Du 2016 tp biomarkers

7 févr. 2012

LiverCenter

Non Invasive Biomarkers in chronic hepatitis C and B

DU 2016

Thierry Poynard +

AP-HP Groupe Hospitalier Pitié Salpêtrière, UPMC Liver Center, Université Paris 6, INSERM U680, Biopredictive France

Serum Biomarker Imaging Biomarker

FibroTest FibroMax

Choice Hepatologist

Epidemiologist GP

FibroScan Aixplorer

2

Biomarkers of liver injury in chronic hepatitis

• Unmet need

• Historic

• Methods and based evidence

• Guidelines in practice

2

FRANCE FIBROSIS ICEBERG

Biopsy: 2% (8 000 /yr) FibroTest: 10% (50 000/yr) Imaging: 10% (50 000/yr)

No-estimate: 78%

0.25 Million Chronic Hepatitis C 0.25 Million Chronic Hepatitis B

USA FIBROSIS ICEBERG

Biopsy: 1% (50 000 /yr) FibroSure: 1% (50 000 /yr) Imaging: 1% (50 000 /yr)

No-estimate: 97%

3 Millions Chronic Hepatitis C 1 Million Chronic Hepatitis B

"META-analysis of histological data in VIRal hepatitis"

Hepatology 1996

METAVIR

THE METAVIR cooperative group. Inter- and intra-observer variation, Hepatology 1995

7 févr. 20127 févr. 2012

Viral necrosis Activity

Fibrosis Steatosis

Alcohol Ash

Nash

Liver Injury

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7 févr. 2012

FibroMAX: HCV-HBV-ALD-NAFLD

ActiTest

FibroTest SteatoTest

AshTest

NashTest

FibroMAX

7

2


• Unmet need

• Historic



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7 févr. 2012

Fibrosis biomarkers: 24 years history

SJG 2008

n=100

n=1 million

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Haptoglobin

Alpha2Macroglobulin

Apolipoprotein A1

Total Bilirubin

Gamma GT

In Situ In Serum: FibroTest

Imbert-Bismut, Lancet 2001

Liver Injury

Activated Stellate CellsFibrotic Matrix

7 févr. 2012

Rational of FibroTest:

• Alpha 2 macroglobulin: key protein for Collagenase metabolism

• Apolipoprotein A1 key protein for Collagen trapping

• Haptoglobin: key protein for binding Free Hemoglobin oxidant

• Total Bilirubin: specific marker of severe late Fibrosis

• Gamma Glutamyl Transpeptidase: sensitive marker of early Fibrosis

• No transaminases: to prevent inflammatory necrosis confusion (ActiTest)

• Proteomic has blindly proved the major diagnostic value of

• Apolipoprotein A1, A2M

• HaptoglobinParadis Cell Mol Biol 1996, Paradis Hepatology 1996, Mathurin Hepatology 1996, Imbert Bismut 2001, Langlois 2006, Watanabe 2009, Ho 2010

2


• Unmet need

• Historic



2

7 févr. 2012

Period 1: 1991-2004 Optimistic

Looking for a fibrosis biomarker with accuracy > 90%

7 févr. 2012

Biopsy =

Gold Standard

Biopsy=

0% False Positive0% False Negative

7 févr. 2012

Liver Injury

Serum biomarker Imaging biomarker

F4

F1

F0

Fibrotic Liver Disease

F2

F3

Hemorrhage Liver failure Cancer

FibroTest OK AUROC >80%

FibroTest OK FibroScan OKAUROC >80%

«Gray Zone»: Biopsy

Imbert Bismut 2001, Castera 2005

7 févr. 2012

Period 2: 2005-2009: Sceptic

Standard statistical methods were inappropriate

Period 3: 2010-2015

New methods

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• Sampling error Bedossa 2003

• Inter-observers variability Rousselet 2005

• Discordance studies Poynard 2004, Halfon 2006

• Prognostic studies Ngo 2006, Vergniol 2011

• Spectrum effect Poynard 2007, Lambert 2008

• Exceeding limits of biopsy Metha 2009

• Biopsy has a gray zone Poynard 2012

• Direct meta-analyses Poynard 201522

8 Key methodological issues:Biopsy is no more a perfect gold standard

Sampling error:AUROCs (F1 vs F2) of Biopsy vs Whole Liver according to length

Bedossa Hepatology 2003

AUROC 15 mm = 0.82 AUROC 25 mm = 0.89

«We showed that with 25-mm long biopsy specimens, only 75% were scored correctly»

