Design, synthesis, and testing of non-nephrotoxic desazadesferrithiocin polyether analogues

Article abstract of DOI:10.1021/jm800154m

A series of iron-clearing efficiencies (ICEs), ferrokinetics, and toxicity studies for (S)-2-(2,4-dihydroxyphenyl)-4,5-dihydro-4-methyl-4- thiazolecarboxylic acid (deferitrin, 1), (S)-4,5-dihydro-2-[2-hydroxy-4-(3,6,9- trioxadecyloxy)phenyl]-4-methyl-4-th

Full text of DOI:10.1021/jm800154m

J. Med. Chem. 2008, 51, 3913–3923
3913
Design, Synthesis, and Testing of Non-Nephrotoxic Desazadesferrithiocin Polyether Analogues
Raymond J. Bergeron,* Jan Wiegand, James S. McManis, Neelam Bharti, and Shailendra Singh
Department of Medicinal Chemistry, UniVersity of Florida, GainesVille, Florida 32610-0485
ReceiVed February 13, 2008
A series of iron-clearing efficiencies (ICEs), ferrokinetics, and toxicity studies for (S)-2-(2,4-dihydroxyphenyl)-
4,5-dihydro-4-methyl-4-thiazolecarboxylic acid (deferitrin, 1), (S)-4,5-dihydro-2-[2-hydroxy-4-(3,6,9-tri-
oxadecyloxy)phenyl]-4-methyl-4-thiazolecarboxylic acid (2), and (S)-4,5-dihydro-2-[2-hydroxy-3-(3,6,9-
trioxadecyloxy)phenyl]-4-methyl-4-thiazolecarboxylic acid (3) are reported. The ICEs in rodents are shown
to be dose-dependent and saturable for ligands 2 and 3 and superior to 1. Both polyether analogues in
subcutaneous (sc) versus oral (po) administration in rodents and primates demonstrated excellent
bioavailability. Finally, in a series of toxicity studies of ligands 1-3, the dosing regimen was shown to
have a profound effect in animals treated with ligand 1. When ligand 1 was given at doses of 237 µmol/
kg/day twice a day (b.i.d.), there was serious proximal tubule damage versus 474 µmol/kg/day once daily
(s.i.d.). With 2 and 3, in iron-overloaded and/or non-iron-loaded rodents, kidney histopathologies remained
normal.
Although iron comprises 5% of the earth’s crust, living
systems have great difficulty in accessing and managing this
vital micronutrient. The low solubility of Fe(III) hydroxide (Ksp
) 1 × 10-39),1 the predominant form of the metal in the
biosphere, has led to the development of sophisticated iron
storage and transport systems in nature. Microorganisms utilize
low molecular weight, virtually ferric ion-specific ligands,
siderophores;2–6 higher eukaryotes tend to employ proteins to
transportandstoreiron(e.g.,transferrinandferritin,respectively).7–9
Humans absorb and excrete only about 1 mg of the metal
daily; there is no effective mechanism for the excretion of excess
iron.10 In humans, nontransferrin-bound plasma iron, a hetero-
geneous pool of the metal in the circulation, unmanaged iron,
seems to be a principal source of iron-mediated organ damage.
Introduction of excess iron into this closed system, whether
derived from transfused red blood cells11–13 or from increased
absorption of dietary iron,14,15 leads to a build up of the metal
in the liver, heart, pancreas, and elsewhere. Such iron accumula-
tion eventually produces (i) liver disease that may progress to
cirrhosis,16–18 (ii) diabetes related both to iron-induced decreases
in pancreatic ꢀ-cell secretion19,20 and increases in hepatic insulin
resistance, and (iii) heart disease, still the leading cause of death
in thalassemia major21–23 and related forms of transfusional iron
overload.
The toxicity associated with excess iron, whether a systemic
or a focal problem, derives from its interaction with reactive
oxygen species, for instance, endogenous hydrogen peroxide
(H2O2).24–27 In the presence of Fe(II), H2O2 is reduced to the
hydroxyl radical (HO•), a very reactive species, and HO-, a
process known as the Fenton reaction. The Fe(III) liberated can
be reduced back to Fe(II) via a variety of biological reductants
(e.g., ascorbate), a problematic cycle. The hydroxyl radical reacts
very quickly with a variety of cellular constituents and can
initiate free radicals and radical-mediated chain processes that
damageDNAandmembranesaswellasproducecarcinogens.25,28–30
The solution to the problem is to remove excess unmanaged
iron.31 In the majority of patients with transfusion-dependent
refractory anemias, treatment with a chelating agent capable of
sequestering iron and permitting its excretion from the body is
the only therapeutic approach available.
