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In a canine study, a vector dose of 2·4 × 10 plaque forming units (pfu) infused into three haemophilia B dogs resulted in transient expression of Factor IX at supraphysiological levels.
As an example, a vector dose of 4×10 VG/kg yielded plasma factor IX protein levels of 9 10% resulting in 55 65% activity in the human factor IX activity assay (Fig. 2A and B).
Haemophilia B canines treated with a vector dose of 1 2 × 10 vg/kg responded with expression of circulating canine Factor IX between 220 ng/ml and 590 ng/ml (4 12%) (Mount et al, 2002).
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Adult mice were treated by subretinal delivery of a vector dose corresponding to 150 ng of p24 protein.
Non-human primates infused through the portal vein with an AAV vector dose of 4 × 10 vg/kg achieved human Factor IX levels of greater than 400 ng/ml (8%) which were sustained for more than 1 year (Nathwani et al, 2002).
Multiple independent adeno-associated virus (AAV) gene therapy clinical trials for hemophilia B, utilizing different AAV serotypes, have reported a vector dose-dependent loss of circulating factor IX (FIX) protein associated with capsid-specific CD8+ T cell (Cap-CD8) elimination of transduced hepatocytes.
IκBα reduced E. coli-induced histologic evidence of lung injury, with the intermediate (1 × 10) vector dose of IκBα again most effective.
We performed subretinal injections of 1 × 10 GC of AAV2/8-RHOK-ZF-R6 in three independent 1-month-old P347S+/− littermates (P30; for a total of 17 animals, 34 eyes: 17 RHOK-ZF-R6 and 17 RHOK-EGFP eyes), with the paired eye of the same animals receiving the same vector dose of AAV2/8-RHOK-EGFP.
To test this, we injected subretinally WT mice with AAV2/8-CMV-EGFP, AAV2/8-CMV-DsRed or a mixture of both vectors (dose of each vector/eye: 3 × 10 GC) and harvested the eyes transduced by single or both vectors 3 weeks post-injection.
To determine the co-transduction efficiency specifically in PR, and to test another red fluorescent reporter possibly more potent than DsRed, we repeated the injection in mice with a mixture of AAV2/8-CMV-EGFP and-RFP vectors (dose of each vector/eye: 1.4 × 10 GC/eye) and analyzed co-transduction on retinal cryosections which allow to unequivocably identify PR from RPE (supplementary Fig S2).
C57BL/6 mice were initially injected subretinally with AAV OZ and dual AAV TS-L and hybrid AK-L vectors (dose of each vector/eye: 1.7 × 10 GC), all encoding EGFP under the transcriptional control of the CMV promoter.
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