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Blood, Vol. 91 No. 4 (February 15), 1998:
pp. 1446-1452
HBED: A Potential Alternative to Deferoxamine for Iron-Chelating
Therapy
By
Raymond J. Bergeron,
Jan Wiegand, and
Gary M. Brittenham
From the Department of Medicinal Chemistry, University of Florida,
Gainesville, FL and the Department of Medicine, MetroHealth Medical
Center, Case Western Reserve University, Cleveland, OH.
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ABSTRACT |
To examine the potential clinical usefulness of the hexadentate
phenolic aminocarboxylate iron chelator
N,N-bis(2-hydroxybenzyl)ethylenediamine-N,N-diacetic acid (HBED) for the chronic treatment of transfusional iron overload, we compared the iron excretion induced by subcutaneous (SC) injection of HBED and deferoxamine (DFO), the reference chelator, in rodents and
primates. In the non-iron-overloaded, bile-duct-cannulated rat, a
single SC injection of HBED, 150 µmol/kg, resulted in a net iron
excretion that was more than threefold greater than that after the same
dose of DFO. In the iron-loaded Cebus apella monkey, a
single SC injection of HBED, 150 µmol/kg, produced a net iron excretion that was more than twice that observed after the same dose of
SC DFO. In patients with transfusional iron overload, SC injections of
HBED may provide a much needed alternative to the use of prolonged
parenteral infusions of DFO.
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INTRODUCTION |
IN PATIENTS WITH transfusional iron
overload, the magnitude of the body iron burden is the principal
determinant of the severity of iron toxicity and clinical
outcome.1,2 During the past 30 years, iron-chelating
therapy with deferoxamine B mesylate (DFO, Fig 1) has
been shown to be a generally safe and efficacious means of controlling
body iron that can prolong survival and prevent or ameliorate organ
dysfunction.3-7 Unfortunately, treatment with DFO is
cumbersome, inefficient, expensive, and unpleasant. Because DFO is
poorly absorbed from the gastrointestinal tract and rapidly eliminated
from the circulation, prolonged parenteral infusion is
needed.8-10 Moreover, DFO is inefficient as an iron chelator; typically only 5% or less of the drug administered binds iron.8,11 Effective therapy usually requires subcutaneous (SC) or intravenous administration by a portable infusion pump for 9 to
12 hours daily. Not surprisingly, almost all patients have difficulty
in complying with such a demanding regimen.
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| Fig 1.
Structures of the iron chelators chosen for evaluation:
deferoxamine B (DFO) and
N,N -bis(2-hydroxybenzyl)ethylenediamine-N,N-diacetic acid dihydrochloride dihydrate (HBED).
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DFO is a bacterial siderophore that is commercially produced by
large-scale fermentation of a strain of Streptomyces
pilosus,12 a method of manufacture that contributes to
the high cost of the drug and to the allergic reaction that some
patients experience, possibly caused by cytokines or other fermentation
products not removed during purification. Almost all patients develop
discomfort or pain at the site of DFO infusion; moreover, some people
are severely allergic to this compound.13-15 A recent study
indicated that, although DFO does not stimulate human basophils to
release histamine, the drug nevertheless induces the nonimmunologic
activation of the dermal mast cells.16 The volunteers in
that study were exposed to DFO for the first time: all developed a
dose-dependent wheal and flare reaction after the intradermal injection
of DFO. The investigators even suggested that DFO could be used as a
positive control in the allergy clinic in intradermal skin tests.
Although allergy with anaphylactic reactions to DFO is
rare,17,18 a variety of neurotoxic19 and other
adverse effects20-23 of DFO have now been recognized,
especially with intensive therapy. Thus, although the experience with
DFO has shown that adequate control of body iron can avert
complications of transfusional iron overload, problems of toxicity in
some patients, the high cost of production, the inefficiency of
chelation, local reactions, and the difficulties with compliance
associated with prolonged parenteral administration have prompted an
ongoing search for safe, inexpensive alternatives to DFO.
