In this issue: Non-coding RNAs as potential laboratory biomarkers

This is a Platinum Open Access Journal distributed under the terms of the Creative Commons Attribution Non-Commercial License which permits unrestricted
non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

How close are miRNAs from clinical practice?

A perspective on the diagnostic
and therapeutic market
Bonneau E.1, Neveu B.2, Kostantin E.3, Tsongalis G.J.4, De Guire V.2
1
Department of Laboratory Medicine, Biochemistry Division, Saint-Eustache Hospital, Québec, Canada
2
Biochemistry Division, Maisonneuve-Rosemont Hospital, Montreal, Québec, Canada
3
Department of Biochemistry, McGill University, Montreal, Québec, Canada
4
Department of Pathology and Laboratory Medicine, Geisel School of Medicine at Dartmouth
and Dartmouth Hitchcock Medical Center, Lebanon, NH, United States of America

ARTICLE INFO ABSTRACT

Corresponding author: The discovery of miRNAs in the mid-90s has changed

De Guire V.
Biochemistry Division
the dogma of gene expression regulation. Currently,
Maisonneuve-Rosemont Hospital miRNAs are the main theme of thousands of publi-
5415 L’Assomption Blvd cations each year and their involvement in human
Montreal, Québec
H1T 2M4
diseases is everyday more deeply understood. With
Canada that being known, what are the actual clinical applica-
E-mail: [email protected] tions of miRNAs and how far are they truly from the
Key words: patients? To address this question, we reviewed the
microRNA, diagnosis, therapeutics, miRNA diagnostic and therapeutic market. With many
cancer, drug development companies developing miRNA panels, the activity is
high in the diagnostic area. Some products, notably
for thyroid cancer (Interpace Diagnostic), are already
available to clinician and covered by major insurance
companies. In comparison, the therapeutic mar-
ket, mainly driven by miRNA mimics and antagomiR
products, is less advanced. Miravirsen (produced by
Roche/Santaris) and RG-101 (produced by Regulus
Therapeutics), designed to treat hepatitis C, are con-
sidered the flagship products of this class of future

Page 114
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

drugs. All of the miRNA-based drugs are cur- Detectable in biopsies and body fluids, miRNAs
rently in clinical trials and none have yet reached are considered as very sensitive and specific
the pharmaceutical breakthrough. However, ac- circulating biomarker (7). The enthusiasm for
quisition of miRNA-based companies by major miRNA in the diagnostics field is reflected by the
pharmas is sending a positive feedback on their number of related publications, reaching around
potentials. With multiple initiatives on their way, 11,000 papers in 2018 (Figure 2A).
the next years will definitely be determinant for On the therapeutic side, polypharmacology
the miRNA market that is still in his infancy. is gaining a lot of interest in the pharmaceuti-
 cal era (8). It is now clear that human diseases
are complex and that deregulation of multiple
INTRODUCTION genes is often needed to transform a normal
For the last thirty years, the fundamental re- cell into a pathological one (9). Furthermore,
search into RNA biology has grown at an expo- redundant cellular pathways can limit efficien-
nential rate. We are now better positioned than cy of monogenic targeting compounds (10).
ever to understand the involvement of RNA in Conversely, the miRNA’s function is by defini-
almost all critical cellular processes. Indeed, for tion based on multitargeting (11). In fact, it is
many years, the number of non-coding RNA well established that these small RNAs recog-
discovered has steadily increased. Hence, it is nize their mRNA targets mainly by the 2nd to the
not surprising that several Nobel prizes were 8th nucleotides of their 5’ end. Mismatches in
awarded for corner stone RNA discoveries, such the 3’ sequence allow one miRNA to specifically
as those won by Cech and Altman in 1989 (RNA bind to hundreds of different mRNAs simultane-
catalytic activities; (1)), Ramakrishnan, Steitz ously regulating their expression (11, 12). It is
and Yonath in 2009 (ribosome structure; (2)), not surprising that these endogenous multitar-
and of most interest for this review, to Fire and geting molecules gained a lot of interest in the
Mello in 2006 (RNA interference; (3)). therapeutic field. In fact, nearly 3,500 studies
were published in 2018 on miRNA-based thera-
Considering the increase in RNA-focused re-
search, one can expect that the advancement peutics (Figure 2B).
of general and specific knowledge about RNA Similarly, a multitude of clinical trials were con-
could result in direct clinical applications. For ducted or are currently underway to test new
example, more than 45,000 studies were pub- miRNA based treatments. This effervescence is
lished in 2017 on RNA (Figure 1A). From these, therefore expressing the evolution of this still
a large proportion of the studies either consid- young and relatively immature field of utilizing
ered that their work could contribute to the the miRNA as a therapeutic tool.
diagnosis or the treatment of disease (about With thousands of academic publications each
13,000 and 10,000, respectively; Figure 1B). year, how far from patients are miRNAs? To an-
From the multitude of RNA discoveries, one of swer this question and bring a different angle,
the most important was the discovery of RNA we reviewed the market of the diagnostic and
interference by Fire and Mello and miRNAs by therapeutic applications of miRNAs. With a lot of
Ambros and colleagues (4, 5). Thousands of companies offering miRNA-profiling services, our
these small RNAs of approximately 20 nucleo- focus was to highlight the ones providing specific
tides in length have been identified in humans expression panels for a given clinical application.
so far and are conserved across all species (6). On the therapeutic side, multiple clinical trials

