Primary endpoint data review concluded no further patients required.

TITLE Rifaximin in Fatty Liver Disease (RiFL) DESIGN Open-label pilot study HYPOTHESIS Reduction in gut flora by the antibiotic Rifaximin reduces hepatic inflammation in Non-Alcoholic Steatohepatitis (NASH).

AIMS To provide proof-of-concept data on the therapeutic potential of gut flora modification in NASH OUTCOME MEASURES

Primary:

• Change in serum ALT from baseline by 25 IU/L or to within normal range after 6 weeks of Rifaximin therapy

Secondary:

• Change in intrahepatic triglyceride, estimated by in vivo proton magnetic resonance spectroscopy (1H MRS)
• Change in hepatic insulin resistance, estimated by the hyperinsulinaemic euglycaemic clamp
• Changes to the faecal bacterial microbiome assessed by faecal DNA pyrosequencing and fluorescent in-situ hybridisation (FISH)
• Differences in urinary metabolic profiles as assessed by high-resolution proton nuclear magnetic resonance spectroscopy

POPULATION Patients with biopsy-confirmed non-alcoholic steatohepatitis and persistently raised serum aminotransferase levels

TREATMENT The non-absorbable antibiotic Rifaximin DURATION This was an open-label study of Rifaximin (Normix, Alfa Wasserman S.p.A, Bologna, Italy) 400mg twice daily for six weeks followed by a further six weeks observation period during which patients received standard care.

STUDY OBJECTIVES The primary endpoint was change in ALT after 6 weeks of Rifaximin. Secondary endpoints were change in hepatic lipid content and insulin sensitivity measured with a hyperinsulinaemic euglycaemic clamp.

STUDY DESIGN This was an open-label study of Rifaximin (Normix, Alfa Wasserman S.p.A, Bologna, Italy) 400mg twice daily for six weeks followed by a further six weeks observation period during which patients received standard care. Compliance with treatment was checked by collection of empty blister packs. Subjects were asked to provide a structured dietary and lifestyle history as previously described (Williams HR, Cox IJ, Walker DG, North BV, Patel VM, Marshall SE, Jewell DP, et al. Characterization of inflammatory bowel disease with urinary metabolic profiling. Am J Gastroenterol 2009;104:1435-1444). The primary endpoint was change in ALT after 6 weeks' Rifaximin therapy. Secondary endpoints were change in hepatic and whole-body insulin sensitivity assessed by the two-stage hyperinsulinaemic euglycaemic clamp and change in hepatic triglyceride content assessed by proton nuclear magnetic resonance spectroscopy at 6 weeks from baseline. Serum ALT, biochemistry and anthropometrics were also measured at 12 weeks to look for longer-term effects. Stool microbiota, urinary metabolic profile and serum cytokine profile were measured before and after intervention.

PARTICIPANT ENTRY

INCLUSION CRITERIA Male and female patients were eligible for inclusion if aged between 18 and 70 years with non-alcoholic steatohepatitis histologically-proven, as evidenced by the presence of all of: steatosis, hepatocyte ballooning and lobular inflammation, and scored according to Kleiner(18) by a single experienced histopathologist (RDG) within the previous year, with or without mild to moderate fibrosis (stage 0-3/4) and with persistently elevated alanine aminotransferase (ALT) values on at least two occasions in the three months prior to recruitment.

EXCLUSION CRITERIA Patients were excluded if there was histological evidence of cirrhosis; hepatic decompensation; regular alcohol consumption exceeding 14 units/week (16g ethanol/day) for a woman or 21 units/week (24g ethanol/day) for a man; evidence of viral, autoimmune or other metabolic liver disease on a chronic liver disease screen; a history of malignancy or systemic inflammatory conditions; myocardial infarction or cerebrovascular events in the preceding 6 months; a history of bariatric surgery, blind loop or short bowel; use of any treatment known or suspected to change bowel flora within 3 months of enrolment; initiation or major dose change of metformin, thiazolinediones, biguanides, statins, fibrates, anti-obesity medications or insulin within 3 months of enrolment.

Inclusion Criteria:

• Subject has provided written informed consent prior to screening
• Men and women aged 18-70 years
• With non-alcoholic steatohepatitis histologically-proven, as evidenced by the presence of all of: steatosis, hepatocyte ballooning and lobular inflammation, and scored according to Kleiner(18) by a single experienced histopathologist (RDG) within the previous year, with or without mild to moderate fibrosis (stage 0-3/4)
• With persistently elevated alanine aminotransferase (ALT) values on at least two occasions in the three months prior to recruitment

Exclusion Criteria:

• NAFLD with cirrhosis (fibrosis score 4)

• Other causes of chronic liver disease

  • Viral hepatitis (HBV, HCV negative)
  • Alcohol intake >14units/week (women) or >21units/week (men)
  • Haemachromatosis (abnormal transferrin saturation, haemochromatosis genotyping)

• Evidence of hepatic decompensation

  • Ascites
  • Hepatic encephalopathy
  • Abnormal total bilirubin (except patients with Gilbert's syndrome), albumin, prolonged prothrombin time, low platelets)
  • Oesophageal or gastric varices

• Moderate or severe renal dysfunction (CKD3+, estimated GFR <60ml/min/1.73m2)

• Hepatocellular carcinoma

• Primary metabolic causes of hepatic steatosis (e.g. familial hypertriglyceridaemia, abetalipoproteinaemia)

• Other malignancy

• Pregnant or lactating women or women of childbearing potential unwilling/unable to use adequate contraceptive methods

• Systemic inflammatory conditions

  • Arthritis
  • Connective tissue disorders
  • Inflammatory bowel disease

• Myocardial infarction within 6 months

• Stroke within 6 months

• Bariatric surgery/ blind loop/ short bowel

• Treatment known/suspected to change gut flora (e.g. systemic antibiotics, colestyramine, lactulose, polyethylene glycol) within 3 months

• Treatment with drugs known to cause hepatic steatosis (e.g. corticosteroids, HAART, amiodarone, high dose oestrogens, tamoxifen) within 3 months

• Initiation or major dose change of metformin, thiazolidinediones, biguanides, statins, fibrates, anti-obesity medications or insulin within 3 months of enrolment

• Patients with allergy to Rifaximin or Rifamycin

• Patients with a cardiac pacemaker, history of penetrating eye injury, metal foreign body or any other contra-indication to MRI scanning, as specified in the local MRI safety checklist

• Any other clinical, social or psychological issues which, in the opinion of the investigators may preclude satisfactory completion of the study protocol

12 weeks