Most efforts to develop substitutes for DFO for iron-chelating therapy
have concentrated on bidentate or tridentate iron-chelating agents that
remain active after oral administration.24,25 Because of
concerns that these partial ligands might exacerbate iron toxicity and
the realization that some patients may have difficulty in taking an
oral formulation three or more times daily, we have examined the
possibility of the administration of a hexadentate iron chelator by SC
injection. The polyanionic amine
N,N-bis(2-hydroxybenzyl)ethylenediamine-N,N-diacetic acid26 (HBED; Fig 1) is a synthetic hexadentate ligand
that, like DFO, forms a 1:1 complex with iron with high affinity and selectivity. HBED and its dimethyl ester prodrug (DMHBED) have both
been thoroughly investigated in rodents.27-29 Both of the drugs looked very promising in this model when administered orally, and
not unexpectedly, these findings generated a great deal of excitement.
The availability of an orally active iron chelator seemed imminent.
Unfortunately, the rodent findings were not mirrored in higher animals.
In previous studies in the iron overloaded Cebus apella model,
an excellent predictor of how a chelator will perform when administered
to humans, we showed that while the iron clearing efficiency of
SC-administered DFO30 given at a dose of 150 µmol/kg was
5.5% ± 0.9%, the efficiencies of orally (PO)
administered HBED or DMHBED,31 also dosed at 150 µmol/kg,
were significantly lower: 0.5% ± 0.5% and 1.5% ± 0.6%,
respectively. Not surprisingly, HBED was also a disappointment when
given orally to patients; 32-33 the limited iron excretion
that resulted is insufficient for use of this agent in the treatment of
transfusional iron overload. The result of oral administration of
DMHBED has not yet been evaluated clinically, but the primate findings
suggest that the iron excretion to be expected, although greater than
that with HBED, will still be only about one third of that produced by
DFO.
We reconsidered the potential therapeutic usefulness of HBED given
parenterally after recalling that intraperitoneal injection of the drug
in the hypertranfused rat produced an iron excretion that was twofold
to threefold greater than that following injection of
DFO.34 Accordingly, in addition to the preceding studies, we also performed preliminary investigations in the primates into the
iron clearing properties of SC-administered HBED and DMHBED. In these
studies, HBED and DMHBED were given to the monkeys SC at a dose of 150 µmol/kg. Whereas the efficiency of SC DMHBED was only 2.4% ± 1.1% (range, 1.0% to 3.4%), the efficiency of SC-administered HBED
was 12.6% ± 3.9% (range, 9.1% to 16.8%), an efficiency more
than twice that of DFO given SC and approximately six times that of
DMHBED administered SC at an equimolar dose. On the basis of this
initial finding, we have now thoroughly explored the iron-clearing
properties of SC-administered HBED in the Cebus monkey model.
In this report, we have compared the iron excretion induced DFO with
that induced by HBED, each administered SC to rats and monkeys to
assess the possibility that SC HBED might provide an alternative to DFO
infusions in patients with transfusional iron overload.
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MATERIALS AND METHODS |
Materials.
Deferoxamine B in the form of the methanesulfonate salt, Desferal, was
obtained from Ciba-Geigy Ltd, (Basel, Switzerland). HBED
dihydrochloride dihydrate was obtained from Strem Chemical Co
(Newburyport, MA). Cremophor RH-40 was obtained from BASF
(Parsippany, NJ). Sprague-Dawley rats were purchased from Charles River
(Wilmington, MA). C apella monkeys were obtained from World
Wide Primates (Miami, FL). All reagents and standard iron solutions
were obtained from Aldrich Chemical Co (Milwaukee, WI). Nalgene
metabolic cages, rat jackets, and fluid swivels were purchased from
Harvard Bioscience (South Natick, MA). Intramedic polyethylene tubing
was obtained from Fisher Scientific (Pittsburgh, PA). Atomic absorption
measurements were made on a Perkin-Elmer model 5100 PC (Norwalk, CT).
Ultrapure salts were obtained from Johnson Matthey Electronics
(Royston, UK). Imferon, an iron dextran solution, was obtained from
Fisons (Bedford, MA). All hematological and biochemical
studies30 were performed by Allied Clinical Laboratories
(Gainesville, FL).