Page 115
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

Figure 1 Number of studies published in PubMed per year

for: (A) RNA studies; (B) RNA diagnostic and therapeutic studies

Page 116
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

Figure 2 Number of studies published in PubMed per year

for: (A) miRNA studies and (B) miRNA diagnostic and therapeutic studies

Page 117
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

are currently ongoing and we focused on the deal by Genoptix failed. Interestingly, Interpace
products that are the most advanced in the field. Diagnostics acquired most of the equipment
through a bankruptcy auction and hired some of
miRNA-BASED DIAGNOSTICS Rosetta Genomics employers.
Applications in the cancer field Interpace Diagnostics (NASDAQ: IDXG), is based
in New Jersey and is a molecular diagnostic test-
Since their initial discovery in 1993, miRNA have
ing company that is offering personalized medi-
showed a great diagnostic potential by being as-
cine strategies for the diagnosis of thyroid and
sociated with various diseases (5, 7). Since then,
pancreatic cancer. Interpace acquired a solution
the number of publications on the diagnostic
developed by Asuragen combining ThyraMIR®, a
potential of miRNA grew almost exponentially
miRNA classifier, and ThyGeNEXT®, an oncogene
(Figure 2B), which attracted numerous compa-
panel for thyroid cancer stratification (Table 1).
nies to develop new miRNA-based diagnostic
Initial validation was completed by Asuragen
tools. To our knowledge, the first company fo-
in 2015, over 12 endocrinology centers across
cusing on miRNA-based diagnosis assays was
the U.S. and 638 surgical and fine needles aspi-
Rosetta Genomics (NASDAQ: ROSG), an Israeli
rations (FNA) biopsies were analyzed (16). The
company incorporated in early 2000. In part-
combination of ThyraMIR® and ThyGeNEXT® of-
nership with Precision Therapeutics, a person-
fers an interesting alternative as 15-30% of stan-
alized cancer therapy company, they launched
dard cytological evaluations fail to discriminate
in 2012 miRview™ mets a miRNA panel allowing
benign from the malignant lesions (17, 18). The
the identification of cancers of unknown or un- ThyraMIR® classifier includes the quantification
certain primary origin (CUP) (Table 1). of 10 miRNAs: miR-29b-1-5p, miR-31-5p, miR-
These cancers account for up to 15% of newly 138-1-3p, miR-139-5p, miR-146b-5p, miR-155,
diagnosed cancer in the U.S. every year (13, 14). miR-204-5p, miR-222-3p, miR-375 and miR-551b-
This CUP classifier was able to identify 42 dif- 3p (Table 1).
ferent tumor types using microarray that mea- This panel was trained using 240 well-charac-
sures the expression levels of 64 miRNAs. The terized, surgically resected, benign or malignant
miRviewTM mets panel was able to identify ac- thyroid lesions. A validation set of 54 indepen-
curately 90% of the 509 validation sample set. dent resected thyroid tissues and 235 preopera-
The assay also showed 88% correspondence tive thyroid FNAs was then used for threshold
with the patient’s clinicopathological evaluation optimization (16). Based on this study, Interpace
(14). Based on this success, Rosetta Genomic Diagnostic claims a Negative Predictive Value
introduced a new product called RossettaGX of 94%, a Positive Predictive value of 74% and
Reveal™ (Reveal) in 2016. a reduction of 85% of unnecessary surgeries.
This new miRNA classifier relied on qRT-PCR to Interpace Diagnostic is CLIA certified and CAP ac-
differentiate between benign or indeterminate credited, but both tests are not FDA approved.
thyroid nodules using FNA cytology smears. Availability of ThyraMIR® through Labcorp, was
Reveal’s performance was validated using a mul- announced on January 12, 2016. Interpace also
ticenter retrospective cohort of 189 FNA smears received Medicare coverage in 2016 covering
and achieved a negative predictive value of 91%, over 50 million patients across the United States.
a sensitivity of 85% and a specificity of 72% (15). Most recently, in November 2018, the Blue
Unfortunately, the company declared bankrupt- Cross Blue Shield and the U.S. Federal Employee
cy in May 2018 after a $10 million acquisition Health Benefit Program have agreed to include

Page 118
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

Table 1 Currently active companies working on miRNA-based diagnostics

Development
Companies Product Targeted miR Disease type Reference
phase

miR-29b-1-5p,
miR-31-5p,
miR-138-1-3p,
miR-139-5p Thyroid and
Interpace pancreatic
ThyraMIR/ miR-146b-5p, thygenext-
Diagnostics/ cancer Available
ThyGENX miR-155, thyramir.com
Asuragen
miR-204-5p,
miR-222-3p,
miR-375 and
miR-551b-3p

Rosetta rosettagenomic
Identify
Genomics/ .com
miRview mets miRNA library tumor origin Available
Precision
of cancer oncotest.co.il
Therapeutics

Genoptix Reveal miRNA library Thyroid Available genoptix.com

Panel of 19
TAmiRNA OsteomiR Osteoporosis Available tamirna.com
miRNAs

Panel of 11 Cardiovasc.
TAmiRNA ThrombomiR Available -
miRNAs Disease

Multiple
Hummingbird Panels Pre-Clinical & hummingbird-
- (cancers,
Diagnostics (unknown) Phase 1 diagnostics.com
brain, heart)

