A universal catalytic hairpin assembly system for direct plasma biopsy of exosomal PIWI-interacting RNAs and microRNAs

PIWI-interacting RNAs (piRNAs) are a complex class of small non-coding RNAs which specifically interact with the PIWI protein to play important roles in germline development and somatic tissues. Aberrant expressions of piRNAs have been recently found in a variety of malignant tumors and associated with cancer hallmarks. However, current methods of analyzing piRNAs are limited to reverse transcription quantitative polymerase chain reaction and next generation sequencing. In this study, we have developed a universal catalytic hybridization assembly system (uniCHA) to quantify piRNAs as well as microRNAs. The system simply comprises two universal hairpin DNA strands and one starting hairpin DNA which can be tailored by a simple rule to bind different piRNA and miRNA targets. The uniCHA system was proved to be able to analyze various piRNAs and miRNAs at the same reaction condition with low leakage and high sensitivity of pM level. With this system, we have detected piR-651 and miR-1246 in 106 particles μL-1 MCF-7 cell-secreted exosomes, and successfully performed a direct plasma biopsy to diagnose breast cancer with sensitivity and specificity both at 100% in cohorts of 21 breast cancer patients and 13 healthy controls. This universal biosensing system provides a simple and efficient strategy in analyzing multiple piRNA/miRNA biomarkers in complicated biological samples, indicating its potential of clinical application in cancer diagnostics.