The iron-chelating agents that are now in use or that have
been clinically evaluated32 include desferrioxamine B mesylate
(DFOa), 1,2-dimethyl-3-hydroxypyridin-4-one (deferiprone,
L1),33–36 4-[3,5-bis(2-hydroxyphenyl)-1,2,4-triazol-1-yl]benzoic
acid (deferasirox, ICL670A),37–40 and the desferrithiocin (DFT)
analogue, (S)-2-(2,4-dihydroxyphenyl)-4,5-dihydro-4-methyl-4-
thiazolecarboxylic acid [deferitrin (1), Table 1]. DFO, a hexa-
coordinate hydroxamate iron chelator produced by Streptomyces
pilosus,41 is not orally active and is best administered sc by
continuous infusion over long periods of time,11,42 a patient
compliance issue.11,43 Deferiprone, an orally active bidentate
chelator, is less efficient than sc DFO at removing iron.33–36
Whereas the orally active tridentate chelator deferasirox has now
been approved by the FDA, it did not demonstrate noninferiority
to DFO.37–40 Furthermore, Novartis has recently (December,
2007) updated the prescribing information40 for deferasirox:
“There have been postmarketing reports of hepatic failure, some
with a fatal outcome, in patients treated with Exjade. Most of
these events occurred in patients greater than 55 years of age.
Most reports of hepatic failure involved patients with significant
comorbidities, including liver cirrhosis and multi-organ failure.”
In addition, the prescribing information was also changed in
April, 2007 to reflect renal toxicity: “Cases of acute renal failure,
some with a fatal outcome, have been reported following the
postmarketing use of Exjade (deferasirox). Most of the fatalities
occurred in patients with multiple co-morbidities and who were
in advanced stages of their hematological disorders.”40 Finally,
ligand 1 is an orally active tridentate DFT analogue whose
clinical development was recently halted. Although deferitrin
was well tolerated in patients at doses of 5, 10, or 15 mg/kg/
day once daily for up to 12 weeks, unacceptable renal toxicity
was observed in three patients after only 4-5 weeks of treatment
when the drug was given at a dose of 25 mg/kg/day (12.5 mg/
kg b.i.d.).44
* To whom correspondence should be addressed. Phone: (352) 273-7725.
Fax: (352) 392-8406. E-mail: rayb@ufl.edu. Address: Raymond J. Bergeron,
Ph.D., Box 100485 JHMHSC Department of Medicinal Chemistry, Uni-
versity of Florida, Gainesville, Florida 32610-0485.
a Abbreviations: b.i.d., twice a day; DFO, desferrioxamine B mesylate;
DFT, desferrithiocin [(S)-4,5-dihydro-2-(3-hydroxy-2-pyridinyl)-4-methyl-
4-thiazolecarboxylic acid]; ICE, iron-clearing efficiency; s.i.d., once a day.
10.1021/jm800154m CCC: $40.75
2008 American Chemical Society
Published on Web 06/06/2008

3914 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 13
Bergeron et al.
Table 1. Iron-Clearing Activity of Desferrithiocin Analogues Given Orally to Non-Iron-Overloaded Rodents
a The efficiency of each compound was calculated by subtracting the iron excretion of control animals from the iron excretion of the treated animals. The
number was then divided by the theoretical output; the result is expressed as a percent. b The compounds were given as their sodium salts, prepared by the
addition of 1 equiv of NaOH to a suspension of the free acid in distilled water. c Data are from ref 45. d The ligand was solubilized in distilled water. e Data
are from ref 46.