CogniMIR Panel (unknown) Alzheimer

DiamiR Phase 1 diamirbio.com
Others Panel (unknown) Brain diseases

Panel with miR

Mirnext Heart failure Development hmirnext.com
423-5p

Quanterix/
DestiNA Simoa miR-122 Liver toxicity Pre-Clinical quanterix.com
Genomics

Page 119
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

the ThyGeNEXT®/ThyraMIR® combined tests for through a Small Business Innovation Research
their 5.3 million beneficiaries. (SBIR) phase II grant of $1.5M from the National
Hummingbird Diagnostics (formerly known as Institute on Aging (NIA) of the National Institutes
Comprehensive Biomarker Center) was found- of Health (NIH) in 2014 and 2015. More recent-
ed in 1998 in Heindelberg, Germany, and has ly, in March 2017, the NIA of the NIH awarded
now extended to Boston, Massachusetts, in DiamiR another SBIR Phase IIB grant of $2.75M
the United States. Operating as a subsidiary of over three years to further support the develop-
Febit Holding, this company is hands-on in the ment of their branded lead product, CogniMIR™
development of novel miRNA signatures in liq- (Table 1). CogniMIR™ is currently in clinical trial
uid biopsies for early detection of various dis- testing for early detection of Alzheimer’s dis-
eases, ranging from cancer (Non-small-cell lung ease at the presymptomatic, mild cognitive im-
carcinoma, melanoma, breast cancer), to neu- pairment and dementia stages. Using different
rodegenerative (multiple sclerosis, Alzheimer, brain-derived miRNA signatures, DiamiR also
Parkinson), cardiovascular (acute myocardial expects to test for Parkinson’s disease, fronto-
infarction and heart failure) and inflammatory temporal degeneration, and amyotrophic lat-
bowel disease (19-21). With its DIN EN ISO/IEC eral sclerosis. However, those products are still
17025:2005 accreditations for RNA (including in the validation process.
miRNA) extraction and microarray services TAmiRNA is another European leader in miRNA
(Agilent Certified Service Provider), the com- diagnostics that was founded in 2013 as a spinoff
pany profiled more than 7,000 disease-related of two Austrian companies, BOKU and Evercyte.
body fluid samples so far. The bioinformatics This R&D company develops and offers validat-
and statistical processing of those large expres- ed microRNAs panels as additional tools for the
sion data led to the identification of multiple dis- diagnostic of age-related disorders. Funded by
ease-related miRNA panels (Table 1). Although AWS Seedfinancing and EU Horizon2020 pro-
none of these are currently commercially avail- grams, TAmiRNA demonstrated the clinical util-
able, Hummingbird Diagnostics has 17 granted ity of their licensed miRNAs as biomarkers in os-
patents in the field of whole blood expression teoporosis (26). OsteomiR™ is their lead product
profiling. The clinical validation of their miRNA intended to provide the risk of a first fracture in
signatures for early diagnostic use are ongoing female patients with postmenopausal osteopo-
with the funding received through their partici- rosis and type-2 diabetes (27-29). The integra-
pation in three European FP7-funded consortia tion of the expression level of 19 blood-circulat-
(BestAgeing, RiskyCAD and EURenOmics). ing miRNAs gives a calculated fracture-risk index
that could be used for preventive therapy and
Applications in age-related diseases treatment follow-up. Similarly, TAmiRNA also
DiamiR is located in Monmouth Junction, New proposes the ThrombomiR™ panel (11 miRNAs)
Jersey and published their first article describing to assess platelet function and the ToxomiR™
the use of miRNA biomarkers in mild cognitive panel (19 miRNAs) to evaluate the toxicity oc-
impairment in 2012 (22). DiamiR have since pub- curring in various tissues (Table 1). Kits based
lished several articles with a similar scope, which on primer-coated qPCR plates can be either
is the use of miRNA as markers of neurodegen- purchased (except ToxomiR™) or samples can
erative and neurodevelopmental disorders (23- be directly processed by TAmiRNA, from extrac-
25). This work was funded in part by the Michael tion to data analysis. By starting a partnership
J. Fox Foundation for Parkinson’s Research and agreement with SimplicityBio in February 2017,

