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Overexpression of MMP9 in colonic
epithelium mediates protection in colitis
associated cancer
PALLAVI GARG
Ph.D.
Disclosures
Nothing to Disclose
Matrix Metalloproteinases (MMPs)
Are a family of zinc-dependent proteolytic
enzymes with the ability to degrade
extracellular matrix substrates such as
collagen, gelatin and proteoglycans
Colitis Associated Cancer (CAC) &
Colorectal Cancer
•
•
•
•
•
CAC is an important complication of Ulcerative Colitis or colonic
Crohn’s Disease that results in significant morbidity and mortality
It causes 1/6 of all deaths in patients with ulcerative colitis
Colon cancer develops in flat dysplastic tissue among IBD
individuals
CAC is often multiple, anaplastic, broadly infiltrating, rapidly
growing and occurs at a younger age
The pathogenesis of CAC is not known
MMP9 and its role in acute colitis
•
•
MMP9 is not expressed in normal tissues and is one of the
predominant MMPs that is up-regulated in several animal
models of colitis and human IBD
MMP9 mRNA and protein levels are increased in the inflamed
colonic mucosa of patients with IBD
•
MMP9 activity correlates with active inflammation
•
MMP9 activates Notch1 signaling in colonic epithelium
Castaneda et al, 2005, Gastroenterology
Garg et al, 2007, Gastroenterology
Role of MMP9 in CAC
• MMP9-/- mice show increased susceptibility to CAC
• CAC in MMP9-/- mice is associated with decreased apoptosis
compared to WT mice
• CAC in MMP9-/- mice showed decreased levels of p21WAF1/Cip1
and p53
• MMP9 mediates its tumor suppressive role via activation of
Notch1 signaling
Garg et al, 2010, Cancer Research
Garg et al, 2011, Gastroenterology
Aim
Which MMP9 protects from CAC ?
In vivo model
C57/B6
WT
Water
DSS (3 cycles)
Tg-villin-MMP9
Water
DSS= Dextran Sodium Sulfate (3% in drinking water)
DSS (3 cycles)
In vivo protocol
Days
0
7
14
28
35
49
53
DSS Cycle
DSS Cycle
DSS Cycle
(3rd ) starts
(1st) starts
(2nd ) starts
Recovery
Recovery
Recovery
st
(1 ) starts
(2nd) starts
(3rd) starts
85
sacrifice
Body weight gain
in percentage
Overexpression of epithelial MMP9
exhibited resistance to CAC
*
*
*
1
2
*
3
Tg-villin-MMP9
WT
Sacrifice
DSS
DSS
DSS
cycle 2
cycle 3
cycle 1
Recovery
Recovery
Recovery
cycle 2
cycle 3
cycle 1
Overexpression of epithelial MMP9
exhibited less tumor incidence in CAC
B
Number of polyps
Number of dysplastic lesions
A
WT
CAC
Tg-villin-MMP9
CAC
*
WT
CAC
Tg-villin-MMP9
CAC
Overexpression of epithelial MMP9
exhibited resistance to CAC
X10
WT CAC
Tg-villinMMP9 CAC
X20
Overexpression of epithelial MMP9
showed increased levels of Notch1and p53
in CAC A
Tg-villinMMP9, CAC
WT, CAC
MMP9, 104 kD
β-tubulin
1
2
3
4
5
6
Tg-villinMMP9, CAC
WT, CAC
B
NICD, 80 kD
β-tubulin
1
2
3
4
5
6
Tg-villinMMP9, CAC
WT, CAC
C
p53, 53 kD
GAPDH
1
2
3
4
5
6
Overexpression of epithelial MMP9
showed increased levels of p21WAF1/Cip1and
Bax1 in CAC
A
Tg-villinMMP9, CAC
WT, CAC
p21WAF1/Cip1, 21kD
GAPDH
1
B
2
3
4
5
6
Tg-villinWT, CAC MMP9, CAC
Bax-1, 37 kD
GAPDH
1
2
3
4
5
6
Overexpression of MMP9 in MMP9-/- MEFs
showed increased levels of Notch1 and p53
MMP9-/MEFs
WT
MEFs
MMP9-/-MEFs
+ MMP9
A
MMP-9, 104 kD
B
NICD, 80 kD
GAPDH
1
2
MMP9-/MEFs
3
4
5
WT
MEFs
6
MMP9-/-MEFs
+ MMP9
C
p53, 53 kD
GAPDH
1
2
3
4
5
6
Summary
•Epithelial-derived MMP9 plays a tumor suppressive role
in CAC
• Overexpression of MMP9 in colonic epithelium is associated
with altered levels of p53, Notch1, Bax1 and p21WAF1/Cip1
• Overexpression of MMP9 in MMP9
of NICD and p53
-/-exhibited
increased levels
Molecular mechanism of p53 regulation by
MMP9 via Notch1
Inflammation
nucleus
ARF
+
+
+
Mdm2
ARF
Notch-1
Mdm2
Mdm2
+
p53
cytosol
+
MMP-9
Conclusions
• Despite being a mediator of acute inflammation,
epithelial derived- MMP9 acts as a tumor suppressor
in CAC
• MMP9 mediated p53 activation via Notch1 signaling
is necessary and sufficient for its tumor suppressive
effect in CAC
Acknowledgement
Didier Merlin
Lewins Walter
Crohn’s & Colitis Foundation of America
Career Development Award, (#3057)