute transplant injury, inclusive of biopsy confirmed BK virus nephritis, acute tubular necrosis and acute rejection episodes. There was significantly greater load of PS1/PS2 specific urine dd-cfDNA (4.87±1.22 copies/¯o;g urine creatinine compared to 1.36±1.94 copies/¯o;g urine creatinine in patients without graft dysfunction; p<0.0001). There was a significant increase of cfDNA in the urine collected at acute rejection episode compared to stable graft (10.55±25.43 vs. 1.14±2.05 copies/mg urine creatinine p=0.002) (median for STA=0.26 copies/mg urine creatinine vs median for AR 1.55 copies/mg urine creatinine). The increase in urinary dd-cfDNA in case of renal transplant injury due to other causes of injury such as polyoma virus nephropathy, pyelonephritis, FSGS, and ATN was also significant (13.48±28.27 copies/mg urine creatinine vs 1.11±2.03 copies/mg urine creatinine, p=0.001). A ROC analysis of the data resulted in an AUC of 0.78 with a p value <0.0001. At the threshold of <4.06 copies/mg urine creatinine the PS1/PS2 dd-cfDNA assay provides a specificity of 94% for ruling out any acute transplant injuy.
Conclusion: We have developed and validated a novel methodology for quantitative assessment of urinary dd-cfDNA in renal transplant recipients, without the expense and labor of DNA sequencing, providing a rapid screening outpatient assay for monitoring for acute transplant injury.
To cite this abstract in AMA style:Sigdel T, Tran T, Hsieh S-C, Sarwal M. Urinary Cell-Free DNA Is a Sensitive Marker of Early Renal Transplant Injury [abstract]. Am J Transplant. 2015; 15 (suppl 3). https://atcmeetingabstracts.com/abstract/urinary-cell-free-dna-is-a-sensitive-marker-of-early-renal-transplant-injury/. Accessed November 27, 2020.
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