LIVER FIBROSIS PREVALENCE IN FRANCE
LIVER FIBROSIS PREVALENCE IN FRANCE
"Mortality related to complications of cirrhosis, (hemorrhage, hepatic insufficiency and
 primary liver cancer) is 15,000 per year in France. These mortality increases, despite that
 advanced fibrosis can be identified by non-invasive biomarkers and treated, more than 10
 years before the onset of complications and cancer. The main goals of the FIBROFRANCE project
 which started in 1997 (initially called the MULTIVIRC group) were to demonstrate the
 performance of serum biomarkers in the more frequent chronic liver diseases, to estimate the
 dynamic of fibrosis progression and finally to demonstrate the feasibility of the fibrosis
 screening in French people.

 The different cohorts of the FIBROFRANCE (HCV, HBV, ALD, NAFLD) permitted many publications
 among the 186 publications of our group since 1986 in the field of liver fibrosis. These
 publications included discovery and validation of non-invasive biomarkers (Poynard
 Gastroenterology 1997, Imbert-Bismut Lancet 2001, Poynard BMC Gastro 2007), modelling
 fibrosis progression or regression (Poynard Lancet 1997, Poynard Gastroenterology 2002,
 Poynard J Hepatol 2003) and fibrosis screening (Ratziu APT 2007, Jacqueminet Clin
 Gastrenterol Hepatol 2008). This research was conducted in Pitié-Salpêtrière hospital for the
 biochemical and clinical part in connection with national and international networks. Several
 panels have been identified and the most predictive FibroTest has been patented (US Patent
 Office 6.631.330) and launched in 2002. This is the first fibrosis biomarker available
 worldwide (50 countries including USA as FibroSURE) with more than 1 million prescriptions
 between 2002-2013. FibroTest, has been validated first in hepatitis C and then in hepatitis B
 alcoholic liver disease and metabolic syndrome. Therefore it is now possible to screen
 advanced fibrosis in the 4 most frequent liver diseases: alcohol, hepatitis C and B, and
 metabolic syndrome (diabetes, overweight, and hyperlipemia). For all the patients detected
 there are therapeutic options to cure the fibrosis or to reduce the progression to cirrhosis
 and cancer.

 FibroTest has been recommended as alternative to biopsy in several guidelines (AFEF, APASL,
 EASL and CASLD) and more recently in US overview (Chou Annals 2013). It reimbursed in France
 in chronic hepatitis C. Several factors of fibrosis progression can be present in the same
 subject, i.e. an overweight and an excessive alcohol consumption. Therefore no realistic
 screenng strategy can be conducted without taking into account the Interdependence of the
 different risk factors. Three biomarkers of fibrosis-associated liver injuries have been
 developed and validated in FIBROFRANCE cohorts: SteatoTest for steatosis (Poynard Comp
 Hepatol 2005), NashTest for non-alcoholic steatohepatitis (Poynard EASL 2006), and AshTest
 for alcoholic steatohepatitis (Naveau J Hepatol 2006). For this purpose different cohorts
 already used for diagnostic validation will be followed at long term for prognostic
 validations: FIBROFRANCE-ALD (Naveau Hepatology 2010), FIBROFRANCE-NAFLD including
 dyslipidemia cohort (Ratziu APT 2007) and diabetes cohort (Jacqueminet Clin Gastrenterol
 Hepatol 2008). These cohorts will allow assessing the prevalence of fibrosis and the specific
 risks of fibrosis progression imputable to steatosis and steatohepatitis.
"This is the initial version of the FIBROFRANCE project which strated in 1997. Methods The
 strategy chosen is to screen advanced fibrosis in several groups of patients with different
 levels of risks in order to reconstruct the French prevalence thereafter. A national
 screening will be suggested according to the identified risk factors.

 Rational and prior hypothesis The simple rational concept is to screen patients before
 complications of cirrhosis. (Figure 1).

 Figure 1: Fibrosis progression with five consecutive stages: F0 no fibrosis, F1 mild fibrosis
 without septa, F2 moderate fibrosis with few septa, F3 severe fibrosis with many septa, F4
 cirrhosis.