Page 120
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

development of additional miRNA panels are Quanterix (NASDAQ QTRX) is a biotech compa-
expected. The Swiss-based bioinformatics com- ny founded in 2007 in Lexington, Massachusett.
pany has a robust in silico biomarker identifica- Through the development of their ultra-sensitive
tion pipeline that could accelerate TAmiRNA’s digital biomarker detection technology Simoa®,
main goal in offering advanced miRNA markers Quanterix provides healthcare researchers the
for the diagnosis and prognosis of age-related ability to investigate the continuum of disease
disorders. progression. In March 2018, they announced
a collaborative effort with DestiNA Genomics
Applications in cardiac function to enhance RNA biomarker detection. DestiNA
and liver toxicity was founded in 2010 in Edinburgh, United
Mirnext is a Dutch biomedical company based in Kingdom, where they developed and patented
Amsterdam that is interested in the diagnostic a unique PCR-free, chemical-based technology
potential of miRNAs. It was established in 2014 for the detection and quantification of nucleic
as a new entity of ACS Biomarker, which was acids such as miRNAs, without prior isolation
built on the Galectin-3 biomarker of heart fail- from serum or plasma (33, 34). This highly-
ure (30, 31), now out-licensed to BG Medicine specific nucleic acid detection combined to the
and available in clinics. Mirnext is currently fi- ultra-sensitive Simoa® system provides a solid
nanced through Life Sciences Fund Amsterdam support for disease-related miRNA biomarker
and Limburg Ventures, two venture capital testing. Accordingly, the collaboration’s first ini-
investors of The Netherlands. Similar to ACS tiative was focused on miR-122 as a liver toxicity
Biomarker, Mirnext’s main goal is to identify and marker (35, 36). They demonstrated that their
commercialize biomarkers in the cardiovascular assay detects miR-122 earlier and outperforms
field but with full dedication towards miRNAs. the current protein-based biomarkers in specifi-
Their high-throughput, disease-based miRNA cally detecting and quantifying liver toxicity.
profiling identified, among others, miR423-5p
as a useful marker of heart failure (32). Together miRNA-BASED THERAPEUTICS
with other clinically relevant miRNAs, Mirnext Several pharmaceutical and biotech companies
pursued their validation in large patient cohorts have launched miRNA projects in their develop-
with different cardiovascular diseases includ- ment pipeline (Table 2). Companies are mainly
ing heart failure, coronary artery disease and working on two types of products; miRNA mimics
myocardial infarction. Thus, their miRNA panel and antagomiRs. The miRNA mimics are used
integrates many different disease mechanisms to re-establish the concentration of a specific
useful for the identification and stratification of miRNA suppressed by the evolution of a giv-
those pathologies. In addition to the diagnosis en pathology (37, 38). Inversely, antagomiRs
of heart diseases, Mirnext is aiming to evalu- are used to suppress the function of specific
ate cardiovascular risk profiles (mortality, hos- miRNAs overexpressed and mechanistically in-
pitalization) of the individuals tested as part of volved in a disease (37, 38). In order to allow
their multi-marker heart failure test. A single the development of miRNA therapeutics, sci-
test is expected to provide the clinician exten- entists must address two main challenges: the
sive information on the patient’s cardiovascular stability and delivery. First, RNA molecules are
health to initiate targeted treatments. At the quite unstable because of their 2’-OH chemi-
time of this writing, we were unable to access cal group (39). Therefore, several companies,
the company’s website. such as Dharmacon, BioSyn and GenScript, can

Page 121
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

Table 2 Currently active companies working on miRNA-based therapeutics

Targeted Disease Development

Companies Product Reference
miR type phase

Roche/Santaris Miravirsen miR-122 HCV Phase 2 roche.com

RG-101 miR-122 HCV Phase 2 (hold)

Regulus Alport
RG-012 miR-21 Phase 1 regulusrx.com
Therapeutics syndrome
miR-
RG-125 NASH Phase 1
103/107

MRG-201 miR-29b Fibrosis Phase 2

Lymphoma Phase 1 and

MRG-106 miR-155
MiRagen and leukemia Phase 2 miragentherapeutics
Therapeutics .com
MRG-107 miR-155 ALS Pre-Clinical

MRG-110 miR-92 Ischemia Phase 1

ENGeneIC Mesomir miR-16 Mesothelioma Phase 2 engeneic.com

Abivax ABX464 miR-124 IBD Phase 2 abivax.com

Synlogic Screening synlogictx.com

Opko Screening opko.com

Alnylam
Screening alnylam.com
Pharmaceuticals
Interna interna-
Screening
Technologies technologies.com

Mello Biotech Screening mellobiotech.com

produce natural and chemically modified RNA of these RNAs to the desired site of action
(2’-O-methyl; 2’-OMe, locked nucleic acid; LNA, (39). Therapeutic application requires the cor-
of 2’-fluor; 2’-F, phosphorothioate; PS) to stabi- rect delivery of the RNAs to the targeted organs
lize and reduce the high reactivity of RNA mole- in order to maintain adequate treatment speci-
cules. The other major challenge is the delivery ficity. When a treatment requires a systemic

