Circulating microRNA Biomarkers as Liquid Biopsy for Cancer Patients: Pros and Cons of Current Assays

Abstract

An increasing number of studies have focused on circulating microRNAs (cmiRNA) in cancer patients’ blood for their potential as minimally-invasive biomarkers. Studies have reported the utility of assessing specific miRNAs in blood as diagnostic/prognostic biomarkers; however, the methodologies are not validated or standardized across laboratories. Unfortunately, there is often minimum limited overlap in techniques between results reported even in similar type studies on the same cancer. This hampers interpretation and reliability of cmiRNA as potential cancer biomarkers. Blood collection and processing, cmiRNA extractions, quality and quantity control of assays, defined patient population assessment, reproducibility, and reference standards all affect the cmiRNA assay results. To date, there is no reported definitive method to assess cmiRNAs. Therefore, appropriate and reliable methodologies are highly necessary in order for cmiRNAs to be used in regulated clinical diagnostic laboratories. In this review, we summarize the developments made over the past decade towards cmiRNA detection and discuss the pros and cons of the assays.

Variability in, variability out: best practice recommendations to standardize pre-analytical variables in the detection of circulating and tissue microRNAs

“…This process specifically selects for mature miRNAs and reduces miRNA precursors. One proposal is direct miRNA detection (without extraction) as a superior method because it eliminates miRNA loss during extraction, removes the need to normalize to total RNA, and is easier to perform routinely in a clinical laboratory [9]. Overall, the lack of consensus regarding miRNA extraction highlights the importance of using the same extraction method throughout a study to minimize the introduction of confounding variables.…”

“…Various detection methods [quantitative reverse transcription PCR (qRT-PCR), microarray, RNA sequencing (RNAseq), next generation sequencing (NGS)] are available for miRNA quantification from circulation and tissue alike [1,9,38]. All of these methods require normalization to account and correct for variation across samples.…”

“…When a relatively large number of miRNAs are measured, endogenous control methods could be supplemented with a broad normalization approach such as global median normalization (the average expression of each miRNA relative to itself) [3,5,41]. Normalization to plasma or serum volume can also correct for variations in extraction efficiency since RNA concentration does not appear to correlate with the number of miRNAs detected [3,9,22,35].…”

“…However, studies to date have yielded inconsistent results and clinical assays remain elusive. A number of recent reviews [1][2][3][4][5][6][7][8][9] have thoroughly discussed the challenges and pitfalls involved in accurately quantifying miRNAs, including pre-analytical, analytical, and methodological factors that contribute to result heterogeneity. There is clearly an urgent need for a global consensus on study design and laboratory procedures.…”

Ultra-Fast Next Generation Human Genome Sequencing Data Processing Using DRAGEN<sup>TM</sup> Bio-IT Processor for Precision Medicine

“…Furthermore, fast sequencing turnaround time and roughly $1000 NGS whole genome cost is encouraging more institutes and individuals to opt for NGS based personalized medicine [3]- [8]. However, the "Big Data Bottleneck" is still the largest obstacle to use the NGS-based precision medicine in the real time disease and healthcare management.…”