Characterization of Mutations at the Mouse Phenylalanine Hydroxylase Locus

McDonald, J. David; Charlton, C K

doi:10.1006/geno.1996.4508

Cited by 80 publications

(53 citation statements)

References 22 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Yet, as we and others observed that the commonly used mouse model for PKU, the BTBR-Pah enu2 strain, exhibits limited fertility (D MacDonald, personal communication), we crossed the Pah enu2 allele into the C57Bl/6 background to improve breeding performance. As expected, homozygous C57Bl/6-Pah enu2 or 'PKU' mice carrying the p.F263S missense mutation in the PAH protein 34 resulted in a complete deficiency of hepatic PAH activity (o0.02 mU/ mg; Table 1). In comparison, normal hepatic PAH activity in C57Bl/6 wild-type mice was 3.5070.64 mU/mg protein (see also Table 1).…”

Section: Resultssupporting

confidence: 75%

“…All animal experiments were approved by the State Veterinary Office of Zurich and carried out according to the guideline of the Swiss Law of Animal Protection. The Pah enu2 allele was crossed into C57BL/6 background, and the genotype was identified by PCR and Alw26I restriction endonuclease digestion according to the method published by McDonald et al 34 For gene therapy experiments, 8-10 weeks old adult male or female mice were anesthetized with isoflurane, and rAAV particles, dissolved in standard PBS solution, were injected into portal or tail veins using a 1 ml syringe with 30-gauge needle. For Phe monitoring, blood samples were collected from tail vein on Guthrie filter cards at different time points, and Phe (and Tyr) concentrations were determined by tandem mass spectrometry.…”

Section: Recombinant Aav Construction and Productionmentioning

confidence: 99%

See 1 more Smart Citation

Administration-route and gender-independent long-term therapeutic correction of phenylketonuria (PKU) in a mouse model by recombinant adeno-associated virus 8 pseudotyped vector-mediated gene transfer

2005

View full text Add to dashboard Cite

Phenylketonuria (PKU) is an inborn error of metabolism caused by deficiency of the hepatic enzyme phenylalanine hydroxylase (PAH) which leads to high blood phenylalanine (Phe) levels and consequent damage of the developing brain with severe mental retardation if left untreated in early infancy. The current dietary Phe restriction treatment has certain clinical limitations. To explore a long-term nondietary restriction treatment, a somatic gene transfer approach in a PKU mouse model (C57Bl/6-Pah enu2 ) was employed to examine its preclinical feasibility. A recombinant adenoassociated virus (rAAV) vector containing the murine Pah-cDNA was generated, pseudotyped with capsids from AAV serotype 8, and delivered into the liver of PKU mice via single intraportal or tail vein injections. The blood Phe concentrations decreased to normal levels (p100 mM or 1.7 mg/dl) 2 weeks after vector application, independent of the sex of the PKU animals and the route of application. In particular, the therapeutic long-term correction in females was also dramatic, which had previously been shown to be difficult to achieve. Therapeutic ranges of Phe were accompanied by the phenotypic reversion from brown to black hair. In treated mice, PAH enzyme activity in whole liver extracts reversed to normal and neither hepatic toxicity nor immunogenicity was observed. In contrast, a lentiviral vector expressing the murine Pah-cDNA, delivered via intraportal vein injection into PKU mice, did not result in therapeutic levels of blood Phe. This study demonstrates the complete correction of hyperphenylalaninemia in both males and females with a rAAV serotype 8 vector. More importantly, the feasibility of a single intravenous injection may pave the way to develop a clinical gene therapy procedure for PKU patients.

show abstract

Section: Resultssupporting

confidence: 75%

Section: Recombinant Aav Construction and Productionmentioning

confidence: 99%

Administration-route and gender-independent long-term therapeutic correction of phenylketonuria (PKU) in a mouse model by recombinant adeno-associated virus 8 pseudotyped vector-mediated gene transfer

