Research we would like to help move forward

From research on our radar to research for TMAU.

TMAU International is following several scientifically plausible routes toward restoring functional FMO3 activity: providing FMO3-producing liver cells, delivering a functional FMO3 instruction to a person’s own hepatocytes, and increasing FMO3 production where the gene can still make a working enzyme.

Whilst these technologies have been demonstrated for other conditions, none have yet been investigated for TMAU. We are highlighting them because donations can help us do more than observe progress in other conditions. As funding allows, we intend to work directly with qualified scientists and help fund the TMAU-specific research needed to test these possibilities.

Explore the research Understand TMAU
Abstract illustration of cellular clusters, DNA, and gene-network lines in violet tones.
Early science. Testable directions.
These are cutting-edge research areas

The proposed applications require laboratory testing, independent review, and appropriate ethical and regulatory oversight. Nothing here is medical advice.

Where we want to contribute

Routes toward restoring FMO3 function.

Each area begins with published science, but none are ready for use in TMAU. Our ambition is to help close that gap: bringing patient priorities into study design, finding specialist collaborators, helping shape proposals, and funding proof-of-concept experiments as donations allow. These are early hypotheses, not promises of treatment, but they are grounded in peer-reviewed science and with further research, have the potential to help find a solution to TMAU.

01 Minicircle technologySmall loops of DNA instructions 02 Satellite hepatocytesHealthy liver-cell clusters 03 Regulatory mechanismsThe switches around FMO3
01

Minicircle technology

Could liver cells receive a functional FMO3 expression cassette?

Minicircle DNA is a compact, circular, nonviral gene-delivery vector. Unlike a conventional plasmid, it is stripped of most bacterial backbone sequences and retains primarily an expression cassette: the coding sequence, promoter, and regulatory elements needed to make a selected protein.

Once a minicircle reaches a cell nucleus, it can remain episomal (physically separate from the chromosomes) and be transcribed into RNA, which the cell translates into protein. Avoiding deliberate integration reduces the risk of disrupting the genome. The trade-off to this is that episomal DNA can be diluted or lost when cells divide.

A TMAU-specific minicircle could theoretically carry functional human FMO3 cDNA under liver-selective regulatory elements. If a safe delivery system placed it into enough hepatocytes, those cells could begin producing active FMO3. This would be gene augmentation: supplying a working instruction without repairing the existing gene.

Proposed FMO3 architecture

A compact FMO3 expression cassette. The central challenge is targeted delivery to enough hepatocyte nuclei, followed by safe and durable FMO3 production.

What the studies explored

In PKU mice, a synthetic hepatocyte-specific cassette produced PAH in 14–20% of hepatocytes and maintained metabolic correction for more than a year. A later design combined codon-optimized PAH cDNA, a truncated intron, and a native liver promoter to lower the dose. More than 95% of vector and activity disappeared after liver regeneration, confirming the durability trade-off of episomal DNA. Both used experimental hydrodynamic delivery, which is not a clinical human delivery method.1,2

Why we want to help fund the next step

These studies show that a nonviral DNA cassette can produce a functional metabolic enzyme in liver cells. We intend to pursue opportunities to fund FMO3-specific vector design, expression testing, and delivery research, beginning in human hepatocyte or liver models before appropriate preclinical studies.

What TMAU research needs to solve

Writing an FMO3 cassette is not the main obstacle: safe, efficient delivery to enough human hepatocytes is. Research must establish liver targeting, dose, persistence, repeat dosing, immune effects, and physiological expression. It must also confirm that the protein folds, binds its required cofactors, localizes correctly within hepatocytes, and measurably reduces TMA. Solving those problems could create a nonviral route to restoring FMO3 activity without permanent genome modification.

02

Satellite hepatocytes

Could transplanted hepatocytes provide a working source of FMO3?

Hepatocytes are the principal metabolic cells of the liver and the normal site of FMO3 expression. Satellite-hepatocyte technology aims to establish a small, functioning graft of liver cells at another location in the body, rather than replacing the whole liver.

The INSITE platform combines primary human hepatocytes and supportive fibroblasts with injectable gelatin methacryloyl (GelMA) hydrogel microspheres. Under pressure, the packed microspheres shear-thin and flow through a needle; after delivery they jam into a porous scaffold. That scaffold localizes the cells, permits nutrient exchange, and allows host blood vessels to grow into the graft. Ultrasound guides placement and can track the tissue afterward.

For TMAU, the proposed application is direct: a vascularized graft containing hepatocytes that express functional FMO3 could encounter trimethylamine (TMA) in the circulation and convert it to trimethylamine N-oxide (TMAO). The living graft, not free enzyme, could provide an ongoing source of FMO3 activity.

A simple picture of the idea
FMO3-functional hepatocytes Cells selected or engineered to produce active FMO3.
A vascularizable niche GelMA microspheres localize the cells and support blood-vessel ingrowth.
Potential TMA clearance Circulating TMA could reach the graft and be oxidized by FMO3.

What the study explored

In mice, primary human hepatocytes delivered in the INSITE scaffold remained localized, recruited vasculature, and secreted human liver proteins for the eight-week study period. This established a cell-engraftment platform. It did not test FMO3 or TMAU.3

Why we want to help fund the next step

It offers a plausible way to add metabolically active, FMO3-producing cells without replacing the liver or permanently altering a patient’s genome. We want to help fund studies that test TMA clearance and establish how much functioning graft tissue is needed.

What TMAU research needs to solve

The next experiments could use FMO3-expressing cells in TMAU-relevant laboratory and animal models, measuring whether the graft retains mature adult FMO3 expression and alters TMA-to-TMAO flux. Researchers must also establish cell dose, durability, vascularization, immune compatibility, cell sourcing, and safe control or removal of the graft. If those challenges can be solved, this could restore missing metabolic function without whole-organ transplantation.