A previous investigation focused on the design of desfer-
rithiocin analogues that balance the lipophilicity/toxicity rela-
tionship while iron-clearing efficiency (ICE) is maintained in
hopes of eliminating nephrotoxicity.45 This study led to the less
lipophilic, more water-soluble ligand, the polyether (S)-4,5-
dihydro-2-[2-hydroxy-4-(3,6,9-trioxadecyloxy)phenyl]-4-methyl-
4-thiazolecarboxylic acid (2).45 A subsequent study assessing
the impact of introducing the 3,6,9-trioxadecyloxy group at
various positions of the desazadesferrithiocin aromatic ring46
revealed the 3′-polyether analogue [(S)-4,5-dihydro-2-[2-hy-
droxy-3-(3,6,9-trioxadecyloxy)phenyl]-4-methyl-4-thiazolecar-
boxylic acid (3, Table 1) to also be an orally active iron
chelator.46
The ICE of 1 in a non-iron-overloaded rodent model after
oral (po) administration at 300 µmol/kg was shown to be 1.1
( 0.8% (Table 1).45,47 Polyether 2, in which a 3,6,9-trioxadecyl
group was fixed to the 4′-hydroxy of 1, performed significantly
better at 300 µmol/kg, with an ICE of 5.5 ( 1.9% when
administered po (p < 0.003 vs 1).45 The ICE of the 3′-polyether
3, again at 300 µmol/kg, was impressive, 10.6 ( 4.4% in rodents
(Table 1).46 The efficiency of 1 given po to iron-loaded primates
at a dose of 150 µmol/kg was found to be 16.8 ( 7.2% (Table
2).47 The corresponding polyether 2, given po at the same dose,
performed very well in primates with an efficiency of 25.4 (
7.4%.45 Polyether 3 given po at 75 µmol/kg, presented an ICE
of 24.5 ( 7.6%.46
new synthetic schemes described below now make ligands 2
and 3 more readily accessible and make further biological
evaluation possible.
Results and Discussion
Synthesis. The synthesis of 2 has recently been reported by
this laboratory.45 Alkylation of the 4′-hydroxyl of the isopropyl
ester of 1 using tri(ethylene glycol) monomethyl ether under
Mitsunobu conditions (diisopropyl azodicarboxylate and tri-
phenylphosphine in THF), filtration, and chromatography gave
the isopropyl ester of 2 in 76% yield. Saponification of the ester
furnished 2 in 95% yield, providing an overall yield of 72%.
In the present work, selective alkylation of ethyl ester 448
was accomplished by heating tosylate 5 (1.3 equiv) and
potassium carbonate (2.1 equiv) in acetone, providing masked
chelator 6 in 84% yield (Scheme 1). Cleavage of ethyl ester 6
as before afforded 2 in 93% yield. The new route to ligand 2
proceeds in greater overall yield, 78% vs 72%; moreover,
attachment of the polyether chain in Scheme 1 employs
inexpensive reagents without formation of triphenylphosphine
oxide and diisopropyl 1,2-hydrazinedicarboxylate, simplifying
purification.
Ligand 3 has also been synthesized in this laboratory.46 (S)-
2-(2,3-Dihydroxyphenyl)-4,5-dihydro-4-methyl-4-thiazolecar-
boxylic acid, which was made in 88% yield from amino acid
cyclization with the appropriate nitrile,49 was converted to its
ethyl ester in 98% yield. However, the two remaining steps to
chelator 3 proceeded in only 15% yield. The polyether chain
was appended to the 3′-hydroxyl under Mitsunobu conditions,
producing the ethyl ester of 3 in 25% yield. Ester hydrolysis
While earlier studies carried out in rodents clearly demon-
strated 2 to be less nephrotoxic than the parent drug 1,45 the
toxicity profile of 3 was not evaluated, nor were any
dose-response studies carried out with any of the ligands to
assess how such manipulation impacts on ICE and toxicity. The

Non-Nephrotoxic Desazadesferrithiocin Polyether Analogues
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 13 3915
Table 2. Iron-Clearing Activity of Desferrithiocin Analogues Given Orally and Subcutaneously to Rodents and Primates
a In the rodents, the efficiency of each compound was calculated by subtracting the iron excretion of control animals from the iron excretion of the treated
animals. The number was then divided by the theoretical output; the result is expressed as a percent. b In the primates, the efficiency was calculated by
averaging the iron output for 4 days before 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. c The compounds were given as their sodium salts, prepared by the addition of 1 equiv
of NaOH to a suspension of the free acid in distilled water. d Data are from ref 45. e The ligand was solubilized in distilled water. f Data are from ref 46.
Scheme 1. Synthesis of 2a
Scheme 2. Synthesis of 3a
a Reagents and conditions: (a) K2CO3 (2.1 equiv), acetone, 84%; (b) 50%
NaOH (13 equiv), CH3OH, then 1 N HCl, rt, 16 h, 93%.
furnished 3 in 60% yield after purification on a reverse phase
column, providing an overall yield of 13%.
A more efficient synthesis of 3 is presented in Scheme 2.
The less hindered phenolic group of 2,3-dihydroxybenzonitrile
(7)49 was alkylated with tosylate 5 (1.0 equiv) and sodium
hydride (2.0 equiv) in DMSO at room temperature,50 generating
8 in 70% chromatographed yield. Thus the triether chain has
been attached in nearly three times the yield compared to the
Mitsunobu coupling while avoiding troublesome byproducts.
