CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara...
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Transcript of CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara...
![Page 1: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/1.jpg)
CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN
HEAD AND NECK CANCER
Devathri NanayakkaraEskitis Institute for Drug Discovery
Griffith University
![Page 2: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/2.jpg)
Head and neck cancer
![Page 3: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/3.jpg)
Head and neck cancer
Sixth most common cancer
![Page 4: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/4.jpg)
Head and neck cancer
Sixth most common cancer
Five year survival rate after diagnosis, 50%
![Page 5: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/5.jpg)
Head and neck cancer
Sixth most common cancer
Five year survival rate after diagnosis, 50%
Drug resistance
Early tumors - asymptomatic
![Page 6: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/6.jpg)
Head and neck cancer
Sixth most common cancer
Five year survival rate after diagnosis, 50%
Drug resistance
Early tumors - asymptomatic
“Need to study the underlying molecular pathways unveiling potential detection markers and drug targets”
![Page 7: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/7.jpg)
In a recent study,
• Characterized the somatic mutation landscape of OSCC-GB
![Page 8: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/8.jpg)
![Page 9: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/9.jpg)
- Found Five new genes associated with OSCC-GB
![Page 10: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/10.jpg)
- Among them was USP9X- 22% harboured copy number loss and truncating
mutations - Role as tumor suppressor
![Page 11: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/11.jpg)
USP9X• Deubiquitylating enzyme– Family of cysteine and metalloproteases
![Page 12: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/12.jpg)
USP9X• Deubiquitylating enzyme– Family of cysteine and metalloproteases
http://legacy.butler.edu/biology/faculty-staff/research-interests/jkowalski/research-in-the-kowalski-lab/
![Page 13: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/13.jpg)
Murtaza et al, 2015
![Page 14: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/14.jpg)
USP9X in cancer
![Page 15: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/15.jpg)
USP9X in cancer
![Page 16: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/16.jpg)
USP9X in cancerBreast cancer
Colorectal cancer
Bladder cancer
Oral cancer
Brain cancer
Pancreatic cancer
Prostate cancer
Lung cancer
Lymphoma
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In this study,
Aims :• To evaluate the role of USP9X in an in vitro
system• To elucidate the molecular mechanisms USP9X
is involved in
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How?• In vitro cell lines – SCC15, CAL27, FaDu and Detroit 562
![Page 19: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/19.jpg)
Immunoblotting to probe for USP9X expression
All 4 cell lines express USP9X
SCC1
5
FaD
u
Det
roit
562
CAL2
7
USP9X290 kDa
β tubulin51 kDa
![Page 20: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/20.jpg)
• Knockdown approach – siRNA
![Page 21: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/21.jpg)
Immunoblotting to probe for USP9X protein levels 72 h after siRNA treatment
USP9X is efficiently knocked down in all four cell lines
NT USP9X NT USP9X NT USP9X NT USP9X
SCC15 CAL27 FaDu Detroit 562
siRNA
USP9X290 kDa
Β tubulin51 kDa
![Page 22: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/22.jpg)
• Effect on cell aspects– Cell proliferation• CyQUANT Assay
![Page 23: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/23.jpg)
48H 72H 96H 120H 144H0
2
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NT siRNA
USP9x siRNA
Nor
mal
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fluor
esce
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48H 72H 96H 120H 144H0
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NT siRNAUSP9X siRNA
Nor
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esce
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NT siRNAUSP9X siRNA
Nor
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fluor
esce
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48H 72H 96H 120H 144H0
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NT siRNAUSP9X siRNA
Nor
mal
ised
fluor
esen
ce
time time
time time
In absence of USP9X, a decrease in cell numbers was observed
CyQUANT Analysis of cell proliferation following siRNA treatment to knockdown USP9X
SCC15
Detroit 562
CAL27
FaDu
*
*
* * *
*
*
*
* P value < 0.05
![Page 24: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/24.jpg)
• Decrease in cell numbers,– Apoptosis?
