Universal noninvasive detection of solid organ transplant rejection

Abstract

It is challenging to monitor the health of transplanted organs, particularly with respect to rejection by the host immune system. Because transplanted organs have genomes that are distinct from the recipient's genome, we used high throughput shotgun sequencing to develop a universal noninvasive approach to monitoring organ health. We analyzed cell-free DNA circulating in the blood of heart transplant recipients and observed significantly increased levels of cell-free DNA from the donor genome at times when an endomyocardial biopsy independently established the presence of acute cellular rejection in these heart transplant recipients. Our results demonstrate that cell-free DNA can be used to detect an organ-specific signature that correlates with rejection, and this measurement can be made on any combination of donor and recipient. This noninvasive test holds promise for replacing the endomyocardial biopsy in heart transplant recipients and may be applicable to other solid organ transplants.next-generation sequencing | noninvasive diagnosis | acute rejection

Extracellular RNA profiles with human age

“…The discovery of circulating cell‐free DNA (extracellular DNA; exDNA) and RNA (extracellular RNA; exRNA) in human body fluids, including serum, has sparked great interest in whether these molecules are functional, have regulatory effects, or can be used as markers of disease. For example, exDNA found in maternal serum has led to significant advancements in prenatal testing and diagnosis (Chiu & Lo, 2013; Fan, Blumenfeld, Chitkara, Hudgins & Quake, 2008; Fan et al., 2012; Lo et al., 1997), and for prediction of heart transplant rejection in adults (De Vlaminck et al., 2014; Snyder, Khush, Valantine & Quake, 2011). …”

Cell-Free DNA: An Upcoming Biomarker in Transplantation

“…In the past 20 years, ddcfDNA quantification techniques have been investigated which differ in terms of applicability, sensitivity, turnaround times, and instrumentation costs ( Table 2). While amplification of Y-chromosome genes in the recipient's plasma and urine has been widely used in gender-mismatched transplantations (21,22,30,31,(34)(35)(36)(37)(38)(39), this strategy is limited to about 25% of the transplant population. A more universal approach is the use of informative genetic polymorphisms whereby a particular allele differs or is absent in the donor genome compared to the recipient genome; in addition to the quantification of donor-specific human leukocyte antigen deoxyribonucleic acid (HLA DNA) (40) and copy number deletion polymorphisms (41), single nucleotide polymorphisms (SNPs) can be targeted to determine the percentage of ddcfDNA by counting donor and recipient's bases at informative SNP sites with digital droplet polymerase chain reaction (ddPCR) (42)(43)(44) or massive parallel shotgun sequencing (MPSS) (34,45).…”

“…Low fraction of total cfDNA (0.06-3.5%) (34,42,45) qPCR or ddPCR of Y-chromosome genes (22,34,(37)(38)(39) (56) cfDNA isolation:…”

Numerous uncharacterized and highly divergent microbes which colonize humans are revealed by circulating cell-free DNA

“…The existence of circulating nucleic acids in blood has been known since the mid-20th century (10), but only in the last few years has the advent of high-throughput sequencing led to clinical diagnostics based on these nucleic acids [also known as cell-free DNA (cfDNA) or RNA], including detecting fetal abnormalities (11), transplanted organ rejection events (12,13), and signatures of cancers (14). It is not only human cells that shed their nucleic acids into the blood: DNA from plant-based foods has been detected (15), and other life forms such as viruses, bacteria, and fungi release their DNA and RNA into the blood, a phenomenon which has been exploited to determine the presence of infectious disease (12,16) and to measure alterations of the virome due to pharmacological immunosuppression (17).…”