MicroRNAs: from
oncogenesis
comprehension to
promising arsenal of
cancer therapies
Martin Simard Ph.D.
Université Laval
Centre de Cancérologie (CHUQ)
microRNAs: A new class of regulatory molecules
•Small non-coding RNAs (21 to 23
nucleotides long)
•First discovered in Caenorhabditis elegans
•Found
in
nearly
all
eukaryotes
more than 1,000 different microRNAs in
human
•Act “negatively” in gene expression and
control cellular timing
•Recent estimation support that more than
60% of the human coding genome is
How microRNAs are produced?
•
•
•
•
Mainly produced by RNA
polymerase II.
1st maturation in the
nucleus
2nd maturation in the
cytoplasm
Nature Reviews MCB, 2008
Active complex: miRISC
How microRNAs work?
•
•
•
Partial complementarity with 3’UTR regions
(position 2 to 8 critical)
Cooperative effect
Abrogate protein synthesis
Nature Reviews MCB, 2008
One microRNA may regulate up to 100 different
genes!
Precise expression profile
miR-1-1
miR-1-2
Zhao et al, Nature 2005
Pena et al, Nature Methods 2009
Implication in cell differentiation
Stefani and Slack, Nature Reviews MCB 2008
Precise expression profile: haematopoiesis
Gangaraju and Lin, Nature Reviews MCB 2009
The loss of microRNAs lead to fatality
Zhao et al,
Cell 2007
Loss of miR-1-2
leads to
overproduction
of muscle cells
Ventura et al,
Cell 2008
Loss of miR-1792 cluster is
embryonic lethal
microRNAs and Cancer
microRNAs are frequently located in
altered genomic regions associated
to various cancers
Calin et al, PNAS 2004
microRNAs and Cancer
Different expression profile
between healthy and cancer
tissue samples
Lu et al, Nature 2005
microRNAs and Cancer
• Oncogenes
Oncogene 2006
microRNA as Oncogene : miR-10b
•
•
•
Overexpression increases
angiogenesis
Induces metastasis formation
Correlation between miR-10b
overexpression and metastatic
tumours caused by breast cancer
Ma et al, Nature 2007
microRNAs and Cancer
• Oncogenes
• Tumour suppressors
Oncogene 2006
microRNA as Tumour suppressor : let-7
•
•
•
let-7 level is altered in lung cancer
let-7 controls RAS expression
HMGA2 is another target of let-7
Mayr et al, Science 2007
Johnson et al, Cell 2005
microRNA as Tumour suppressor : miR-335 and miR-126
•
•
•
Re-establishing their expressions
diminish the metastasis formation
miR-335 controls cell migration
miR-126 controls cell proliferation
Tavazoie et al, Nature 2008
Implication of the p53
p53
miR-34
BCL2
Survival
CDK6
CDK4
CyclinE2
Cell proliferation
He et al. Nature 2007 (+4)
Implication of the p53
p53
miR-34
BCL2
CDK6
CDK4
CyclinE2
miR-16-1
miR-145
miR-143
Survival
Cell proliferation
He et al. Nature 2007 (+4)
Suzuki et al. Nature 2009
microRNAs and Cancer: more and more
examples
Mutation or epigenetic changes can lead to:
• Deletion
of microRNA
miR-15a,-miR16-1 silencing of microRNA locus
• Epigenetic
miR-29, miR203
• Point
mutation affecting a microRNA or microRNA precursor
miR-15a,-miR16-1
• miR-17~92,
Genomic amplification
or
translocation
of
microRNA
locus
miR21
• Loss of epigenetic silencing of microRNA locus
• Point mutation in the microRNA targets
• Rearrangement
of 3’UTR
HMGA2
Spizzo et al,
Cell 2009
microRNA as a Cancer therapy: miR-26a
Decrease in tumour mass
Kota et al. Cell, 2009
New therapies: Controlling microRNA expression
Science, 2008
Controlling microRNA expression: different methods
Garzon et al, Nature Reviews Drug Discovery, 2010
Why microRNA are functionally important in
primary cancers?