Plain-language overview

MIT News: injectable satellite livers
03

Regulatory mechanisms

Could insufficient FMO3 production be safely increased?

FMO3 output depends on more than its protein-coding sequence. Promoters and enhancers regulate transcription. Liver-enriched transcription factors such as HNF4α, C/EBPβ, and NF-Y bind regulatory DNA. Hormonal and metabolic signals influence those factors. Additional cellular processes determine how much functional enzyme remains.

This matters greatly for people whose FMO3 coding sequence can make functional enzyme but whose hepatocytes produce too little of it. An inducer therapeutic (a small molecule or other intervention that safely activates a validated regulatory pathway) might enable the person’s own liver to make more functional FMO3.

A proposed intervention pathway

Regulatory DNAPromoters and enhancers
Safe inductionA selective regulatory pathway
FMO3 transcriptionMore FMO3 RNA in hepatocytes
TMA oxidationMore functional enzyme converts TMA

An inducer could only help where the FMO3 produced is functional. The aim would be selective, measurable induction in the right biological subgroup, not indiscriminate activation.

What the research explores

Cell, liver-sample, animal, and genetic studies show that FMO3 transcription responds to a layered control system. HNF4α, NF-Y, C/EBPβ, USF1, and YY1 emerged as candidate transcriptional regulators. Steroid and glucocorticoid signalling altered expression in specific models. NRF2 was tested but did not appear indispensable.4–14

Why we want to help fund the next step

For individuals with inadequate FMO3 expression rather than a protein-damaging loss-of-function variant, increasing production of their own working enzyme could be more direct than replacing cells or delivering a new gene. We want to fund work that identifies the relevant mechanisms and screens selective inducers.

What TMAU research needs to solve

Researchers need patient-relevant hepatocyte models, assays that measure TMA-to-TMAO conversion rather than expression alone, and a way to identify who could benefit. Candidate inducers must raise functional FMO3 by a useful amount without disrupting drug metabolism or wider physiology. For the right subgroup, a safe, selective inducer could let existing hepatocytes supply more enzyme.

References

Online citations for the research discussed on this page.

  1. Viecelli HM, Harbottle RP, Wong SP, et al. Treatment of phenylketonuria using minicircle-based naked-DNA gene transfer to murine liver. Hepatology. 2014;60(3):1035–1043.
  2. Grisch-Chan HM, Schlegel A, Scherer T, et al. Low-dose gene therapy for murine PKU using episomal naked DNA vectors expressing PAH from its endogenous liver promoter. Molecular Therapy – Nucleic Acids. 2017;7:339–349.
  3. Kumar V, Yun J, Elledge SK, et al. Image-guided injectable niche for hepatocyte transplantation. Cell Biomaterials. 2026;2:100378.
  4. Koukouritaki SB, Poch MT, Cabacungan ET, McCarver DG, Hines RN. Discovery of novel FMO3 single nucleotide polymorphisms and functional analysis of upstream haplotype variants. Molecular Pharmacology. 2005;68(2):383–392.
  5. Klick DE, Hines RN. Mechanisms regulating human FMO3 transcription. Drug Metabolism Reviews. 2007;39(2–3):419–442.
  6. Klick DE, Shadley JD, Hines RN. Differential regulation of human hepatic FMO3 by C/EBPβ liver inhibitory and liver activating proteins. Biochemical Pharmacology. 2008;76(2):268–278.
  7. Shimizu M, Murayama N, Nagashima S, Fujieda M, Yamazaki H. Complex mechanism underlying transcriptional control of the haplotyped FMO3 gene in Japanese. Drug Metabolism and Pharmacokinetics. 2008;23(1):54–58.
  8. Nagashima S, Shimizu M, Yano H, et al. Inter-individual variation in FMO3 in livers from Japanese: correlation with hepatic transcription factors. Drug Metabolism and Pharmacokinetics. 2009;24(3):218–225.
  9. Shimizu M, Allerston CK, Shephard EA, Yamazaki H, Phillips IR. Relationships between FMO3 genotype and trimethylaminuria phenotype in a Japanese population. British Journal of Clinical Pharmacology. 2014;77(5):839–851.
  10. Esposito T, Varriale B, D’Angelo R, Amato A, Sidoti A. Regulation of FMO3 gene expression by steroids in mice and humans. Hormone Molecular Biology and Clinical Investigation. 2014;20(3):99–109.
  11. Rudraiah S, Gu X, Hines RN, Manautou JE. NRF2 is not indispensable for human hepatic FMO3 gene expression in HepG2 cells. Toxicology in Vitro. 2016;31:54–59.
  12. Ganapathy T, Yuan J, Ho MYM, et al. Adipocyte FMO3-derived TMAO induces WAT dysfunction and metabolic disorders by promoting inflammasome activation in ageing. Nature Communications. 2025;16:8873.
  13. Chen B, Chen M, Pan X, et al. Upregulation of hepatic FMO3 by increased corticosterone via glucocorticoid receptor contributes to gestational diabetes mellitus. Drug Metabolism and Disposition. 2025;53(9):100133.
  14. Chen B, Chen M, Pan X, et al. Corrigendum to the 2025 FMO3–corticosterone article. Drug Metabolism and Disposition. 2026;54(1):100225.

Reference 14 is the publisher’s correction to reference 13 and is included for transparency.

Help turn a hypothesis into treatment

TMAU-specific research will not advance through observation alone.

TMAU International intends to be an active research partner: working with specialist laboratories, shaping questions around the needs of people with TMAU, seeding proof-of-concept work, and co-funding larger projects. No result can be guaranteed, but donations can give researchers the time, materials, models, and collaborative support required to find out what is possible.

Help fund TMAU research Why donations matter