Cannulation of bile duct in rats.
Male Sprague-Dawley rats averaging 400 g were housed in Nalgene plastic
metabolic cages during the experimental period and given free access to
water. The animals were anesthetized using sodium pentobarbital (55 mg/kg) given intraperitoneally. The bile duct was cannulated, using
22-gauge polyethylene tubing, about 1 cm from the duodenum. The cannula
was inserted about 2 cm into the duct, and once bile flow was
established, the cannula was tied snugly in place. A skin tunneling
needle was inserted from the shoulder area around to the abdominal
incision. The cannula was threaded through the needle until it emerged
from the shoulder opening. The cannula was then passed from the rat to
the swivel inside a metal torque-transmitting tether, which was
attached to a rodent jacket around the animal's chest. The cannula was directed from the rat to a Gilson micro fraction collector (Middleton, WI) by a fluid swivel mounted above the metabolic cage. This system allowed the animal to move freely in the cage while continuous bile
samples were being collected. Bile samples were collected at 3-hour
intervals for 24 hours. Urine samples were taken every 24 hours. Sample
collection and handling were as previously described.35
Iron loading of C apella monkeys.
After intramuscular anesthesia with ketamine, an
intravenous infusion was started in a leg vein. The iron
dextran was added to approximately 90 mL of sterile normal saline and
administered to the animals by slow infusion at a dose of 200 to 300 mg
of iron per kilogram of body weight over 45 to 60 minutes. Two to three
infusions, separated by between 10 and 14 days, were necessary to
provide about 500 mg of iron per kilogram of body weight. After administration of iron dextran, the serum transferrin iron
saturation rose to between 70% and 80%. The serum half-life of iron
dextran in humans is 2.5 to 3.0 days.36 We waited at least
20 half-lives, 60 days, before using any of the animals in
experiments evaluating iron-chelating agents.
Iron-balance studies in C apella monkeys.
Seven days before the administration of the drug, the animals were
placed in metabolic cages37 and started on a low-iron liquid diet.30 The monkeys were maintained on the low-iron
liquid diet for the duration of the experiment. They were given food according to their body weight, and intake was very carefully monitored.
Three days before drug administration, day 2 to day 0, baseline iron
intake and output values were measured. This same measurement was made
for day +1 to day +3. The total amount of iron intake was compared with
the total amount of iron excreted.
Primate fecal and urine samples.
Fecal and urine samples were collected at 24-hour intervals. The
collections began 4 days before the administration of the test drug and
continued for an additional 5 days after the drug was given. Fecal
samples were assayed for the presence of occult blood, weighed, and
mixed with distilled deionized water before autoclaving for 30 minutes.
The mixture was then freeze-dried, and a known portion of the powder
was mixed with low-iron nitric acid and refluxed for 24 hours. Once any
particulate matter in the digested samples was removed by
centrifugation, iron concentrations were determined by flame atomic
absorption. Monkey urine samples were acidified and reconstituted to
initial volume after sterilization, if necessary.
Drug preparation and administration.
DFO was administered to the rats at a dose of 150 µmol/kg in 40%
Cremophor RH-40. HBED was administered SC and PO to the rats at a dose
of 150 µmol/kg in a phosphate buffer.
In the primates, DFO was administered SC in sterile water for injection
at a dose of 150 µmol/kg, whereas HBED was first dissolved in a
phosphate buffer and given SC in 40% Cremophor RH-40 at a dose of 75 or 150 µmol/kg. In addition, to more closely mimic clinical
applications in patients, HBED was also given to the monkeys SC at a
dose of 150 µmol/kg in a phosphate buffer; no Cremophor vehicle was
used.
Calculation of iron chelator efficiency.
The efficiency of each chelator was calculated on the basis of a 1:1
ligand-iron complex. In the monkeys the numbers were generated by
averaging the iron output for 4 days before the administration of the
drug, subtracting these numbers from the 2-day iron clearance after the
administration of the drug, and then dividing by the theoretical
output; the result is expressed as a percent. The efficiencies in the
rodent model were calculated by subtracting the iron excretion of
control animals from the iron excretion of treated animals. This number
was then divided by the theoretical output; the result is expressed as
a percent.