7 févr. 2012

F0 F1 F2 F3 F4

Inter-Observers variability:Biopsy has lower inter-observers concordance for intermediate stages

Rousselet, Hepatology 2005

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Discordances studies: independent endpoints

• 537 prospective cases hepatitis C

• 154 (29%) discordances FibroTest/Biopsy

• Error attributable

• To FibroTest: 2%

• To Biopsy: 18%

25

Poynard Clin Chem 2004, Halfon AJG 2006

F4.1

F1

F0

F2

F3

7 Stages Presumed by Biomarkers

Decompensated

F4.2

F4.3

Varices

FibroTest

0.48

0.74

0.85

0.95

TE

7.1

9.5

20

50

CHC Poynard J Hepatol 2014 CHB Poynard J Hepatol 2014

12.5

0.58

F4.1

F1

F0

F2

F3

7 Stages Presumed by Biomarkers

Decompensated

F4.2

F4.3

Varices

FibroTest

0.48

0.74

0.85

0.95

TE

7.1

9.5

20

50

CHC Poynard J Hepatol 2014 CHB Poynard J Hepatol 2014

12.5

0.58

25

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29

3/7 key methodological issues not well understoodBiopsy is no more a perfect gold standard

F4

F1

F0


F2

F3

DANA=4

DANA=Difference between Advanced and non-advanced fibrosis stages

Obuchowski measure=AUROCs Pair-wise comparison between all stages

Black and White Spectrum

FibroTest AUROC=0.98

F4

F1

F0


F2

F3

DANA=1



Gray Spectrum


F4

F1

F0


F2

F3

DANA=2.5




Standard Spectrum

7 févr. 2012

Hazardous AUROC Scores for defining test performance:

AUROC Score* Biopsy (length) FibroTest (Spectrum)

0.90-1.00 Excellent 100mm F1 vs F2 F0 vs F4

0.80-0.90 Good 25 mm F1 vs F2 F01 vs F234

0.70-0.80 Fair 5 mm F1 vs F2 F0 vs F2

0.60-0.70 Poor 5 mm F0 vs F1 F1 vs F2

0.50-0.60 Fail

*Sebastiani CCLM 2011, Bedossa Hepatology 2003, Poynard Clin Chem 2007

7 févr. 2012

Hazardous AUROC Scores for defining test performance:

AUROC Score* FibroTest Spectrum DANA

0.90-1.00 Excellent 50% F0 vs 50% F4 4

0.80-0.90 Good F01 vs F234 20% each stage 2.5

0.70-0.80 Fair 50% F0 vs 50% F2 2

0.60-0.70 Poor 50% F1 vs 50% F2 1

0.50-0.60 Fail

Poynard Clin Chem 2007, Lambert Clin Chem 2008,

7 févr. 2012

Hazardous Tables due to Spectrum Effect (October 2012)

Ochi Hepatology 2012

Real-time tissue elastography cut-off values by stage in the training set were 2.47 for F1, 2.67 for F2, 3.02 for F3, and 3.36 for F4. Usingthese cut-off values, the diagnostic accuracy of hepatic fibrosis in the validation set was 82.6%-96.0% in all stages.

The area under the receiver operating characteristic curve of elastic ratio better correlated than serum fibrosis markers in both early and advanced fibrosis stages.

Conclusion: Real-time tissue elastography is useful in evaluating hepatic fibrosis and PH in patients with NAFLD. (HEPATOLOGY 2012;1271-1278)

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29


Using 25 mm liver biopsy a perfect market cannot be validated

Black shading represents the set of conditions under which the AUROC values exceed what has already been observed

Metha J Hepatol 2009

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Exceeding limits of biopsy: >90% accuracy is impossible for advanced fibrosis

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«Comparison of 8 diagnostic algorithms for liver fibrosis in hepatitis C: New algorithms are more precise and entirely non-invasive».