 Advanced fibrosis is defined by presence of stages F2-F3-F4. The aim of the study is to
 demonstrate that the screening is possible at stage F2 many years before complications. From
 previous modeling (Figure 2) we suggested that an efficient screening must start around the
 age of 40 years for the majority of high-risk group and around 30 years old in HIV coinfected
 patients.

 Population included Five major groups and 15 cohorts of different populations at risk will be
 included. Five groups at high risk will be included: patients with hepatitis C, B, heavy
 alcohol drinkers and patients with metabolic risk factors diabetics and hyperlipemics. These
 populations were chosen because a blood sample can be easily available. The higher number
 will permit to reconstruct the French prevalence taking into account the major
 characteristics of subjects included (age, gender, socio economic status) and the known
 prevalence of some risk factors (HBV and HCV prevalence, diabetes, hyperlipemia and heavy
 drinker's prevalence).

 Characteristics of subjects and recorded items The following characteristics will be recorded
 in all studies: age, gender, high-risk factor, fibrosis stage and activity grade. In all
 FibroFrance ANR cohorts the following factors are recorded and will be used in the
 sensitivity analysis of prevalence estimates: socio-economic status, body mass index, area of
 residence, alcohol consumption, metabolic syndrome factors, cholesterol, triglycerides,
 glucose.

 All data will be recorded anonymously with a code number and declared to the CNIL.

 Endpoints The main endpoints are the French prevalence of advanced fibrosis and cirrhosis.
 The estimates will be calculated according to age and gender adjusted on the absence or
 presence of high risk factors in the French population. Non-high risk group represents in
 France 48,000,000 people and the high risk group 12,000,000 people. The non-high risk group
 will be derived from the estimates in blood donors, free screening center, and social
 security center after exclusion of the subjects with high-risk factors which are assessed in
 these three cohorts: heavy drinkers, HBV, HIV, diabetes, hyperlipemia.

 The final target for a national screening will take into account the relative risks
 attributed to the predicted steatosis and steato-hepatitis. Estimates of steatosis and
 steatohepatitis will be secondary endpoints.

 Biomarkers FibroTest-ActiTest, SteatoTest, NashTest, AshTest will be performed in the
 reference laboratory (GHPS) using the recommended pre analytical and analytical procedures.
 Results will provide the estimated stages and grades and if the results profile is at risk of
 false positive and negative. All patients with advanced fibrosis identified with FibroTest
 will be contacted by the Liver reference center to organize the management. Elastometry
 (FibroScan) ultrasonography will be performed for all advanced fibrosis and endoscopy if
 cirrhosis predicted.

 Statistical analysis Prevalence will be expressed as mean with 95% confidence interval.
 Sensitivity analyses will be performed to assess the impact of the following factors on the
 prevalence estimates: socio economic status, body mass index, area of residence, non-heavy
 drinkers (<50g/day), metabolic syndrome factors (ATP III definition) in univariate and
 multivariate analysis using multiple regression analysis, together with age and gender.

 The feasibility of the project has been checked in patients with chronic hepatitis C, B as
 well as in patients with alcoholic and non alcoholic liver steatosis (see Fibrochure in annex
 for details and references).

 The risk of false positive or false negative of FibroTest (FT), has been also checked on
 large populations of hepatitis C and in blood donors.

 These studies allow us to expand the screening to community based populations. Blood donors
 (BD) Security algorithms (SA) were used at the time of calculating FT to alert users and
 correct a potential error of data entry for each 6 FT components in different cohorts. One
 SA1 detects abnormal values (AV) outside the 98% percentiles. Another SA2 detects, HR profile
 of false positive (HR-FP) or false negative (HR-FN) serum, defined as a serum for which the
 switch of one component by the median value of this particular component, is associated with
 a dramatic change in the FT, greater than +0.30.

 A total of 954 Blood Donors (HBV, HCV, HIV neg) were prospectively included; BMI, daily
 alcohol consumption and treatment were assessed. FT was performed on fresh serum according to
 analytical recommendations.

