hepatic fibrosis basics to ongoing trials

1. HEPATIC FIBROSIS
Dr Srujan kumar
PGY 1 Gastro enterology
Gandhi Hospital

2. HEPATIC FIBROSIS
DEFN: Reversible wound healing response
characterised by accumulation of extracellular
matrix/scar
follows chronic, but not selflimited
liverdisease.

3. Natural history
Rapid fibrosers and slow fibrosers:
variable progression of fibrosis was first
highlighted in patients with Hepatitis C
Study emphasized the influence of both
modifiable and non modifiable factors

8. Cellular and molecular features
Between the sinusoid and hepatocytes
fenestrated endothelial cells line a basement
membrane which seperates the sinusoidal
lumen from space of Disse
Nutrients and other molecules reach the basal
surface of hepatocyte by passing through the
fenestrae of sinusoidal wall and across space
of Disse- This process is impaired by cellular
and matrix changes in liver injury

10. Cellular and molecular features
ECM COMPOSITION: Normal and scar tissue
Normal liver has connective tissue matrix
which includes type 4 /nonfibrillary collagen
Glycoproteins(fibronectin,laminin) and
proteoglycans(Heparan sulphate)

11. After Hepatic injury there is 3 to 8 fold increase in ECM
composed predominantly of high density interstitial fibril
forming collagens types 1 and 3 rather than type4 as well as
cellular fibronectin,hyaluronic acid and other matrix
proteoglycans and glycoconjugates
Gradual accumulation of type 1 collagen results from inceased
synthesis and reduced degradation :Hallmark of fibrogenesis

12. Stellate cell Activation: A central feature
Hepatic stellate cell/ HSC is the principal cell involved in
fibrogenesis
In the normal liver these HSC have intracellular droplets
containing vit.A .They contain 40-70%of body stores of
retinoids
In it’s quiscent stage the HSC produces predominantly
type4 collagen- characteristic of normal basement membrane
Activation : refers to phenotypic changes occuring in HSC
with injury

13. Phenotypic changes in HSC in Activation stage
• Loss of retinoid droplets
• Cellular proliferation
• Increased endoplasmic reticulum
• Increased contractility with expression of smooth muscle
specific a-Actin and secretion of cytokines and
chemokines
This phenotypic switch is also characterized by
production of type 1 collagen, the high density interstitial
collagen characteristic of cirrhotic liver as well as matrix
degrading enzymes

15. Stellate cell activation occurs in two stages :
INITIATION
PERPETUATION
Initiation refers to early events including rapid changes in
gene epression and a cellular phenotype
that renders HSCs responsive to cytokines and other
stimuli
Perpetuation involves cellular events that amplify the
activated phenotype through enhanced cytokine
expression and responsivess and acquisition of features
critical to development of fibrosis

17. characteristics of activated
HSCs.

19. Diagnosis and Assesment
• Accurate assesment of fibrosis is essential to guide
management and predict prognosis in patients with
chronic liver disease
1)Invasive (Histology of liver biopsy)- Gold std for
quantifying fibrosis
2)Non invasive : serology and Radiology tests

20. Three methods of staging in widest use are:
The Ishak score
Metavir score
Desmet/scheur staging system

26. Non invasive assesment
Direct markers : reflect ECM turnover
procollagen type 1carboxy terminal peptide(picp): increased
in cirrhosis.not accurate in ALD,HCV
Procollagen type 3(PIIINP) : serum levels corelate with
histologic stage of inflmmation,fibrosis in ALD,VIRAL
HEPATITIS,PBC
TYPE 1 ANDTYPE 4 COLLAGEN : increaed in all types of
fibrosis. Levels of type 1 corelated wih fibrosis score

27. Laminin : serum levels increase with peri sinusoidal fibrosis
co relate with severity of fibrosis,hepatitis,CTP ,HVPG
Hyaluronic acid:Have greatest predictive accuracy for advanced
fibrosis
YKL 40/CHONDREX: 38KD Glycoprotein with function in
remodelling and degradation of ECM.Levels corelate with degree of
fibrosis
Matrix metalloproteinase/mmp 2,3,9: only MMP 2 corelate with degree
of fibrosis
TIMP 1&2 : 100 %sensitivity for diagnosis in cirrhosis but low
specificity
CYTOKINES AND CHEMOKINES : TGF ALPHA,BETA,PDGF
Levels corelate with severity of liver dysfunction

28. Panels of direct markers of fibrosis
Direct markers have been combined into panels to predict
fibrosis.The panels may also include indirect markers
FIBROSPECT II :serum hyaluronic acid,TIMP
alpha 2macroglobulin
sn 77% sp73% in a study of 402pts with HCV
SHASTA : serum hyaluronic acid,serum AST
Albumin level
Evaluated in pts with hiv/hcv co infection
Cutoff value <0.30(sn 88%, npv>94%) for significant fibrosis

