1. Why Animal Models of Human Disease?
Gregory J. Gores, M.D.

2. Pros Cons Recapitulate disease process Allow genetic manipulation
Allow pharmacologic studies
Necessary for FDA approval processes
Cons
Disease processes in animals and humans can be strikingly different
Few large animal models due to expense
Drug metabolism is different
Models are time limited (days or months) vs. humans (years)

3. Ideal Animal Model of PSC Would Display
Inflammatory bowel disease
Large bile duct strictures
Progression over time
Loss of bile ducts
Cirrhosis
Develop bile duct malignancy
Unfortunately, the ideal animal model of PSC does not exist!

4. Animal Models of Inflammatory Bowel Disease
Toxic injury of the intestinal lining
Genetic models – IL10 genetic deficiency
Immune cell manipulation
Genetic
Adaptive transfer
None of these models develop PSC!

5. What are we doing at Mayo Clinic?
Acute model of PSC in mice
Mdr2-/- chronic models of PSC in mice
Genetic models of bile duct cancer in mice

6. SMAC MIMETIC ANIMAL MODEL
BV6 (0.1 mg/100 L)
Saline (100 L)
CBD clamp
CBD clamp,
Gallbladder
Injection
CBD clamp
release
sacrifice
45 min
Day 5

7. SM Intra-biliary Injection Results in a PSC-like Phenotype
Sirius red
αSMA
Mac2
Saline M5
BV6
100 µm
ALT
Alk. Phos
Bile Acids
Total Bilirubin
1400 *
700 14 *
Saline *
250 *
1200
600 12
BV6
200
1000
500 10
units/L
800
units/L
400
umol/L
150 8
mg/dL
600
300 6
100
400
200 4
200 50
100 2
saline
BV6
saline
BV6
saline
BV6
saline
BV6
(N=8)
(N=8)
* p<0.05

8. Cholangiographic Evidence of Intrahepatic Bile Duct Alterations in Mice Injected with the SM
saline
BV6

9. CONCLUSIONS & WORKING MODEL
Cholangiocyte
apoptosis
macrophage
cIAPs
stellate cell
Fibro-inflammatory
cholangiopathy
cholangiocyte
Pro-inflammatory
Cytokines & Chemokines

10. Mdr2 Knock Out Mouse as a Model of PSC
Defect in cannalicular phospholipid transport
PSC-like cholangiopathy in the mouse
Obliterative biliary fibrosis
Primary dysfunction is in hepatocytes, abnormal bile composition  cholestasis and chronic biliary inflammation and fibrosis
Lazaridis & LaRusso, NEJM 2018
Lazaridis & LaRusso, Mayo Clin Proc 2016
Mariotti et al, Biochim Biophys Acta Mol Basis Dis 2018

11. Reichert MC and Lammert F. Seminars in Liver Disease 2018

13. Cenicroviroc – CCR2/CCR5 Inhibitor
Oral drug which blocks monocyte/macrophage recruitment to the liver
In clinical trials for PSC NCT
Completed enrolling (25 patients)
24 week study

14. BV6
BV6 + cenicriviroc (CVC)
CD68
C57BL/6 mice 5 days after intrabiliary BV6 injection, ± CVC (15 mg/kg, s.c., for 5 days)
CVC: CCR2/CCR5 antagonist

15. CCR2 inhibition “Cenicriviroc” Cholangiocyte insult CCL2
Monocyte-derived
macrophages
Circulating
monocytes
CCR2 inhibition
“Cenicriviroc”
Bile duct
Fibro-inflammatory
cholangiopathy

16. Ductular Reaction Aberrant reparative response to liver injury1
Comprise of reactive or activated cholangiocyte-like cells2
Proliferative
Pro-inflammatory
Pro-fibrotic
Form ductules and extend into liver parenchyma
Progenitor characteristics
Cholangiocyte or hepatocyte derived based on injury model
EpCAM, SOX9, CK7 and CK19, (also markers of cholangiocytes) are used to identify ductular reactive cells in cholestasis
Systemic injection of the cytokine TWEAK induces ductular reaction in normal mice3
Sato et al, Hepatology 2018
Banales et al, Nat Rev Gastroenterol Hepatol 2019
Bird et al, PNAS 2013
PanCK
SOX9
The left most panel is the schematic from the first slide showing a normal bile duct and these ductules with irregular or obliterated lumens and dyspmorphic cholangiocytes. This is reflected in the immunohistochemistry images taken from one of my study mice. And the point here is that you have ductules and then you have these cells tracking into the parenchyma.

17. Question: Does MCL-1 inhibition reduce ductular reaction and fibrosis in vivo?
Serum analysis
Immunohistochemistry
SOX9, PanCK, αSMA
Sirius red, Masson-Trichrome staining
S63845
40 mg/kg, IV, 5 days
Mdr2-/-

18. S63845 reduces fibrosis: Mdr2-/- mice
Vehicle
S63845
Trichrome
α-SMA
Reduction in fibrosis correlates with attenuated activation of fibroblasts

19. Animal Models of Cholangiocarcinoma

20. Summary
Animal models of PSC are important to understand mechanisms of disease and test preclinical therapies
PSC Partners Seeking a Cure has helped support critical research
THANKS FOR EVERYTHING YOU DO TO RAISE AWARENESS OF PSC AND SUPPORT RESEARCH RELEVANT TO THIS DISEASE

21. Acknowledgements Gores lab members Greg Gores Steve Bronk
Eugenia Guicciardi
Petra Hirsova
Tomohiro Katsumi
Anuradha Krishnan
Ayano Niibe
Genome Analysis Core
Biomedical Statistics and Informatics
Ying Li
Optical Microscopy Core