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Page 6 of 18 Happel et al. J Cancer Metastasis Treat 2020;6:32 I http://dx.doi.org/10.20517/2394-4722.2020.71

LIQUID BIOPSY AND EXRNA
ExRNAs and EVs are among the liquid biopsy analytes that have demonstrated potential as cancer
biomarkers due to their function, availability in most body fluids, and ability to be collected in a non-
invasive manner, allowing frequent and longitudinal sampling. In cancer research, there is substantial
[18]
evidence to support the use of exRNA for both diagnostic and prognostic purposes . Differential
expression of cellular and extracellular miRNAs has been associated with a wide range of human
[34]
diseases . While exRNA can include many diverse types of RNA species (as discussed above), most
studies investigating the use of exRNAs as biomarkers have focused on miRNAs since its expression
[35]
patterns are unique to individual tissues and differ between cancer and apparently uninvolved tissues . Y
RNA is abundantly expressed in multiple body fluids and increased levels of Y RNA have been observed
in the circulation of cancer patients [5,36] . These observations have triggered interest in the potential use of
Y RNA as a biomarker for cancer and other diseases. However, many other types for exRNAs are being
[18]
explored as potential biomarkers, including mRNA mutations and other non-coding RNAs .

There are opportunities for exRNA to be developed into reliable biomarker tests for cancer detection
using liquid biopsy samples, since exRNA is remarkably stable and resists degradation mediated by
ribonucleases [15,37] . The discovery of stable RNA or exRNA outside of cells is continuously changing
the fundamental understanding of intercellular signalling and of the role RNA may play in cell-to-cell
communication and other complex biological processes. Because of their relative stability within vesicles
or in association with RNPs and LPPs, and, marked differences between exRNAs secreted by apparently
normal and tumour cells, exRNA molecules have high potential for development as biomarkers of various
[39]
[38]
cancers including lung , breast and prostate cancers [40,41] .
ADVANTAGES OF EXRNA IN LIQUID BIOPSY
The remarkable stability and relatively non-invasive access of different exRNA molecules makes them an
interesting class of biomarkers. The stability of exRNAs have been tested ex vivo under various conditions
[42]
including freeze-thaw cycles, extreme pH values, and storage at room temperature (RT) . Examination
of exRNA from CSF or blood in the diagnosis of glioma revealed that the EV number and morphology
[43]
remained largely unchanged if CSF was stored at RT . The total RNA and representative miRNA levels
were well-preserved under this condition for up to a week, and a single cycle of freezing and thawing did
not significantly alter EV number, morphology, RNA content, or miRNA levels, confirming its stability
at RT. These findings demonstrated stability and the incredible ease and speed of obtaining specimens
for testing compared to conventional biopsy. Measuring circulating RNA as liquid biopsy is a reliable
alternative to conventional biopsies, offering a potentially cheaper, reliable, and non-invasive way of
monitoring cancer development, progression, and remission.

Most of the tests for disease diagnosis and monitoring used in clinics are based on specific protein
concentration changes in body fluids. In comparison to protein-based biomarkers, exRNA has several
advantages including easier developed assays with specificity, and an amplifiable detection signal.
Measuring low abundance RNA in biofluids also circumvents the inherent obstacle of high protein
concentration and complexity in human body fluids in liquid biopsy.

In various diseases, normal EV cargo contents change as diseases initiate, and progress, altering the types
[44]
of proteins and RNAs that are packaged. These changes are rapid and quantitative . Therefore, the exRNA
profile of an individual may provide a snapshot of their health. Real-time changes in expression of exRNA
offer prognostic values in predicting disease outcomes, monitoring treatment response and assessing
[45]
treatment risk .