Assessment of Gut Microbiota-Derived Amino Acid Metabolite Production in Patients With MASLD (MASLD-GUT)

Evaluation of the Production of Amino Acid-Derived Metabolites by the Gut Microbiota of Patients With Metabolic Dysfunction-Associated Steatotic Liver Disease (MASLD)

Metabolic dysfunction-associated steatotic liver disease (MASLD) encompasses a spectrum of liver disorders ranging from simple steatosis-a relatively benign and non-progressive condition-to metabolic dysfunction-associated steatohepatitis (MASH), characterized by hepatocellular inflammation. MASLD is now the leading cause of chronic liver disease worldwide, affecting approximately one in three adults, particularly those with obesity or type 2 diabetes.

Recent studies have highlighted a strong interconnection between the gut microbiota, the liver, metabolism, and the immune system, collectively referred to as the gut-liver axis. Alterations in the gut microbiota are observed at all stages of MASLD, and several microbial metabolites-such as trimethylamine, bile acids, short-chain fatty acids, and ethanol-have been implicated in disease progression.

Emerging evidence points to a role for gut-derived metabolites of tryptophan (Trp) and phenylalanine (Phe), including phenylacetic acid (PAA), 3-(4-hydroxyphenyl)-lactate (HPL), and phenyllactate (PL). These compounds have been associated with the severity of MASLD, particularly with hepatic steatosis and fibrosis. Elevated plasma levels of aromatic amino acids (AAAs), such as L-phenylalanine and L-tyrosine, are also correlated with increased hepatic fat content.

A newly identified Phe-derived metabolite, N-acetyl-phenylalanine (NAPA), together with PAA, HPL, and PL, has been shown to correlate with hepatic steatosis. These metabolites can induce steatosis both in vitro and in vivo, acting through the disruption of endoplasmic reticulum-mitochondria interactions. They therefore represent potential new therapeutic targets.

These four metabolites of interest (NAPA, PAA, HPL, PL) can be produced both by gut bacteria and through endogenous human metabolism. Positive correlations between plasma NAPA concentrations and specific bacterial species have been observed, although the responsible taxa remain to be identified.

HYPOTHESIS

We hypothesize that the gut microbiota of MASLD patients produces aromatic amino acid-derived metabolites, contributing to the elevated plasma concentrations observed in these patients

Two complementary strategies will be used : Human Microbiota Culture and Fecal Microbiota Transplantation

Study Overview

• Status: Not yet recruiting
• Conditions: Metabolic Dysfunction-Associated Steatohepatitis (MASH); MASLD - Metabolic Dysfunction-Associated Steatotic Liver Disease
• Intervention / Treatment: Procedure: stool sampling, dietary assessment, and collection of blood and urine samples

• Study Type: Interventional
• Enrollment (Estimated): 24
• Phase: Not Applicable

Contacts and Locations

• Study Contact: Name: Cyrielle CAUSSY, MD, PhD; Phone Number: +33 4 78 86 44 48; Email: cyrielle.caussy@chu-lyon.fr
• Study Contact Backup: Name: Dominique DELAUNAY, PhD; Phone Number: +33 4 72 11 00 64; Email: dominique.delaunay@chu-lyon.fr

Study Locations

• France
  • Pierre-Bénite, France, 69495
    • Centre Hospitalier Lyon Sud Service Endocrinologie, Diabète et nutrition
    • Contact: Cyrielle CAUSSY, MD, PhD; Phone Number: +33 4 78 86 44 48; Email: cyrielle.caussy@chu-lyon.fr

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: Adult; Older Adult
• Accepts Healthy Volunteers: Yes

Description

Inclusion Criteria:

• For patients and healthy volunteers:

  • Age between 18 and 80 years.
  • Non-diabetic participant (fasting blood glucose < 1.26 g/L, or absence of insulin therapy or oral antidiabetic medication).
  • Body mass index (BMI) between 18 and 30 kg/m².
  • For women of childbearing potential, use of at least one recognized effective contraceptive method.
  • Very regular bowel movements every 24 to 48 hours.
  • Participant living within 100 km of the Lyon Sud Hospital Center (CHLS).
  • Participant willing to participate in the study and providing written informed consent.
  • Participant affiliated with the general French Social Security system or an equivalent scheme.

• For patients:

  • Presence of MASLD according to the definition of the European Association for the Study of the Liver (EASL), defined by hepatic steatosis on imaging performed within the previous year (abdominal ultrasound, abdominal CT scan, magnetic resonance imaging, or controlled attenuation parameter (CAP) measured by FibroScan) and at least one cardiometabolic criterion among: dyslipidemia, arterial hypertension, overweight (BMI ≥ 25 kg/m²), impaired glucose tolerance, or type 2 diabetes.
  • No history of liver transplantation.
  • No excessive alcohol consumption (less than 20 g/day for women and less than 30 g/day for men).
  • Patient followed in the Endocrinology, Diabetes and Nutrition Department at Lyon Sud Hospital.