 Mean age of pts was 50 yrs, 45% female. SA2 detected 3,965 tests (5.05%) with HR profile. The
 most frequent AV observed was haptoglobin <0.12 g/L in 3,562 pts (4.53%), but among them
 there was only 614 HR-FP cases (0.78%) suspect of hemolysis, for which the other components
 were not concordant in favor of significant fibrosis. HR-FP due to possible Gilbert Syndrome
 was observed in 921 (1.17%). Mean age of BD was 36 yrs, 49% female, BMI 23.4 Kg/M2; 265 (28%)
 received treatment: 125 (13%) oestro-progestative (OP), 20 (2%) hypolipemiant, 16 (2%)
 anti-histaminic, 11 (1%) thyroid hormone, 11 benzodiazepine (1%); 203 (21%) BD were totally
 abstinent, 650 (68%) drunk less than 10g, 101 between 10-50g/day and none drunk more. SA2
 detected 29 (3%) HR profile including 9 Gilbert (0.9%), and 4 HR-FP with hemolysis profile
 (0.4%). Among the 925 interpretable BD, the median FT was 0.08 (range 0.01-0.46), AT 0.06
 (0.01-0.66). No subjects had severe fibrosis F3F4 or F2, and 909 (98.3%) had none or minimal
 fibrosis (FT<0.32). FT-AT were not associated with age, weight, BMI and treatments other than
 OP: FT, lower in 125 women treated by OP (median 0.0490; 95% CI 0.044-0.058) than in non
 treated (0.0659; 0.064-0.072; P=0.01). Mean age of BD was 36 yrs, 49% female, BMI 23.4 Kg/M2;
 265 (28%) received treatment: 125 (13%) oestro-progestative (OP), 20 (2%) hypolipemiant, 16
 (2%) anti-histaminic, 11 (1%) thyroid hormone, 11 benzodiazepine (1%); 203 (21%) BD were
 totally abstinent, 650 (68%) drunk less than 10g, 101 between 10-50g/day and none drunk more.
 SA2 detected 29 (3%) HR profile including 9 Gilbert (0.9%), and 4 HR-FP with hemolysis
 profile (0.4%). Among the 925 interpretable BD, the median FT was 0.08 (range 0.01-0.46), AT
 0.06 (0.01-0.66). No subjects had severe fibrosis F3F4 or F2, and 909 (98.3%) had none or
 minimal fibrosis (FT<0.32). FT-AT were not associated with age, weight, BMI and treatments
 other than OP: FT, lower in 125 women treated by OP (median 0.0490; 95% CI 0.044-0.058) than
 in non treated (0.0659; 0.064-0.072; P=0.01).

 Diabetics (DI) Mortality related to cirrhosis is increasing in patients with
 insulino-resistance factors. Type 2 diabetic patients (DI) vs non-DI have higher risk of
 non-alcoholic fatty liver disease (NAFLD) and liver cancer death.

 411 Consecutive DI seen in a diabetes unit, (HCV HBV neg), were prospectively included, as
 well as a prospective control group of blood donors (BD). FT was performed blindly to any
 clinical or biological data, according to analytic recommendations with security algorithms
 permitting to exclude high-risk (HR) profiles of false negative (FN) and positive FT (FP).
 Analysis used univariate comparison with BD matched for gender, age, alcohol consumption and
 BMI, and multivariate logistic regression.

 Among DI 53% were male, mean age 55yrs, 66% DI type 2; 80% had fasting glucose ≥6.1mmol/L,
 67% (257/382) had glycohemoglobin (glyHb) ≥7.5%, 43% (164/382) cholesterol ≥2.0 g/L, 29%
 (111/382) triglycerides ≥1.5g/L, 50% had BMI ≥27, 48% had arterial hypertension, 72% had no
 alcohol consumption, 26% a consumption ≤50g/d, and 2% >50 g/d. BD were younger (36yr), had
 lower BMI and alcohol consumption was similar.

 F2F3F4 were identified by FT in 41/411 (10%) of DI, including 16 F2, 12 F3 and 13 F4, vs none
 F2F3F4 among the 925 BD (P<0.0001).

 Among DI type 2, 11% had F2F3F4 vs 2% type 1 (p=0.01). DI was the only independent
 significant factor associated with F2F3F4 in multivariate regression analysis (P<0.0001).
 After exclusion of 93 DI not matched because of higher age than BD, there were still 8.8%
 (28/318) F2F3F4 in DI versus 0% in BD (P<0.0001). Among the 41 DI F2F3F4, 22 had ALT<50IU/L,
 10 GGT <50 IU/L, 10 both normal, 19 both elevated and 31 one. Only 2 DI had overt
 clinical-biological signs of cirrhosis.

