Biomedical Engineering Reference
In-Depth Information
insulin glargine and insulin detemir, which are used to
improve glycemic control in patients with diabetes mellitus.
However, these analogs fail to provide stable basal glycemia
for more than a few hours and are associated with hyper-
glycemic and hypoglycemic excursions [43,67]. Albulin
was generated by fusing single-chain insulin (containing
the B and A chains of mature human insulin linked by a
dodecapeptide linker) to the N-terminal end of rHA [43].
The resulting fusion gene was expressed in either yeast or
Chinese hamster ovary (CHO) cells. In normoglycemic
mice, the elimination half-life of subcutaneous administered
albulin was 7 h compared to about 10min with recombinant
insulin. In vivo studies in severely hyperglycemic diabetic
mice indicated that albulin was capable of maintaining a
near-normoglycemic state for long periods of time. A single
subcutaneous injection of albulin normalized blood glucose
levels in a relatively constant and sustained manner for up to
24 h [43]. In addition, albulin had a lower affinity for the
human insulin receptor and insulin growth factor 1 (IGF-1),
suggesting that it may be less mitogenic compared to
recombinant insulin.
10.3.2.4 Butyrylcholinesterase and Cocaine Hydrola-
se
Albumin has been fused to butyrylcholinesterase
(BChE) and its potent analog cocaine hydrolase with the
aim of improving the pharmacokinetic properties of these
proteins.
BChE is used to prevent nerve toxicity caused by expo-
sure to organophosphorus compounds used in various chem-
ical weapons and pesticides. An albumin-BChE fusion
protein was developed to overcome some of the limitations
of PEGylation [34]. The sequence encoding rHA was fused
to the 3
0
end of the human BChE sequence, using a glycine/
serine linker (Gly
6
Ser). The construct was transfected into
baby hamster kidney cells, and transgenic mice and goats
were generated. In vivo, the fusion protein had a considera-
bly longer half-life in pigs (32 h) than did recombinant
human BChE (3 h) and retained much of its biological
activity, according to the results of nerve binding and
inhibition studies. Notably, the albumin-BChE fusion pro-
tein was successfully expressed in goat milk, offering a
potential method for large-scale production and stockpiling
of BChE for the prevention and treatment of organo-
phosphorus compound toxicity.
A BChE mutant designed to elevate the rate constant for
cocaine hydrolysis (called cocaine hydrolase) is a more
potent inhibitor of cocaine effects than is BChE, but its
effects are also limited by a short half-life. An albumin-
cocaine hydrolase fusion protein (Albu-CocH) was therefore
developed [35]. After a single injection into rats, the elim-
ination half-life of albumin-cocaine hydrolase was 8 h,
suggesting ample stability for short-term applications and
the potential for long-term use [36]. The fusion protein was
found to block cocaine toxicity and improve relapse behav-
ior in rats exposed to cocaine. Specifically, it prevented
seizures in rats that were given typically lethal doses of
cocaine, and rescued animals after convulsions had com-
menced. It also blocked reinstatement behavior in drug-
seeking rats that had self-administered cocaine, but it had no
effect on amphetamine-primed reinstatement, locomotor
behavior, or food-rewarded behavior. These results suggest
that albumin-cocaine hydrolase may have applications in not
only the acute setting of cocaine overdose but also the
chronic setting to help prevent recidivism.
10.3.2.3 Growth Hormone
Human growth hormone
(hGH) is used to treat adult and pediatric GH deficiency,
AIDS-associated wasting in adults, Turner syndrome, and
chronic renal insufficiency [29]. hGH is a 191-amino-acid
pituitary hormone that stimulates the production and release
of IGF-1. The hormone has a very short half-life of less than
20min in vivo and therefore requires frequent administration
[68]. Compliance can therefore be an issue, particularly in
children. The fusion protein Albutropin
1
was generated by
fusing the N-terminus of recombinant hGH to the C-termi-
nus of albumin. The protein was expressed in S. cerevisiae
and purified from the supernatants obtained. Following
subcutaneous injection in rats, clearance of Albutropin
was more than twofold slower than that of hGH, and the
volume of distribution was higher [29]. In GH-deficient
hypophysectomized rats, body weight increased signifi-
cantly when Albutropin was given daily, every other day,
or every 4 days; these effects on body weight were greater
than those of hGH when given at equimolar doses at
the same schedule. A marked benefit over hGH was seen
when Albutropin was given every 4 days: with this schedule,
Albutropin increased body weight and bone growth, whereas
hGH had no effect. In monkeys, the half-life of Albutropin
was sixfold longer than that of hGH. In addition, its mean
residence time was 5- to 10-fold greater, and clearance was
eightfold slower. The improved pharmacokinetic properties
translated into improved pharmacodynamics: a single sub-
cutaneous dose of Albutropin maintained IGF-1 levels above
baseline for 7 days in a manner similar to daily dosing with
hGH for 7 days. Thus, Albutropin may provide the same
efficacy as does hGH while allowing a reduced dosing
frequency [29].
10.3.2.5 B-Type Natriuretic Peptide
B-type natriuretic
peptide (BNP) is used in the treatment of decompensated
congestive heart failure (CHF) but has a short half-life,
thereby limiting its use to the acute setting and necessitating
continuous intravenous infusion [28]. An albumin-BNP
fusion protein (AlbuBNP; Cardeva
TM
) was generated using
a mammalian expression system (human embryonic kidney
(HEK)-293 cells) [28]. In vitro, albumin-BNP has the same
maximal bioactivity as BNP in terms of activating cyclic
guanosine monophosphate (cGMP). In a hypertensive rat
model, a single injection of albumin-BNP reduced systolic
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