![Page 25: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/25.jpg)
No elevation in apoptosis detected upon depletion of USP9X
β tubulin 51 kDa
Cleaved PARP-1 89 kDa
siRNA NT USP9X NT USP9X NT USP9X NT USP9X NT USP9X NT USP9X NT USP9X NT USP9X NT USP9X NT USP9X NT USP9X NT USP9X
time 48h 96h 144h 48h 96h 144h 48h 96h 144h 48h 96h 144h
SCC15 CAL27 FaDu Detroit 562
Immunoblotting for cleaved PARP-1
![Page 26: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/26.jpg)
Decrease in cell numbers in absence of USP9X prompts a role of a
![Page 27: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/27.jpg)
Decrease in cell numbers in absence of USP9X prompts a role of a “tumor promoter”
![Page 28: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/28.jpg)
Decrease in cell numbers in absence of USP9X prompts a role of a “tumor promoter”• Contradicts predicted oncosupressive role• Context specific
![Page 29: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/29.jpg)
Decrease in cell numbers in absence of USP9X prompts a role of a “tumor promoter”• Contradicts predicted oncosupressive role• Context specific
– Pancreatic cancer
![Page 30: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/30.jpg)
Decrease in cell numbers in absence of USP9X prompts a role of a “tumor promoter”• Contradicts predicted oncosupressive role• Context specific
– Pancreatic cancer
![Page 31: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/31.jpg)
Decrease in cell numbers in absence of USP9X prompts a role of a “tumor promoter”• Contradicts predicted oncosupressive role• Context specific
– Pancreatic cancer
Tumor suppressor
![Page 32: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/32.jpg)
Decrease in cell numbers in absence of USP9X prompts a role of a “tumor promoter”• Contradicts predicted oncosupressive role• Context specific
– Pancreatic cancer
Tumor suppressor
![Page 33: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/33.jpg)
Decrease in cell numbers in absence of USP9X prompts a role of a “tumor promoter”• Contradicts predicted oncosupressive role• Context specific
– Pancreatic cancer
Tumor suppressor
Tumor promotor
![Page 34: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/34.jpg)
To further confirm,• Overexpression of USP9X
![Page 35: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/35.jpg)
To further confirm,• Overexpression of USP9X
Usp9x cDNA
![Page 36: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/36.jpg)
To further confirm,• Overexpression of USP9X
Usp9x cDNA
Linearized the plasmid
Lipofectamine 2000
After 3 days: antibiotic selection(12 days)
To establish stable cell lines,
![Page 37: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/37.jpg)
USP9X is ectopically expressed in all four cell lines
V5SC
C15
CAL2
7
FaDu
Detr
oit 5
62
scc1
5
CAL2
7
FaDu
Detr
oit 5
62
pDEST51 USP9X pDEST51
β tubulin
Immunoblotting to detect ectopic expression of USP9X
![Page 38: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/38.jpg)
48h 72h 96h 120h 144h0
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pdest51pdest51 USP9X
Nor
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fluor
esen
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pdest51pdest51 USP9X
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pdest51pdest51 USP9X
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fluor
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48h 72h 96h 120h 144h02468
101214161820
pdest51pdest51 USP9X
Nor
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fluor
esen
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Ectopic USP9X protein expression increased cell proliferation
CyQUANT Analysis of cell proliferation following ectopic expression of USP9X
SCC15
Detroit 562
CAL27
FaDu
*
* *
*
* *
**
*
*
* *
*
* P value < 0.05
![Page 39: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/39.jpg)
Cell numbers are directly proportional to level of USP9X protein
![Page 40: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/40.jpg)
Cell numbers are directly proportional to level of USP9X protein has an oncogenic role
![Page 41: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/41.jpg)
Molecular mechanism regulated by USP9X
![Page 42: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/42.jpg)
Molecular mechanism regulated by USP9X
Murtaza et al, 2015
![Page 43: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/43.jpg)
Molecular mechanism regulated by USP9X– mTOR– Wnt – Notch
• Regulates cell proliferation• Known USP9X substrates
![Page 44: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/44.jpg)
Molecular mechanism regulated by USP9X?– mTOR– Wnt – Notch
CyclinD1, c-MYC, HES1
![Page 45: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/45.jpg)
Molecular mechanism regulated by USP9X?– mTOR– Wnt – Notch
Quantitate the RNA levels by qPCR
CyclinD1, c-MYC, HES1
![Page 46: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/46.jpg)
48H 72H 96H 120H 144H0
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USP9x siRNA
Nor
mal
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fluor
esce
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48h 72h 96h 120h 144h0
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pdest51pdest51 USP9X
Nor
mal
ised
fluor
esen
ce
RNA extraction
cDNA
qPCR
![Page 47: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/47.jpg)
Fold change of target genes 144 h after knockdown of USP9X
SCC15 CAL27
FaDu Detroit 562
![Page 48: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/48.jpg)
Fold change of target genes 144 h after knockdown of USP9X
SCC15 CAL27
FaDu Detroit 562
![Page 49: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/49.jpg)
Fold change of target genes 144 h after knockdown of USP9X
SCC15 CAL27
FaDu Detroit 562
48H 72H 96H 120H 144H0
5
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NT siRNAUSP9X siRNA
Nor
mal
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fluor
esce
nce
![Page 50: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/50.jpg)
SCC15 CAL27
FaDu Detroit 562
Fold change of target genes 144 h after ectopic expression of USP9X
![Page 51: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/51.jpg)
• Consistently HES1 expression correlated with cell proliferation measured by CyQUANT assay
• USP9X seems to positively regulate notch pathway
![Page 52: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/52.jpg)
• Consistently HES1 expression correlated with cell proliferation measured by CyQUANT assay
• USP9X seems to positively regulate notch pathway
Hypothesis: USP9X regulates proliferation of head and neck cancer cells through notch pathway
![Page 53: CONTEXT SPECIFIC ROLE OF DEUBIQUITYLASE ENZYME, USP9X, IN HEAD AND NECK CANCER Devathri Nanayakkara Eskitis Institute for Drug Discovery Griffith University.](https://reader035.fdocuments.in/reader035/viewer/2022062718/56649eab5503460f94bb1377/html5/thumbnails/53.jpg)
Conclusions• USP9X depletion caused a decrease in cell
proliferation• Ectopic expression of USP9X led to increase in
cell proliferation• USP9X positively regulates Notch pathway
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ACKNOWLEDGEMENT
StephenGeorge
Nicholas SaundersWood lab members
Funding: Griffith University
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THANK YOU!