Haematologica 2000;85:supplement to no. 10 - Supplements ...

Haematologica 2000; 85(suppl. to n.10)
p. 103-107
Immunetolerance:
Molecular and biological aspects
Animal models to explore mechanisms of tolerance induction to
FVIII: SCID mice and SCID-FVIII-KO mice
JEAN GUY GILLES, BEATRIJS VANZIELEGHEM, JEAN-MARIE SAINT-REMY
Center for Molecular and Vascular Biology, Katholieke Universiteit Leuven, Leuven, Belgium
Type A hemophilia is an inborn disease of
coagulation. It is characterized by a deficiency
or dysfunction of factor VIII (FVIII)
and affects 1 in 10,000 males. The clinical manifestations
of the disease include bleeding in
muscles, joints, and soft tissues, and are optimally
treated by infusions of FVIII. However, such
treatment results in the development of antibodies
towards FVIII (usually described as
inhibitors), in at least 15 to 25% of the cases. 1,2
Autoantibodies are also described in some
patients with an annual incidence of 1/5 million
people and remain the most common cause of
antibody-mediated inhibition of the coagulation
system. 3 The occurrence of such anti-FVIII antibodies
is a severe, life-threatening complication
which practically precludes efficient FVIII replacement
therapy. Present treatments, including regular
infusion of high doses of FVIII, 4 administration
of corticosteroids and cyclophosphamide
alone or in combination with infusions of large
doses of FVIII concentrates, 5 IgG immunoadsorption
or infusions of IV Ig, 6 are of limited efficiency
and extremely expensive. 7
Alternative methods of specific treatment are
required. Therefore, efforts must be devoted to
improving our understanding of reasons as to
why anti-FVIII antibodies are formed and the
mechanisms of production of such antibodies.
An animal model that can be used to reconstitute
and analyze a human immune response
towards FVIII was considered as optimal. Severe
combined immunodeficient mice (SCID mice)
or variants of this strain have been chosen for
this purpose. The repopulation of these animals
with immunocompetent cells of a hemophilia A
patient or of healthy donors enables us to
reconstitute a human immune response.
Such a model should enable us to study factors
governing FVIII immunogenicity, the physiology
of the anti-FVIII immune response, namely
the molecular mechanisms leading to the
development and maintenance of anti-FVIII
Correspondence: Jean Guy Gilles, PhD, Center for Molecular and
Vascular Biology, Katholieke Universiteit Leuven, Campus Gasthuisberg,
O&N, Herestraat 49, B-3000 Leuven, Belgium. Phone: international
+32-16-346018; Fax: international+32-16-345990; E-mail:
jeanguy.gilles@med.kuleuven.ac.be
antibodies, and finally to explore the potentialities
of new forms of immunotherapy.
These studies should yield benefits not only
for hemophilia A patients, but could also result
in a significant reduction in the cost related to
the treatment of patients with anti-FVIII antibodies.
SCID mouse model
SCID mice exhibit an immunodeficiency
owing to a defect in their mature B-(no surface
Ig) and T-(no functional TCR) lymphocytes. 8
Detailed analysis has disclosed an aberration of
V(D)J recombination in such cells and a high
sensitivity to ionizing radiation. The SCID-related
gene codes for a DNA-dependent protein
kinase catalytic subunit and resides in the centromeric
region of chromosome 16. 9 Because
of their severe immune deficiency, such mice are
able to accept xenotransplants and can therefore
be reconstituted with peripheral blood
mononuclear cells (PBMC) isolated from hemophilia
A patients or healthy donors. 10
SCID mice and hemophilia A patients
Some hemophilia A patients produce anti-FVIII
antibodies upon FVIII infusion. The strength of
the immune response is unpredictable, varying
from one patient to another, but is also related to
the immunogenicity of the FVIII concentrates.
There is no current method to evaluate these critical
parameters prior to the first FVIII infusion.
The SCID mice model was evaluated to this end.
We reconstituted mice with immunocompetent
cells from two hemophilia A patients with a stable,
high level of FVIII inhibitors.
Mice were then immunized with two different
FVIII preparations, a recombinant or a plasmaderived
FVIII. All mice produced a significant
amount of human IgG antibodies and the presence
of specific anti-FVIII antibodies was detected
in each serum sample by using a direct binding
ELISA. Mice injected with the FVIII preparation
produced at last 4-fold more specific antibodies,
including inhibitors, than control animals.
The level of total IgG produced by each
group of mice depended on cell origin, but also
on the FVIII concentrate used (Table 1). These
Haematologica vol. 85(supplement to n. 10):October 2000