Editor’s Note: Metabolism-related fatty liver disease (MAFLD) has become the leading cause of global chronic liver disease, yet there are currently no targeted and effective treatments. In recent years, the development of new drugs for MAFLD and metabolism-related fatty liver hepatitis (MASH) has been a mix of optimism and concern. The 2023 American Association for the Study of Liver Diseases (AASLD) Annual Meeting has selected six studies on novel MAFLD therapies from numerous abstract submissions. “Novel Therapeutic Approaches for MASH” will showcase relevant content, and interested readers can tune in on November 13th at 8:30 AM local time.
01 Deep Phenotypic Analysis of TM6SF2 Using a Genomic Prioritization Approach to Characterize Liver Steatosis, Lipid Traits, and Metabolic Risk Factors
Background:
An unbiased “genomic prioritization” approach may enhance understanding of common coding and predicted loss-of-function (pLOF) variants associated with non-alcoholic fatty liver disease (NAFLD). This study, led by Huang H from the University of Pennsylvania, aimed to unveil the functional impact of pathogenic variants in TM6SF2 using this method.
Methods:
Exon sequencing data from 44,297 patients in the University of Pennsylvania Medical School Biobank (PMBB) were analyzed to investigate the correlation of 121 synonymous missense variants and 12 pLOF variants with ICD-10 coded NAFLD, non-alcoholic steatohepatitis (NASH), and hepatocellular carcinoma (HCC). Serum parameters, liver fat scores, and imaging results were further analyzed after adjusting for age, gender, body mass index (BMI), ancestral principal components, and PNPLA3 rs738409: G carriage. Results were replicated in the UK Biobank (UKB), and the ColabFold Alphafold2 notebook with MMseqs2 was used for computer predictions of TM6SF2 structure.
Results:
E167K homozygotes increased the risk of diagnosed NAFLD (P < 0.0001, OR: 4.8), NASH (P < 0.0001, OR: 5.1), and HCC (P = 0.001, OR: 1.77). E167K heterozygotes showed a similar trend limited to NAFLD and NASH. Additionally, L156P heterozygotes increased the risk of diagnosed NAFLD (P < 0.001, OR: 2.16) and NASH (P < 0.0001, OR: 2.10), while only L156P homozygotes increased the risk of HCC (N = 2, P < 0.0001, OR: 532.0). These associations were successfully replicated in the UKB population. Additional imaging diagnoses in the PMBB cohort, coupled with elevated liver fat scores derived from CT, reinforced these associations in E167K and L156P carriers (P < 0.01).
Metabolomic analysis revealed lower circulating total cholesterol, triglycerides, fatty acids, and total choline in E167K and L156P carriers (after Bonferroni correction, P < 0.0001). Computer predictions confirmed that both mutations lead to structural disruption of the EXPERA domain, resulting in the loss of TM6SF2 protein function. The study further investigated pLOF variants and identified a premature stop codon (W35X) associated with an increased risk of diagnosed NAFLD (P = 0.03, OR: 15.9), NASH (P = 0.01, OR: 23.4), and elevated liver fat scores (P < 0.001), reinforcing the hypothesis in line with E167K and L156P.
Conclusion:
This study confirms the functional loss impact of E167K and L156P on protein structure, leading to liver disease, and identifies a premature stop codon with potential therapeutic efficacy for NAFLD.
02 Retatrutide, a Triple Hormone Receptor Agonist, Resolves Hepatic Steatosis in Over 85% of MASLD Subjects
Research Background:
Retatrutide (RETA; LY3437943), a triple hormone receptor agonist, stimulates glucagon-like peptide-1 receptor (GLP-1), glucose-dependent insulinotropic peptide receptor (GIP), and glucagon receptor (GCG). It is under investigation for the treatment of obesity. A 48-week Phase 2 obesity study demonstrated weight reductions of 22.8% and 24.2% with RETA doses of 8mg and 12mg, respectively. Dr. Sanyal AJ from Virginia Commonwealth University presented findings on RETA’s impact on hepatic fat in subjects with metabolic associated fatty liver disease (MASLD) and its correlation with metabolic parameters.