Boursier et al, Hepatology 2012

7 févr. 2012

Misleading presentation using biopsy as Gold-Standard

Boursier Hepatology 2012

Mathematically impossible with biopsy as «Gold Standard

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Review of tests by Gebo, Hepatology 2002

« These panels of tests may have the greatest value in predicting fibrosis or cirrhosis »

« Biochemical tests were best at predicting no or minimal fibrosis, or at predicting advanced fibrosis/cirrhosis, and were poor at predicting intermediate levels of fibrosis »

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FibroTest/FibroSure has a Gray Zone

Biopsy has a Gray Zone

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Review of fibrosis tests by Nguyen, Hepatology 2011

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Liver Biopsy Analysis Has a Low Level of Performance for Diagnosis of IntermediateStages of Fibrosis

The gray anatomy of 27,869 virtual biopsies and 6,500 patients

Poynard Clin Gastro Hepatol 2012 Poynard, BMC 2005, J Hepatol 2011

7 févr. 2012

The gray zone of liver biopsy: 27,864 virtual biopsies

Area Fibrosis (Log)

25 mm Liver Biopsies

Poynard Clin Gastro Hepatol 2012

7 févr. 2012

The gray zone of liver biopsy: 27,864 virtual biopsies


Area Fibrosis (Log)

25 mm Liver Biopsies

Lower gray zone of FibroTest relative to biopsy

Lower gray zone F2vsF1 for FibroTest vs Biopsy

58% lower F2vsF1 vs F1vsF0 41% lower F2vsF1 vs F4vsF3.

Biopsyn=27,864

Fibrotestn=6500


7 févr. 2012

Biopsy is no more a perfect gold standard

FibroTest and Elastography have similar performance

2006: Approval Markers French Health Authorities HCV2011: Guidelines EASL 2011

7 févr. 2012

Period 2: 2005-2009: Sceptic

Standard statistical methods were inappropriate

Period 3: 2010-2015

New methods

(c) BioPredictive 2008 - All Rights Reserved - No reproduction without written permission

Benefit/Risk must be evaluated for each change in the formula:

High Risk False Positive Negative

5/954 (0.52%)


38/7494 (0.51%)

FibroTest Global Quality Estimates

High Risk False Positive Negative 3349/345,695 (0.97%)


491/24,872 (1.97%)

FibroScan (Roulot et al 2008) >7.1 kPa= 12.6%: False Positives ?

Poynard BMC Gastro 2011, Roulot J Hepatol 2008

(c) BioPredictive 2008 - All Rights Reserved - No reproduction without written permission

One Test, One formula

360,000 FibroTest for Quality Control

Risk of False positive/negative of FibroTest

• Tertiary center: 1.97%

• HIV co-infection: 1.77%

• Sub-Saharan origin: 2.61%

7 févr. 2012

Which Fibrometer for patients with Hepatitis C ? Too many variants = Risk of false positive

FibroMeter Variant Year Components

FM-1G 2005 PLT, PI, AST, A2M, HA, Urea, Age

FM-2G V* 2008 + Gender

FM-3G 2008 Switch GGT/HA

FM-3G+ (CirrhoMeter) 2009 New formula for cirrhosis

FM-HICV 2010 AST, A2M, PI

CSF-Index 2011 Combined with LSM

SF-Index 2011 Combined with LSM

C-Index 2011 Combined with LSM

*ONLY one ( FM-2G V) is approved by Haute Autorité de Santé

PLT: platelet counts, PI prothrombin index, AST aspartate amino transferase, A2M alpha2 macroglobulin, HA hyaluronic acid

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Biopsy vs Serum marker Main advantages/disadvantages

Serum Marker FibroTest

Less accurate for intermediate stages

No grey zone relatively to biopsy

Fibrosis only ActiTest/SteatoTest

Delays result proprietary tests 1-48h

False positive/hemolysis/inflammation/Gilbert

Yes but 0.97% (3349/345695; 0.94-1.00)

Nguyen Hepatology, 2011 Poynard BMC Gastro 2011

7 févr. 2012

Period 3: 2010-2015

Welcome in a world without perfect Gold Standard

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Gold Standard

25 mm Biopsy 0%False PositiveFalse Negative

7 févr. 2012

Truth in the Absence of

Gold Standard

25 mm Biopsy 25%False PositiveFalse Negative

7 févr. 2012

Area of fibrosis estimated by biopsy according to its length (mm) in subjects scoring METAVIR F0 (no fibrosis) on large surgical section.

