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Page 8 of 18 Happel et al. J Cancer Metastasis Treat 2020;6:32 I http://dx.doi.org/10.20517/2394-4722.2020.71

ExRNA as cancer biomarker
Medical oncologists have been using cancer biomarker tests to guide molecularly targeted therapies to
achieve better therapeutic outcomes. In this regard, developing biomarkers and biomarker tests based
on exRNA is relevant and important in the context of precision cancer therapy. Investigators have been
assessing the current state-of-the-art methods for body fluid sample collection, exRNA isolation, and
analysis, with exRNA biomarker discovery as the goal. This data has been unified in a report on the current
[49]
state of knowledge of exRNA isolation and analysis techniques . To avoid loss of potential biomarkers,
investigators have been using comprehensive methods, such as qRT-PCR and cutting-edge platforms for
RNA sequencing, rather than selection methods for specific RNA species.

[50]
ExRNAs have already begun to demonstrate their utility as clinical biomarkers. A study by McKiernan et al.
reported the development of a urine exosome-based non-invasive gene expression assay that discriminates
[51]
high-grade from low-grade prostate cancer and benign disease. In another study Li et al. identified
and validated a panel of salivary exRNA biomarkers for potential use in screening and risk assessment
for gastric cancer. Using salivary gland secretions, investigators have identified 30 mRNA and 15 miRNA
candidates whose expression patterns were associated with the presence of gastric cancer . These exRNA
[51]
biomarkers were identified and validated with credible clinical performance for non-invasive detection of
gastric cancer. Another recent study reported analysis of ctDNA and exRNA for monitoring tumour burden
[52]
and therapeutic response in patients with multiple myeloma . This exploratory analysis has provided
evidence of ctDNA for predicting disease outcome and the utility of exRNA as a biomarker of therapeutic
response in multiple myeloma. It has been reported that an exosome-based detection of EGFR T790M in
plasma from non-small cell lung cancer patients (NSCLC) may benefit from ALK (anaplastic lymphoma
kinase) inhibitor therapy whose tissue samples are not available or who are unable or unwilling to undergo
repeat biopsy . To address this need, Exosome Diagnostics developed an assay (ExoDx Lung-ALK) in
[53]
a CLIA certified laboratory to isolate and analyse exosomal RNA from blood samples enabling sensitive,
accurate and real-time detection of EML4-ALK mutations in patients with NSCLC.

Exosome Diagnostics has also developed a qPCR-based test (ExoDx EGFR) that interrogates mutations
within the EGFR gene in NSCLC. The assay uses plasma derived exosomal RNA/DNA and cfDNA to detect
EGFR mutations to inform clinical management [53,54] . Castellanos-Rizaldos and colleagues compared this
assay to the FDA approved companion diagnostic, cobas® EGFR Mutation Test v2 (Roche), that detects
defined mutations within the EGFR gene from plasma cfDNA liquid biopsy samples of NSCLC patients
and found increased sensitivity and specificity using the ExoDx EGFR assay which they attributed to the
[53]
exRNA-based assay design .

EXRNAS AS BIOMARKERS OF CLINICAL SIGNIFICANCE IN CANCER
To demonstrate the potential of exRNA and exosomes as clinical biomarkers, we mined data from
current clinical trials exploring the utility of these liquid biopsy analytes in cancer. The ClinicalTrials.gov
is a database for publicly and privately supported research studies conducted around the world. As of
May 11th, 2020, there are 45 clinical trials on ClinicalTrials.gov that focus on the use of exRNA and
exosomes as clinical biomarkers in cancer . The search results are summarized in Figure 3A-C. While a
[55]
vast majority of the clinical trials are taking place in the U.S., there are many trials in other countries as
well, including China, Italy, and Spain [Figure 3A]. These clinical trials span a large variety of cancer types
[Figure 3B]. Lung and Prostate cancers are the most common disease models exploring the use of exRNA
and exosomes as clinical biomarkers, as both cancer types are the focus of seven clinical studies. Overall,
there is a large number of different cancer types represented in this data set. Fourteen out of the twenty
different cancer types are the focus of one or two clinical studies, demonstrating the utility of exRNA
and exosomes as clinical biomarkers. Blood is the primary biofluid utilized in these studies while urine is
also a common source of biofluid used for liquid biopsies [Figure 3C]. The combination of blood/serum/