1. Serum fibrosis markers are associated with liver disease progression in non-responder patients with chronic hepatitis C Robert J Fontana, Jules L Dienstag, Herbert L Bonkovsky, Richard K Sterling, Deepa Naishadham, Zachary D Goodman, Anna S F Lok, Elizabeth C Wright, Grace L Su, the HALT-C Trial Group Gut 2010;59:1401~1409 R1 Chihyeok Oh

2. Introduction The morbidity & mortality a/w HCV infection increased in patients with cirrhosis, decompensation and hepatocellular carcinoma (HCC) The Liver biopsy –Recommended in Management guidelines –to grade and stage the severity of liver disease and assist with decision making –But, the risks and sampling variability of liver biopsy –accurate and reliable non-invasive means are needed Numerous biochemical indices, serum fibrosis marker algorithms and liver elasticity measurements : not been tested or validated in a large group of patients with CHC SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

3. Administration of ‘maintenance’ interferon to patients with CHC who failed to respond to a course of antiviral treatment –may slow the rate of liver disease progression Hepatitis C Antiviral Longterm Treatment against Cirrhosis (HALT-C) Trial –Peginterferon Vs. no additional treatment : similar outcome –if serum fibrosis marker levels would correlate with initial disease severity and disease progression over time

4. In our analysis – ① pretreatment serum hyaluronic acid (HA) ② tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) ③ YKL-40 ④ N-terminal peptide of procollagen type 3 (PIIINP) levels ⇔ baseline disease severity ⇔ Active fibrogenesis & fibrolysis Hypothesis –These analytes level will be increased in clinical or histological disease progression patient

5. Methods - Patient population Randomised HALT-C Trial patients –detectable serum HCV RNA and bridging hepatic fibrosis (ie, Ishak fibrosis score ≥3) or cirrhosis on liver biopsy obtained within 12 months of enrolment –had failed to achieve a sustained virological response to a prior course of peginterferon and ribavirin SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

6. The ‘lead-in phase’ of the study retreated with peginterferon alfa-2a and ribavirin for 24 weeks –Subjects who remained viraemic at week 20 were eligible for randomisation at week 24 –Subjects treated with at least 12 weeks of peginterferon and ribavirin without viral clearance –Subjects with undetectable HCV RNA at week 20 –Subjects who experienced an on-treatment breakthrough or post-treatment virological relapse following the responder arm

7. Methods - Laboratory and clinical assessment(1) Lifetime alcohol consumption Routine baseline laboratory values –AST, ALT, albumin, bilirubin and platelet count Insulin resistance A baseline liver biopsy –scored for the degree of hepatic fibrosis and inflammation defined by the Ishak scoring system Laboratory and clinical assessment –every 3 months To screen for HCC –annual liver ultrasounds, serum AFP(every 3 months) SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

8. Methods - Laboratory and clinical assessment(2) Clinical end points –CTP (Child-Turcotte-Pugh) score to ≥7 on two separate 3 months apart –variceal bleeding, ascites, SBP, hepatic encephalopathy, HCC or death histological progression –defined as a ≥2 point increase in the Ishak fibrosis score SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

9. Methods - Serum fibrosis marker assays TIMP-1 –normal range : 80~500 ng/ml –Quantikine, R & D Systems, Minneapolis,Minnesota, USA YKL-40 –normal range : 24~125 mg/l –Metra YKL-40, Quidel, San Diego, California, USA PIIINP –normal range : 2~4 mg/l –UniQ, Orion Diagnostica, Espoo, Finland Serum HA levels –normal range : 10~100 ng/ml –automated liquid-phase immunoassay using the LiBASys Analyser (Wako Diagnostics, Richmond, Virginia, USA) SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

10. Methods - Statistical analyses Cox proportional hazards regression –Time to first clinical outcome Complementary log-log regression analysis –Time to histological progression AIC (Akaike information criterion) statistic –Compare the goodness of fit of individual models with each other Patient –Low, medium(75 th percentile), high risk(90 th percentile) group SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

11. Results - Overall study population SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

12. Results - Baseline predictors of clinical outcomes CTP score increasen=40 HCCn=7 Ascites n=32 Encephalopathy n=15 Variceal bleeding n=8 Death n=24 SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

13. Results - Multivariate models of clinical outcomes On univariate analysis –Baseline PIIINP levels were significantly associated with clinical outcomes In a multivariate model –only baseline bilirubin, INR and albumin levels remained associated in a multivariate model –TIMP-1, HA, YKL-40, bilirubin, INR and albumin levels All four baseline serum fibrosis marker levels entered together with other significant parameters –Clinical risk score=0.883×(total bilirubin)+0.809 (INR >1.0)- 1.63×(albumin)+0.89× (log YKL-40) –low (score less than 2.5), medium (score between 2.5 and 1.7) and high risk (score greater than 1.7) SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

14. Results - Serum fibrosis marker levels over time Serum fibrosis marker levels in HALT-C patients with and without clinical progression. SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

15. Results - Peginterferon treatment and serum fibrosis marker levels Serum fibrosis marker levels over time by treatment group. SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

16. Results - Baseline predictors of histological progression SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

17. Results - Multivariate models of histological progression baseline YKL-40 and TIMP-1 levels did not remain associated with histological progression both baseline PIIINP and HA levels remained significantly associated with histological progression The risk of fibrosis progression is estimated as risk = -0.3285×platelets+0.8831×logHA SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder

18. Conclusion Pretreatment serum fibrosis marker levels are significantly increased in patients with CHC at risk of clinical and histological disease progression. If validated in additional cohorts, measurement of these markers could help identify patients with CHC who would benefit from more frequent and intensive monitoring. SerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responderPatientsWithChronicHepatitisCSerumFibrosisMarkersAreAssociatedWithLiverDiseaseProgressionInNon-responder