Area of fibrosis >5.3%: 16.3% false positives 20mm biopsy for diagnosis of advanced fibrosis >16.5%: 0.3% false positives 20mm biopsy for diagnosis of cirrhosis.

Cirrhosis

Advanced fibrosis

Poynard J Hepatol 2012

7 févr. 2012

Poynard J Hepatol 2011

Truth

FibroTest FibroScan

5-30 mm Biopsy

ALT

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Distribution of 1893 subjects according to the 16 possible combinations of the 4 tests' results: presumed advanced fibrosis (present=1) or not (=0)

16 combinations of 4 tests results Number of subjects

FibroTest LSM ALT Biopsy Observed Expected by model

0 0 0 0 621 615.5

0 0 0 1 186 191.1

...

1 1 1 1 276 277.0

Poynard, J Hepatol 2011

7 févr. 2012

FibroTest Se LSM Se Biopsie Se

Performance for Cirrhosis: Sensitivity

The standard cutoffs: 0.74 FibroTest, 14.5 kPa Stiffness


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FibroTest Sp LSM Sp Biopsy Sp

Performance for Cirrhosis: Specificity

The standard cutoffs: for cirrhosis 0.74 for FibroTest, and 14.5 kilo-Pascal for stiffness (LSM)


7 févr. 2012

SWE Fibrotest 1 TE-M Fibrotest 2 TE-XL FibroTest 3


Performances for diagnosis of Cirrhosis (HCV, HBV, NAFLD, ALD) of FibroTest, and Elastography: Transient M-XL probes and Share Wave

Latent Class Model: Best model for FibroTest with TE-XL or SWE (Likelihood ratio test 5.5, 6.9)

n = 322 simultaneous reliable tests

2


• Unmet need

• Historic



2


FibroTest FibroMax

Choice Hepatologist

Epidemiologist GP

FibroScan SWE Aixplorer

Competitors

65

• AFEF, HAS

• EASL, AASLD

• WHO

Guidelines: HCV and HBV

7 févr. 20127 févr. 2012

FibroTest APRI FIB4TE

Imbert-Bismuth Lancet 2001

Sandrin Ultra Med Biol

2003

Wai Hepatology

2003

Sterling Hepatology

2006

• Applicability

• Variability

• False positive if activity

• False negative for cirrhosis

Fibroscan limitations

7 févr. 2012

Pitfalls of Fibroscan

3.1% Failures and Unreliable results 15.8%

23 sept. 2014

Performances for cirrhosis diagnosis

FibroTest Fibrosure Transient elastography

AUROC* 0.86 (0.71-0.92) 0.94 (0.93-0.95)

Applicability >95% 80 %

Afdhal, JVH Nov 2013 Chou, Ann Int Med 2013

Not in intention to diagnose

7 févr. 2012

Oliveri WJG 2008

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Choice of FibroScan Cutoffs

Castera 2005, Ketanneh 2007 Roulot 2008

For F2: 7.1 or 8.8 kPa ? Patients: false negatives ? Low negative predictive value

Healthy volunteers: 7.1 kPa 12.6% false positives ?

For screening 7.1 kPa ?

For patients 8.8 kPa ?

No rationale for changing cutoff according to liver disease

F2 8.8 kPa F4 14.5 kPa

F4 0.73

F2 0.48

Poynard PlosOne 2008

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Elasto-FibroTest® 1289 patients with CHC and 604 healthy volunteers

• For the diagnosis of cirrhosis Elasto-FibroTest has significantly higher performances than FibroTest or Fibroscan alone.

• For the diagnosis of advanced fibrosis (F234) no improvement in performance has been observed vs FibroTest alone, when a method without gold standard was used.

