3.4. Emerging Advanced Therapeutic Medicinal Products: Gene
therapy for haemophilia
Gene replacement therapies represent a frequently submitted category of
therapeutic for orphan designation [19]. In this form of ATMPs, a
functional copy of a defective gene in the target condition, which is
either absent or expressed as a non-functional protein is substituted by
a fully functional gene which is inserted using a viral vector thereby
offering a highly effective means for overcoming diseases, such as
haemophilia. The gene correction in haemophilia is well suited because
it is associated with a well understood mutated gene sequence which can
be substituted by a corrected DNA sequence leading to the endogenous
production of a functional factor VIII or IX [5]. This would
therefore alleviate the need for exogenous sources of the defective
clotting factors.
An important characteristic of haemophilia treatment is that there is no
need to normalize circulating clotting factor levels to obtain a
therapeutic effect since a slight increase in plasma clotting factor
levels (above 1%) is sufficient to decrease the risk of morbidity and
mortality. As a consequence, a small increase in clotting factor levels
by gene therapy can greatly improve the clinical symptoms. In addition,
therapeutic efficacy of GTMPs can be determined by clear-cut clinical
endpoints such as circulating clotting factor levels and bleeding
frequency [20]. Thus, gene therapy could provide a continuous source
of clotting factor from a single treatment.
Initial clinical gene therapy studies using integrating retroviral,
adenoviral, and non-viral (ex vivo ) approaches were associated
with transient low-level factor expression [21-22].
This led to a shift toward usage of ‘non-integrating’ recombinant
adeno-associated viral (AAV) vectors. AAV vectors are derived from
wild-type AAV [23], a member of the parvovirus family. Wild-type AAV
is non-pathogenic, weakly immunogenic, and replication-deficient,
requiring a helper virus for replication. AAV vectors can deliver a
therapeutic transgene cassette up to 5 kb into both dividing and
nondividing cells. The DNA sequences carried by recombinant AAV vectors
are stabilised predominantly in an episomal form so that long-term
expression can occur only with delivery into long-lived, post mitotic
cell types; the vector DNA integrates at a very low frequency and is
typically lost from replicating cells. The recombinant vector has
tropism for a range of target tissues including the liver, cell types in
the retina and the central nervous system, skeletal muscle, and cardiac
muscle, among others. AAV vectors are the most frequently used viral
vectors for gene therapy for Haemophilia (90%), followed by lentiviral
vectors (10%).
Table 4 provides a detailed list of GTMPs for Haemophilia, which were
granted an orphan designation by European Commission.
FVIII and FIX synthesis primarily takes place in the liver, from where
the proteins can easily enter the bloodstream. This makes hepatocytes a
suitable target for gene therapy in haemophilia.
Over the years, hepatic in vivo gene transfer using adeno-associated
viral (AAV) vectors has shown the best success in preclinical and
clinical studies, with several clinical studies for both hemophilia A
and B enrolling patients for phase 3 trials. We analyzed the studies
present in the Clinical trial.gov and in the EU Clinical Trials Register
repositories [24-25].
Many AAV vectors have progressed into clinical studies for both
haemophilia A and B. The ClinicalTrials.gov database presently lists 29
active gene therapy clinical trials for haemophilia, 12 clinical trials
to evaluate different AAV-based GTMPs for haemophilia B and 11 trials to
evaluate AAV-FVIII gene therapy for haemophilia A (Table 5), with some
overlap, as the same vector is being evaluated in both phase 1/2 and
phase 3 studies.
Of the ongoing Haemophilia A and B AAV clinical trials, there are
remarkable successes. The trial sponsored by St. Jude Children’s
Research Hospital haemophilia B patients, using an scAAV2/8-LP1-hFIXco
vector, was the first long-term success story for haemophilia gene
therapy [26].
However, an important limitation to successful AAV-base gene therapy is
the pre-existing humoral capsid immunity. Immune recognition by
cytotoxic CD8+ T cells or antibody responses to the
vector capsid, the transgene product, or both can compromise the
therapeutic expression of the transgene. Wild-type AAV infection occurs
during childhood, and thus patients may develop neutralising antibiotics
(nAb) that prevent gene transfer with AAV vectors. Cross-reactivity with
multiple AAV serotypes has also been reported. As a result, many
patients have been excluded from recent studies. The main toxicity
observed in AAV-based clinical studies has been dose-related elevation
of liver transaminase following vector infusion. Within some studies,
this coincides with the demonstration of cell-mediated AAV capsid
immunity. Most studies have used either early intervention or
prophylaxis with corticosteroids in order to protect transduced
hepatocytes. Although the majority of episodes have been managed
effectively with this approach, some episodes have been associated with
partial or complete loss of transgene expression, despite intervention
[26].
Due to the pre-existing AAV humoral immunity and the potential loss of
vector transduction with time, there is for the need of other viral and
non-viral-based vectors for transgene delivery. Lentiviral vectors may
complement the therapeutic reach of AAV vectors due to lower incidence
of pre-existing humoral immunity and greater packaging capacity.
Pre-clinical studies using in-vivo or ex-vivo stem cell
(haematopoietic or induced pluripotent) transduction or blood outgrowth
endothelial cells have been recently reviewed [27-28]. Two phase 1
studies are registered with plans to evaluate ex-vivo lentiviral
stem cell transduction for FVIII (NCT03818763) or FVIII/FIX
(NCT03217032/NCT03961243).