Statistical analysis.
Data are presented as the mean ± the standard error of the mean.
For comparisons of the means of two groups, the two-sample t-test (without the assumption of equality of variances) was
used for analyzing the rodent data, whereas the primate data were
analyzed using a paired t-test. All tests were two-tailed, and
a significance level of P < .05 was used.
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RESULTS |
Chelator-induced iron excretion in rats.
These studies were conducted in non-iron-overloaded,
bile-duct-cannulated rats. The chelators were administered at a dose of 150 µmol/kg body weight. Groups of rats were given DFO by SC injection (n = 6) or HBED PO by gavage (n = 4) or SC injection (n = 3).
Figure 2 shows the time course of the mean
biliary iron excretion induced by the chelators in each group of rats,
expressed as µg Fe/kg body weight. The peak amounts of iron excreted
with SC HBED were more than twofold greater than the peak iron
excretion after either SC DFO or HBED given by gavage (P < .05 at 3 hours and P < .01 at 6 hours). The iron excretion
induced by the chelators in each group of rats is shown in
Fig 3 expressed as the net mean amount of
iron excreted in the urine and in the bile (µg Fe/kg body weight) and
as the efficiency of iron chelation (net iron excretion/total
iron-binding capacity of chelator administered, expressed as a
percent).
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| Fig 2.
Time course of mean biliary iron excretion in normal rats
after administration of DFO by SC injection and after administration of
HBED by gavage or by SC injection. Both chelators were given at a dose
of 150 µmol/kg body weight. The peak amounts of iron excreted with SC
HBED (*) were more than twofold greater than the peak iron excretion
after either SC DFO or HBED given by gavage (P < .05 at 3 hours and P < .01 at 6 hours).
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| Fig 3.
Mean net iron excretion in normal rats after
administration of DFO by SC injection and after administration of HBED
by gavage or by SC injection. Excretion is shown as µg Fe/kg body
weight on the scale of the left vertical axis and as efficiency of
chelation (net iron excretion/total iron-binding capacity of chelator
administered, expressed as a percent) on the right vertical axis. Both
chelators were given at a dose of 150 µmol/kg body weight.
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Subcutaneous DFO induced the excretion of 209 ± 59 µg Fe/kg body
weight and was found to have an efficiency of 2.5% ± 0.7% (range, 1.7% to 3.7%), with about 74% of the chelator-induced iron excreted through the bile and about 26% through the urine. Compared to SC DFO, HBED given PO by gavage resulted in a twofold greater iron excretion of 436 ± 176 µg Fe/kg body weight
(P < .02); the efficiency was 5.2% ± 2.1% (range, 3.5%
to 8.3%), with about 97% of the chelator-induced iron excretion
through the bile and about 3% through the urine. HBED given to the
rodents by SC injection was more than three times as effective in
inducing iron excretion as DFO administered SC (P < .001),
inducing the excretion of 679 ± 8 µg Fe/kg body weight with an
overall efficiency of 8.1% ± 0.1% (range, 7.9% to 8.2%). About
83% of the iron was excreted in the bile and 17% in the urine. No
untoward side effects were noted in the rodents.
Chelator-induced iron excretion in C apella monkeys.
The studies were conducted with C apella monkeys who had
previously been administered intravenous iron dextran to provide about
500 mg of iron per kilogram body weight, as described in Materials and
Methods. Groups of monkeys (6 in each group) were given SC injections
of either DFO or HBED. DFO in aqueous solution was given SC at a dose
of 150 µmol/kg and induced the excretion of 435 ± 115 µg Fe/kg
body weight and was found to have an efficiency of 5.1% ± 1.3%
(range, 3.3% to 6.6%), with about 65% of the chelator-induced iron
excretion in the stool and about 35% in the urine. In our initial
experiments, the HBED was dissolved in a phosphate buffer and
administered in 40% Cremophor RH-40 (a polyethoxylated castor oil used
as a vehicle for compounds with poor aqueous solubility), because of
the low water solubility of the HBED dihydrochloride dihydrate.