 Conclusion: The prevalence of advanced fibrosis, estimated by FibroTest, is very high (10%)
 in diabetics patients followed in a tertiary center. This study strongly suggests that
 non-invasive biomarkers could be very useful for the screening of advanced fibrosis in type 2
 diabetic patients to prevent liver mortality.

 Patients with hyperlipemia (HL) A consecutive cohort of HL, (HCV,HBVneg), were prospectively
 followed in a lipid center and the sera (stored at -80) were retrospectively analyzed; a
 control group of blood donors (BD) was prospectively included. FT was performed blindly to
 any clinical or biological data, and according to analytic recommendations including security
 algorithms permitting to exclude extreme values and high-risk profiles of false negative and
 positive (HR).

 A total of 2,834 HL were included (51% female, median age 49 years); 83% had cholesterol
 >200mg/dl, 94% LDL-C >100mg/dl, 21% HDL-C >70mg/dl, 16% triglycerides >200mg/dl. 36% BMI >27,
 32% insulin >10mIU/ml, 39% arterial hypertension, 15% HOMA >3.8 and 13.2% had fasting glucose
 >6mmol. GGT or ALT were >50 IU/L in 458 HL (24%).

 F2F3F4 were identified by FT in 53/1909 (2.8%) HL, including 31 F2, 14 F3, 3 F3-F4 and 4 F4
 versus no (0%) among the 925 BD (P<0.0001).

 Among the 53 F2F3F4 42 had ALT<50IU/L, 27 GGT <50 IU/L, 24 both normal, 8 both elevated and
 29 at least one. Factors significantly (p<0.01) associated with fibrosis in univariate
 analysis were higher age, BMI, triglycerides, uricemia, insulinemia, and lower HDL
 cholesterol. Drugs and alcohol consumption were not associated with fibrosis. In multivariate
 logistic regression insulin [Odds ratio 43.7, CI (1.5; 61.0); P=0.001] was the most
 significant risk factor. In patients with insulinemia >10mIU/ml the prevalence of F2F3F4 was
 9.7%.

 Conclusion: The prevalence of advanced fibrosis estimated by FibroTest, is high (2.8%) in HL
 followed in a tertiary center. This study strongly suggests that non-invasive biomarkers
 could be very useful for the screening of advanced fibrosis in high-risk groups such as
 hyperlipemic patients to prevent liver mortality.

 Strategy for the management of subjects identified as possible advanced fibrosis For each
 subject with possible advanced fibrosis, information will be given by the physicians in
 charge of the cohort. These subjects could have the routine assessment of advanced liver
 fibrosis, including the elastometry (Fibroscan) available in the reference centre (Hepatology
 Department of Groupe Hospitalier Pitié Salpêtrière) and the other independent signs of
 cirrhosis (ultrasonography and endoscopy).

 Calendar This project can be conducted in 3 years.

 Interdisciplinarity According to the wide population exposed to the risk of advanced
 fibrosis, steatosis and steatohepatitis, this project is only possible if several medical
 disciplines are involved.

 As described in table 2 and 3, this project involves four different reference centers: Viral
 Hepatitis Center (Thierry Poynard), Diabetes Center (Andre Grimaldi), Lipid Center (Eric
 Bruckert), Alcoholic Center (Sylvie Naveau), two Prevention Centers (CPAM and CDAG), one
 Anesthesiology department (Pierre Coriat) and one Transfusion Unit (Anne Mercadier).

 Originality Our group has now the opportunity to demonstrate the feasibility of such
 screening for the first time in the world. We can do that because of the new non invasive
 biomarkers and the large number of subjects consulting in the different participating
 centers. The prevalence of advanced fibrosis and asymptomatic cirrhosis, steatosis and
 steatohepatitis will be established in France and similar studies could thereafter be applied
 in other countries.

 "
Inclusion Criteria:

 -Patients Exposed to fibrosis risk factors (HBV, HCV, ALD, NAFLD) or healthy volunteers

 Exclusion Criteria:

 -Non reliable fibrosis estimate, follow up shorter than months, acute liver diseases