A Triple-Mutant Mouse Model Reproducing Both Classic and Intermediate Maple Syrup Urine Disease

This invention is a genetically engineered mouse strain carrying a targeted knockout of the branched-chain keto acid dehydrogenase (BCKDH) E2 subunit gene combined with a tetracycline-regulated human E2 transgene, producing animals that model both the lethal classic and the milder intermediate forms of Maple Syrup Urine Disease. It gives researchers, for the first time, a viable, breedable small-animal platform for studying MSUD pathology and testing gene- and cell-based therapies that was previously unavailable.

Description

Unmet Need - Prior to this strain, no practical small-animal model existed that accurately reproduced human MSUD biochemistry and survival patterns, since the only available large-animal (calf) and ENU-mutant mouse models either diverged pathologically from human disease or did not target the actual deficient enzyme.

The strain was built by first disrupting the E2 (Dbt) subunit gene of BCKDH in embryonic stem cells, replacing part of exon 4 and all of exon 5 with a selectable marker cassette, then generating chimeric and subsequently heterozygous knockout mice. Homozygous knockout animals completely lack BCKDH enzyme activity and E2 protein, leading to markedly elevated circulating branched-chain amino acids and neonatal lethality that closely parallels classic MSUD in humans.

To create a surviving model of the milder intermediate disease form, these knockout mice were crossed with animals carrying a liver-specific tetracycline-controlled transactivator transgene and a second transgene expressing a modified human E2 cDNA under tetracycline-responsive control. The resulting triple-mutant mice express just 5–6% of normal BCKDH activity in the liver, enough to prevent neonatal death but not enough to normalize blood amino acid levels giving a stable, breedable model of intermediate MSUD that mirrors the biochemical hallmarks seen in human patients.

Applications

- Preclinical testing of gene therapy approaches for MSUD
- Evaluation of cell-based therapies, including hepatocyte or stem cell transplantation, for metabolic liver disease
- Research tool for studying branched-chain amino acid metabolism and related enzyme deficiency disorders
- Platform for thiamin-responsiveness studies in MSUD
- Model system for broader studies of mitochondrial multi-subunit enzyme disorders

Advantages

- Closely replicates the biochemical and survival phenotypes of both classic and intermediate human MSUD
- Provides a viable, fertile, breedable model (unlike the lethal classic-only knockout)
- Targets the actual enzyme deficient in human disease (BCKDH E2), unlike earlier BCAT-mutant mouse models
- More practical and accessible than large-animal (calf) MSUD models
- Already deposited and available for distribution through an established mouse repository

Invention Readiness

The strain has been fully developed and biochemically characterized, with published data confirming BCKDH enzyme activity, E2 protein expression, and blood amino acid profiles in knockout and triple-mutant animals. The mice are already available through an external mouse strain repository. No further regulatory approval, human testing, or independent trials are required to advance the model, though additional transgenic lines could be screened to identify strains more responsive to tetracycline-based regulation.

IP Status

Research Tool

Related Publication(s)

Homanics, Gregg E., et al. "Production and characterization of murine models of classic and intermediate maple syrup urine disease." BMC medical genetics 7.1 (2006): 33. https://doi.org/10.1186/1471-2350-7-33