Page 122
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

delivery through intra-venous injection, the de- with Ionis Pharmaceuticals and GSK (Table 2). RG-
livery strategies are either passive or active (40, 101 is an N-acetyl-D-galactosamine- conjugated
41). The passive strategy utilizes the tendency of RNA antagomiR that also targets miR-122 in
several organs, like the liver, the spleen and the HCV infected hepatocytes (45). RG-101, like
lymph nodes to internalize accumulated parti- Miravirsen, shows considerable efficacy with
cles. Using this non-specific approach, designed patients displaying undetectable HCV RNA lev-
nanoparticles or liposome-like particles incorpo- els (45). However, some serious adverse events
rating RNAs can be targeted to these organs (40, of severe jaundice were recently declared in
41). Inversely, the active strategies combine the a clinical trial and the FDA put the studies on
RNA or the particle with a specific molecule that hold until the situation is clarified. It is worth
will bind to the cells of interest and will be endo- mentioning that Regulus was also working on
cytosed (40, 41). These structural and delivery RG-125 (also described as AZD4076), an an-
challenges, albeit being constantly addressed by tagomiR targeting miR-103/107, in phase 1 clin-
new design strategies, still complicate the devel- ical trial for treatment of nonalcoholic steato-
opment of miRNA therapeutics. hepatitis (NASH; Identifier NCT02612662 and
NCT02826525) as well as RGLS5040, an anti-
Applications in liver disease miR-27 aiming to reduce cholestatic diseases.
Among the most advanced products, there is However, development of these latter two were
Miravirsen (or SPC3649), an antagomiR target- recently suspended.
ing miR-122. Santaris Pharma initially devel-
oped this drug candidate before Roche acquired Applications in fibrotic disease
the company in 2014 to expand its RNA thera- Regulus has also worked with Genzyme (Sanofi)
peutic research and development department to test the efficacy of RG-012, an antagomiR
(Table 2). Miravirsen is a locked nucleic acid against miR-21, which reduces the fibrogenesis
(LNA) containing phosphorothioate modifica- of organs associated with Alport syndrome (46).
tions. MiR-122, is known to be essential in the This is an X-linked disease and is characterized by
life cycle of hepatitis C virus (HCV) expressed in kidney disease, hearing loss and visual impair-
the liver (42, 43). Reducing the activity of miR- ment caused by mutations of the genes encod-
122 in the context of HCV infection is impor- ing type-IV collagen (47). The use of a modified
tant. In fact, miR-122 is a host factor that binds single-stranded antagomiR with phosphorothio-
to the 5’-UTR region of the HCV genome and ate, 2’-O-methoxyethoxy and constrained ethyl
enhances its transcription (43, 44). In phase 1 modifications showed an important improve-
clinical trials, some patients who received high ment in the survival of a Alport mouse model
doses of Miravirsen in monotherapy resulted in with a reduction of kidney disease progression
undetectable HCV RNA levels (43, 44). Because (46). Despite interesting results, the phase 1
Miravirsen is a modified RNA (LNA and phos- clinical trial of RG-012 has recently been discon-
phorothioate), it naturally accumulates in the tinued mid-2018 because of the reorganization
liver and does not require special delivery strat- between Regulus and Sanofi (Clinical trial identi-
egy. Miravirsen is currently undergoing multiple fier NCT03373786).
phase 2 clinical trials. Another promising company is MiRagen Thera­
Another product was developed to target peutics (NASDAQ: MGEN), based in Boulder,
miR-122, RG-101, and is produced by Regulus Colorado. First, the company developed MRG-
Therapeutics (NASDAQ: RGLS) in collaboration 201, also known as Remlarsen, a miRNA mimic

Page 123
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

that aims to restore the levels of miR-29b, which amyotrophic lateral sclerosis (ALS) but has not
is a negative regulator of the extracellular matrix yet entered clinical trials. In an ever-growing
deposition processes. The miR-29 family (miR- pipeline, they also work on MRG-110 in col-
29a/b/c) is constantly downregulated in fibrotic laboration with Servier. This LNA antagomiR
diseases. MRG-201 is an LNA RNA mimic that targets miR-92 in order to treat ischemic con-
is administered by intradermal injection and ditions such as heart failure (48). Its phase 1
the phase 2 clinical trial is currently underway clinical trial is currently recruiting (Identifier
(Identifier: NCT03601052). Remlarsen could vir- NCT03603431).
tually be used for the treatment or prevention Pharmaceutical and biotech companies are
of pathological cutaneous fibrosis, as well as of heavily engaged in developing successful prod-
other fibrotic diseases, including idiopathic pul- ucts and RNA biologics are closer than ever to
monary fibrosis (48). entering the market. Another indicator of this
effervescence is the acquisition of RNA-based
Applications in cancer
companies by pharmaceuticals giants. Santaris
While some companies are having great suc- Pharma was acquired by Roche in 2014, SiRNA
cesses, others struggle to positively impact pa- Therapeutics by Merck in 2007, followed by
tient outcomes. This was the case of MiRNA the acquisition of this division by Alnylam
Therapeutics (NASDAQ: MIRN) and a miRNA Pharmaceuticals in 2014, and more recently
mimic, MRX34. MiR-34, is a well characterized MiRNA Therapeutics by Synlogic Inc. However,
tumor suppressor downregulated in a broad even more companies are currently testing new
range of cancers (49-51). MRX34 was delivered miRNA therapeutics. For example, ENGeneIC is
as a double stranded RNA encapsulated into a currently designing and producing Mesomir, a
liposome-formulated nanoparticle. Preclinical miRNA mimic that aims to replace miR-16, a tu-
studies were promising when used in several mor suppressor that is reduced in cases of can-
cancer types such as renal cell carcinoma, acral cer, such as malignant pleural mesothelioma
melanoma and hepatocellular carcinoma (52). (54). It successfully completed phase 1 clinical
However, the FDA halted their phase 1 clinical trial and will soon start phase 2 (55). On an-
trial when many immune-related serious ad- other hand, Abivax produces ABX464, a small
verse events leading to death were registered. molecular compound that triggers the increase
It reached a point where MiRNA Therapeutics of miR-124 to reduce the symptoms of inflam-
reduced its staff before Synlogic Inc (NASDAQ: matory bowel disease for patients refractory to
SYBX) finally acquired it in 2017. anti-TNF biologics and corticosteroids. It is cur-
MiRagen Therapeutics is actively developing rently in preparation for a phase 2b clinical trial
MRG-106, also known as Cobomarsen, an LNA for ulcerative colitis and phase 2a for Crohn’s
antagomiR that targets miR-155. This miRNA is disease.
involved in the differentiation and proliferation Finally, a multitude of companies work in pre-
of blood and lymphoid cells. Cobormarsen is clinical and large screening studies to identify
actually involved in phase 1 trials (Identifier potential biologic miRNA such as Opko with
NCT02580552) and phase 2 clinical trials their CURNA program, Alnylam Pharmaceuticals,
(Identifier NCT03713320), with the goal of Interna Technologies and Mello Biotech. These
treating certain types of lymphoma and leu- companies could therefore increase, in the next
kemia (53). Similarly, MRG-107 also targets several years, the number of miRNA therapeutics
miR-155 to alleviate symptoms associated with being tested or entering the market.