Cyclocondensation of nitrile 8 with (S)-R-methyl cysteine (9)
in aqueous CH3OH buffered at pH 6 completed the synthesis
of 3 in 90% yield. Since the unusual amino acid 9 was not
introduced until the last step of Scheme 2, the carboxyl group
did not require protection. The overall yield to 3 is 63%, much
higher than from the previous route.
Chelator-Induced Iron Clearance and Iron Clearing
Efficiency in Non-Iron-Overloaded Rodents: Dose Response
Studies. Because there is a limited amount of chelatable iron
available in an animal at any given time, the iron clearance,
and therefore iron-clearing efficiency of a ligand, is saturable.
The key to managing this phenomenon can be found in the
ferrokinetics and the dose-response properties of the ligand.
a Reagents and conditions: (a) 60% NaH (2.0 equiv), DMSO, 70%; (b)
CH3OH (aq), pH 6, 70 °C, 16 h, 90%.
In this regard, the dose-response along with the corresponding
ferrokinetics of ligands 1-3 given po were evaluated in the
non-iron-overloaded, bile duct-cannulated rodent model. Table
1 probably best illustrates the significance of the differences in
saturation properties between the three ligands. Four sets of
numbers are provided: the ligand dose, the theoretical amount
of iron this dose should clear if it were 100% efficient, the actual
quantity of iron cleared, and ICE. Recall that ICE is calculated
by dividing the actual amount of iron cleared by a given
compound by the theoretical amount that should be cleared. For
example, the theoretical iron excretion of 1, 2, or 3 administered
at a dose of 300 µmol/kg is 8.38 mg Fe/kg (Table 1). When the
dose is reduced to 150 µmol/kg, the theoretical iron excretion
likewise decreases, to 4.19 mg Fe/kg, and when the dose is

3916 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 13
Bergeron et al.
Figure 3. Biliary ferrokinetics of 2 vs 1 in bile duct-cannulated rats.
The compounds were administered po.
Figure 1. Dose-response curve: ICE for compounds 1-3 administered
po in bile duct-cannulated rats.
Figure 4. Biliary ferrokinetics of 3 vs 1 in bile duct-cannulated rats.
The compounds were administered po.
Figure 2. Biliary ferrokinetics of 1 in bile duct-cannulated rats. The
compound was administered po.
kg of 2, the biliary ferrokinetics curves show that essentially
all the deferration has ended at 12 h (Figure 3). In each instance,
the iron was found largely in the bile (>95%).
further dropped to 50 µmol/kg, the theoretical iron clearance is
now only 1.40 mg Fe/kg (Table 1).
The parent drug 1 was given orally at 30051 or 150 µmol/kg
to bile duct-cannulated rodents. Iron clearance was minimal at
each dose, 0.089 ( 0.068 mg Fe/kg and 0.061 ( 0.073 mg
Fe/kg, respectively (Table 1). Not surprisingly, the ICE values
were poor in each instance (<2%), and both were within
experimental error (Figure 1). Because of the significant serum
albumin binding of 1 in rodents, unlike what was seen with the
4′-polyether 2, the saturable nature of iron clearance is likely
overshadowed.46 At 300 and 150 µmol/kg, the iron clearance
of 1 was virtually over by 9-12 h (Figure 2). At both doses,
>98% of the ligand-induced iron excretion occurred in the bile.
When 2 was given orally to rodents at a dose of 300 µmol/
kg,45 the drug induced the excretion of 0.459 ( 0.161 mg Fe/
kg and had an ICE of 5.5 ( 1.9% (Figure 1). When the dose of
the chelator was reduced by half to 150 µmol/kg, the rats
excreted a similar amount of iron, 0.471 ( 0.176 mg Fe/kg (p
> 0.05). However, because the theoretical amount of iron is
also proportionally lower, the ICE doubled, 11.2 ( 4.2% (Figure
1). Finally, when 2 was given po to the rats at a dose of 50
µmol/kg, the drug induced the clearance of 0.303 ( 0.050 mg
Fe/kg (Table 1) and had an ICE of 21.7 ( 3.5% (Figure 1).
This means that in reducing the dose from 300 to 50 µmol/kg,
the ICE increased by nearly 4-fold, from 5.5 ( 1.9% (300 µmol/
kg) to 21.7 ( 3.5% (50 µmol/kg). At 300, 150, and 50 µmol/
When rodents were given 3 po at 300 µmol/kg (Table 1),
the drug caused the excretion of nearly twice as much iron as
2, 0.887 ( 0.367 mg Fe/kg and had an ICE of 10.6 ( 4.4%46
(Figure 1). Iron clearance persisted much longer for 3 than for
2 and was not complete until nearly 24 h postdrug (Figure 4).
