Hyperammonemia is a clinical problem with severe consequences to the central nervous system. It is usually caused by liver disease, inherited metabolic disorders and various toxins. The main source of ammonia production is the bowel where it is generated and then diffuses into the portal blood. If the liver is unable to convert ambiguous ammonia to urea due to liver disease or an inherited enzymatic defect, the ammonia enters the systemic circulation exerting toxicity on the brain. The goal of this project is to develop a novel form off therapy for hyperammonemia aimed to act within aimed the intestinal environment. This approach will deliver into the bowel's lumen bacterial genes over-expressing arginine biosynthesis enzymes so that large amounts of arginine can be synthesized. Our hypothesis is that the expressed enzymes, residing within an ammonia rich environment, will trap ammonia and covert it to arginine rendering it non toxic. Escherichia coli strains incorporating large amounts of nitrogen into arginine have already been successfully engineered. These bacteria will be delivered to the bowel of a hyper- ammonemic animal model, the spf-ASH mouse. We will study the viability, colonization and anatomical distribution of the engineered bacteria in the bowel. The metabolic effects of this treatment of this treatment on the host with respect to ammonia metabolism will be investigated. Various regulated promoters will be tested for optimal expression of arginine biosynthesis genes in the bowel's environment As the anaerobic bacteria Bacteroides fragilis is orders of magnitudes more numerous within the colonic bacterial flora than E. coli, it is our ultimate vehicle for bacterial therapy. We are currently completing the identification of arginine biosynthesis genes and their regulation in this organism. B. fragilis will be engineered to over-produce arginine and will then be used to trap ammonia in the intestine of the spf-ASH mouse. The expression of arginine of arginine genes in the transformed B. fragilis will be investigated and their hyperammonemic mice as with engineered E. coli. The long term goal of this project is to test this therapy in humans with hyperammonemia after its efficacy and safety have been demonstrated in laboratory animals. There are many other inherited and acquired conditions which may be amenable to bacterial therapy. This project will provide better understanding of the promise and challenges of such therapeutic approach.

Project Start
2001-05-29
Project End
2001-12-31
Budget Start
Budget End
Support Year
7
Fiscal Year
2001
Total Cost
Indirect Cost
Name
University of Minnesota Twin Cities
Department
Type
DUNS #
168559177
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
Ou, Li; Przybilla, Michael J; Whitley, Chester B (2018) Metabolomics profiling reveals profound metabolic impairments in mice and patients with Sandhoff disease. Mol Genet Metab :
Ou, L; Przybilla, M J; Whitley, C B (2018) SAAMP 2.0: An algorithm to predict genotype-phenotype correlation of lysosomal storage diseases. Clin Genet 93:1008-1014
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Ou, Li; Przybilla, Michael J; Whitley, Chester B (2017) Proteomic analysis of mucopolysaccharidosis I mouse brain with two-dimensional polyacrylamide gel electrophoresis. Mol Genet Metab 120:101-110
Verhaart, Ingrid E C; Robertson, Agata; Wilson, Ian J et al. (2017) Prevalence, incidence and carrier frequency of 5q-linked spinal muscular atrophy - a literature review. Orphanet J Rare Dis 12:124
Ou, Li; Przybilla, Michael J; Koniar, Brenda L et al. (2016) Elements of lentiviral vector design toward gene therapy for treating mucopolysaccharidosis I. Mol Genet Metab Rep 8:87-93
Aronovich, Elena L; Hackett, Perry B (2015) Lysosomal storage disease: gene therapy on both sides of the blood-brain barrier. Mol Genet Metab 114:83-93
Satzer, David; DiBartolomeo, Christina; Ritchie, Michael M et al. (2015) Assessment of dysmyelination with RAFFn MRI: application to murine MPS I. PLoS One 10:e0116788

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