HBED-Cremophor was administered by SC injection at doses of 75 or 150 µmol/kg. At the dose of 75 µmol/kg, HBED-Cremophor induced the
clearance of 793 ± 410 µg/kg of iron and had an
efficiency of 18.4% ± 9.1% (range, 7.4% to 27.8%). Most of the
iron (92%) was excreted in the feces, whereas 8% was excreted in the
urine. At a dose of 150 µmol/kg, HBED-Cremophor induced the clearance of 1,349 ± 475 µg/kg, an efficiency of 16.1% ± 5.6% (range, 9.3% to 23.0%). Once again, the majority of the iron,
90%, was excreted in the feces; 10% was found in the urine.
Finally, to more closely mimic potential clinical applications, HBED
has also been given SC to the same group of monkeys used in the
preceding experiments at a dose of 150 µmol/kg in a phosphate buffer;
no Cremophor vehicle was used. Once again, subcutaneous HBED induced
the excretion of about twice as much iron as DFO, 899 ± 193 µg
Fe/kg body weight (P < .001), and was found to have an
efficiency of 10.7% ± 2.3% (range, 8.3% to 13.8%), with about 92% of the chelator-induced iron excretion in the stool and about 8%
in the urine. At the dose of 150 µmol/kg, no significant difference (P > .2) was found between the mean net iron excretion
induced by HBED prepared in phosphate buffer or in Cremophor.
Figure 4 shows the mean iron excretion
induced by DFO given SC in an aqueous solution and HBED administered SC
in a buffer at a dose of 150 µmol/kg body weight. The data are
expressed as the mean net amount of iron excreted in the urine and in
the feces (µg Fe/kg body weight) and as the efficiency of iron
chelation. For convenience, Fig 4 also shows, marked with an asterisk,
the results of our previously published study of the oral
administration of HBED in a phosphate buffer to a group of
C apella monkeys with a similar magnitude of iron
overload.31 Oral HBED induced the excretion of only 50 ± 44 µg Fe/kg body weight and was found to have an efficiency of
0.5% ± 0.5% (range, 0.1% to 1.1%), with about 56% of the
chelator-induced iron excretion in the stool and about 44% in the
urine. No adverse effects of chelator administration were noted in the
monkeys; all hematologic and biochemical tests remained within normal
ranges.
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| Fig 4.
Mean net iron excretion in C apella monkeys with
iron overload (see text) after administration of DFO by SC injection
and of HBED by SC injection. Excretion is shown as µg Fe/kg body
weight on the scale of the left vertical axis and as efficiency of
chelation (net iron excretion/total iron-binding capacity of chelator
administered, expressed as a percent) on the right vertical axis. Both
chelators were given at a dose of 150 µmol/kg body weight. For
convenience, the result of our previously published study31
of the oral administration of HBED in phosphate buffer to C
apella monkeys with a similar magnitude of iron overload is also
shown; this previous result is indicated by an asterisk.
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Primate iron balance studies.
The results of these studies clearly indicate that both DFO and HBED
can hold the monkeys in a negative iron balance (Table 1). Iron balance was calculated by comparing the amount of iron absorbed by the untreated animals over a 3-day period (day 2 to day 0)
with the amount of iron absorbed by treated animals over a 3-day period
(day +1 to day +3). Net iron balance = dietary iron intake (urinary + fecal iron excretion); animals in a negative iron balance are excreting
more iron than they are absorbing. Monkeys treated with 150 µmol/kg
of DFO on the average excreted 278 ± 185 µg/kg more iron than
they absorbed (P < .0001), whereas animals treated with 75 µmol/kg of HBED excreted 606 ± 406 µg/kg more iron than they
absorbed (P < .005). Finally, animals given HBED SC at the
same molar quantity as DFO excreted 1,141 ± 456 µg/kg more than
they absorbed when the compound was dosed with the Cremophor vehicle
(P < .001) and 689 ± 158 µg/kg more than they absorbed
(P < .001) when the drug was given without the Cremophor vehicle. As was observed with the urinary and fecal iron clearance data, the animals given HBED SC at a dose of 150 µmol/kg had a negative iron balance that was significantly greater than that seen
with DFO, 4.1 times greater when the compound was dosed with the
Cremophor vehicle (P < .001), and 2.5 times greater when the drug was given without the Cremophor vehicle (P < .01). At
the dose of 150 µmol/kg, no significant difference (P > .1)
was found between the iron balance observed with HBED prepared in a
phosphate buffer versus HBED given in the Cremophor vehicle.
