AAV8-mediated gene therapy prevents induced biochemical attacks of acute intermittent porphyria and improves neuromotor function - PubMed (original) (raw)

AAV8-mediated gene therapy prevents induced biochemical attacks of acute intermittent porphyria and improves neuromotor function

Makiko Yasuda et al. Mol Ther. 2010 Jan.

Abstract

Acute intermittent porphyria (AIP), an autosomal dominant hepatic porphyria due to half-normal hydroxymethylbilane synthase (HMB-synthase) activity, is manifested by life-threatening acute neurological attacks that are precipitated by factors that induce heme biosynthesis. The acute attacks are currently treated with intravenous hemin, but a more continuous therapy is needed, particularly for patients experiencing frequent attacks. Thus, a recombinant AAV8-based serotype vector expressing murine HMB-synthase driven by liver-specific regulatory elements was generated and its effectiveness to prevent the biochemical induction of an acute attack was evaluated in an AIP mouse model. Intraperitoneal administration of the adeno-associated viral (AAV) vector resulted in a rapid and dose-dependent increase of HMB-synthase activity that was restricted to the liver. Stable expression of hepatic HMB-synthase was achieved and wild-type or greater levels were sustained for 36 weeks. When heme synthesis was periodically induced by a series of phenobarbital injections, the treated mice did not accumulate urinary delta-aminolevulinic acid (ALA) or porphobilinogen (PBG), indicating that the expressed enzyme was functional in vivo and prevented induction of the acute attack. Further, rotarod performance and footprint analyses improved significantly. Thus, liver-directed gene therapy provided successful long-term correction of the hepatic metabolic abnormalities and improved neuromotor function in the murine model of human AIP.

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Figures

**Figure 1

Dose-response of hepatic HMB-synthase expression. AIP mice were administered 3.8 × 1011, 5.7 × 1011, or 7.6 × 1011 drp of rAAV2/8-HMBS and the hepatic HMB-synthase activities were determined 1 week later. Black bars represent HMB-synthase activities detected in the AIP (T1/T2) mice, whereas the white bar represents that of saline-treated wild-type (wt/wt) mice. Data are presented as mean + SD (n = 4). AIP, acute intermittent porphyria.

**Figure 2

Tissue specificity of rAAV2/8-HMBS-mediated HMB-synthase expression. Various tissues from saline- and rAAV2/8-HMBS-treated AIP mice were assayed for HMB-synthase activity (right) and vector copy number (left). The data presented are means + SD (n = 4). Note that vector copy number is shown on a logarithmic scale. AIP, acute intermittent porphyria.

**Figure 3

Time course of rAAV2/8-HMBS-mediated hepatic HMB-synthase expression. AIP mice were sacrificed 1, 2, 6, 12, 24, and 36 weeks after vector administration and the hepatic HMB-synthase activities were determined. Results are shown as mean ± SD. Of note, the HMB-synthase activity at week 0 represents baseline activity in the AIP mice. The area between the broken lines represents the range of hepatic HMB-synthase activity detected in wild-type mice (n = 5). AIP, acute intermittent porphyria.

**Figure 4

Urinary porphyrin precursor levels following phenobarbital injections. After collection of two baseline urine samples (days 1 and 2), increasing doses of phenobarbital (110, 120, 125, 130 mg/kg/day) were administered to wild-type (triangles) and saline- (squares) and rAAV2/8-HMBS-treated AIP mice (circles) for four consecutive days, as indicated by the arrows, and the (a) ALA and (b) PBG levels were determined in 24 hour urines. Results from a representative phenobarbital induction performed 22 weeks after vector administration are shown. For each time-point, the means and standard deviations are presented (n = 5). AIP, acute intermittent porphyria; ALA, δ-aminolevulinic acid; PBG, porphobilinogen.

**Figure 5

Effect of rAAV2/8-HMBS therapy on neuromotor function. (a) Seven month old male wild-type (wt/wt) and saline- and rAAV2/8-HMBS-treated AIP (T1/T2) mice were evaluated for their performance on a rotarod rotating at 16 rpm, for a maximal time of 180 seconds. The results are expressed as means and standard deviations (n = 5). (b) Gait patterns were analyzed in nine month old mice by footprint analysis. Representative footprints of wild-type and saline- and rAAV2/8-HMBS-treated AIP mice are shown. LSLs were measured and statistically evaluated as described in the “Materials and Methods” section (n = 4–6). AIP, acute intermittent porphyria; LSL, left stride length.

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AAV8-mediated gene therapy prevents induced biochemical attacks of acute intermittent porphyria and improves neuromotor function - PubMed (original) (raw)