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.
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.
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.
The immune response against therapeutic clotting factors VIII and IX (FVIII and FIX) is a major adverse event that can effectively thwart their effectiveness in correcting bleeding disorders.
Approximately 6-39% of the platelets expressed FIX in the transduced recipients, which was sufficient to rescue the bleeding diathesis in FIX(null) mice in tail clipping models.
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.
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.
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.
Different kinds of mutations, mostly point mutations, in the coagulation factor IX (FIX) gene F9 result in a recessive X-linked bleeding disorder known as haemophilia B.
Administration of only 1 x 10(10) scAAV particles led to expression of hFIX at supraphysiologic levels (8I U/mL) and correction of the bleeding diathesis in FIX knock-out mice.
Deficiencies of coagulation factors other than factor VIII and factor IX that cause bleeding disorders are inherited as autosomal recessive traits and are rare, with prevalences in the general population varying between 1 in 500 000 and 1 in 2 million for the homozygous forms.
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.
Deficiencies of coagulation factors (other than factor VIII and factor IX) that cause a bleeding disorder are inherited as autosomal recessive traits and are generally rare, with prevalences in the general population varying between 1 : 500 000 and 1 : 2 000 000.
Furthermore, when a linear human factor IX expression cassette was delivered to factor IX-deficient mice, sustained serum concentrations of more than 4 microg/ml (80% of normal) of the human clotting factor and correction of the bleeding diathesis were obtained.