•There
are tumours-specific microRNA signatures
which accurately distinguish different sub-types of
cancers
•Modulation
of microRNAs in cancer cell lines can
directly regulates fundamental behaviours of
cancer cells such as proliferation and apoptosis
•Many
microRNAs deregulated in cancers have
been shown to control oncogenes, tumour
suppressors and signalling pathway components
as direct targets
microRNAs and Cancer: new biomarkers
Nature Biotechnology 2008
microRNAs and Cancer: new biomarkers
PNAS, 2007
Using ovarian cancer as a model, we demonstrate that expression of let-7 and HMGA2 is a
better predictor of prognosis than classical markers such as E-cadherin, vimentin, and Snail.
These data identify loss of let-7 expression as a marker for less differentiated cancer.
British J. of Surgery, 2009
microRNAs and Cancer: new biomarkers to
predict drug resistance
Clinical Cancer Research, 2009
Why microRNA are excellent biomarkers?
•Extremely stable in fluids as well as on formalinfixed paraffin-embedded tissue
•Expression profile correlates well between fresh
and formalin-fixed paraffin-embedded samples
•Resistant to degradation
Why microRNA are excellent biomarkers?
Weber et al, Clinical chemistry, 2010
Our microRNA profiling platform
TaqMan based quantitative RT-PCR
Our microRNA profiling platform
•Extremely sensitive (from 10 cells)
•Great specificity
•No hybridization required
•Easily go from profiling to
validation
let-7a
let-7b
UGA GGU AGU AGG UUG UAU AGU
U UUG UGU GGU
UGA GGU AGU AGG
let-7d
U UUG UAU GGU
UGA GGU AGU AGG
U
AGA GGU AGU AGG UUG CAU AGU
let-7e
UGA GGU AGG AGG UUG UAU AGU
let-7c
microRNAs are associated to various diseases
• Viral infections
-
Viruses encodes microRNAs that target viral mRNAs
to regulate various stages of the viral life cycle
-
Viral microRNAs suppress expression of specific host
genes
-
Viral infections induce expression of host microRNAs
that inhibits expression of cellular genes
-
Upon viral infections, host cells express specific
microRNA that suppress viral mRNA expression
• Cardiac, immune, neurological and metabolic
disorders
To summarize
microRNAs: A new class of regulatory molecules
•Small non-coding RNAs (21 to 23 nucleotides
long)
•First discovered in Caenorhabditis elegans
More than
found in 142
•Found
in 17,000
nearlymicroRNAs
all eukaryotes
species!
•microRNA as new biomarkers
•Modulation of microRNA expression: new
therapeutic strategy
Fundamental research:
Questions to be answered
• What are the targets of microRNAs?
• How microRNA expression is
regulated?
• How microRNAs regulate gene
expression?
• Which are the cellular factors implicated
in the microRNA pathway?
Translational research:
What we can do as collaborators
Implication of miR-204 in pulmonary arterial
hypertension
A collaboration with Sebastien Bonnet
us
microRNA profiling
from tissues
you
Identification of
microRNA targets
Functional test in
animal models
Journal of Experimental Medicine, in revision
Take home message
Stop thinking about DNA...
... think about RNA!
Acknowledgments
The Lab
Collaborators
Guillaume Jannot
Samir Bouasker
Gyorgy Hutvàgner, Dundee University
Jean-Yves Masson, Université Laval
Sébastien Bonnet, Université Laval
Ivan Robert Nabi, UBC
Dinshaw Patel, SloanKettering
Eric Miska, Cambridge University
Michael Hengartner, Zurich University
Craig Mello, U. Mass Med. School
Victor Ambros, U. Mass, Med. School
Alejandro Vasquez
Gabriel Bossé
Irfan Syed Bukhari
Johannie Ducharme
Sandra Piquet
Isabelle Banville
Marie-Eve Boisvert
Evelyne Rondeau
Nellie Giguère
Eric Paquet, CRC