Research Methods:
Adult subjects aged 18-75 with a body mass index (BMI) ≥30, or BMI ≥27 kg/m2 with ≥1 weight-related comorbidity, excluding type 2 diabetes patients, were randomized into RETA (1, 4, 8, or 12 mg) or placebo groups. The treatment involved subcutaneous injections once a week for 48 weeks. The MASLD subgroup included participants with liver fat content ≥10% assessed by MRI proton density fat fraction (PDFF). The primary outcome was the change in liver fat content at 24 weeks (LF CFB), with additional outcomes including LF CFB at 48 weeks and the proportion of subjects achieving liver fat content <5%. The study also explored the relationship between LF CFB and changes in weight, waist circumference, and fasting metabolic biomarkers.
Research Results:
Among 338 participants, 98 (46.9% female) joined the MASLD subgroup. At baseline, mean age was 46.6 years, mean BMI was 38.4 kg/m2, mean waist circumference was 118.3 cm, mean alanine transaminase (ALT) was 35.9 IU/L, mean aspartate transaminase (AST) was 25.4 IU/L, fibrosis-4 (FIB4) was 0.79, and enhanced liver fibrosis (ELF) score was 8.1.
Baseline liver fat content across treatment groups ranged from 15.6% to 21.0%. At 24 weeks, mean relative LF CFB (%) was -42.9% (RETA 1mg), -57.0% (4mg), -81.4% (8mg), -82.4% (12mg), and +0.3% (placebo). At 48 weeks, mean relative LF CFB was -51.3% (1mg), -59.0% (4mg), -81.7% (8mg), -86.0% (12mg), and -4.6% (placebo) (all P<0.001 vs. placebo).
At week 48, the proportions of subjects achieving liver fat content <5% were 57% (1 mg), 29% (4 mg), 89% (8 mg), 93% (12 mg), and 0% (placebo) (all P<0.001 vs. placebo). Changes in ALT and AST were inconsistent compared to placebo. At week 48, the decrease in relative LF CFB was significantly correlated with changes in weight and waist circumference relative to baseline (r=0.774 and 0.588, both P<0.001), demonstrating a nonlinear relationship between LF CFB and weight change, with maximal liver fat reduction occurring at approximately 20% weight loss (P=0.002). RETA doses ≥4 mg improved insulin sensitivity, reflected in fasting insulin (range: -37.3% to -70.9%) and HOMA2-IR (insulin; -35.8% to -69.3%) reductions compared to placebo, while adiponectin increased (29.8% to 99.3%) (P<0.05).
By week 24, RETA doses ≥4 mg significantly altered biomarkers of lipid storage and metabolism compared to placebo (P<0.05), including triglycerides (TG; range -35.4% to -40.0%), leptin (-29.0% to -55.8%), and FGF-21 (-52.2% to -65.7%) reductions, and an increase in beta-hydroxybutyrate (BOHB; 78.0% to 181.2%). At 24 and 48 weeks, the reduction in relative liver fat content was significantly linearly correlated with changes in liver volume, TG, insulin, HOMA2-IR, adiponectin, leptin, and FGF-21 relative to baseline (all P<0.05), but not with BOHB.
Research Conclusion:
Treatment with RETA 8mg and 12mg resolved hepatic steatosis in over 85% of MASLD subjects. The reduction in liver fat content showed a linear correlation with improvements in insulin sensitivity and metabolic markers. The maximum reduction in liver fat content was achieved at approximately 20% weight loss.
