Factors influencing in vivo transduction by recombinant adeno-associated viral vectors expressing the human factor IX cDNA

Abstract

Long-term expression of coagulation factor IX (FIX) has been observed in murine and canine models following administration of recombinant adeno-associated viral (rAAV) vectors into either the portal vein or muscle. These studies were designed to evaluate factors that influence rAAV-mediated FIX expression.

Major role of local immune responses in antibody formation to factor IX in AAV gene transfer

“…12 In this study, we used hF.IX as the transgene product expressed from an AAV vector. We know from previous studies that mice are capable of forming T-and B-cell responses if this neo-antigen is introduced by gene transfer.…”

“…injection of vector in animals that lacked tolerance to the F.IX transgene product such as mice with F.IX gene deletion, dogs with F.IX nonsense mutation, or animals that received transfer of a F.IX transgene that was not species-specific. 8,10,12,14 Use of a muscle-specific promoter did not prevent antibody formation to F.IX. 15 Anti-F.IX formation upon muscle gene delivery is due to an adaptive, CD4 + T-celldependent immune response which is absent in CD4-or Rag1-deficient mice.…”

“…6 However, several studies demonstrated formation of antibodies to the F.IX transgene product after AAV-mediated gene transfer. [7][8][9][10][11][12] The risk for such immune responses depended on the underlying F.IX mutation, route of vector administration, vector dose and serotype. 13 Inhibitor formation to F.IX was most frequently observed after intramuscular (i.m.)…”

In utero administration of Ad5 and AAV pseudotypes to the fetal brain leads to efficient, widespread and long-term gene expression

“…45 The self-complimentary AAV2/5-GFP and AAV2/8-GFP were produced using the methods previously described by Nathwani et al 46 Vector genomes were calculated using the previously described slot-blot analysis with supercoiled plasmid DNA as standards. 47 The University of Pennsylvania Vector Core Facility supplied the self-complimentary AAV2/ 9-GFP virus and the details of vector production, titration, and quality control can be found on their website (http://www.med.upenn.edu/gtp/ vectorcore/). The titres of the viral preparations used in the study were: Ad5 ¼ 2 Â 10 11 viral genomes per ml, and AAV2/5, AAV2/8 and AAV2/ 9 ¼ 7 Â 10 11 viral genomes per ml.…”

Gene transfer vector biodistribution: pivotal safety studies in clinical gene therapy development

“…Distribution for this latter transgene was studied by PCR after injection via tail or portal veins of 10 11 or 5 Â 10 10 vg. 37 After tail vein injection, vector DNA was detected in the liver and spleen only. After portal vein injection, the liver and spleen were also positive, but the latter at the highest dose only.…”