Gene Therapy for Haemophilia A.

Haemophilia A is an x-linked, life threatening bleeding disorder arising from defects in the
coagulation factor VIII (FVIII) gene. Current treatment for haemophilia A, the commonest
inherited bleeding disorder (prevalence of 1 in 5000 individuals) consists of life-long,
2-3X/week, intravenous injection of clotting factor concentrates, which is demanding,
exceedingly expensive not widely available nor curative. In contrast, gene therapy offers the
potential of a cure for haemophilia A as illustrated by our first unequivocal success in a
related condition, haemophilia B. In that study the investigators showed that a single
intravenous administration of a serotype 8 based adeno-associated virus, (AAV8) vector
encoding the factor IX (FIX) gene resulted in stable (>6 years) therapeutic expression of FIX
without long-lasting toxicity. The investigators plan to use the same AAV8 platform to
evaluate a novel FVIII expression cassette, AAV2/8-HLP-FVIII-V3, in patient with haemophilia
A. Extensive preclinical studies demonstrate that AAV2/8-HLP-FVIII-V3 leads to long-term,
endogenous expression of FVIII in mouse and non-human primate models without toxicity even
when fifty-fold higher doses than the proposed starting clinical trial dose were used.
Therefore, an open label, Phase I/II dose escalation study entailing a single systemic
administration of AAV2/8-HLP-FVIII-V3 in adults (>18 years of age) with severe haemophilia A
who have baseline factor FVIII levels of <1% of normal has been designed to establish safety
and efficacy of our approach. Dosing will begin at 6x10^11 vector genome (vg)/kg progressing
sequentially to 2x10^12vg/kg and ultimately 6x10^12vg/kg in the absence of toxicity. A
minimum of 2 patients will be recruited at each dose with a possibility of expanding the dose
cohort to a maximum of 6 patients based on safety and efficacy. The study duration for each
patient will be 15 years after vector infusion.

Inclusion Criteria:

- Adult males, aged 18 years or over, with a confirmed diagnosis of severe Haemophilia A
defined as baseline plasma FVIII levels of <1% of normal as assessed by a validated
one-stage clotting assay or a chromogenic assay, resulting from intron 22 inversions,
intron 1 inversions, splice-site mutations, small deletions/insertions and missense
mutations.

- A severe bleeding phenotype as defined by at least one of the following:

1. On prophylaxis for a history of bleeding, or

2. On demand therapy with a current or past history of 4 or more bleeding
episodes/year, or

3. Evidence of chronic haemophilic arthropathy (pain, joint destruction, and loss of
range of motion)

- Received treatment with human FVIII concentrates with at least >50 exposure days;

- Able to give full informed consent and able to comply with all requirements of the
trial including 15-year long-term follow-up;

- Willing to practice barrier contraception after vector administration until at least
three consecutive semen samples are negative for vector sequences (may be for 2-3
months).

Exclusion Criteria:

- Presence of neutralising anti-hFVIII antibodies (inhibitor, determined by the Bethesda
inhibitor assay) at the time of enrolment or a previous history of hFVIII inhibitor;

- Severe haemophilia A patients with large deletions (multiple exons) and nonsense
mutations of the F8 gene.

- Use of investigational therapy for haemophilia within 30 days before enrolment;

- Subjects with active hepatitis B or C, and HBsAg or HCV RNA viral load positivity,
respectively or currently on antiviral therapy for hepatitis B or C. (Negative viral
assays in two samples, collected at least six months apart, will be required to be
considered negative. Both natural clearers and those who have cleared HCV on antiviral
therapy are eligible).

- Serological evidence of HIV-1 who have CD4 counts ≤ 200/mm3. Subjects who are
HIV-positive and stable, with an adequate CD4 count (> 200/mm3) and undetectable viral
load measured twice in the six months prior to enrolment, on an antiretroviral drug
regimen are eligible to enrol.

- Evidence of liver dysfunction (persistently elevated ALT >1.5X upper limit of normal);

- Uncontrolled glaucoma, diabetes mellitus, or hypertension (systolic BP consistently
≥140 mmHg or diastolic BP consistently ≥90 mmHg);

- History of malignancy;

- Suspicious Lung lesions on CT scan that raise the possibility of cancer or
premalignant pathology

- Presence of liver abnormality that is suspicious of malignancy on screening liver
ultrasound

- Patients with uncontrolled cardiac failure or unstable angina;

- Detectable neutralising anti-AAV8 antibodies

- Received an AAV vector, or any other gene transfer agent in the previous 6 months

- History of active tuberculosis, fungal disease or other chronic infection

- Subjects who are unwilling to provide the required semen samples XVI. Poor performance
status (WHO score >1) XVII. Previous history or family history of venous or arterial
thromboembolism