03 Relationship between Non-Invasive Measurements and Histological Response in Non-Alcoholic Steatohepatitis (NASH) with Fibrosis Patients: 52-Week Data from the Phase III MAESTRO-NASH Trial
Research Background:
The MAESTRO-NASH trial (NCT03900429) is an ongoing 54-month, randomized, double-blind, placebo-controlled Phase III trial evaluating the efficacy of Resmetirom in patients with histologically confirmed non-alcoholic steatohepatitis (NASH) with fibrosis. 966 patients with biopsy-confirmed NASH were randomly assigned to receive daily treatment with 80 mg Resmetirom, 100 mg Resmetirom, or placebo in a 1:1:1 ratio. Tissue endpoints were assessed at 52 weeks, and the dual primary endpoints at week 52 were NASH resolution with no worsening of fibrosis (non-response, NR) or a decrease of at least 1 point in fibrosis with no worsening of NAFLD Activity Score (NAS) (partial response, FR).
Research Methods:
Adults with ≥3 metabolic risk factors, liver stiffness ≥8.5 kPa, liver fat content ≥8%, biopsy-confirmed NASH with F1B-F3 fibrosis, and NAS score ≥4 were eligible for the MAESTRO-NASH trial. The study assessed the relationship between non-invasive measurements and histological response (NR and/or FR) in the Resmetirom 80mg, Resmetirom 100mg, and placebo groups.
Research Results:
Patients with biopsy-confirmed NASH and fibrosis had high metabolic risks, including obesity (mean BMI 36), type 2 diabetes (70%), hypertension (78%), and a 10-year atherosclerotic cardiovascular disease (ASCVD) risk score >14. Baseline FibroScan Vibration-Controlled Transient Elastography (VCTE) values for the Resmetirom 80mg, Resmetirom 100mg, and placebo groups were 13.3 (6.8), 13.6 (7.1), and 12.9 (5.6) kPa, respectively. The baseline Enhanced Liver Fibrosis (ELF) score for all fibrosis groups was 9.8 (0.87), and FIB-4 for all dose groups was 1.3.
In the paired biopsy populations for 80 mg and 100 mg doses, MRI-PDFF scores decreased on average by 42% and 52%, respectively. In the Resmetirom 80mg or 100mg treatment groups, the proportions of subjects achieving NR (i.e., no fibrosis worsening) were 28% and 38% higher than the placebo group, and the proportions achieving FR (i.e., no NAS score worsening) were 17% and 18% higher than the placebo group when MRI-PDFF scores decreased by ≥30%. In Resmetirom-treated patients with an increase in sex hormone-binding globulin (SHBG) ≥120%, the proportions achieving NR were 34% and 37% higher, and those achieving FR were 22% and 20% higher, in the 80mg and 100mg groups, respectively, compared to the placebo group.
In the Resmetirom 100mg group, 96%, 88%, and 92% of NR, FR, and NR and/or FR responders had a ≥30% reduction in MRI-PDFF (histological response threshold). Among Resmetirom-treated patients with no NR or FR and a ≥30% decrease in MRI-PDFF, half showed a reduction in NAS score of ≥2 points. In biopsies, NR was correlated with FR (r2=0.30). Other factors associated with NR and FR in the Resmetirom 100mg group included PDFF (r2=0.39, 0.23), ALT reduction (r2=0.20, 0.24), and a decrease in liver volume (r2=0.25, 0.18). AST and FibroScan CAP showed weaker correlations. The correlations for the Resmetirom 80mg group were similar. LDL-C reduction was unrelated to NR or FR. Although Resmetirom treatment reduced Magnetic Resonance Elastography (MRE kPa), Fibroscan VCTE, and ELF, these reductions were not associated with NR or FR.
Research Conclusion:
Under both Resmetirom treatment doses (80mg and 100mg), NASH resolution and fibrosis reduction were associated with a ≥30% decrease in MRI-PDFF and a ≥120% increase in SHBG. Additional analyses, including artificial intelligence-based histological response assessments, are ongoing.