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DISCUSSION |
Previous efforts to develop alternatives to DFO for iron-chelating
therapy have focused almost exclusively on identifying agents that
remain active after oral administration. DFO (Mr 657) is a
hexadentate ligand (Fig 1); one molecule of DFO binds a single atom of
iron and the chelate (feroxamine) is virtually inert biologically. Because absorption across the gastrointestinal tract is generally better for compounds of lower molecular weight (Mr < 350),38,39 most of the efforts to develop orally active
agents have been devoted to smaller chelators (Mr 100 to
250) that are bidentate (2,3 dihydroxybenzoic acid, hydroxypyridones)
or tridentate (pyridoxal isonicotinoyl hydrazone,
desferrithiocin). However, recent theoretical considerations, laboratory studies in vitro and in vivo, and clinical experience have raised concerns that, in some circumstances, these incomplete ligands might exacerbate iron-related tissue damage. To
occupy the six coordination sites of a single atom of iron, one
molecule of a hexadentate (eg, DFO or HBED), two molecules of a
tridentate, or three molecules of a bidentate chelator are needed. At
low concentrations of bidentate or tridentate chelators relative to the
available iron, partially liganded forms of iron (bound to only one or
two molecules of bidentate or tridentate ligands) appear in which the
unoccupied coordination sites are either open or are occupied by
another readily dissociable ligand such as water. Although iron is
poorly soluble under physiological conditions, bidentate or tridentate
chelators can greatly increase the solubility of iron, in part through
the production of partially liganded chelates. Theoretically, these
partially liganded forms of iron may catalyze the formation of hydroxyl
radicals or other reactive oxygen species, in which rapid and
nonspecific reactivity may be particularly injurious.40
Laboratory studies have confirmed that bidentate or tridentate
chelators may increase both the availability and reactivity of iron for
participation in free radical-mediated lipid
peroxidation,41,42 which accumulating evidence implicates in the pathogenesis of hepatic fibrosis and other tissue
damage.43,44 In addition, studies in an animal model of
human iron overload45 have found that the combination of
iron overload and treatment with a bidentate hydroxypyridone chelator
(1,2-diethyl-3-hydroxypyridin-4-one; CP94) produced a worsening of
hepatic fibrosis and increased cardiac iron accumulation.46
A closely related orally active bidentate hydroxypyridone, deferiprone
(1,2-dimethyl-3-hydroxypyridin-4-one; CP20; L1), is now in
clinical trials.47 Recently, evidence of exacerbation of
hepatic fibrosis was found in a group of patients who have had
long-term treatment with deferiprone.48 Overall, these
theoretical concerns and the related clinical and laboratory findings
have created caution about the sole reliance on the development of
orally active bidentate and tridentate iron-chelating agents as
potential alternatives to parenteral DFO.
HBED (Mr 388) is a phenolic aminocarboxylate chelator that
was first synthesized by Martell et al some three decades
ago26 and, like DFO, forms a 1:1 hexadentate complex with
ferric iron. In rats, the LD50 (PO or intraperitoneally) is
in excess of 800 mg/kg,49 and the compound was originally
chosen for further development as an iron-chelating agent after studies
in rodents suggested that it was well absorbed from the
gastrointestinal tract and remained active as an iron chelator after
oral administration.34,50 The results of the studies in
rats without iron overload that are reported in this paper confirm that
an oral dose of HBED of 150 µmol/kg is about twice as effective as
the same dose of DFO given by SC injection. Although rodents provide a
valuable first-line animal screen, allowing for a rapid and inexpensive
way to identify and discard chelators that are ineffective in vivo,
there is no strict correspondence between the effectiveness of a
chelator in rodents and that in humans. Therefore, for the secondary
screening of iron-chelating agents intended for clinical use, we have
developed and validated an iron loaded C apella monkey model.