Page 124
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

CONCLUSION 5. Lee RC, Feinbaum RL, Ambros V. The C. elegans heter-

ochronic gene lin-4 encodes small RNAs with antisense
Enthusiasm, promise and hope are evident in complementarity to lin-14. Cell. 1993;75(5):843-54.
the miRNA industry. As described, multiple com- 6. Tetreault N, De Guire V. miRNAs: their discovery, bio-
panies are dedicating significant efforts and re- genesis and mechanism of action. Clinical biochemistry.
2013;46(10-11):842-5.
sources to develop miRNA-based products. The
diagnostic field is definitely the most advanced 7. De Guire V, Robitaille R, Tetreault N, Guerin R, Menard
C, Bambace N, et al. Circulating miRNAs as sensitive and
with some miRNA panels already offered to cli- specific biomarkers for the diagnosis and monitoring of
nicians and covered by major insurance com- human diseases: promises and challenges. Clin Biochem.
panies. However, considering the thousands of 2013;46(10-11):846-60.
publications in this area, miRNAs as diagnostic 8. Proschak E, Stark H, Merk D. Polypharmacology by De-
products can still be considered in their infancy. sign: A Medicinal Chemist’s Perspective on Multitargeting
Compounds. J Med Chem. 2018.
On the therapeutic side, despite the potentials,
the miRNA-based therapeutic breakthroughs 9. Boran AD, Iyengar R. Systems approaches to polyphar-
macology and drug discovery. Curr Opin Drug Discov De-
have not arrived yet. Recently, an analytical vel. 2010;13(3):297-309.
model based on technological growth metrics
10. Nussinov R, Tsai CJ, Jang H. A New View of Pathway-
showed that miRNAs still require time to reach Driven Drug Resistance in Tumor Proliferation. Trends
the maturity point needed to yield a significant Pharmacol Sci. 2017;38(5):427-37.
number of products that could enter the mar- 11. Lim LP, Lau NC, Garrett-Engele P, Grimson A, Schel-
ket (56). For this reason, most of the technolo- ter JM, Castle J, et al. Microarray analysis shows that
gies discussed are currently in clinical trials. some microRNAs downregulate large numbers of target
mRNAs. Nature. 2005;433(7027):769-73.
The development and commercialization of
12. Lewis BP, Burge CB, Bartel DP. Conserved seed pair-
new diagnostic and therapeutic tools is defi- ing, often flanked by adenosines, indicates that thou-
nitely a long process. Considering the first evi- sands of human genes are microRNA targets. Cell.
dence of the involvement of miRNA in human 2005;120(1):15-20.
disease in 2002 and the first detection of miR- 13. Greco FA, Hainsworth JD. Introduction: unknown pri-
NAs in blood in 2008, only a decade later, tre- mary cancer. Semin Oncol. 2009;36(1):6-7.
mendous progress and effort has been made to 14. Meiri E, Mueller WC, Rosenwald S, Zepeniuk M, Klinke
bring these small RNAs from the bench to the E, Edmonston TB, et al. A second-generation microRNA-
based assay for diagnosing tumor tissue origin. Oncolo-
bedside (57, 58). gist. 2012;17(6):801-12.
15. Lithwick-Yanai G, Dromi N, Shtabsky A, Morgenstern
REFERENCES S, Strenov Y, Feinmesser M, et al. Multicentre validation
of a microRNA-based assay for diagnosing indeterminate
1. North G. Nobel prizes: chemistry. RNA’s catalytic role.
Nature. 1989;341(6243):556. thyroid nodules utilising fine needle aspirate smears.
Journal of clinical pathology. 2017;70(6):500-7.
2. Service RF. Chemistry Nobel. Honors to researchers
who probed atomic structure of ribosomes. Science (New 16. Labourier E, Shifrin A, Busseniers AE, Lupo MA,
York, NY). 2009;326(5951):346-7. Manganelli ML, Andruss B, et al. Molecular Testing for
miRNA, mRNA, and DNA on Fine-Needle Aspiration Im-
3. Bernards R. [The Nobel Prize in Physiology or Medicine proves the Preoperative Diagnosis of Thyroid Nodules
for 2006 for the discovery of RNA interference]. Nederlands With Indeterminate Cytology. J Clin Endocrinol Metab.
tijdschrift voor geneeskunde. 2006;150(52):2849-53. 2015;100(7):2743-50.
4. Fire A, Xu S, Montgomery MK, Kostas SA, Driver SE, 17. Kato MA, Buitrago D, Moo TA, Keutgen XM, Hoda RS,
Mello CC. Potent and specific genetic interference by Ricci JA, et al. Predictive value of cytologic atypia in inde-
double-stranded RNA in Caenorhabditis elegans. Nature. terminate thyroid fine-needle aspirate biopsies. Ann Surg
1998;391(6669):806-11. Oncol. 2011;18(10):2893-8.