67

Poynard, CRHG 2012

8-week $ 63,000 12-Week $ 94,500 24-week $ 189,000

$ 1125 per pill

FibroTest used to identify cirrhosis at baseline or for follow-up

Number of Studies Quality Consistency Precision Strength of

evidence

32 Fair High High High

Chou, Ann Int Med 2013

FibroTest is the most validated test in Chronic Hepatitis C: Strength-of-evidence domains and overall ratings

Results:

Area under the ROC curves:

Significant fibrosis 0.84 (0.78 – 0.88) Cirrhosis 0.87 (0.85 – 0.90)

Am J Gastro 2013

APRI has lower performance than Fibrotest

due to its high variability

1. Analytical variability: ULN-AST definitions

2. Interaction with non-fibrosis features

• Activity and AST

• Steatosis and AST

Impact on performance: Obuchowski measure

Change in AUROCs between fibrosis stages

ULN 26 IU/L ULN >=30 IU/L

APRI 0,862 0,820

FibroTest 0,867 0,867

Significance 0,30 <0,0001

High variability of APRI associated with ULN definitions:

Impact on diagnosis performance

• Range of AST-ULN in controls: 26-49 IU/L

• According gender, BMI and cholesterol

• Fibrosis prevalence in CHC: spectrum effect

• Clinically significant fibrosis (F2F3F4): 35-69%

• Cirrhosis (F4): 11-32%

Higher Diagnostic and Prognostic performance of FibroTest versus APRI in CHC

Poynard, Ann Int Med 2013

5 years prognostic value in chronic hepatitis B FibroTest better biomarker

de Ledinghen APT 2013

LSM Not in intention to diagnose

10 year Prognostic value FibroTest versus TE n= 272445% CHC, 24% CHB, NAFLD 10%, ALD 6%

Biomarker

Analytic variability

Risk false positive due to

ActivityFasting Applicability Investment Cost

FibroTest < 7% no (ActiTest) no >95% 0 38€-200$

FibroScan Inter Observer yes yes 80 % 60,000€ 200,000$ 38€-350$

APRIHazard of AST

Upper Limit Normal

yes no ? 0 7€-40 $

Castera Hepatology 2010, Afdhal JVH 2013, Chou Ann Int Med 2013, Poynard BMC Gastro 2011, Poynard Ann Int Med 2013

Pro and Con of the three «Standard of Care» biomarkers

2


• Unmet need

• Historic



2

2016

Direct comparisons between APRI, FIB4, FibroTest and TE n=185 comparisons: 99 Fibrosis n=12,725 / 86 Cirrhosis n=10,929

Houot APT 2016

FT better than TE

AUROC + 0.06

Fibrosis

Cirrhosis

FT = TE

AUROC + 0.00

13

Three reasons to assess fibrosis stages in 2016

•Expensive IFN-free regimen, priority to severe fibrosis

•Cirrhosis could need longer treatment

•Viral cure is not fibrosis cure

Cohen, Science 2013, Afdahl NEJM 2014, Poynard J Hepatol 2013

FibroTest similar to biopsy for estimating fibrosis progression

Progression to cirrhosis in 2472 patients

Biopsy FibroTest

Poynard et al, J Hepatol 2012

Response is associated with longer treatment in cirrhosis stage* vs non-cirrhosis in experienced Genotype 1, treated by Sofosbuvir-Ledispasvir:

80

85

90

95

100

12 weeks 24 weeks

Cirrhosis No Cirrhosis

Afdahl NEJM 2014*cirrhosis stage defined by biopsy or FibroTest

n=22 n=22n=87 n=87

23 sept. 2014

Survival without liver complications

n = 933NS

SVR n=43 HCC1 CholangiocarcinomaAll F4 before SVR2 F2 after


Cirrhosis regression in SVR n=24/43 ( 56%)

FibroTest = 0.74


Cirrhosis Occurrence in SVR n=15/128 (12%)

FibroTest = 0.74


Fibrosis Progression in 13 HBV Sustained Virological Responders with occurrence of HepatoCellular Carcinoma at 10 years

FibroTest = 0.74 = F4


F1: Subsaharan female, BMI 37 kg/m2

F4

F1

F0

France: 12,000,000 at Risk100%

5%

Death 15,000/year0.1%

Biomarker10% F2

F3

n= 1,016,557 (100 %) ActiTest

Poynard BmjOpen 2015

Fibrosis density Birth-Year and Gender

Other

USA

France

n= 470,762


FibroTest FibroMax

Choice Hepatologist

Epidemiologist GP

FibroScan Aixplorer

Du 2016 tp biomarkers

Health & Medicine