At a dose of 150 µmol/kg po, the rats excreted a similar amount
of iron to what was excreted at 300 µmol/kg, 0.782 ( 0.121
mg Fe/kg (p > 0.05); the ICE nearly doubled (18.7 ( 2.9%,
Figure 1). Iron excretion was essentially back to baseline in
18 h (Figure 4). Finally, 3 given po at a dose of 50 µmol/kg
caused the excretion of 0.289 ( 0.062 mg Fe/kg (Table 1); the
ICE was 20.7 ( 4.4% and deferration was complete by 15 h.
In every instance, the majority of iron cleared was seen in the
bile (>95%). Unlike with the 4′-polyether 2 in which the ICE
doubled between the 150 µmol/kg dose and 50 µmol/kg doses,
the change for 3′-polyether 3 was within error (Figure 1). This
is in keeping with the idea that the iron-clearing capacity of 2
becomes saturated more quickly than with 3.
There are three notable differences between the 3′ and 4′-
polyether analogues: (i) although 4′-polyether 2 induced defer-
ration is essentially over at 12 h at all doses, the induced iron
clearance by the 3′-polyether 3 persists much longer; (ii) DFT
analogue 3 causes the excretion of twice as much iron as 2 when
given po at doses of 300 or 150 µmol/kg; and (iii) ICE versus

Non-Nephrotoxic Desazadesferrithiocin Polyether Analogues
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 13 3917
Table 3. Toxicity Studies of Desferrithiocin Analogues in Rodents
a Mild renal toxicity. b Study described in ref 52. c Significant renal toxicity. d Study described in ref 45. e No renal toxicity. f Study described in ref 53.
dose-response curves indicate 2-promoted metal clearance
quickly becomes saturated, while 3 is likely to be effective at
a wider range of concentrations.
When 1 was given to primates at 150 µmol/kg either po or
sc45 (Table 2), the ICE numbers were within error 16.8 ( 7.2%
(po) and 15.9 ( 2.7% (sc, p > 0.05). Polyether 2 at the same
dose, 150 µmol/kg, either po45 (25.4 ( 7.4%) or sc45 (30.4 (
7.2%), had ICE values also within error (p > 0.05). We
previously reported the ICE of 3 given po at a dose of 75 µmol/
kg to be 24.5 ( 7.6%.46 In the current study, the dose was
increased to 150 µmol/kg and given to the primates either po
or sc. The ICEs were found to be within error of each other
(Table 2), 23.0 ( 4.1% (po) and 21.5 ( 3.2% (sc, p > 0.05).
These numbers are in keeping with the idea of high oral
bioavailability of all three drugs (Table 2).45
Finally, the ferrokinetics curves help to define potential
multiple dosing scenarios. For example, because of the pro-
longed iron excretion observed with 3 at 150 or 300 µmol/kg
(Figure 4), it is not unreasonable to expect that this drug could
be given only once daily. However, the more rapid return to
baseline iron excretion levels observed with the parent 1 or the
4′-polyether 2 may require administering a second dose 12 to
15 h after the initial exposure (Figures 2 and 3). On the basis
of the renal damage in rodents described below for b.i.d. versus
s.i.d. dosing of 1 and the nephrotoxicity seen in the patients in
the clinical trials given the drug b.i.d.,44 this kind of multiple
exposure regimen is not possible with 1.
Iron-Clearing Efficiency in Non-Iron-Overloaded Rodents
and Iron-Loaded Primates: Oral versus Subcutaneous Adminis-
tration. The ICE values for a single ligand given either po or sc
to non-iron-overloaded, bile duct-cannulated rodents are almost
always within experimental error of each other. When 1 was
given to rats at a dose of 300 µmol/kg either po or sc45 (Table
2), the ICE numbers were very low and within error 1.1 ( 0.8%
(po) and 1.1 ( 0.6% (sc, p > 0.05). The corresponding polyether
2 at the same dose, 300 µmol/kg, either po (5.5 ( 1.9%) or sc
(8.7 ( 2.6%)45 had ICE values that were also within error (p
> 0.05). Finally, the ICE values of 3 given to the rats po or sc
at a dose of 300 µmol/kg were found to be within error of each
other (Table 2), 10.6 ( 4.4%45 (po) and 13.4 ( 4.5% (sc, p >
0.05). These numbers are in keeping with the idea of high oral
bioavailability of all three drugs (Table 2). This phenomenon
seems relatively consistent throughout the series of desferrithio-
cin analogues and was also observed in the iron-loaded
primates.45
Toxicity Evaluation in Rodents. In previous investigations
we evaluated the toxicity of ligands 152 and 245 when given
orally by gavage s.i.d. to non-iron-overload rats at a dose of
384 µmol/kg/day (equivalent to 100 mg/kg of the DFT sodium
salt) for 10 days (Table 3). All of the rats survived the exposure
period. Extensive tissues were sent out for histopathological
analysis; all 2-treated kidneys were normal. However, the
kidneys of the 1-treated rodents displayed mild damage to the
proximal tubules.52 In the current study, ligand 3 was given
under the same experimental conditions. All of the rats survived
the exposure period. Extensive tissues were sent out for
histopathological analysis; no drug-related abnormalities were
found.
