1. FT in diagnostic of HCV FibroTest in the diagnosis of HCV Publications on diagnostic performance

2. FT in diagnostic of HCV 1. Diagnosis and clinical options 2. First validation: Prospective Study 3. FibroTest in histological changes 4. FibroTest in HCV carriers with mixed cryoglobulinemia systemic vasculatis 5. Comparison and combination with other methods 6.Meta-analysis In this Presentation

3. FT in diagnostic of HCV 95% 5% (For liver injuries Assessment) Diagnosis and clinical options Positive serology Poynard et al, Comp Hepatol 2004

4. FT in diagnostic of HCV First validation study

5. FT in diagnostic of HCV Imber-Bismut et al, Lancet 2001 Biochemical markers of liver fibrosis in patients with hepatitis C virus infection: a prospective study (n=205) - The top and bottom of the box are the 25th and 75th percentiles. - The length of the box is thus the IQR. The line through the middle of the box is the median. - The upper error bar is the largest observation 75 th percentile plus 1·5IQR. - The lower error bar is the smallest observation that is 25th percentile minus 1·5IQR. - Bonferroni allpairwise multiple comparison; p<0·0001 between all stages. Results  high negative predictive value  (>90% certainty of absence of F2, F3, or F4) was obtained for scores from zero to 0·20.  Of these 119 patients with low scores (35% of total) there were 13 false negatives: four F2, A0; six F2, A1; three F2, A2.  high positive predictive value  (>90% certainty of presence of F2, F3, or F4) was obtained for scores from 0·80 to 1·00.  Of these 50 patients with high scores (15% of total), there were five false positives: two F1, A1, and three F1, A2.

6. FT in diagnostic of HCV Imber-Bismut et al, Lancet 2001 ROC curves of fibrosis indices combining five, six, or ten biochemical factors, and age and sex Results  The ROC curves for the three indices were very similar  The areas (SD) under the curves did not differ:  5 factors: 0·851 (0·370)  6 factors: 0·847 (0·370)  10 factors: 0·837 (0·370) ConclusionsA combination of basic serum markers could be used to substantially reduce the number of liver biopsies done in patients with chronic HCV infection

7. FT in diagnostic of HCV FibroTest in histological changes

8. FT in diagnostic of HCV D’Arondel JVHep 2006 Utility of biomarkers to estimate the dynamics in the histological response in HCV to pegylated-IFN alpha-2b and ribavirin n=96 Results  Baseline F2F3F4 sustained viral responders (n=22)  FibroTest -ActiTest decrease (p<0.0001)  This improvement was significant even in virologic non- responders. Conclusions FibroTest and ActiTest are sensitive markers to histological changes during and after HCV treatment

9. FT in diagnostic of HCV Poynard et al, Hepatology 2003 Biochemical Surrogate markers of liver fibrosis and activity in a randomized trial of Peginterferon Alfa-2B and ribavirin Patients and methods  208 patients treated with IFN alpha & ribavirin 1.5mcg/kilo, 3/week during 48 weeks  144 patients treated with IFN alpha 3mU, 3/week during 48 weeks  Follow up at 24 weeks after treatment  Assessment with FibroTest ActiTest and biopsy at baseline and after treatment ResultsSee next slide Conclusion  FT could be used as surrogate marker for liver biopsy in patients with chronic hepatitis C, both for initial evaluation and for follow-up  Due to liver biposy risks and limitation and the improvmentof nin invasive markers of fibrosis, liver biopsy should no longer be mandatory

10. FT in diagnostic of HCV Poynard et al, Hepatology 2003 - Results Results FibroTest vs METAVIR at Baseline FibroTest vs METAVIR at follow up ActiTest vs METAVIR at Baseline ActiTest vs METAVIR at follow up FibroTest vs Knodell score at Baseline FibroTest vs Knodell score at follow up ActiTest vs Knodell score at Baseline ActiTest vs Knodell score at follow up

