du 2015 poynard biomarkers

7 févr. 2012

LiverCenter

Non Invasive Biomarkers in chronic hepatitis C and B

!DU 2015

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

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Viral necrosis Activity

Fibrosis Steatosis

Alcohol Ash

Nash

Liver Injury

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FibroMAX: HCV-HBV-ALD-NAFLD

ActiTest

FibroTest SteatoTest

AshTest

NashTest

FibroMAX

7

2


• Unmet need

• Historic



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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



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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

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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

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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

12.5

0.58

CHC Poynard J Hepatol 2014 CHB Poynard J Hepatol 2014

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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

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Hazardous Tables due to Spectrum Effect (1)

Interpretation of AUROC

AUROC Score* Biopsy length FibroTest and Spectrum

0.90-1 Excellent 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

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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. Using these 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

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

Area Fibrosis (Log)

25 mm Liver Biopsies

Poynard Clin Gastro Hepatol 2012

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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


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Biopsy is no more a perfect gold standard!

FibroTest and Elastography have similar performance!!!!!

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

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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

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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

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Truth in the Absence of

Gold Standard

25 mm Biopsy 25%False PositiveFalse Negative

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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 Advanced Fibrosis: Sensitivity

The standard cutoffs: 0.48 FibroTest, 8.8 kPa Stiffness


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FibroTest Sp LSM Sp Biopsy Sp

Performance for Advanced Fibrosis: Specificity



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FibroTest Se LSM Se Biopsie Se

Performance for Cirrhosis: Sensitivity



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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

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Period 3: 2010-2015!Improving serum biomarker

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HCV-GenoFibroTest: Liver injury, Virus Resistance, Host Genes for treatment Response and Tolerance

88

ActiTest

FibroTest SteatoTest

IL28B

HCV-GenoFibroTest

Viral Load

Viral Resistance

ITPA

UGT1A1

Genotype

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Period 3: 2010-2015!Combining serum and imaging biomarkers

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• 11 Published studies

• n=2,260

• Standardized AUROC

• Advanced Fibrosis

• 0.89 (0.84-0.95)

Friedrich Rust et al Gastroenterology 2008, Poynard et al SJG 2008

79

Elastography

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Oliveri WJG 2008

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Pitfalls of Fibroscan

3.1% Failures and Unreliable results 15.8%

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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

A la Parisienne FibrotestFirst Line

If not interpretableBiopsy

FibroTest ActiTest

If not interpretableFibroscan

98%

<1%

2%

Prevalence

Serum biomarker

FibroTest

Imaging biomarker

FibroScan

Elasto-FibroTest

Poynard, CRHG 2012

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Elasto-FibroTest®

• 1289 patients with CHC and 604 healthy volunteers

• Appropriate methods

• Obuchowski measures

• Methods without Gold Standard

66

Poynard, CRHG 2012

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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

7 févr. 2012

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

2

FibroTest strengths

1.Same algorithms and cutoffs for all chronic liver diseases: chronic hepatitis C (CHC) and B (CHB), Alcoholic Liver Disease ALD, and Non-Alcoholic Fatty Liver Disease (NAFLD)

2.Most validated fibrosis biomarker (CHC, CHB, ALD, NAFLD), including repeated biopsies, prognostic studies, meta-analyses, guidelines, and cost-effectiveness

3.Combined with biomarker of Activity (ActiTest), Steatosis (SteatoTest), Alcoholic Hepatitis (AshTest) and Non-Alcoholic Steato-Hepatitis (NashTest), IL28B (GenoFibroTest) and elastography (ElastoFibroTest).

4.Security algorithms to prevent false positive/negative (Gilbert, Hemolysis,) and possible analytical errors

5.>95% applicability with more than 1 million prescriptions,

6.Used for inclusion of patients in phase 3 trials of DAA

2

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

Fibrosis Cirrhosis

Number studies 25 11

Number patients 9549 6893

AUROC 0.79 (0.70-0.89) 0.86 (0.71-0.92)

Applicability >95% >95%

Afdhal, JVH Nov 2013 Chou, Ann Int Med 2013

FibroTest performances forclinically significant fibrosis and cirrhosis in CHC

Competitors

• Transient elastography first generation: Fibroscan

!

• Aspartate amino transferase - Platelet Ratio Index (APRI)

84

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

Fibrosis F2F3F4 F4 All stages

Method AUROC (95%) AUROC Obuchowski

FibroTest 0.83 (0.81-0.85) 0.88 (0.87-0.91) 0.89 (0.88-0.89)

FibroScan 0.69 (0.67-0.71) 0.77 (0.76-0.79) 0.74 (0.74-0.75)

P value <0.01 <0.01 <0.01

FibroScan: 7.1 kPa cutoff for F2: 12.6% false positives ?!Poynard CRHG 2011, Roulot J Hepatol 2008

Higher performance of FibroTest vs FibroScan FF3F4 and F4 in «intention to diagnose» 1893 patients with chronic hepatitis C and 604 healthy volunteers

Competitors

• Transient elastography first generation: Fibroscan

!

• Aspartate amino transferase - Platelet Ratio Index (APRI)

89

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

Clin Res Hepatol Gastro 2014

High variability of APRI: methods

• Overview literature of AST-ULN: 26-49 IU/L

• 7521 healthy volunteers

• 393 blood donors

• 1651 patients with CHC, biopsy, FibroTest, APRI

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%

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

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

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

EASL Recommendations on Treatment of Hepatitis C, April 2014

13

Three reasons to assess fibrosis stages in 2014

!

•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 = 933!NS

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

Standard of Care in 2014

FibroTest, not perfect....but...

The most validated biomarker of liver fibrosis and activity!!

Better performance than APRI!

Better than FibroScan in intention to diagnose and for screening in general population

F4

F1

F0

France: 12,000,000 at Risk100%

5%

Death 15,000/year0.1%

Biomarker10% F2

F3

du 2015 poynard biomarkers

Health & Medicine

practice27 fvr

million7 fvr

millions chronic hepatitis

false negative7 fvr

new methods197 fvr

fibrosis biomarkers

yr imaging

yr fibrotest