897. Recombinant AAV2/8 Vector-Mediated Gene Therapy Corrects Hyperphenylalaninemia in Murine PKU

Abstract

Top of pageAbstract Novel recombinant adeno-associated virus vectors pseudotyped with serotype 8 capsid (rAAV2/8) have recently shown exciting promise as effective liver-directed gene transfer reagents. We have designed and produced an rAAV2/8 vector containing the mouse phenylalanine hydroxylase (PAH) cDNA under the transcriptional control of a strong liver promoter (LSP) and have administered this vector to hyperphenylalaninemic Pahenu2 mice, a model of human PKU. Our hypothesis was that this vector would produce sufficient hepatocyte transduction frequency and PAH activity to correct blood phenylalanine (Phe) levels in murine PKU. Recombinant AAV2/8 vector was injected via the portal vein into five adult (four male and one female) hyperphenylalaninemic Pahenu2 homozygous mice (preinjection Phe = 2108 |[plusmn]| 102 |[mu]|M, normal = 145 |[plusmn]| 45 |[mu]|M) and evaluated the effect upon serum Phe levels weekly. Each animal received 2.5 X 1011 rAAV2/8 vector genomes via portal vein injection. A sixth Pahenu2 mouse (female, preinjection Phe = 3579 |[mu]|M) received only saline as a control, and serum Phe levels remained elevated in this animal (4174 |[mu]|M ten weeks following injection). Administration of rAAV2/8 yielded a dramatic and stable reduction in serum Phe to normal levels by 1-2 weeks following injection. At ten weeks following injection, serum Phe levels remained normal (157 |[plusmn]| 32 |[mu]|M) and were significantly reduced in comparison to pretreatment levels (p < 0.001). Phenylalanine clearance measured after parenteral Phe challenge significantly increased following rAAV2/8 treatment. Hyperphenylalaninemia-associated hypopigmentation was also completely corrected. Liver PAH activity was 6-10% of wild type levels in three mice that underwent liver biopsy at 8-10 weeks postinjection. Vector copy number, integration status of the vector genome, biodistribution, and long-term stability of expression will be evaluated. Recombinant AAV2/8-mediated, liver-directed gene therapy is a promising novel treatment approach for PKU and allied inborn errors of metabolism.