11. FT in diagnostic of HCV FibroTest in HCV carriers with mixed cryoglobulinemia systemic vasculatis

12. FT in diagnostic of HCV Sene D. et al, Clin Biochem 2006 Biochemical markers of liver fibrosis and activity as non invasive alternatives to liver biopsy in patients with CHC and mixed cryoglobulinemia vasculatis (MCV) Patients and methods  238 patients with HCV (50% male, mean age: 57y, 56% genotype1, 32% genotype 2-3, 12% genotype 4-5  Performed tests: Liver biopsy, FibroTest, Apri, Forns index  Systemic vasculatis defined by the presence of skin purpura, and rheumatological, neurological or renal involvement.  52% had serum cryoglobulin, 27% had serum inflammation and 11% had hemolysis. Results FT/AT vs Biopsy  FibroTest versus biopsy  AUROC for F2F3F4 vs F0F1 was 0.83(0.04),  without a difference between patients with and those without systemic vasculitis [0.81 (95%CI = 0.75-0.90) vs 0.85 (95%CI=0.75-0.90); p = 0.65]  correlated by METAVIR  ActiTest versus biopsy  ActiTest assessed by the AUROC (SE) for A2A3 vs A0A1 was 0.77 (0.05)  slightly higher but without significative difference between patients with and those without systemic vasculitis [0.89 (95%CI 0.73-0.97) vs 0.71 (95%CI=0.60-0.80); p = 0.055]  correlated by METAVIR

13. FT in diagnostic of HCV Sene D. et al, Clin Biochem 2006 Results FT/AT vs other biomarkers AUROC  FibroTest: AUC= 0,83  APRI: AUC= 0,73P=0,01  Forns index: AUC= 0,77P=0,054  Age-platelet index: AUC= 0,0,72P=0,01  Platelet count: AUC= 0,66P≤10-3 Results FT unaffected by vasculatis Conclusion  FT is a reliable non invasve alternative to liver biopsy in HCVwith systemic vasculitis or serum non-septic inflammation and is more reliable than other non-invasive fibrosis markers (Forns, APRI, HA, age-platelet).  Diagnostic value of FT in patients with HCV-mixed cryoglobulinemia vasculitis similar to that of patients without this condition and to those found in previous studies Vasculatis No vasculatis

14. FT in diagnostic of HCV Comparison and combination with other methods Hyaluronic acid, Historical index, Apri, FirboScan

15. FT in diagnostic of HCV FibroTest versus other non invasive markers Poynard et al, Comp Hepatol 2004 Overview of the diagnostic value of biochemical markers of liver fibrosis (FibroTest, HCV FibroSure) and necrosis (ActiTest) in patients with chronic Hepatitis C 16 HCV publications: FT= Alternative to Biospy 1,570 subject data base New Analysis/ Summary FibroTest in different HCV genotypes Genotypes: 1, 2, 3, 4- 5-6 FibroTest versus… APRI, Forns, Hyaluronic acid Control group: 300 blood donors Study Design

16. FT in diagnostic of HCV FibroTest versus other non invasive markers Poynard et al, Comp Hepatol 2004 - Results Conclusions Better AUROC for FibroTest  Lower than random 0.50 value (upper panel) (P < 0.001).  Higher then AUROCs of other fibrosis markers (lower panel) (P < 0.05). Diagnostic Value of FibroTest versus other non invasive markers

17. FT in diagnostic of HCV FibroTest versus other non invasive markers Poynard et al, Comp Hepatol 2004 - Results Conclusion  No significant difference in the AUROC of FibroTest and ActiTest in HCV according to HCV genotype or viral load.  FibroTest-ActiTest is a good surrogate to liver biopsy for the assessment of HCV related liver fibrosis AUROCs of FibroTest for the diagnosis of significant fibrosis, according to HCV genotypes. AUROCs of ActiTest for the diagnosis of significant necrosis, according to HCV genotypes. AUROCs of FibroTest for the diagnosis of significant fibrosis, according to serum HCV viral load. AUROCs of ActiTest for the diagnosis of significant necrosis, according to serum HCV viral load.

18. FT in diagnostic of HCV FibroTest versus Biopsy Myers R.P et al, American Journal of Gastro 2002 Biochemical markers of liver Fibrosis: a comparison with historical features in patients with chronic Hepatitis C Patients and methods  211 patients, 52% male, median age of 28y, median duration of infection 18y  Untreated HVC patients  Biopsy and FibroTest tested for the diagnosis of clinically significant fibrosis (F2-F4) Results FibroTest vs METAVIRHistorical index vs METAVIRAUROC F2-F4AUROC A2-A3/ F2-F4 Conclusion  FibroTest accurately predicts significant fibrosis in HCV infected patients  Markers used are widely available  Represents the most discriminative tool available for non-invasive prediction of fibrosis in HCV  More accurate than historical features, the addition of which to the existing index was not helpful Historical indexFibroTestHistorical indexFibroTest