04 Itfg1: A Promising Target for Enhancing Liver Regeneration and Treating Chronic Liver Disease
Research Background:
The liver possesses significant regenerative capacity; however, under conditions of chronic liver injury, this ability can be depleted, leading to the accumulation of fibrotic tissue and ultimately resulting in end-stage liver disease. Targeting regenerative interruptions to enhance endogenous regenerative capacity represents a novel therapeutic approach. Researchers established an in vivo functional gene screening mechanism to identify such regenerative interruptions, with Integrin-β-like 1 (Itfg1) emerging as a prominent target. It has been demonstrated that liver cell-specific knockdown of Itfg1 can enhance the liver’s endogenous regenerative capacity and counteract chronic liver disease.
Research Methods:
In a thioacetamide (TAA)-induced chronic liver disease model, researchers conducted in vivo functional gene RNA interference (RNAi) screening. Two independent short hairpin RNAs (shRNAs) targeting Itfg1 were significantly enriched. Subsequently, the impact of Itfg1 knockdown on the proliferation of mouse and human liver cells was tested in vitro. Additionally, using a fumarylacetoacetate hydrolase (FAH) knockout mouse model, the researchers assessed accelerated liver regeneration driven by Itfg1 knockdown, enhanced regeneration after partial hepatectomy, and reduced fibrosis under conditions of chronic liver injury, including “Western diet”-driven non-alcoholic fatty liver disease (NAFLD). To elucidate the mechanisms behind Itfg1 inhibition driving accelerated liver regeneration, in-depth transcriptomic and proteomic analyses were performed.
Research Results:
In immortalized mouse and human liver cells, Itfg1 knockdown accelerated cell proliferation and wound healing in vitro. In vivo knockdown of Itfg1 in liver cells enhanced their ability to repopulate in the livers of FAH-deficient mice. Moreover, in fully regenerated mice, each liver cell expressing shRNA targeting Itfg1 exhibited accelerated liver regeneration after partial hepatectomy. The enhanced regenerative capacity also mitigated chronic injuries induced by TAA and “Western diet.” In the NAFLD model, Itfg1 knockdown not only reduced fibrosis but also decreased fat deposition. Transcriptomic and proteomic analyses indicated that Itfg1 knockdown affected fatty acid metabolism, MAPK and AKT pathways.
Research Conclusion:
This study identifies and validates Itfg1 as a target for enhancing the liver’s endogenous regenerative capacity. Augmenting endogenous regenerative capacity may alleviate chronic liver disease. The research findings are currently being translated into RNAi-based therapeutic approaches.
Study 05: Empagliflozin Equally Effective in Reducing Liver Fat Content in T2DM and Non-Diabetic Patients
Research Background:
This study aimed to investigate the impact of empagliflozin on liver fat content in both Type 2 Diabetes Mellitus (T2DM) patients and non-diabetic individuals. Additionally, the study aimed to explore the relationship between the reduction in liver fat content and other metabolic effects of empagliflozin.
Research Methods:
A total of 30 T2DM patients and 27 non-diabetic participants were randomly assigned in a 2:1 ratio to receive either empagliflozin or a matching placebo in a double-blind fashion. Participants underwent an oral glucose tolerance test (OGTT) and magnetic resonance spectroscopy analysis of liver fat content before and after the treatment. The study also measured liver glucose production using 3H-glucose infusion at the start of the treatment.
Research Results:
In both T2DM and non-diabetic subjects, empagliflozin led to an absolute decrease in liver fat content by 2.75% and 1.93%, respectively, while the placebo group showed an increase of 0.9% and 0.8% (both p < 0.05). The reduction in liver fat content was significantly influenced by baseline liver fat content and was closely correlated with weight loss (r=0.53, p < 0.001) and improved insulin sensitivity induced by empagliflozin (r=-0.51, p < 0.001). The reduction was not influenced by decreases in fasting blood glucose or increased liver glucose production.