29. Indirect markers
Reflect alteration in hepatic function
AST to PLATELET RATIO INDEX(APRI):
meta analysis of 40 studies found that for
predicting significant fibrosis(F2 to F4) ,APRI
cutoff of 0.7 had sensitivity of 77% and sp
72%
APRI appears most useful for excluding
significant fibrosis in HCV

30. Fibotest/fibrosure and Actitest
Fibrotest : alpha2 macroglobulin,alpha2
globulin(haptoglobin),gamma globulin,apolipoprotein
A1,GGT and Total bilirubin
Sensitivity for detection of significant fibrosis is 60to 75% and
sp 80-90%
Actitest is a modification of above that incorporates ALT and
reflects both fibrosis and necroinflammatory activity
Metanalysis including 1570pts concluded that these tests were
reliable alternatives to Liver biopsy in pts with chronic HCV

31. HEPA SCORE:Bi ,GGT,Hyaluronic acid,Alpha2
macroglobulin,Age ,Sex
AST/ALT ratio: more than 1 suggests cirrhosis
utility is uncertain due
to inconsistent results

32. Other indirect markers
• FIB 4 INDEX: combines biochemical values(Plt count,ALT and
AST) and age.
useful in pts with HCV,NAFLD
Values have been associated with risk of development of HCC in
those who consume alcohol(>1.75)
NAFLD FIBROSIS SCORE: Age,BMI,Blood
glucose,Aminotransferase levels,PLT count,Albumin
sn 43%,sp96% for advanced fibrosis(F3 to F4)
PGA INDEX: PT index,GGT,Apolipoprotein A1
Validated in pts of ALD
Accuracy for detection of cirrhosis is 66to 72%

33. • Fibro index(plt count,AST,gammaglobulin)
accuracy is still being determined
• FORNS INDEX(age,GGT,cholesterol,plt)
performance characteristics similar to those seen with APRI
• FIBROMETER: Plt count,PT index,AST,
Alpha2macro globulin
hyaluronic acid,BUN,age
performed well in predicting severe fibrosis in
chronic viral hepatitis

34. • BARD score: Developed to predict fibrosis in
NAFLD
BMI, AST/ALT, DM
Sudies showed ppv and Npv of 69 and 96% for
advanced fibrosis

35. Radiologic tests
Radiologic methods are emerging as promising tools
USG based elastography( most studied radiologic test)
Magnetic resonance elastography(MRE)
Acoustic radiation force impulse imaging(ARFI)
Cross sectional imaging

38. Magnetic resonance elastography

39. MR Elastography
• Detecting any fibrosis (F1): cutoff 3.45kpa
• Detecting significant fibrosis(F2): cutoff 3.66kpa
• Detecing advanced fibrosis(F3)optimal cutoff
4.11kpa
• Cirrhosis(F4): optimal cutoff 4.71kpa
Higher technical success and diagnostic accuracy
compared to US elastography
Advantage of scanning entire liverand thus doesnot
depend on acoustic window
May also detect lesions with in liver .eg: HCC

40. Acoustic radiation force impulse imaging
(ARFI)
Another USG based approach for estimating liver
stiffness
Uses short duration high inensity acoustic pulses to
produce mechanical excitation in tissue.This results in
localized displacement and shear wave propogation
Velocity of these waves corelate with degree of fibrosis
Cutoff values: 1.44m/s ( F2 ) 1.9m/s (F4)

43. Treatment of Fibrosis

46. Inhibition of Hepatic Injury
• PAN-CASPASE INHIBITOR: IDN-6556
attenuated hepatic injury and fibrosis in mice
• OCADEIC ACID protects in part from liver
fibrosis by protecting hepatocytes from injury
• ANTI OXIDANTS: regulating ROS is a promising
strategy of liver fibrosis therapy
• PPARδ agonist protects hepatocytes from cell
death by reducing ROS generation of
hepatocytes, leading to less liver fibrosis

47. Nuclear Receptors
• PPAR𝛾 and Farnesoid X receptor (FXR), that play
an important role in HSC regulation
• PPAR𝛾 ligands/agonists, PPAR𝛾 expression is
restored, and HSC activation and collagen
expression are reduced in vitro
• obeticholic acid, semisynthetic FXRagonist,
showed improvement of the histological and
biochemical markers, ameliorated fibrosis,
inflammation, and steatosis in NASH patients
• Obeticholic acid :clinical trials for long-term
treatment of cholestatic liver diseases

48. registered clinical trials
(Clinicaltrials.gov)

49. registered clinical trials
(Clinicaltrials.gov)

50. Inhibition of Inflammation
• Serum amyloid P (SAP) or pentraxin-2, a member of the pentraxin
family, is a 27-kDa protein that is produced by the liver, secreted
into the blood, and circulates as stable 135-kDa pentamers .
• SAP reduces
1. neutrophil adhesion to ECM proteins
2. inhibits the differentiation of monocytes into fibrocytes
3. decreases profibrotic macrophages
4. activates the complement pathway
5. promotes phagocytosis of cell debris.
• SAP reduces bleomycin-induced lung fibrosis .
• Injection of SAP into humans, mice and rats has no toxic effects.

