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

The peptide kinin, after binding to its specific
receptor, triggers a broad spectrum of biological effects
such as vasodilatation, increase in vascular permeability,
smooth muscle contraction and relaxation, and electrolyte
and glucose transport; it plays an important role in
homeostasis of blood pressure, sodium excretion in the
kidney, and inflammatory disorders. It is produced from a
larger oligopeptide precursor, kininogen, after cleavage
by a specific protease called kallikrein. Low levels of this
protease in urine have been associated with hypertension.
Repeated oral administration of swine pancreatic
kallikrein can lower the blood pressure of hypertensive
patients albeit in a temporal manner.
Delivery of a 5.6 kb genomic clone or of a 834-bp
cDNA clone encoding the kallikrein gene under control of
the albumin promoter, CMV, RSV, or metallothionein
promoters into the portal vein or tail vein of spontaneously
hypertensive rats resulted in significant reduction of their
blood pressure for about 5-6 weeks (Chao et al, 1996).
Intravenous injection of an adenoviral vector containing
the human tissue kallikrein gene under the control of a
CMV promoter, into spontaneously hypertensive rats
caused a sustained delay in the increase in blood pressure
from day 2 to day 41 post-injection. The therapeutic effect
was a result of transfection of the human gene into several
rat tissues and human tissue kallikrein mRNA was
detected in the liver, kidney, spleen, adrenal gland, and
aorta (Jin et al, 1997).
Treatment of hypertension with gene therapy has also
been attained by transfer of the human tissue kallikreinbinding
protein (HKBP) or kallistatin, a serine proteinase
inhibitor (serpin). Transgenic mice overexpressing rat
kallikrein-binding protein are hypotensive; kallistatin may
function as a vasodilator in vivo. Delivery of the human
kallistatin cDNA under control of the RSV 3' LTR in an
adenoviral vector into spontaneously hypertensive rats by
portal vein injection resulted in a significant reduction of
blood pressure for 4 weeks; human kallistatin mRNA was
detected in liver, spleen, kidney, aorta, and lung (Chen et
al, 1997).
Blood pressure is also controlled by other factors such
as by the endothelium-derived nitric oxide (NO) in
peripheral vessels. Transfer of the human endothelial NO
synthase (eNOS) gene to spontaneously hypertensive rats
gave a continuous supply of eNOS which caused a
significant reduction of systemic blood pressure for 5 to 6
weeks; the effect continued for up to 10 weeks after a
second injection (Lin et al, 1997).
Angiotensinogen, a substrate for angiotensin I
generation, is mainly produced in the liver, and is a unique
component of the renin-angiotensin system. Mutations in
the angiotensinogen gene are associated with
hypertension. It is unclear whether circulating
angiotensinogen is a rate-limiting step in blood pressure
regulation. Transfer of antisense oligonucleotides against
Gene Therapy and Molecular Biology Vol 1, page 123
123
rat angiotensinogen into the rat liver via the portal vein
diminished the expression of hepatic angiotensinogen
mRNA and resulted in a transient decrease in plasma
angiotensinogen levels in spontaneously hypertensive rats
from day 1 to day 7 after the injection. Liposomes were
used for the transfer of oligonucleotides containing viral
agglutinins to promote fusion with target cells. This
treatment resulted in a decrease in plasma angiotensin II
concentration; transfection of sense and scrambled
oligonucleotides did not show any changes in plasma
angiotensinogen level, blood pressure, or angiotensinogen
mRNA level (Tomita et al, 1995).
The renin-angiotensin system plays an important role
in blood pressure regulation; Phillips and coworkers
(1997) have targeted the renin-angiotensin system at the
level of synthesis (angiotensinogen, AT) and the receptor
(AT1 receptor). Antisense oligonucleotides to AT1receptor
mRNA and to angiotensinogen mRNA reduced
blood pressure. The cDNA for the AT1 receptor was
inserted in the antisense direction under control of CMV
promoter in AAV (which was the system of choice among
adeno, retrovirus, naked DNA and liposomes tested) and
injected either directly in the hypothalamus (1 µL) or in
the lateral ventricles (5 µL). A prolonged decrease in
blood pressure in spontaneously hypertensive rats was
achieved via delivery of antisense DNA for AT1-R
causing a significant reduction in AT1 receptors. After a
single injection there was a significant decrease of blood
pressure (approximately 23 +/- 2 mm Hg) for up to 9
weeks (Phillips, 1997; Phillips et al, 1997).
Blood vessels of spontaneously hypertensive rats were
shown to be associated with sub-physiological amounts of
endothelial basic FGF (bFGF); this decrease correlated
both with hypertension and with a decrease in the
endothelial content of nitric oxide synthase. As a
consequence, transfer of the bFGF gene corrected
hypertension, restored the physiological levels of bFGF in
the vascular wall, significantly enhanced the number of
endothelial cells with positive immunostaining for nitric
oxide synthase, and ameliorated endothelial-dependent
responses to vasoconstrictors (Cuevas et al, 1996).
Gene therapy of hypertension has been achieved via
transfer of the atrial natriuretic peptide (ANP) gene to
genetically hypertensive rats; chronic infusion of ANP has
been shown to cause natriuresis, diuresis, and hypotension
in rats and humans. Intravenous delivery of the human
ANP gene fused to the RSV 3'-LTR (shown to be
expressed in heart, lung, and kidney) caused a significant
reduction of systemic blood pressure in young
hypertensive rats (4 weeks old), and the effect continued
for 7 weeks; a maximal blood pressure reduction of 21
mm Hg in young hypertensive rats was observed 5 weeks
after injection along with significant increases in urinary
volume and urinary potassium output (Lin et al, 1995).