Page 125
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

18. Bongiovanni M, Spitale A, Faquin WC, Mazzuc- 29. Kocijan R, Muschitz C, Geiger E, Skalicky S, Baierl A,
chelli L, Baloch ZW. The Bethesda System for Report- Dormann R, et al. Circulating microRNA Signatures in
ing Thyroid Cytopathology: a meta-analysis. Acta Cytol. Patients With Idiopathic and Postmenopausal Osteo-
2012;56(4):333-9. porosis and Fragility Fractures. J Clin Endocrinol Metab.
19. Fehlmann T, Backes C, Alles J, Fischer U, Hart M, Kern 2016;101(11):4125-34.
F, et al. A high-resolution map of the human small non- 30. Amin HZ, Amin LZ, Wijaya IP. Galectin-3: a novel bio-
coding transcriptome. Bioinformatics (Oxford, England). marker for the prognosis of heart failure. Clujul medical.
2018;34(10):1621-8. 2017;90(2):129-32.
20. Keller A, Fehlmann T, Ludwig N, Kahraman M, Laufer 31. Gehlken C, Suthahar N, Meijers WC, de Boer RA. Ga-
T, Backes C, et al. Genome-wide MicroRNA Expres- lectin-3 in Heart Failure: An Update of the Last 3 Years.
sion Profiles in COPD: Early Predictors for Cancer De- Heart failure clinics. 2018;14(1):75-92.
velopment. Genomics, proteomics & bioinformatics.
2018;16(3):162-71. 32. Tijsen AJ, Creemers EE, Moerland PD, de Windt LJ,
van der Wal AC, Kok WE, et al. MiR423-5p as a circulating
21. Kahraman M, Roske A, Laufer T, Fehlmann T, Backes C,
biomarker for heart failure. Circ Res. 2010;106(6):1035-9.
Kern F, et al. MicroRNA in diagnosis and therapy monitor-
ing of early-stage triple-negative breast cancer. Scientific 33. Bowler FR, Diaz-Mochon JJ, Swift MD, Bradley M. DNA
reports. 2018;8(1):11584. analysis by dynamic chemistry. Angewandte Chemie.
22. Sheinerman KS, Tsivinsky VG, Crawford F, Mullan MJ, 2010;49(10):1809-12.
Abdullah L, Umansky SR. Plasma microRNA biomark- 34. Marin-Romero A, Robles-Remacho A, Tabraue-Chavez
ers for detection of mild cognitive impairment. Aging. M, Lopez-Longarela B, Sanchez-Martin RM, Guardia-
2012;4(9):590-605. Monteagudo JJ, et al. A PCR-free technology to detect
23. Sheinerman KS, Tsivinsky VG, Abdullah L, Crawford F, and quantify microRNAs directly from human plasma.
Umansky SR. Plasma microRNA biomarkers for detection The Analyst. 2018;143(23):5676-82.
of mild cognitive impairment: biomarker validation study. 35. Venkateswaran S, Luque-Gonzalez MA, Tabraue-
Aging. 2013;5(12):925-38.
Chavez M, Fara MA, Lopez-Longarela B, Cano-Cortes V, et
24. Sheinerman KS, Toledo JB, Tsivinsky VG, Irwin D, al. Novel bead-based platform for direct detection of un-
Grossman M, Weintraub D, et al. Circulating brain-en- labelled nucleic acids through Single Nucleobase Label-
riched microRNAs as novel biomarkers for detection and ling. Talanta. 2016;161:489-96.
differentiation of neurodegenerative diseases. Alzheim-
er’s research & therapy. 2017;9(1):89. 36. Rissin DM, Lopez-Longarela B, Pernagallo S, Ilyine H,
Vliegenthart ADB, Dear JW, et al. Polymerase-free mea-
25. Sheinerman K, Tsivinsky V, Mathur A, Kessler D, Shaz surement of microRNA-122 with single base specificity
B, Umansky S. Age- and sex-dependent changes in levels using single molecule arrays: Detection of drug-induced
of circulating brain-enriched microRNAs during normal liver injury. PloS one. 2017;12(7):e0179669.
aging. Aging. 2018;10(10):3017-41.
37. Metias SM, Lianidou E, Yousef GM. MicroRNAs in clin-
26. Walter E, Dellago H, Grillari J, Dimai HP, Hackl M. Cost- ical oncology: at the crossroads between promises and
utility analysis of fracture risk assessment using microR- problems. Journal of clinical pathology. 2009;62(9):771-6.
NAs compared with standard tools and no monitoring in
the Austrian female population. Bone. 2018;108:44-54. 38. Farooqi AA, Fayyaz S, Shatynska-Mytsyk I, Javed
Z, Jabeen S, Yaylim I, et al. Is miR-34a a Well-equipped
27. Weilner S, Skalicky S, Salzer B, Keider V, Wagner M, Swordsman to Conquer Temple of Molecular Oncology?
Hildner F, et al. Differentially circulating miRNAs after re-
Chemical biology & drug design. 2016;87(3):321-34.
cent osteoporotic fractures can influence osteogenic dif-
ferentiation. Bone. 2015;79:43-51. 39. Haussecker D. Current issues of RNAi therapeutics
28. Heilmeier U, Hackl M, Skalicky S, Weilner S, Schro- delivery and development. Journal of controlled re-
eder F, Vierlinger K, et al. Serum miRNA Signatures Are lease : official journal of the Controlled Release Society.
Indicative of Skeletal Fractures in Postmenopausal Wom- 2014;195:49-54.
en With and Without Type 2 Diabetes and Influence 40. Burnett JC, Rossi JJ. RNA-based therapeutics: cur-
Osteogenic and Adipogenic Differentiation of Adipose rent progress and future prospects. Chemistry & biology.
Tissue-Derived Mesenchymal Stem Cells In Vitro. Jour- 2012;19(1):60-71.
nal of bone and mineral research : the official journal of
the American Society for Bone and Mineral Research. 41. Peer D, Lieberman J. Special delivery: targeted thera-
2016;31(12):2173-92. py with small RNAs. Gene therapy. 2011;18(12):1127-33.