51. Inhibition of Activation of HSCs
• Fresolimumab (GC1008) is a human anti-TGF-
β1 monoclonal antibody that neutralizes all
isoforms of TGF-β
• TGF-β transduces its signal to target genes
through the ALK5 Ser/Thr kinase receptor.
GW6604 , an ALK5 inhibitor, inhibits the
transcription and deposition of ECM and
improves the deterioration of liver function in
mice

52. • Integrin αvβ1 expressed on activated HSCs.
Inhibitor of αvβ1 prevents TGF-β1 activation
and inhibits experimental liver fibrosis
• Lysophosphatidic acid (LPA) is a lipid mediator,
which is produced mainly by activated
platelets. LPA1R antagonists showed anti-
fibrotic effect on models of liver fibrosis

53. registered clinical trials
(Clinicaltrials.gov)

54. Inhibition of Proliferation of HSCs
• Inhibitors of receptor tyrosine kinase and Ser/Thr
kinase also demonstrate anti-fibrosis effects
• tyrosine kinase inhibitor sorafenib :HCC, and
sunitinib can improve experimental hepatic
fibrosis, inflammation, and angiogenesis
• SiRNA melastatin 7, a nonselective cation
channel with protein serine/ threonine kinase
activity, attenuates TGF-β1-induced expression of
myofibroblast markers, increases the ratio of
MMPs/TIMPs, and decreases the phosphorylation
of Smad2 and 3 associated collagen production

55. • Hepatic nuclear factor kappa B inducing kinase, a
Ser/Thr kinase – target for liver fibrosis therapy
• RAS pathway in HSCs induces ROS and
accelerates hepatic fibrosis , accelerates
inflammation, tissue repair and fibrogenesis by
production of angiotensin II
• HALT-C cohort study did not show any
antifibrogenic effects of ACEi/ARB for chronic
hepatitis C patients
• Activation and proliferation of HSCs require NOX/
ROS signaling.

56. registered clinical trials
(Clinicaltrials.gov)

57. registered clinical trials
(Clinicaltrials.gov)

59. NOX
• HSCs express 3 NOX isoforms, NOX1, NOX2, and NOX4.
• NOX2 :phagocytic cells use to produce ROS to kill bacteria,
not a good target for anti-fibrotic therapy.
• HSCs from p47 phox -deficient mice (without a regulatory
component of NOX) fail to generate ROS in response to Ang
II, PDGF, leptin, or apoptotic bodies, and p47 phox -
deficient mice demonstrate reduced liver fibrosis after BDL
or the hepatotoxin CCl4 .
• NOX1 and NOX4 are expressed as activated HSCs, but only
at very low levels in the uninjured liver.
• GKT137831, a potent dual NOX1/ NOX4 inhibitor,
attenuates ROS production and inhibits activation of HSCs
and experimental liver fibrosis

61. Promotion of Apoptosis of Activated
HSCs
• CB1 and CB2 receptors are increased in liver
fibrosis.
• CB1 agonists activate HSCs to myofibroblasts.
• CB1 receptor agonists, Rimonabant, inhibit and
reverse experimental liver fibrosis.
• MMP: Monoclonal Anti-TIMP1 Ab partially
reverses established CCl4-induced fibrosis
• a decrease in TIMP-1 protein levels correlated
with decreased numbers of HSCs

62. Promotion of HSC Inactivation
• Inactivation of HSCs is associated with the re-
expression of lipogenic genes PPAR-γ, Insig1,
and CREBP.
• PPAR-γ is reported to be important for
maintaining and for re-establishing the
quiescent phenotype

63. Inhibition of Deposition of Type I
Collagen
• In liver fibrosis, ↑ type I collagen . cross-linking of
type I collagen is also increased , enzyme
lysyloxidase-like-2 (LOXL2).
• inhibition of LOXL2 by a monoclonal antibody
(AB0023) reduces the production of cytokines,
attenuates TGF-β signaling, and inhibits the
activate fibroblasts (Barry-Hamilton, 2010).
• Similar to AB0023, another humanized
monoclonal LOXL2 antibody (GS-6624) is in
randomized, double blind, phase II clinical trials
to treat NASH and PSC

64. registered clinical trials
(Clinicaltrials.gov)