The tolerability of 153 and 245 were also previously evaluated
when the compounds were given to the rodents for a much
longer time period, 30 days. Because of the mild renal toxicity
noted when 1 was given at 384 µmol/kg/day for 10 days,52 we
took the added precaution of iron overloading the rats54,55 prior
to the 30-day exposure. Because there were no drug-related
abnormalities noted with 2 in the 10-day study, these animals
were not iron loaded. The rats were given the chelators at a

3918 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 13
Bergeron et al.
dose that, in primates, causes the excretion of 450 µg Fe/kg.
This is the suggested iron clearance required to keep a
thalassemia patient in negative iron balance.56 Accordingly, rats
were given 1 orally by gavage once daily for 30 days at a dose
of 119 µmol/kg/day;53 rats were treated with 2 at a dose of 63.3
µmol/kg/day.45 All of the rats survived the dosing period.
Extensive tissues were sent out for histopathological analysis;
no drug-related abnormalities were found.
In the current study, ligand 3 was administered to the rats
orally by gavage once daily for 28 days. Because there were
no drug-related abnormalities noted with either the 10-day
evaluation of 3 or in rodents treated with 2 for 30 days,45 we
decided to administer drug 3, not just at the “usual” dose, the
dose that causes the excretion of 450 µg Fe/kg in the primates
(65.7 µmol/kg), but at higher doses as well. The higher doses
were chosen in a deliberate attempt to induce renal toxicity.
Accordingly, the compound was dosed at 75, 150, or 300 µmol/
kg/day to iron-loaded animals (350 mg Fe/kg) as well as to
rats with normal iron stores. Note that these doses are ap-
proximately 1, 2, and 4 times the dose necessary to excrete 450
µg/Fe/kg in the primates. There were six rodents per group.
Unloaded and iron-loaded rats served as age-matched controls.
All of the rats survived the dosing period. Although extensive
tissues were taken, due to the number of animals involved in
this study (48 rats) and the expense of performing the histo-
pathologies, we elected to send out only the kidney and bone
marrow. Neither renal toxicity nor any evidence of bone marrow
toxicity was observed in any of the rodents at any of the doses
studied. Bone marrow samples were checked because of
previous reports of hydroxypyridones33–36 and deferasirox,40
causing agranulocytosis.
Figure 5. Renal perfusion. Control (panel A), 1 237 µmol/kg b.i.d. ×
7 d (panel B), 1 474 µmol/kg s.i.d. × 7 d (panel C), 2 237 µmol/kg
b.i.d. × 7 d (panel D), 3 237 µmol/kg b.i.d. × 7 d (panel E).
Magnification ) 400×.
It is important to reiterate the ICE values in the non-iron-
overloaded rodents for these three ligands: the ICE of 1 given
po at a dose of 300 µmol/kg is 1.1 ( 0.8%; the ICEs of the
polyethers given po at the same dose are 5.5 ( 1.9% for the
4′-polyether 2, while the ICE of the 3′-polyether 3 is 10.6 (
4.4%.45 This means that under comparable dosing regimens,
we are removing significantly more iron with either of the
polyether analogues than with the parent 1, 5 times as much
iron with ligand 2 and 9.6 times as much with analogue 3. A
similar scenario occurs when the ligands are given po at a dose
of 150 µmol/kg: the ICE of the parent 1 is 1.5 ( 1.7%, while
the ICEs for the 4′-polyether 2 and 3′-polyether 3 are 11.2 (
4.2% and 18.7 ( 2.9%, respectively. Therefore, at a dose of
150 µmol/kg, analogue 2 causes the excretion of 7.5 times more
iron than the parent 1, while 3 is 12.5 times more effective than
the parent 1 (Table 1). Thus, the nephrotoxicity observed with
1 is not likely related to its global iron clearing properties.