Research Conclusion:
Empagliflozin is equally effective in reducing liver fat content in both T2DM and non-diabetic patients. The decrease in liver fat content is unrelated to plasma glucose concentration but is closely associated with weight loss and improved insulin sensitivity.
Study 06: Saroglitazaar Improves Liver Hardness and Enzymes in Non-Alcoholic Steatohepatitis (NASH) Patients
Research Background:
While progress has been made in understanding the pathophysiology of Non-Alcoholic Steatohepatitis (NASH), there is still no proven drug therapy that effectively improves outcomes. Peroxisome Proliferator-Activated Receptors (PPARs) play a crucial role in metabolic homeostasis and inflammation, with PPAR gene knockout mice being susceptible to NASH. Saroglitazaar, a dual agonist of PPAR-α and PPAR-γ, has been approved for treating diabetes-related lipid abnormalities. Although approved for NASH patients in India, data on its impact on liver fibrosis improvement is awaited.
Research Methods:
This prospective, randomized, open-label clinical trial was conducted at a tertiary medical center from July 2021 to December 2022. Ethical approval was obtained, and the trial was registered with CTRI (CTRI/2021/07/034946). Biopsy-confirmed NASH patients were randomly assigned to receive either Saroglitazaar 4 mg OD (Group I) or Saroglitazaar 4 mg OD + Vitamin E 400 IU OD (Group II) for six months. The primary endpoints were NAS score, liver stiffness measurement (LSM) values, and improvement in liver enzyme (SGPT). Secondary endpoints included improvement in body mass index (BMI), serum triglycerides (TG), and cholesterol (Chol) levels.
Research Results:
Out of 53 enrolled patients, two in Group I were lost to follow-up, leaving 25 in Group I and 26 in Group II for analysis. Both groups exhibited comparable baseline characteristics. In terms of primary endpoints, LSM improvement was significant in Group I (9.1 to 7.0 kPa, p = 0.03) and Group II (7.9 to 7.1 kPa, p = 0.04), as was SGPT improvement in Group I (101 to 81 U/L, p = 0.02) and Group II (98 to 75 U/L, p = 0.04). There was no change in NAS score in both groups from baseline to 24 weeks. Regarding secondary endpoints, both Group I (234 to 167 mg/dL, p = 0.003; 234 to 199 mg/dL, p = 0.04) and Group II (223 to 188 mg/dL, p = 0.04; 229 to 189 mg/dL, p = 0.007) showed a significant reduction in triglycerides and total cholesterol, with no change in BMI.
Research Conclusion:
Saroglitazaar monotherapy effectively improves LSM and SGPT values in NASH patients. However, no improvement in NAS score was observed within 24 weeks.
Reference:
[1] Huang H, Vitali C, Phillips MC, et al. Deep Phenotyping of TM6SF2 to characterize hepatic steatosis, plasma lipid traits, and metabolic risk factors using a genome-first approach. AASLD 2023. Oral 147.
[2] Sanyal AJ, Frias JP, Thomas MK, et al. Triple hormone receptor agonist retatrutide resolves steatosis in >85% of subjects with MASLD and obesity in association with improved metabolic health. AASLD 2023. Oral 148.
[3] Loomba R, Schattenberg JM, Taub RA, et al. Relationship of non-invasive measures with histological response in patients with nonalcoholic steatohepatitis and fibrosis: 52-week data from the phase 3 maestro-NASH trial. AASLD 2023. Oral 149.
[4] Iakovleva V, Potapova A, Ong A, et al. Itfg1: a promising target for enhanced liver regeneration and chronic liver disease treatment. AASLD 2023. Oral 150.
[5] Abdelgani S. Empagliflozin is equally effective in reducing liver fat content in T2DM patients and in non-diabetic individuals, a randomized trial. AASLD 2023. Oral 151.
[6] Gupta T, Kaushik P, Mittal R. Saroglitazaar is effective in improving liver stiffness measurement and liver enzymes in nonalcoholic steatohepatitis. AASLD 2023. Oral 152.
