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Haemophilia gene therapy: progress and obstacles

Presented by
Prof. Amit Nathwani, University College London, United Kingdom
ISTH 2020
Review study
The future of gene therapy for haemophilia patients looks bright. Prof. Amit Nathwani (University College London, United Kingdom) presented a review of promising gene therapy trial results in haemophilia B and A [1].

Among vectors of viral origin, and as far as is known to date, adeno-associated virus (AAV) vectors have the best safety profile. “AAV vectors can integrate into the host DNA but at frequencies that are quite low,” Prof. Nathwani explained. However, this integration has only been shown in animal studies until today. A single administration of AAV vectors leads to stable long-term expression of transgenic protein.

Haemophilia B: fewer factor concentrates necessary

AAV comes in a variety of different serotypes (e.g. AAV2, AAV5, AAV8, AAV9) and selection of the most suitable capsid is required. For example, Prof. Nathwani and colleagues selected an AAV8-based capsid for their gene therapy programme in the B-AMAZE study, as an initial seroprevalence study showed that there was a lower immune response against AAV8 in comparison with AAV2.

In the B-AMAZE trial, the investigators developed a unique liver-specific factor IX (FIX) expression cassette, small enough to accommodate the self-complementary genome (see Figure). The vector makes its way to the liver by taking advantage of the strong tropism the capsid for this organ, thereby ensuring that transgenic FIX is synthesised at the natural site for FIX synthesis, leading to appropriate glycosylation of the transgenic protein.

Figure: FIX-expressing AAV vector used in the phase 1/2 haemophilia B gene therapy trial B-AMAZE. Adapted from [1]

Prof. Nathwani and his team introduced this FIX-expressing vector in 10 patients with haemophilia B. Patients revealed significant changes in bleeding phenotypes, shifting from severe to mild, and a dose-dependent increase in FIX levels, which is still stable after 10 years. "In all 10 patients, we observed an 87% drop in bleeding rates. Additionally, we noticed a 66% reduction in FIX concentrate usage,” he said. No long-term toxicities or inhibitor formation were observed. The only AAV-related adverse event was an asymptomatic rise in ALT levels, which typically took place between 2 to 26 weeks after gene therapy. The investigators observed transaminitis in 4 of the 6 patients treated with high dosages but not in any patient treated at the low-dose levels. This complication has been reported with all AAV serotypes that have been administered systemically, regardless of the target disease.

Haemophilia A: a paradigm shift?

With the results of the B-AMAZE study in mind, Prof. Nathwani questioned if the same AAV approach can be used for the more complicated and more common haemophilia A. He explained that gene therapy programmes started in haemophilia B and not in haemophilia A because factor VIII (FVIII) cDNA is large, difficult to package, and expression of the FVIII gene leads to poor expression profiles when compared with other proteins of similar size (e.g. FVII). For successful gene therapy, researchers had to overcome these obstacles. They succeeded in creating small codon optimised FVIII expression cassettes in which the B domain -which is not required for FVIII/cofactor activity- is deleted. "By taking out the B-domain, we remained just within the packaging limits of AAV vectors,” Prof. Nathwani explained.

In mice, codon optimisation of FVIII cDNA resulted in a 10-fold higher expression [2]. In patients with severe haemophilia A, FVIII levels rose to a peak of 64 IU/dL 1 year after administration of a single dose of a codon-optimised recombinant (r)AAV5 capsid pseudotyped vector made using baculovirus/SF9 cells in 2 doses (i.e. 6×1013 vg/kg and 4×1013 vg/kg) [3,4]. A 96% reduction in bleeding rates was observed in the 6×1013 vg/kg dose group, the other dose showed similar efficacy with 92% reduction. Many of the patients did not bleed at all, following a single administration of the rAAV5 vectors. None of the patients developed neutralising antibodies to FVIII and the only adverse event was asymptomatic grade 1 transaminitis. FVIII gene therapy might be licensed towards the end of this year for patients with severe haemophilia A.

Prof. Nathwani concluded that “gene therapy can be life-changing,” and that AAV-mediated gene therapy has the potential to shift the paradigm in the treatment for haemophilia patients.

  1. Nathwani A. SOA01.01, ISTH 2020 Virtual Congress, 12-14 July 2020.
  2. Mcintosh J et al. Blood 2013;121:3335-3344.
  3. Pasi KJ, et al. Abstract LB 01.2. ISTH 2019 Congress, 6-10 July 2019; Melbourne, Australia.
  4. Pasi KJ, et al. New Engl J Med 2020;382:29-40.


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