at 25 mg/kg/day (12.5 mg/kg/dose) was associated with unac-
ceptable renal toxicity.44
In the current study, because of the apparent increase in renal
toxicity observed in patients treated with deferitrin (1) twice
daily versus once daily, we elected to determine if this damage
could be reproduced in the rodents. To allow for a head-to-
head comparison of the impact of b.i.d. versus s.i.d. dosing on
nephrotoxicity, we elected to repeat our previously published
b.i.d. dosing regimen.45 Accordingly, one group of rodents
received 1 at a dose of 237 µmol/kg b.i.d. (474 µmol/kg/day)
for 7 days; an additional group received the drug once daily at
the same total dose, 474 µmol/kg/day, for 7 days. In addition,
rodents were given 3 administered orally by gavage under the
same dosing regimen, 237 µmol/kg b.i.d. (474 µmol/kg/day),
for 7 days. Untreated animals served as age-matched controls.
The rodents were anesthetized one day postdrug, and the kidneys
were perfusion-fixed with glutaraldehyde in Tyrodes buffer.
Thick sections (500 nm) were prepared for light microscopy.
The results of the renal perfusion studies demonstrate
profound differences in toxicity between the polyethers 2 and
3 and the parent ligand 1 and also show a dramatic reduction
in toxicity when 1 was given once daily versus twice daily.
Under light microscopy, the proximal and distal tubules of the
untreated rats show normal tubular architecture (Figure 5A).
The kidneys of rodents treated with the parent 1 given twice
daily at 237 µmol/kg/dose (474 µmol/kg/day) for 7 days show
heavy vacuolization and thinning of the apical membranes
(Figure 5B). These results are virtually identical to what we
had published previously.45 However, when 1 was given once
A Comparison of the Impact of 1, 2, and 3 on Rodent
Kidneys. In a previous investigation, ligands 1 and 2 were
administered orally by gavage to rats twice daily at a dose of
237 µmol/kg/dose (474 µmol/kg/day) for 7 days.45 This dose
is equivalent to 120 mg/kg of the parent 1. Untreated rats served
as age-matched controls. One day postdrug, the rodents were
anesthetized and the kidneys were perfusion-fixed; one kidney
from each rat was dissected. While there was little difference
in the renal structural architecture between the untreated animals
and the 2 treated rats, kidneys from rodents given 1 b.i.d. showed
regional, moderate to severe vacuolization in the proximal
tubules, a loss of the brush border, and tubular extrusions toward
the lumen.45 In addition, the distal tubules displayed moderate
to severe vacuolization.45 Recall that although deferitrin was
generally well tolerated when given to patients at doses of 5,
10, or 15 mg/kg once daily, administering the drug twice daily

Non-Nephrotoxic Desazadesferrithiocin Polyether Analogues
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 13 3919
Table 4. Tissue Distribution of Desferrithiocin Analogues in Rodentsa
daily at the same total dose, 474 µmol/kg/day for 7 days,
although there was some vacuolization of the proximal tubule
cells (Figure 5C), the damage was much less severe than when
the drug was given twice daily (Figure 5B). Interestingly, and
much to our surprise, besides some inclusion bodies seen when
either 2 or 3 were given at 237 µmol/kg b.i.d. for 7 days, there
was little if any damage to the proximal tubules (parts D and E
of Figure 5, respectively). This allows for tremendous flexibility
in dosing schedules with the polyethers.
liver
1
2
3
time (nmol/g wet weight) (nmol/g wet weight) (nmol/g wet weight)
1 h (sc)
1 h (po)
80 ( 7
131 ( 9
339 ( 70
159 ( 16
318 ( 46
234 ( 58
2 h (sc)
2 h (po)
48 ( 20
73 ( 13
132 ( 32
144 ( 18
193 ( 17
151 ( 41
4 h (sc)
4 h (po)
25 ( 4
44 ( 5
67 ( 12
35 ( 14
168 ( 17
87 ( 18
Tissue Distribution of Desferrithiocin Analogues in Rodents.
