01. Gene therapy Boulikas.pdf - Gene therapy & Molecular Biology

Infection of arthritogenic splenocytes from DBA/1
mice transferred to SCID mice with a recombinant
retrovirus carrying the TGF-β1 gene was effective in
lowering inflammation of joints with already established
arthritis and inhibiting the spreading of the disease to other
joints in mice (Chernajovsky et al, 1997).
D. Direct gene delivery for RA
A retroviral vector based on a murine leukemia virus
was used to deliver the human growth hormone and lacZ
genes to the synovium of the rabbit knee to test the
efficacy of gene transfer and to develop an approach for
the gene therapy of RA (Ghivizzani et al, 1997).
An effective treatment of arthritis is via eliminating
most or all of the activated synovial cells. The death factor
Fas/Apo-1 and its ligand (FasL) play pivotal roles in
maintaining self-tolerance and immune privilege; Fas is
expressed constitutively in most tissues and is
dramatically upregulated at the site of inflammation.
Unlike Fas, however, the levels of FasL expressed in the
arthritic joints are extremely low, and most activated
synovial cells survive despite high levels of Fas
expression. Delivery of the FasL gene via a replicationdefective
adenovirus by injection into inflamed joints
conferred high levels of FasL expression, induced
apoptosis of synovial cells, and ameliorated collageninduced
arthritis in DBA/1 mice (Zhang et al, 1997).
A strategy was developed for inhibiting T lymphocyte
retention and activation within the rheumatoid synovium
(Chen et al, 1995). The inflamed synovium in RA is
infiltrated by lymphocytes and monocytes, a process
mediated by the enhanced binding of the very late antigen-
4 (VLA-4) to vascular cell adhesion molecule-1 (VCAM-
1) expressed on microvascular endothelial cells; VLA-4
binding appears to play a role in T cell retention and
activation within the inflamed synovial membrane.
Therefore, blocking of VLA-4 binding by utilizing a
soluble congener of the VCAM-1 molecule might be of
therapeutic efficiency for RA. Adenoviral infection of
human synoviocytes carrying the cDNA for a secreted
form of VCAM-1 (sVCAM-1) showed secretion of
transgenic sVCAM-1 by ELISA of tissue culture
supernatants. In vivo, transgenic sVCAM-1 expression
was determined by immunohistochemical analysis and in
situ hybridization of synovial tissue, and secretion of
transgenic sVCAM-1 was demonstrated by ELISA of tidal
knee lavage fluid. The results showed that recombinant
adenovirus can mediate the expression of a biologically
active sVCAM-1 by synoviocytes in vivo and suggested
that this strategy may be useful for inhibiting T
lymphocyte retention and activation within rheumatoid
synovium. Ex vivo studies using this strategy have not
been reported.
Boulikas: An overview on gene therapy
114
XXXV. Adenosine deaminase (ADA)
deficiency and severe combined
immunodeficiency (SCID)
Severe combined immunodeficiency is secondary to
the deficiency in adenosine deaminase; the enzyme is
involved in purine catabolism. The syndrome is
characterized by defective B and T cell function caused by
the large amounts of deoxyadenosine which is
preferentially converted into the toxic compound
deoxyadenosine triphosphate in T cells disabling the
immune system (see Blaese et al, 1995 and the references
cited therein). Affected individuals experience recurrent
infections and the disease is usually fatal unless affected
children are kept in isolation. One therapeutic approach
has been to partially reconstitute the immune system by
bone marrow transplantation from a human leukocyte
antigen (HLA)-identical sibling donor (Hirschorn et al,
1981). An enzyme replacement therapy consisted of
introducing bovine ADA enzyme conjugated with
polyethylene glycol (PEG-ADA) in order to increase the
circulation time of the ADA enzyme in the blood and
other extracellular fluids (Hershfield et al, 1987).
The first person to be treated ex vivo was a 4-year-old
suffering with ADA deficiency in 1990. Protocol #2
treating ADA deficiency with autologous lymphocytes
transduced ex vivo with the ADA gene was the second
RAC-approved protocol. Because of this innovative work,
the US Patent Office has issued in 1995 a patent covering
all ex vivo gene therapy to French Anderson, Steven
Rosenberg, and Michael Blaese.
From 1990-1992, a clinical trial was initiated using
retrovirus mediated transfer of the 1.5 kb ADA gene
cDNA to T cells from two children with severe combined
immunodeficiency following multiple transplantations of
ex vivo modified blood cells; the integrated ADA gene
was expressed for long periods (Blaese et al, 1995;
Bordignon et al, 1995). The success of this ex vivo
approach probably arose from that the ADA genecorrected
T cells acquired a survival advantage compared
with uncorrected cells when transplanted into
immunodeficient but ADA normal BNX mice (Ferrari et
al, 1991) and humans (Kohn et al, 1995).
Three neonates with ADA deficiency have been
successfully treated later with hematopoietic stem CD34 +
cells, isolated from their umbilical cord blood, transduced
with the ADA gene under control of the LTR of the
MoMuLV using retroviral vectors, followed by autologous
transplantation (Kohn et al, 1995).
Ex vivo studies have shown correction of the severe
combined immunodeficiency in ADA-deficient mice by
transfer of human peripheral blood lymphocytes
transduced with a retroviral vector in cell culture carrying
the ADA gene; the injected human cells survived for long
times in mice and restored the immune functions (presence