Prevention of Liver Fibrosis to Cirrhosis Hongqun Liu University of Calgary

Categories Eliminate the causative agent(s) of liver injury; Directly downregulate HSC activation; Reduce inflammation or modulate inflammatory cells; “Hepatoprotection” to reduce hepatocyte injury; Inhibit fibrogenesis; Neutralize proliferative, fibrogenic, contractile and/or proinflammatory factors; Induce apoptosis of activated HSC; Increase the degradation of extracellular matrix (ECM).

Quiescent HSC Injury (inflammation) Activated HSC Proliferation Matrix synthesis Apoptosis

Anti-fibrosis Ideal drugs: “Anti-fibrosis” Anti-inflammation Anti-angiogenesis

Anti-inflammatory drugs Corticosteroids Useful for autoimmune hepatitis. But long termadministration is not recommended. Interleukin 10 Shifts cytokines from proinflammatory to anti-inflammatory, decreases TNF α, IL-1 and 2 and IFN γ. However, increases viral load and therefore, not suiltabefor viral hepatitis. Anti-TNF α Effective in cell culture of fibroblasts and HSC, not human

Antioxidants “in vitro” and in experimental animals; limited evidence in humans. N-acetyl-cysteine (NAC): in cell cultures and inexperimental animals, its antifibrosing properties islimited in humans Cu-Zn superoxide dismutase: Emerit mentioned theclinical trail in 2006 but so far no further document. Curcumin and silymarin: natural products, antioxidant and inflammation (experimental)

HSC proliferation TGF β (Transforming Growth Factor beta) PDGF (Platelet derived growth factor) EGF (Epidermal Growth Factor), CTGF (Connective Tissue Growth Factor), VEGF (Vascular Endothelial Growth Factor), IGF (Insulin-like Growth Factor), bFGF (basic Fibroblast Growth Factor), RAAS (renin-angiotensin-aldosterone system) TGF α ( Transforming Growth Factor alpha) Leptin, endothelin, thrombin, IL-1, endocannabinoids

TGFβ The most powerful fibrogenic factor Inducing hepatic oval cells to HSC (You hong) Inducing fibrogenic related gene transcriptions Favoring HSC activation Reducing ECM degradation Side effects of TGFβ blockage: Affecting the tissue and organ architecture such as bone integrity; excessive inflammation.

Collagen Synthesis: 4-prolylhydroxylase. Safironil. Activated in liver, experimentally useful, not in human. Secretion: colchicin, inconclusive in human Formation of cross-linking among collagen molecules: D-Penicillamine good for Wilson’s disease.

HSC inactivation PPARγ (Peroxisome Proliferator-Activated Re- ceptors): PPARγ ligands reduce activation of HSC, intensity of fibrosis, expression of TGFβ and proin- flammatory cytokines TNFα and IL-6, and increases PPARγ expression in HSC Retinoic acid ( in cell culture only, not animal and human) IL-10 Trans-resveratrol, Pentoxifylline inhibits HSCs proliferation

HSC apoptosis Fas ligand and TNFα, P53, BCL2/Bax, NK and NKTγδ cells, MMPs, IFNγ RAIL receptor 2 (TNF-related apoptosis- induced ligand) (TRAIL-R2) Inhibition of PI 3-K/Akt signaling pathway induces apoptosis in HSC (Jiang) NF-κB knockdown enhances hepatic stellate cell apoptosis (Jiang)

Drugs for HSC apoptosis Sorafenib (Tumor) J Hepatol. 2010;53:132-44J Hepatol. reduces HSC proliferation; induces apoptosis. downregulates Cyclin D1 and Cyclin-dependent kinase 4 (Cdk-4), Bcl-2/Bax, collagen synthesis. ERK, Akt phosphorylation increases Fas, Fas-L, and Caspase-3, increases MMPs/TIMPs. Nilotinib (Leukemia) J Hepatol. 2011;55:612-25J Hepatol Ursolic acid (tumor) J Hepatol. 2011;55: J Hepatol. Gliotoxin (immunosuppressant) J Hepatol. 2007; 47:

Is the fibrosis/cirrhosis reversible? Animal experiments CCL4, fibrosis/cirrhosis disappears spontaneously when CCl4 discontinue Bile duct ligation, liver will be back to when bile duct ligation is released. Human Spontaneous regression of liver fibrosis when causative agent of disease is eliminated

Hurdles for clinical validation 1.Slow progression of liver fibrosis 2.Lack of sensitive and specific markers to evaluate progression and regression. 3.Well-stratified, high number of patients, long term treatment.

Well-stratified (1) Recent fibrosis, as characterized by the presence of thin reticulin (collagen) fibres, often in the presence of a diffuse inflammatory infiltrate, appears to be fully reversible; Histopathological evidence of significant fibrosis (F ≥2) incurs a high risk for progression to cirrhosis and is thus an indication for anti- fibrotic treatment.

Well-stratified (2) Patients suitable for the anti-fibrogenic clinical trials: Rapidly progressing fibrosis such as in HCV re- infection after liver transplant or in HCV–HIV coinfection; 1–2 years trial; NASH, Cholestatic CLD such as PBC, PSC or pediatric biliary liver diseases; Nonresponders to standard antiviral treatment

Well-stratified (3) Long-standing fibrosis, as characterized by thick collagen fibrils embedded in an acellular or paucicellular ECM and consequent decreased expression and/or activity of fibrolytic MMPs, is not easy to reverse

Challenges from basic study to clinical application Time for progression and regression in humans is measurable in years Genetic differences affecting fibrosis progression and regression Insufficient performance of both liver biopsy and noninvasive methods to differentiate within and between a single stage of fibrosis (0-4), Ethical problems to include a placebo group; Lack of data on the efficiency of a given treatment in CLD with different etiologies Difficulty to assess the impact of contributing factors (i.e. obesity/overweight/insulin resistance, alcohol and tobacco consumption Realistic endpoints

Fibrosis “scores” correlated with HVPG, ascites, INR, bilirubin and inversely with albumin; as well as Child-Pugh class 24 As a result of liver injury, quiescent vitamin-A-rich hepatic stellate cells undergo 'activation' towards proliferative, fibrogenic and contractile myofibroblasts excessive amounts of collagens, downregulate matrix metalloproteinases (MMPs) and show an enhanced expression of the tissue inhibitors of the MMPs ( TIMP-1 & - 2). During resolution of liver injury, hepatic stellate cells might revert to a quiescent phenotype and/or be selectively cleared by apoptosis. Scott L Friedman : Nat Clin Pract Gastroenterol Hepatol 1(2),2004. Hepatic stellate cell activation 24