INTESTINAL TM6SF2 PROTECTS AGAINST NON-ALCOHOLIC STEATOHEPATITIS THROUGH GUT-LIVER AXIS

Background: Nonalcoholic steatohepatitis (NASH) is a major cause of liver-related morbidity, mortality, and the leading indication for liver transplant in the US. Fibrosis is the best predictor of clinical outcomes in NASH. Obeticholic acid (OCA), a first-in-class farnesoid X receptor agonist, demonstrated efficacy as an antifibrotic agent in the phase 3 REGENERATE trial in NASH. The goal of the new analysis was to confirm the original 18-month biopsy efficacy results and to provide additional safety data from more than 8000 total subject-years, with nearly 1000 subjects receiving OCA for ≥4 years in the REGENERATE study.

Methods: In this multicenter, randomized, double-blind, placebo-controlled phase 3 study, subjects were randomized 1:1:1 to receive once-daily oral placebo, OCA 10 mg, or OCA 25 mg. Safety data were evaluated from 2477 subjects who received ≥1 dose of the study drug. A consensus method using a panel of 3 pathologists for histologic assessment of liver biopsies per NASH Clinical Research Network criteria was employed to confirm the results of the original 18-month interim efficacy analysis performed by individual readers.

Results: Treatment-emergent adverse events (TEAEs), serious TEAEs, and deaths were balanced across treatment groups (N=2477). Pruritus was the most common TEAE in all 3 treatment groups; its incidence was higher in both OCA groups compared to placebo. For adverse events of special interest, incidence of pancreatitis and hepatic disorders was similar between the OCA and placebo groups. The incidence of gallbladder- and gallstone-related TEAEs (including serious events) was higher in both OCA groups compared to placebo, though most were mild to moderate in severity and managed with temporary drug interruption and usual clinical care (Table 1). In the new analysis of the intent-to-treat population (n=931) using a consensus read method, 22.4% of subjects receiving OCA 25 mg experienced ≥1 stage improvement of fibrosis with no worsening of NASH at month 18 vs 9.6% of subjects receiving placebo (primary endpoint), which was consistent with the original analysis performed by individual readers (Figure 1).

Conclusions: In the ongoing REGENERATE trial, subjects receiving OCA 25 mg were twice as likely to experience ≥1 stage improvement in liver fibrosis with no worsening of NASH at month 18 compared to subjects receiving placebo at 18 months using 2 different histologic methodologies. The confirmed antifibrotic effect in liver histology, together with extended exposure within the largest safety database in NASH to date, demonstrates that OCA 25 mg was generally safe and well tolerated, supporting the long-term use of OCA to treat pre-cirrhotic fibrosis due to NASH.

Background: Transmembrane 6 superfamily member 2 (TM6SF2) is predominately expressed in liver and small intestine, and its loss-of-function variant is linked to increased risk of non-alcoholic fatty liver disease (NAFLD). We aim to investigate the role and mechanism of intestinal TM6SF2 dysfunction in the pathogenesis non-alcoholic steatohepatitis (NASH).
Methods: Mice with systematic, liver-specific, or intestine-specific Tm6sf2 knockout (Tm6sf2^ΔIEC) were fed with normal chow, high fat high cholesterol (HFHC), or choline-deficient high fat diet (CD-HFD) for 2 months. Gut microbiota and metabolites were profiled by metagenomic and metabolomic analysis. To determine the role of gut microbiota of Tm6sf2^ΔIEC mice, fecal microbiota transplantation was performed in germ-free mice. Tm6sf2^ΔIEC mice were co-housed with wildtype (WT) mice to evaluate the therapeutic potential of microbiota manipulation.
Results: Hepatocyte-specific Tm6sf2 deletion in mice induced spontaneous hepatic steatosis. while systematic Tm6sf2 ablation showed pronounced hepatic steatosis with necroinflammation, implying that extra-hepatic Tm6sf2 deletion could also contribute to NASH progression. Given that intestine has high endogenous TM6SF2 expression, we established the Tm6sf2^ΔIEC mice. Tm6sf2^ΔIEC mice fed with normal chow spontaneously developed NASH with increased hepatic triglyceride, lipid peroxidation, enhanced pro-inflammatory cytokines and NF-κB. Intestinal Tm6sf2 knockout accelerated NASH development in mice fed HFHC or CD-HFD. The gut barrier in Tm6sf2^ΔIEC mice was impaired with increased permeability and abnormalities of intestinal junctions, suggesting that loss of intestinal TM6SF2 might deregulate the gut-liver axis. Indeed, integrative metagenomic and metabolomic analysis unveiled the enrichment of gut pathogenic bacteria in Tm6sf2^ΔIEC mice that was correlated with increased lysophosphatidic acid (LPA) in the portal vein, leading to LPA accumulation in liver of Tm6sf2^ΔIEC mice. Moreover, LPA promoted lipid accumulation and pro-inflammatory cytokines secretion in hepatocytes. Intestine cells isolated from Tm6sf2^ΔIEC mice secreted more free fatty acids compared to the intestine cells isolated from WT mice, which could impair gut barrier in mice. Notably, transplanting stool from Tm6sf2^ΔIEC mice to germ-free mice induced steatohepatitis with microbial dysbiosis and increased LPA level in portal vein in recipient mice; whereas co-housing of Tm6sf2^ΔIEC mice with WT mice significantly restored gut barrier function, and ameliorated hepatic lipid accumulation and inflammation in Tm6sf2^ΔIEC mice.
Conclusion: Intestinal TM6SF2 deficiency induces gut dysbiosis and barrier dysfunction, leading to increased translocation LPA to liver to promote NASH progression. Microbiota manipulation may be an effective strategy for the prevention of NASH driven by down-regulation of TMFSF2.