J Korean Med Sci.  2008 Oct;23(5):877-883. 10.3346/jkms.2008.23.5.877.

Protective Effect of Recombinant Adeno-Associated Virus 2/8-Mediated Gene Therapy from the Maternal Hyperphenylalaninemia in Offsprings of a Mouse Model of Phenylketonuria

• Jung SC
• Park JW
• Oh HJ
• Choi JO
• Seo KI
• Park ES
• Park HY

• KMID: 1783076
• DOI: http://doi.org/10.3346/jkms.2008.23.5.877

Abstract

Phenylketonuria (PKU) is an autosomal recessively inherited metabolic disorder caused by a deficiency of phenylalanine hydroxylase (PAH). The accumulation of phenylalanine leads to severe mental and psychomotor retardation, and the fetus of an uncontrolled pregnant female patient presents with maternal PKU syndrome. We have reported previously on the cognitive outcome of biochemical and phenotypic reversal of PKU in a mouse model, Pah(enu2), by the AAV serotype 2-mediated gene delivery of a human PAH transgene. However, the therapeutic efficacy had been limited to only male PKU mice. In this study, we generated a pseudotyped recombinant AAV2/8-hPAH vector and infused it into female PKU mice through the hepatic portal vein or tail vein. Two weeks after injection, complete fur color change to black was observed in female PKU, as in males. The PAH activity in the liver increased to 65-70% of the wild-type activity in female PKU mice and to 90% in male PKU mice. Plasma phenylalanine concentration in female PKU mice decreased to the normal value. In addition, the offsprings of the treated female PKU mice can rescue from the harmful effect of maternal hyperphenylalaninemia. These results indicate that recombinant AAV2/8-mediated gene therapy is a potential therapeutic strategy for PKU.

Keyword

Phenylketonurias; Hyperphenylalaninemia; Mice; Dependovirus; Gene Therapy

MeSH Terms

Animals
Cell Line
Dependovirus/*genetics
Female
Gene Therapy/*methods
Gene Transfer Techniques
Humans
Male
Mice
Mice, Transgenic
Phenylketonurias/*genetics/*therapy
Recombinant Proteins/metabolism
Sex Factors
Time Factors

Figure

• 

  Fig. 1 (A) PAH enzyme activity in the livers of Pahenu2 mice six weeks after rAAV-hPAH administration. Enzymatic activity is represented as a percentage of that found in wild-type mice. The dotted bar represents PAH activities in homozygous Pahenu2 mice; the black bars, rAAV2/8-hPAH-injected homozygous female mice; the opened bars, rAAV2/8-hPAH-injected homozygous male mice. (B) Plasma phenylalanine concentration in female Pahenu2 mice after rAAV2/2-hPAH or aAAV2/8-hPAH treatment. The dotted bars represent plasma phenylalanine concentrations in homozygous female Pahenu2 mice 2 weeks after treatment; the black bars, 6 weeks after treatment; the open bars, 6 months after treatment. The dotted line A represents the recommended maximum value by the National Society for PKU (U.K.); dotted line B, plasma concentration in wild-type control mice. Values are presented as the mean±SD. *, p<0.001 vs. untreated Pahenu2 mice (n=3 for each experiment).

• 

  Fig. 2 Fur color change to black in rAAV2/8-hPAH-treated Pahenu2 mice. (A) Female Pahenu2 mice infused with 2×1012 viral particles through the portal vein displayed pigmentation six weeks after injection. (B) Pregnant female Pahenu2 mice administered with 3×1012 viral particles through the tail vein exhibited the fur color change four weeks after injection.

• 

  Fig. 3 Analysis of offspring of homozygous PKU female mice. Mean values of crown-rump length (A), weight (B), and spontaneous abortion rates (C) of the fetuses by maternal status. The dotted bars represent offspring of wild type mothers; the black bars, offspring of untreated homozygous mothers; the opened bars, offspring of rAAV2/8-hPAH-injected homozygous mothers. *p<0.001 vs. wild type and untreated mothers, treated and untreated mothers (n=38 for fetuses from 4 wild type mothers, n=29 for fetuses from 5 untreated mothers, and n=21 for fetuses from 3 treated mothers).

• 

  Fig. 4 (A) Pregnant female Pahenu2 mice administered with 3×1012 viral particles through the tail vein exhibited the fur color change on day 18.5 gestation. Genotyping of fetuses from treated mother was confirmed by sequencing (n=10). (B) Pregnant female Pahenu2 mice on day 18.5 gestation and fetuses from untreated mother. Sequencing was performed for genotyping of fetuses (n=2).

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