Page 126
eJIFCC2019Vol30No2pp114-127
 Bonneau E., Neveu B., Kostantin E., Tsongalis G.J., De Guire V.
How close are miRNAs from clinical practice? A perspective on the diagnostic and therapeutic market

42. Gebert LF, Rebhan MA, Crivelli SE, Denzler R, Stoffel 51. Misso G, Di Martino MT, De Rosa G, Farooqi AA, Lom-
M, Hall J. Miravirsen (SPC3649) can inhibit the biogenesis bardi A, Campani V, et al. Mir-34: a new weapon against
of miR-122. Nucleic acids research. 2014;42(1):609-21. cancer? Mol Ther Nucleic Acids. 2014;3:e194.
43. Lindow M, Kauppinen S. Discovering the first mi- 52. Ling H, Girnita L, Buda O, Calin GA. Non-coding RNAs:
croRNA-targeted drug. The Journal of cell biology. the cancer genome dark matter that matters! Clinical
2012;199(3):407-12. chemistry and laboratory medicine. 2017;55(5):705-14.
44. Janssen HL, Reesink HW, Zeuzem S, Lawitz E, Rodri- 53. Seto AG, Beatty X, Lynch JM, Hermreck M, Tetzlaff M,
guez-Torres M, Chen A, et al., editors. A randomized, dou- Duvic M, et al. Cobomarsen, an oligonucleotide inhibi-
ble-blind, placebo (plb) controlled safety and anti-viral tor of miR-155, co-ordinately regulates multiple survival
proof of concept study of miravirsen (MIR), an oligonu- pathways to reduce cellular proliferation and survival in
cleotide targeting miR-122, in treatment naïve patients cutaneous T-cell lymphoma. British journal of haematol-
with genotype 1 (gt1) chronic HCV infection. Hepatology; ogy. 2018;183(3):428-44.
2011: WILEY-BLACKWELL COMMERCE PLACE, 350 MAIN 54. Reid G, Pel ME, Kirschner MB, Cheng YY, Mugridge N,
ST, MALDEN 02148, MA USA. Weiss J, et al. Restoring expression of miR-16: a novel ap-
45. Baek J, Kang S, Min H. MicroRNA-targeting thera- proach to therapy for malignant pleural mesothelioma.
peutics for hepatitis C. Archives of pharmacal research. Annals of oncology : official journal of the European Soci-
2014;37(3):299-305. ety for Medical Oncology. 2013;24(12):3128-35.
46. Gomez IG, MacKenna DA, Johnson BG, Kaimal V, 55. van Zandwijk N, Pavlakis N, Kao SC, Linton A, Boy-
Roach AM, Ren S, et al. Anti-microRNA-21 oligonucle- er MJ, Clarke S, et al. Safety and activity of microRNA-
otides prevent Alport nephropathy progression by stimu- loaded minicells in patients with recurrent malignant
lating metabolic pathways. The Journal of clinical investi- pleural mesothelioma: a first-in-man, phase 1, open-
gation. 2015;125(1):141-56. label, dose-escalation study. The Lancet Oncology.
2017;18(10):1386-96.
47. Watson S, Bush JS. Alport Syndrome. StatPearls. Trea-
sure Island FL: StatPearls Publishing LLC.; 2018. 56. Beierlein JM, McNamee LM, Ledley FD. As Tech-
nologies for Nucleotide Therapeutics Mature, Products
48. Gallant-Behm CL, Piper J, Lynch JM, Seto AG, Hong SJ, Emerge. Molecular therapy Nucleic acids. 2017;9:379-86.
Mustoe TA, et al. A microRNA-29 mimic (remlarsen) re-
presses extracellular matrix expression and fibroplasia in 57. Calin GA, Dumitru CD, Shimizu M, Bichi R, Zupo S,
the skin. The Journal of investigative dermatology. 2018. Noch E, et al. Frequent deletions and down-regulation
of micro- RNA genes miR15 and miR16 at 13q14 in
49. Bouchie A. First microRNA mimic enters clinic. Nature chronic lymphocytic leukemia. Proc Natl Acad Sci U S A.
biotechnology. 2013;31(7):577. 2002;99(24):15524-9.
50. Adams BD, Parsons C, Slack FJ. The tumor-suppressive 58. Lawrie CH, Gal S, Dunlop HM, Pushkaran B, Liggins AP,
and potential therapeutic functions of miR-34a in epithe- Pulford K, et al. Detection of elevated levels of tumour-
lial carcinomas. Expert opinion on therapeutic targets. associated microRNAs in serum of patients with diffuse
2016;20(6):737-53. large B-cell lymphoma. Br J Haematol. 2008;141(5):672-5.

Page 127
eJIFCC2019Vol30No2pp114-127