19. FT in diagnostic of HCV Sebastiani et al, J Hepatol 2006 HEPATITIS C (AUC) HEPATITIS C with NALT (AUC) APRIFTAPRIFT >F2 0.690.810.690.70 F4 0.610.71 Stepwise combination algorithms of non-invasive markers to diagnose significant fibrosis in chronic hepatitis C Conclusions  Fibrotest presents with the best accuracy in all the subgroups of patients with chronic liver disease  Combination of markers should reduce the need for liver biopsy and ultimately health expenses IN >F2FibroTestAPRI Classified cases %10051 SE %6583.5 SP %80.677.1 PPV%8086.6 NPV%66.772.5 Accuracy%72.681.2

20. FT in diagnostic of HCV Sebastiani et al, J Hepatol 2006 – Safe Biopsy Sequential Algorithms for Fibrosis Evaluation (SAFE BIOPSY) Stepwise modelling aimed to achive accuracy> 95% For significant fibrosisFor cirrhosis

21. FT in diagnostic of HCV Sebastiani et al, J Hepatol 2006 – Biopsy and cost reduction Sequential Algorithms for Fibrosis Evaluation (SAFE BIOPSY) INTERIM ANALYSIS ON 2035 HCV CASES SAFE BIOPSY for SIGNIFICANT FIBROSIS SAFE BIOPSY for CIRRHOSIS Accuracy (%)9093 Saved biopsies (%) 4782 Saved cost (%)4580 SAFE biopsyFibropaca algorithm Leroy algorithm >F2 F4 >F2 APRI + Fibrotest sequential APRI + Fibrotest + Forns in parallel APRI + Fibrotest in parallel Accuracy (%) 9091.287.69493.5 Saved biopsies (%) 43.879.151.776.229.2 Reduced cost (%) 52.270.934.659.115

22. FT in diagnostic of HCV FibroTest versus Glycomics and FibroScan - Castera et al, Gasteroenterol Clin Biol 2005 “Clinical glycomics”: independant validation and comparison with Fibroscan and FibroTest for the assessment of fibrosis in CHC Patients and methods  211 CHC patients (57 males, mean age 53)  Exams (on same day): Liver biopsy, FirboScan, FibroTest and profile of glycosylation of serum proteins to obtain the GlycoMarker (GM) Results ( Auroc) Conclusion  FibroTest had an excellent diagnostic values (0.84 for F≥2, 0.92 for F≥3, 0.88 for F=4) compared to  other non-invasive methods like Fibroscan  to “Clinical Glycomics”  to a combination of both

23. FT in diagnostic of HCV FibroTest versus FibroScan - Castera et al, Gastroenterology 2005 Prospective comparison of transient elastography, FibroTest, APRI, and liver biopsy for the assessment of fibrosis in chronic hepatitis C. ConclusionsRecommend algorithm (from author)  From this first study, Fibroscan seemed able to assess liver fibrosis with a performance similar to that of FibroTest.  The combined use of Fibroscan and FibroTest to evaluate liver fibrosis could avoid a biopsy procedure in most patients with chronic hepatitis C

24. FT in diagnostic of HCV Multi center validation study: FibroTest versus HA - Poynard et al, J viral Hepat 2002 Biochemical markers of liver fibrosis in patients infected by hepatitis C virus: longitudinal validation in a randomized trial (n=165) Patients and methods  244 patients from 15 university hospitals, Positive HCV serology, never treated with elevated ALT  Group 1: 3mU IFN alpha thrice weekly (24 weeks)  Group 2: 6mU IFN alpha daily for 12 days and weekly for 22 weeks  Liver biopsies performed before and 72 weeks after  Comparison Biopsy, FibroTest and hyaluronic acid Results FibroTest vs METAVIRHyaloronic acid vs METAVIRAUROC FT (0,74)AUROC HA (0,65) Conclusion  Validation in an external population  Greater diagnostic value than HA  FT could be used as surrogate marker of the impact of HCV treatment on fibrosis progression

25. FT in diagnostic of HCV Meta analysis

26. FT in diagnostic of HCV Meta-analysis – Poynard et al, clin chem 2007 FibroTest Meta-Analysis 30 Published Studies 6.378 Patients 2001-2006 AUROC=0.84 (0.83-0.86) for F2F3F4 The best you can obtain with 20mm biopsy is 0.90 Bedossa 2003