This resulted in sustained supraphysiologic FIX activity (400%), correction of the bleeding diathesis at clinically relevant, low vector doses (5 × 10(10) vector genomes [vg]/kg) that are considered safe in patients undergoing gene therapy.
Correction of the hemophilic coagulopathy by sustained expression of FIX, reduction of bleeding events, and a comprehensive assessment of the humoral and cell-mediated immune responses to the expressed transgene and recombinant AAV vector are all feasible end points in these dogs.
Adeno-associated viral (AAV) gene transfer of coagulation factor IX to skeletal muscle and liver of murine and canine models of hemophilia has resulted in sustained systemic expression and, in several studies, in complete cure of the bleeding disorder.
Intraperitoneal delivery of AAV8/ Factor IX (hF.IX) during weeks 1-4 of life, over a 20-fold dose range, directed stable hF.IX expression, correction of coagulopathy in F.IX-null hemophilia B mice, and induction of tolerance to hF.IX; however, only primary injection at 1-2 days of life enabled increasing AAV8-mediated hF.IX expression after re-administration, due to the absence of anti-viral capsid antibodies.
It is unclear why this kindred does not exhibit a bleeding tendency but it may correlate with a FXI-like antigen and factor IX binding activity expressed on platelets.
At the time of testing, patient HB530 was a 17-year-old post-puberty male with a persistent, clinically severe bleeding disorder and markedly reduced plasma procoagulant factor IX activity (< 1%).
Although patients with severe hemophilia (i.e., with FVIII:C and FIX:C levels <1IU/dL) are generally those with the most severe bleeding phenotype, it is common experience that a variable proportion of them experiences a milder bleeding tendency.
In further studies using the Chapel Hill strain of hemophilia B dogs, we demonstrate for the first time FIX expression and partial correction of the bleeding disorder following i.v. administration of an AAV vector.
In hemophilic dogs, a dose of rAAV that was approximately 1/10 per body weight that given to mice resulted in 1% of normal canine factor IX levels, the absence of inhibitors, and a sustained partial correction of the coagulation defect for at least 8 months.
The observed bleeding tendency is related to this--compared to the other vitamin K dependent factors (FII, FVII, FX)--excessively and disproportionately low level of FIX.