2005

View full text Add to dashboard Cite

show abstract

“…More successful was the chemical mutagenesis of a BTBR-mouse strain using the alkylating agent N-ethyl-N-nitrososurea (ENU), resulting in the isolation of a hyperphenylalaninemic mouse with a mutation called Pah enu2 [22]. The Pah enu2 allele was genetically mapped to the Pah locus, and sequence analysis revealed a misssense mutation at exon 7, a region that encodes the active site of the PAH enzyme, which is by far the most frequent mutation site for PKU in human [23]. Besides hyperphenylalaninemia, this mouse displays many symptoms found in human PKU patients, including slow growth, small head, hypopigmentation, behavior disturbances, and maternal PKU [22,24], and numerous studies from genetics and biochemistry have demonstrated the reliability of this animal model for human PKU [25].…”

Section: Pku Animal Modelsmentioning

confidence: 99%

State-of-the-art 2003 on PKU gene therapy

Ding

Harding

Thöny

2004

Molecular Genetics and Metabolism

View full text Add to dashboard Cite

Phenylketonuria (or PKU) is a well-known and widespread genetic disease for which many countries perform newborn screening, and life-long dietary restriction is still the ultimate and effective therapy. However, the diet is complicated, unpalatable, and expensive. The long-term effects of diet discontinuation in adults, except for the serious adverse effects of maternal hyperphenylalaninemia upon the developing fetus, have not been systematically studied, but congnitive decline and neurologic abnormalities have been anecdotally reported. Thus, alternative approaches for PKU therapy, including gene therapy, must be further explored. Here we summarize past present nonviral and viral gene transfer approaches, both in vitro studies and preclinical animal trials, to delivering the PAH gene into liver or other organs as potential alternatives to life-long phenylalanine-restricted dietary theraphy.

show abstract

“…15 The resulting MCKPah transgenic/Pah enu2 heterozygous progeny were then bred to Pah enu2 /Pah enu2 mice. In this second generation, 10 mice (designation Tg/Pah enu2 ) out of 52 progeny were both heterozygous for the MCK␤gmPAH transgene and homozygous for the T788C Pah enu2 mutation as verified by mutation-specific PCR analysis.…”

Section: Effect Of Muscle Pah Expression Upon Hyperphenylalaninemic Micementioning

confidence: 99%

Metabolic engineering as therapy for inborn errors of metabolism – development of mice with phenylalanine hydroxylase expression in muscle

Harding¹,

Wild²,

Chang³

et al. 1998

Gene Ther

View full text Add to dashboard Cite

Treatment of many inherited liver enzyme deficiencies muscle. These mice exhibited hyperphenylalaninemia at requires the removal of toxic intermediate metabolites frombaseline, but serum phenylalanine levels decreased sigthe blood of affected individuals. We propose that circulatnificantly when the mice were supplemented with tetrahyding toxins can be adequately cleared and disease phenorobiopterin (BH 4 ), a required cofactor for PAH. This is the type influenced by enzyme expressed in tissues other than first demonstration that a liver-specific enzyme, when the liver. Phenylalanine hydroxylase (PAH) activity was expressed in a heterologous tissue and supplied with constitutively expressed in skeletal and cardiac muscle of necessary cofactors, can effectively clear toxic metabolites transgenic mice which carried the PAH cDNA under the from the circulation of individuals with inherited enzyme transcriptional control of the mouse muscle creatine kinase deficiency. This result suggests that gene therapy targeted promoter. Muscle PAH-expressing mice were bred to liver to heterologous tissues, such as muscle, will be effective PAH-deficient, hyperphenylalaninemic mice to yield proin the treatment of selected inborn errors of metabolism. geny that lack PAH activity in liver but express PAH in

show abstract

Characterization of Mutations at the Mouse Phenylalanine Hydroxylase Locus

Cited by 80 publications

References 22 publications

Administration-route and gender-independent long-term therapeutic correction of phenylketonuria (PKU) in a mouse model by recombinant adeno-associated virus 8 pseudotyped vector-mediated gene transfer

Administration-route and gender-independent long-term therapeutic correction of phenylketonuria (PKU) in a mouse model by recombinant adeno-associated virus 8 pseudotyped vector-mediated gene transfer

State-of-the-art 2003 on PKU gene therapy

Metabolic engineering as therapy for inborn errors of metabolism – development of mice with phenylalanine hydroxylase expression in muscle

Contact Info

Product

Resources

About