MI RNASexpression in oesophagealand gastriccancers · Cech TR et al -PNAS 1979 …ribosomal...
Transcript of MI RNASexpression in oesophagealand gastriccancers · Cech TR et al -PNAS 1979 …ribosomal...
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Matteo FASSANUNIVERSITÀ DEGLI STUDI DI PADOVA, ITALIA
Hot Issues in Molecular PathologyPopper H, Schaff Z, Tímár J
Technology Transfer in Diagnostic Pathology
7th Central European Regional Meeting
Siófok - Hungary, May 14-16 , 2012
MIRNAS expressionin oesophageal and gastric cancers
MIRNAS expressionin oesophageal and gastric cancers
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MULTISTEP MODELOF SCIENTIFIC PARADIGMS
1761 1858 1953
Die ZellularpathologieDe sedibus et causis
morborum per anatomen indagatis
Molecular structure of nucleic acids; a structure
for deoxyribose nucleic acid
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THE CODING-RNA PARADIGM
DNA
Gen
e
AA
CG
GC
TG
CG
CA
DNA strand
UUCC
GGCC
GGGG
GGUU
AACC
CCUU
mRNA
TRANSCRIPTION
protein
Cod
onTRANSLATION
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The begin of a new story…
Holley RW et al - Science 1965
…the first non-coding RNA to be characterised was the alanine tRNA found in baker's yeast…
Cech TR et al - PNAS 1979
…ribosomal ribonucleic acid (rRNA) is the RNA component of the ribosome, the protein manufacturing machinery of all living cells…
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NON-CODING RNAS’ WORLD
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Bartel DP - Cell 2004
Small RNAS’ WORLD
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THE INTERFERENCE ERA
Fire A et al - Nature 1998
…reported a potent gene silencing effect after injecting double stranded RNA into C. elegans…
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ONCOMIRS: the new actors in the cancer scenario
Visone R & Croce CM - Am J Pathol 2009
Baffa R, Fassan M & Croce CM - Laborwelt 2009
Fassan M, Rugge M & Baffa R - Front Biosci 2011
… one-third of protein-coding human mRNAs are susceptible to this complex miRNA regulatory network…
let-7
RAS; C-MYC
Normal Cancer
miRNA as tumor suppressor gene
miR-21
PDCD4
miRNA as oncogene
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PNASHuman microRNA genes are frequently located at fragile sites and genomic regions involved in cancers
Calin GA et al - PNAS 2003 Volinia S et al - PNAS 2005
A microRNA expression signature of human solid tumors defines cancer gene targets
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Iorio MV & Croce CM – J Clin Oncol 2009
Mechanisms of miRNAs’ dysregulation in cancer
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Baffa R, Fassan M, Gardiman M, Rugge M, Croce CM - J Pathol 2009
Fassan M, Palazzo JP, Rugge M, Croce CM - Breast Cancer Res 2009
The use ofFF-PE tissues
The pathologist’s point of view
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PDCD4 nuclear loss inversely correlates with miR-21 levels in colon carcinogenesis
Fassan M, Pizzi M, Mescoli C, Ludwig K, Rugge M - Virchows Arch 2011
Fassan M, Pizzi M, Balistreri M, Rugge M - Dis Esophagus 2011
Programmed cell death 4 nuclear loss and miR-21 or activated Aktoverexpression in esophageal squamous cell carcinogenesis.
PRENEOPLASTIC LESIONS
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Baffa R, Fassan M, Gardiman M, Rugge M, Croce CM - J Pathol 2009
Fassan M, Palazzo JP, Rugge M, Croce CM - Breast Cancer Res 2009
The pathologist’s point of view
The direct visualization in FF-PE tissues - ISH
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MIRNAS IN SITU HYBRIDIZATION
Van Baal J, Fassan M, Rugge M, Siersema P – Gut 2012
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Baffa R, Fassan M, Gardiman M, Rugge M, Croce CM - J Pathol 2009
Fassan M, Palazzo JP, Rugge M, Croce CM - Breast Cancer Res 2009
The pathologist’s point of view
The identification of the target genes - IHC
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+3.00
0.00
-3.00
hsa-miR-205
N IM LG HG BAchsa-let-7c
hsa-mIR-605hsa-miR-100
hsa-miR-203
hsa-miR-99ahsa-miR-23ahsa-miR-147hsa-miR-192hsa-miR-215hsa-miR-560hsa-miR-326hsa-miR-615
Up
Down
13 MIRNAS DIFFERENTLY EXPRESSED IN BE-ONCOGENESIS:
hsa-miR-147 hsa-miR-192 hsa-miR-215 hsa-miR-326 hsa-miR-560 hsa-miR-615-3p
6 UP-REGULATED hsa-let-7chsa-miR-23a hsa-miR-99a hsa-miR-100 hsa-miR-203 hsa-miR-205 hsa-miR-605
7 DOWN-REGULATED
Fassan M, Croce CM, Rugge M – Int J Cancer 2011
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hsa-let-7c TARGETS HMGA2/HMGI-C
� HMGA2/HMGI-C is a direct target of hsa-let-7c;
� HMGA2/HMGI-C is a small non-histone chromosomalprotein that modulates transcription;
� HMGA2/HMGI-C over-expression is a hallmark of variousbenign & malignant human tumors, including Gastric Cancer.
