title: Factor IX variants improve gene therapy efficacy for hemophilia B.
abstract: Intramuscular injection of adeno-associated viral (AAV) vector to skeletal muscle of humans with hemophilia B is safe, but higher doses are required to achieve therapeutic factor IX (F.IX) levels. The efficacy of this approach is hampered by the retention of F.IX in muscle extracellular spaces and by the limiting capacity of muscle to synthesize fully active F.IX at high expression rates. To overcome these limitations, we constructed AAV vectors encoding F.IX variants for muscle- or liver-directed expression in hemophilia B mice. Circulating F.IX levels following intramuscular injection of AAV-F.IX-K5A/V10K, a variant with low-affinity to extracellular matrix, were 2-5 fold higher compared with wild-type (WT) F.IX, while the protein-specific activities remained similar. Expression of F.IX-R338A generated a protein with 2- or 6-fold higher specific activity than F.IX-WT following vector delivery to skeletal muscle or liver, respectively. F.IX-WT and variant forms provide effective hemostasis in vivo upon challenge by tail-clipping assay. Importantly, intramuscular injection of AAV-F.IX variants did not trigger antibody formation to F.IX in mice tolerant to F.IX-WT. These studies demonstrate that F.IX variants provide a promising strategy to improve the efficacy for a variety of gene-based therapies for hemophilia B.
authors.affiliation: The Children's Hospital of Philadelphia, 34th St and Civic Center Blvd, Philadelphia, PA 19104, USA.
authors.forename: Joerg
authors.initials: J
authors.lastname: Schuettrumpf
authors.forename: Roland W
authors.initials: RW
authors.lastname: Herzog
authors.forename: Alexander
authors.initials: A
authors.lastname: Schlachterman
authors.forename: Antje
authors.initials: A
authors.lastname: Kaufhold
authors.forename: Darrel W
authors.initials: DW
authors.lastname: Stafford
authors.forename: Valder R
authors.initials: VR
authors.lastname: Arruda
mesh_terms: D000256:Adenoviridae; D019943:Amino Acid Substitution; D000818:Animals; D005164:Factor IX / Q000235:genetics*; D015316:Genetic Therapy; D002836:Hemophilia B / Q000235:genetics / Q000628:therapy*; D006487:Hemostasis / Q000235:genetics; D006801:Humans; D008099:Liver / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D018482:Muscle, Skeletal / Q000378:metabolism; D009928:Organ Specificity / Q000235:genetics; D017354:Point Mutation
chemical_list: D005164:Factor IX