Abstract Number: PB1087
Meeting: ISTH 2020 Congress
Theme: Hemophilia and Rare Bleeding Disorders » Hemophilia Gene Therapy
Background: Gene therapy using adeno-associated viral (AAV) vectors has demonstrated durable Factor VIII (FVIII) expression in patients and animal models of severe hemophilia A. The mechanisms mediating long-term expression and capsid immune responses are not known.
Aims: To evaluate the site of expression, vector DNA forms and capsid immunogenicity to AAV-FVIII in a hemophilia dog model.
Methods: Hemophilia A dogs received a single canine-FVIII AAV vector infusion (AAV-BDD-cFVIII, AAV2=4, AAV6=3 and AAV8=1). FVIII activity (FVIII:C) was measured using pooled canine plasma as a standard. AAV-BDD-cFVIII vector genomes and BDD-cFVIII mRNA were quantified by qPCR and qRT-PCR. Circular full-length AAV episomes were quantified using droplet-digital PCR. Anti-AAV capsid neutralizing antibody (nAb) titres were measured using a cell-based in-vitro transduction inhibition assay.
Results: Eight dogs received a median dose of 1.25×1013 vg/kg AAV-BDD-cFVIII by portal vein infusion (Table 1). After 10.8 years (8.2-12.0 years), persistent FVIII:C (median one-stage=12.7%, chromogenic=7.2%) was seen in all responding dogs (n=6), with improvement in bleeding phenotype. AAV-BDD-cFVIII DNA was detected in the liver of all dogs (median=30.5 vg/ng), with lower levels in the spleen in 4 dogs (median=1.0 vg/ng). cFVIII mRNA was only detected in the liver of responding dogs (median=7.3 copies/ng). Hepatic cFVIII mRNA copies were significantly associated with FVIII:C. Full-length linear and circular AAV-BDD-cFVIII was detected in the liver of all animals. Anti-AAV capsid nAbs were detected throughout the study: mean earliest available peak post-treatment titre 6485. Most samples demonstrated limited cross-reactivity to other capsids. Although a decline in nAb titre was seen with time (coefficient= -314, p< 0.001), substantial nAbs remained detectable (mean final titre: 809).
Conclusions: Persistent liver-derived FVIII expression was seen 10 years after a single AAV-BDD-cFVIII infusion, with detectable full-length linear and circular AAV-FVIII forms. Although anti-capsid humoral immune responses decrease with time, these remain at levels potentially preventing redosing with the same capsid.
| ID (Sex) | AAV Serotype (dose vg/kg) | Final Chromogenic FVIII:C (years) | Liver BDD-cFVIII vg/ng DNA | Spleen BDD-cFVIII vg/ng DNA | Liver BDD-cFVIII copies/ng mRNA | Spleen BDD-cFVIII copies/ng mRNA | Liver Full-length Circular AAV-FVIII vg/diploid genome | Peak AAV nAb titre (years) | Late AAV nAb titre (years) |
| EL (F) | 2 (6.0e12) | n/a (8.2) | 8.9 | 0.3 | 6.2 | nd | 0.014 | 22068 (3.6) | n/a |
| VC (M) | 2 (1.5e13) | 2.9% (11.0) | 120.4 | nd | 8.3 | nd | 0.096 | 6110 (2.4) | 220 (11.0) |
| ANG (M) | 2 (1.5e13) | <1% (11.4) | 10.7 | 1.0 | nd | nd | 0.007 | 5631 (7.9) | 1445 (11.5) |
| JU (M) | 2 (2.7e13) | 7.2% (11.9) | 50.2 | 1.0 | 15.9 | nd | 0.003 | 5092 (3.2) | 1864 (11.9) |
| ALX (F) | 6 (1.0e13) | 8.6% (10.5) | 36.0 | nd | 26.4 | nd | 0.089 | 4130 (1.5) | 293 (10.5) |
| MZ (M) | 6 (1.0e13) | 7.9% (10.1) | 35.1 | nd | 5.9 | nd | 0.054 | 4805 (1.2) | 634 (10.1) |
| MG (F) | 6 (1.7e13) | <1% (11.3) | 2.8 | nd | nd | nd | 0.002 | 761 (8.0) | 647 (11.5) |
| FLO (F) | 8 (1.0e13) | 1.8% (9.8) | 25.9 | 6.8 | 4.4 | nd | 0.018 | 3280 (1.0) | 559 (10.5) |
[Table 1: FVIII expression, vector genomes and capsid immune response post AAV-FVIII. nd=not detected]
To cite this abstract in AMA style:
Batty P, Sihn CR, Ishida J, Mo AM, Yates B, Brown C, Harpell L, Pender A, Russell C, Sardo Infirri S, Torres R, Winterborn A, Fong S, Lillicrap D. Long-Term Vector Genome Outcomes and Immunogenicity of AAV FVIII Gene Transfer in the Hemophilia A Dog Model [abstract]. Res Pract Thromb Haemost. 2020; 4 (Suppl 1). https://abstracts.isth.org/abstract/long-term-vector-genome-outcomes-and-immunogenicity-of-aav-fviii-gene-transfer-in-the-hemophilia-a-dog-model/. Accessed November 30, 2023.« Back to ISTH 2020 Congress
ISTH Congress Abstracts - https://abstracts.isth.org/abstract/long-term-vector-genome-outcomes-and-immunogenicity-of-aav-fviii-gene-transfer-in-the-hemophilia-a-dog-model/