Bioanalytics – Biomolecular
2019 PharmSci 360
Extracellular DNA (exDNA) and exRNA are traces of nucleic acids released from cells into the extracellular environment. Their use as disease biomarkers has been limited by technical challenges in their isolation caused by abundant RNA- and DNA-degrading enzymes in biofluids. Using combinations of denaturants, reducing agents, proteolysis, and revised organic extraction, we developed an automated, high-throughput approach for recovery of exRNAs and exDNA from the same biofluid sample. We applied this method to characterize exRNAs from hundreds of plasma and serum samples collected from healthy volunteers. We determined the effect of critical clinical parameters such as age and gender. Surprisingly, we encountered one participant with dramatically increased endocrine-origin exRNA contributions stable over 4 years and detectable in all of his samples, thereby demonstrating the robustness of this approach and the clinical potential of circulating RNAs as biomarkers. While plasma exRNA profiles were shown to detect pregnancy or heart and liver tissue injury, the neuroendocrine exRNA alterations observed in our outlier study participant was even more pronounced. We now prepare for sub-fractionation of blood and plasma to identify the cellular or extracellular compartment most enriched for this neuroendocrine signature and define its originating source(s). Our biofluid nucleic acid isolation approaches may also be valuable to study the pharmacology of nucleic acid drugs.