HMGA2/HMGI-C IHCRing chromosome
Differential Diagnosis: Liposarcoma lipoma-like versus lipoma.
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N
20%
60%
100%
N IM LG HG BAc
BARRETT’S ONCOGENESIS: HMGA2 IHC
BAcHG
N
BAc
� Biopsy samples:
N=25, IM=25, LG=16, HG=11, BAc=10
� Score Values: 0= 0-5%; 1= 6-50%; 2= >50%
p < 0.001
Fassan M, Croce CM, Rugge M – Int J Cancer 2011
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Fassan M, Croce CM & Rugge M – WJG 2011
The clinician’s point of view
miRNAs as new prognostic tools
1.0
1.0
0.5
Su
rviv
al p
rob
abil
ity
Time
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1.0
0.5
Surv
ival
pro
babi
lity
Time
Kogo R et al – Clin Cancer Res 2011
…the miR-146a expression status is associated with patients’survival and the presence of lymph node metastases; miR-146ais an independent prognostic factor in GC…
high miR-146a
low miR-146a
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MIR-27a POLYMORPHISM & GASTRIC ATROPHY
Arisawa T et al – Dig Dis Sci 2007
…the miR-27a genome region polymorphism may be an important definitive factor to develop the gastric mucosal atrophy in Japanese…
100
0
50
% o
f gas
tric
atr
oph
y
A/A G/G A/G
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Fassan M, Croce CM & Rugge M – WJG 2011
The clinician’s point of view
miRNAs as non-invasive diagnostic
markers
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MIRNAS AS PLASMA BIOMARKERS IN GC
Tsujiura M et al – BJC 2010
Plasma miRNAs are dysregulated in gastric cancer patients
7.0
3.0
1.0
5.0
.
.
.
.Rel
ativ
e qu
anti
ty
Normal Cancer
7.0
3.0
1.0
5.0
.
.
.
.Rel
ativ
e qu
anti
ty
Beforesurgery
Aftersurgery
miR-21 miR-21
Plasma miRNAs expression after 1 month from surgery
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Fassan M, Croce CM & Rugge M – WJG 2011
The clinician’s point of view
miRNAs as biomarkers of therapeutic response
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MIRNAS AND MULTI-DRUG RESISTANCE
Xia L et al – Int J Cancer 2008
miR-15b and miR-16 modulate MDR by targeting BCL2 in human gastric cancer cells
1
0
0.5
bcl2
rel
ativ
e ex
pre
ssio
n
miR15b miR16
1
0
0.5
bcl2
rel
ativ
e ex
pre
ssio
n
miR15b miR16
miR15b miR16ctrl
bcl2
β-actin
40
0
20A
pop
tosi
s %
3000
0
1500
Cas
pas
eac
tivi
ty
microarray qRT-PCR WB
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Fassan M, Croce CM & Rugge M – WJG 2011
The clinician’s point of view
miRNAs as new therapeutic tools
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MicroRNA Replacement Therapy for miR-145 and miR-33aIs Efficacious in a Model of Colon Carcinoma
polyethylenimine (PEI)-mediated delivery of unmodified miRNAs
miR-145 delivery reduced tumor proliferation and increased apoptosis, with concomitant repression of c-Myc and ERK5
Ibrahim AF et al – Cancer Res 2011
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Beyond miRNAs…the UCRs(ultraconserved regions)
� noncoding RNA (ncRNA) transcripts (longer than 200 bp)
� a subset (n=481) of conserved sequences that are located in both intra- and intergenic regions
� conserved (100%) between orthologous regions of the human, rat, and mouse genomes
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Fassan M & Rugge M – Unpublished data
2.0
0
BM
vs
N
uc.153 uc.472 uc.473
1.0
Human
2.0
0
1.0
BM
vs
N
uc.153 uc.472 uc.473
Mouse (model)2.0
0
BM
vs
N
uc.153 uc.472 uc.473
1.0
Rat (model)
Gastroduodenal anastomosis
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CONCLUSIONS (1/2)
miRNAs expression patterns have a significant impact on human disease
miRNAs are dysregulated in GI diseases: from inflammation to cancer
miRNAs are fundamental in all the cellular processes
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CONCLUSIONS (2/2)
miRNA expression profiling identifies cancer gene targets
Regulating miRNA expression might be a novel strategy for gastrointestinal tumors chemoprevention
miRNAs as novel diagnostic and prognostic tool
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Laboratory of surgical pathology - Surgical pathology & cytopathology unit – University of Padua
EBRA registry (Barrett’s esophagus Veneto region registry; www.esofagodibarrett.org)
Ohio State University - Comprehensive Cancer Center
Istituto Oncologico del Veneto - Surgery & endoscopy units
Funding agencies
…a particular thank to…
Laboratory of molecular pathology - Surgical pathology & cytopathology unit - University of Padua
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