<p>Circulating nucleic acids as biomarkers for allograft injury after solid organ transplantation: current state-of-the-art</p>

Abstract

Immune-mediated injury of a transplanted organ can lead to allograft dysfunction and even patient death. Acute cellular rejection typically occurs within the first months post-transplantation but patients are at life-time risk, particularly if there is medication non-compliance or reduction of immunosuppression due to complications. Therefore, safe and accurate monitoring of the donated organ for signs of rejection is essential for long-term survival of the transplanted organ and recipient. The current gold standard for rejection surveillance is through tissue biopsy and histology, which is costly, invasive, and subjective. Thus, efforts to develop non-invasive methods for the detection of rejection post-transplantation are a priority in the field. The first FDA-approved noninvasive assay, AlloMap, was developed in 2006 and monitored the peripheral expression of 11 genes associated with immune system activation. More recently, there has been a shift towards interrogating the status of the transplanted organ directly. Fragments of genomic DNA are released into the blood during cellular apoptosis and levels of cell-free DNA (cfDNA) have been shown to be elevated in the presence of organ injury, including after transplantation. Since the genomic characteristics of DNA are maintained in cfDNA (eg, sequence variants), this circulating molecule represents a promising organ-specific biomarker for allograft injury. DNA sequence variants have been used to distinguish donor and recipient cfDNA with or without a priori donor genotyping in a variety of solid organs post-transplant. Current research has established the groundwork and future multicenter trials will determine if this novel molecular diagnostic tool represents a viable alternative to tissue biopsy. Other nucleic acid molecules released from the transplanted organ (eg, microRNAs) are presently less well developed in comparison to cfDNA but may also represent potential novel biomarkers. This review summarizes current literature and evaluates the promises and pitfalls of circulating nucleic acids as biomarkers for allograft injury post-transplant.

Advances in personalized medicine and noninvasive diagnostics in solid organ transplantation

“…The dd‐cfDNA has a short half‐life of only 30 minutes providing clinicians with insight to potential allograft damage occurring in real time. Various methods including the “two genomes” model, the “single genome” method, quantitative polymerase chain reaction PCR (qPCR), droplet digital PCR (ddPCR), and next generation sequencing (NGS) can be used to assess presence of dd‐cfDNA 30 . There are currently three commercially available assays in the United States that use NGS to detect dd‐cfDNA in the blood of transplants recipients ‐ AlloSure (CareDx), TRAC (Viracor Eurofins), and Prospera (Natera).…”

The Use of Donor-Derived Cell-Free DNA for Assessment of Allograft Rejection and Injury Status

“…Following organ transplantation, dd-cfDNA circulates in the recipient's blood, and accounts for a relatively small fraction of total cfDNA (recipient plus donor derived) [46]. As cells from the donor allograft degrade, the nucleic acids within become fragmented, resulting in approximately 120-160 base pair pieces of double-stranded dd-cfDNA released into the blood, and cleared from blood by the liver and kidney with half-life of about 30 min [47]. The mechanisms of release of cfDNA into the bloodstream is believed to be a result of several possible mechanisms, including cell death by apoptosis or necrosis, in addition to active secretion by various activated cells of the immune system [48][49][50][51][52].…”

“…dd-cfDNA is typically low in concentration, only a few thousand genomic copies/mL [53], and dd-cfDNA is usually <1% of the total cell-free DNA when there is no active damage to the allograft [46]. However, during allograft rejection, significantly higher amounts of dd-cfDNA are released from the injured allograft into the bloodstream [46,47]. Early rises of total dd-cfDNA levels during acute rejection have been observed in KT recipients [48,54].…”

Identification of Cell-Free DNA Methylation Patterns Unique to the Human Left Ventricle as a Potential Indicator of Acute Cellular Rejection

“…Plasma cell-free DNA (cfDNA) has shown promise as a minimally-invasive biomarker for the detection of rejection after solid organ transplantation although many questions regarding the production and clearance of cfDNA remain unanswered. [1][2][3] To date, the identification of donor-derived cfDNA has been based on the quantification of polymorphic single nucleotide polymorphisms (SNPs) that differ between the donor and recipient with or without donor genotyping. [4][5][6] While the requirement for a priori donor genotyping has limitations associated with cost and feasibility, especially in situations where the donor is deceased, alternative methods are subject to assumptions and estimations of genotype frequencies that may not accurately represent the donor.…”