Abstract

A Novel Extracellular Vesicle Isolation Method Used to Discover Urine Liver Disease Candidate Biomarkers

Shannon Pendergrast, Chief Scientific Officer, Ymir Genomics

Hepatocellular carcinoma (HCC) is the sixth most common cancer worldwide, with increasing incidence, and is the third most common cause of cancer death. The clinical management of patients suffering from HCC, despite the fact that at-risk populations are well defined, is not satisfactory. This is mainly due to the unavailability of current diagnostics to predict and monitor the disease. OHSU and Ymir Genomics LLC have teamed up to develop a high-compliance urinary diagnostic for liver diseases, including HCC. We have developed a new method for the rapid and effective isolation of urine extracellular vesicles (EVs) and integrated that into Mass Spectrometry and miRNA array workflows. We have applied  the workflow to 3 groups of 20 urine samples each from 1) healthy controls, 2) patients with cirrhosis, and 3) and cirrhotic patients with untreated HCC. Mass Spectrometry data shows that this protocol is capable of identifying over 2500 proteins from only 2 mls of urine; far less sample than some previously reported urine EV proteomics studies. Our protocol also yields sensitive miRNA detection from the Firefly miRNA array. Preliminary data establishes proof-of-concept by identifying multiple liver-selective and known liver disease biomarkers in urine EV preps. Several candidate protein and miRNA biomarkers that are potentially capable of distinguishing disease from healthy groups have also been identified. The methods developed in this study are suitable for a larger longitudinal study, as they are already compatible with standard clinical chemistry laboratory assay requirements.  Hopefully, candidates biomarkers identified in this study will provide clinicians with powerful weapons in the fight against HCC.