In previous studies with this model, we found that the iron excretion
induced by PO administration of HBED was only about 10% of that
resulting from the SC administration of an equivalent dose of
DFO31 (Fig 4). Iron balance studies with PO administration
of HBED to human volunteers32,33 have now confirmed the
lack of activity predicted from the primate model. The iron excretion
produced after oral administration of HBED to patients is insufficient for therapeutic use in transfusional iron overload. Nonetheless, no
adverse effects of oral administration of HBED were noted in these
trials.
The potential therapeutic usefulness of HBED given parenterally was
evaluated in the monkeys after recalling that intraperitoneal injection
of the drug in the hypertranfused rat produced an iron excretion that
was significantly greater than that after injection of
DFO.34 Accordingly, we compared the iron excretion induced by SC injection of HBED in rats and monkeys with that induced by SC
injection of DFO. The net iron excretion after SC HBED in the rat was
more than threefold greater than that after DFO (Fig 3). Most
importantly, in the primate model with iron overload, a single SC
injection of HBED produced iron excretion that was at least twice that
observed after SC DFO in the same group of monkeys (Fig 4). The use of
a Cremophor vehicle did not significantly increase HBED-induced iron
excretion.
In addition to enhanced iron excretion, parenteral HBED has some
possible advantages over DFO as a candidate iron-chelating agent for
the chronic treatment of iron overload. For patients allergic to DFO,
HBED, a member of a different family of chelators, would be unlikely to
provoke a similar response. Because HBED is a synthetic product,
problems of local reactions to fermentation products not removed during
purification would be avoided. Finally, HBED, like DFO, protects
against iron-mediated free radical generation.42 The
results reported here strongly suggest that SC injection of HBED could
potentially provide patients with a clinically effective form of
iron-chelating therapy.
In quantitative terms, most patients with thalassemia major require
between 200 and 300 mL/kg body weight per year of blood, an amount
equivalent to about 250 to 400 µg Fe/kg body weight per day. In the
C apella monkey with iron overload, the iron excretion observed
after a single SC injection of DFO, 435 ± 115 µg Fe/kg body weight, is consistent with the established ability of the daily
use of DFO to control body iron. Giving DFO by prolonged SC infusion
would be expected to increase iron excretion, but the clinical
significance of the monkey data may be put into perspective by
considering recent reports indicating that twice-daily SC injections of
DFO may be as effective as the same dose of the drug administered by
prolonged SC infusion.51-52 The results of the primate
studies presented here suggested that, in patients with iron overload, a single SC injection of HBED might produce an iron excretion even
greater than that induced by the same dose of DFO. Although caution is
needed in extrapolating from the primate model to patients with iron
overload, the higher iron excretion observed after a single SC
injection of HBED in a phosphate buffer, 899 ± 193 µg Fe/kg,
suggests that a regimen in which SC HBED was used every other day might
be effective in maintaining iron balance. The possibility of using
higher doses of HBED administered less frequently, eg, once or twice
weekly, also needs examination. Overall, these findings suggest that
preclinical evaluation of parenteral HBED should be completed promptly,
followed by iron balance studies in human volunteers. Subcutaneous
injection of HBED may provide patients with transfusional iron overload
an alternative to the use of prolonged parenteral infusions of DFO for
iron-chelating therapy.
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FOOTNOTES |
Submitted August 21, 1997;
accepted October 2, 1997.
Supported in part by research grants from the National Institutes of
Health, Bethesda, MD (DK49108, AI35827, and HL57607).
Address reprint requests to Raymond J. Bergeron, PhD, Box 100485 JHMHC,
Department of Medicinal Chemistry, University of Florida, Gainesville,
FL 32610.
The publication costs of this article were defrayed in part by page
charge payment. This article must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C. section 1734 solely
to indicate this fact.
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Abstract
Introduction
Abstract