Exclusion Criteria:

• For patients and healthy volunteers:

  • Participant with active inflammatory, infectious, cardiovascular, or neoplastic disease.
  • Participant with a history of colectomy, small bowel resection, or cholecystectomy.
  • Participant who received antibiotics, prebiotics, or probiotics within the past 3 months.
  • Participant using laxatives (more than 2 doses per day over the past 3 months).
  • Participant with chronic constipation.
  • Pregnant, parturient, or breastfeeding women.
  • Participant deprived of liberty by judicial or administrative decision.
  • Participant receiving psychiatric care.
  • Participant institutionalized for reasons other than research.
  • Adult participant under legal protection (guardianship or trusteeship).
  • Participant who does not understand the French language and cannot provide informed consent.
  • Participant already enrolled in a study presenting a conflict of interest with the present study.

• For healthy volunteers

  • Known liver disease
  • No long-term drug medication except oral contraception for women.

Study Plan

How is the study designed?

Design Details

• Primary Purpose: Basic Science
• Allocation: Non-Randomized
• Interventional Model: Single Group Assignment
• Masking: None (Open Label)

Number of Arms

2

Arms and Interventions

Participant Group / Arm	Intervention / Treatment
Other: MASLD Group; Patients with metabolic dysfunction-associated steatotic liver disease Participants in this group are adults (18-80 years) with confirmed MASLD based on recent imaging and at least one associated cardiometabolic risk factor. These patients are already monitored in the Endocrinology, Diabetes, and Nutrition Department at Lyon Sud Hospital.	Procedure: stool sampling, dietary assessment, and collection of blood and urine samples; MASLD group includes individuals receiving routine clinical care for liver-related conditions. During their scheduled medical visits, participants undergo standard clinical assessments and routine blood tests. Additional research-specific procedures-such as stool sampling, dietary assessment, and collection of blood and urine samples-are carried out without altering routine patient management. A fraction of blood sample is used for NAPA measurement. Stool samples are collected at home and returned directly to the Endocrinology, Diabetes and Nutrition Department within 24 hours after collection. Healthy volunteers undergo the same research-specific procedures as patients but exclusively for research purposes, with no routine-care-related assessments.
Other: Healthy Volunteers Group; Healthy volunteers meet the general inclusion criteria shared with the patient group but do not present MASLD or other cardiometabolic diseases and do not have any liver pathology. Their participation serves as a control group for comparison with MASLD patients in the context of the study's biological and metabolic analyses.	Procedure: stool sampling, dietary assessment, and collection of blood and urine samples; MASLD group includes individuals receiving routine clinical care for liver-related conditions. During their scheduled medical visits, participants undergo standard clinical assessments and routine blood tests. Additional research-specific procedures-such as stool sampling, dietary assessment, and collection of blood and urine samples-are carried out without altering routine patient management. A fraction of blood sample is used for NAPA measurement. Stool samples are collected at home and returned directly to the Endocrinology, Diabetes and Nutrition Department within 24 hours after collection. Healthy volunteers undergo the same research-specific procedures as patients but exclusively for research purposes, with no routine-care-related assessments.

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Concentration of NAPA in Stool Culture Medium	Quantification of the metabolite N-acetyl-phenylalanine (NAPA) in the culture medium of stool samples cultivated using the MiPro in vitro human gut microbiota culture system.	Within 30 days after Visit 1 (sample collection and analysis performed at a single time point)

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Concentration of Selected AAA-Derived Metabolites in Stool Culture Medium	Quantification of selected aromatic amino acid (AAA)-derived metabolites in the culture medium of stool samples cultivated using the MiPro in vitro human gut microbiota culture system.	Within 30 days after Visit 1 (sample collection and analysis performed at a single time point)
Gut Microbiota Composition in Stool Samples	Characterization of gut microbiota composition in stool samples who produce NAPA or other metabolites of interest, to identify bacterial species potentially involved in metabolite production. Microbiota profiling will be performed on a stool sample using bacterial DNA sequencing methods (16S rRNA gene sequencing or shotgun metagenomics).	Within 30 days after Visit 1 (sample collection and analysis performed at a single time point)
Concentration of NAPA in plasma	The quantification of NAPA level will be performed on plasma samples	Baseline Visit 1 (sample collection and analysis performed at a single time point)

Collaborators and Investigators

• Sponsor: Hospices Civils de Lyon

Study record dates

Study Major Dates

• Study Start (Estimated): May 1, 2026
• Primary Completion (Estimated): November 1, 2026
• Study Completion (Estimated): November 1, 2026

Study Registration Dates

• First Submitted: December 11, 2025
• First Submitted That Met QC Criteria: December 17, 2025
• First Posted (Actual): December 31, 2025

Study Record Updates

• Last Update Posted (Actual): April 3, 2026
• Last Update Submitted That Met QC Criteria: April 2, 2026
• Last Verified: April 1, 2026

More Information

Terms related to this study

• Keywords: MASLD; MASH; GUT MICROBIOTA; N-acétyl-phenylalanine (NAPA)
• Additional Relevant MeSH Terms: Health Care Quality, Access, and Evaluation; Investigative Techniques; Epidemiologic Methods; Data Collection; Health Care Evaluation Mechanisms; Quality of Health Care; Public Health; Environment and Public Health; Epidemiologic Measurements; Nutrition Assessment
• Other Study ID Numbers: 69HCL25_1015

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No