The differences in behavior, for example, ICE, dose-response,
and ferrokinetics, between the three ligands in rats prompted
us to look more closely at the tissue distribution in rodents. In
a previous investigation, we described the tissue distribution of
the three ligands after they were given to rats sc at a dose of
300 µmol/kg.46 While the results were described in detail in
the previous work, the data are included in Table 4 to allow for
the comparison of how the route of administration (sc versus
po) affects tissue distribution. In the current study, the rodents,
three per group, were given a single 300 µmol/kg dose of the
chelators po. The animals were sacrificed 1, 2, or 4 h postdrug;
the liver, kidney, heart, pancreas, and blood were removed. The
plasma was then separated from the blood cells. All samples
were analyzed by HPLC for the appropriate ligands.
kidney
1
2
3
time (nmol/g wet weight) (nmol/g wet weight) (nmol/g wet weight)
1 h (sc)
1 h (po)
179 ( 4
208 ( 54
252 ( 10
114 ( 16
259 ( 35
150 ( 66
2 h (sc)
2 h (po)
97 ( 50
100 ( 21
41 ( 3
69 ( 21
145 ( 27
48 ( 5
4 h (sc)
4 h (po)
27 ( 7
61 ( 9
34 ( 13
10 ( 1
90 ( 8
15 ( 2
heart
1
2
3
time (nmol/g wet weight) (nmol/g wet weight) (nmol/g wet weight)
1 h (sc)
1 h (po)
13 ( 1
8 ( 1
35 ( 7
17 ( 2
16 ( 1
9 ( 2
In the liver at 1 h postdrug, the relative concentrations of the
chelators given po are 3 > 2 > 1. The concentration of both of
the polyether ligands were similar to each other (p > 0.05) and
were significantly greater than the parent 1, p < 0.04 for 2 and
p < 0.05 for 3. At 2 h postdrug, the order was 3 ≈ 2 > 1.
However, while the concentrations of both of the polyether
ligands were similar to each other (p > 0.05), they were
significantly greater than the parent 1, p < 0.003 for 2 and p <
0.04 for 3. At 4 h, while the 4′-polyether 2 and the parent 1
had levels that were within error of each other, the 3′-polyether
3 had the highest hepatic concentration, p < 0.03 versus 1 and
p < 0.01 versus 2 (Table 4).
2 h (sc)
2 h (po)
5 ( 3
5 ( 1
9 ( 3
9 ( 3
8 ( 1
4 ( 0
4 h (sc)
4 h (po)
not detectable
5 ( 1
not detectable
6 ( 1
not detectable
3 ( 0
pancreas
1
2
3
time (nmol/g wet weight) (nmol/g wet weight) (nmol/g wet weight)
1 h (sc)
1 h (po)
28 ( 13
13 ( 3
30 ( 5
25 ( 14
22 ( 17
9 ( 5
2 h (sc)
2 h (po)
6 ( 1
9 ( 5
30 ( 25
9 ( 2
30 ( 26
24 ( 11
In the kidney at 1 h postdrug, the tissue concentration trend
after po dosing was 1 ≈ 3 > 2. The renal concentration of the
2 was significantly lower than the parent 1 (p < 0.05). At 2 h,
the order was now 1 > 2 > 3. The renal concentration of 3
was significantly lower than 1 (p < 0.02). At 4 h, the level of
the parent 1 in the kidney was significantly greater than either
of the polyether analogues, p < 0.004 versus 2 and p < 0.005
versus 3 (Table 4).
The levels of all three of the chelators in the heart after po
dosing were quite low, <20 nmol/g wet weight at 1 h postdrug
and <10 nmol/g wet weight 2 h postdrug. By 4 h, only the
parent 1 was detectable.
In the pancreas at 1 h postdrug, the relative concentrations
of the chelators given po are 2 > 1 > 3. However, none of the
differences were significant. At 2 h, the pancreatic content of
all three of the ligands are within error of each other. At 4 h
postdrug, only 3 had detectable level.
In the plasma 1 h after po dosing, the relative drug levels are
2 > 1 ≈ 3. The concentration of the 4′-polyether 2 was
significantly greater than both the parent 1 (p < 0.009) and the
3′-polyether 3 (p < 0.02). At 2 h after po dosing, the relative
ligand concentration order was 2 > 3 ≈ 1, although none of
the differences were significant. At 4 h postdrug, the plasma
chelator concentrations were uniformly e5 µM (Table 4).
A comparison of how the tissue distribution of the individual
ligands is affected by route of administration is also notable. In
the case of ligand 1, the liver concentrations were significantly
higher in the po versus sc dosed animals 1 h (p < 0.001) and
4 h (sc)
4 h (po)
6 ( 3
13 ( 5
12 ( 5
6 ( 2
not detectable
not detectable
plasma
time
1 (µM)
2 (µM)
3 (µM)
1 h (sc)
1 h (po)
34 ( 7
31 ( 11
140 ( 29
64 ( 7
46 ( 3
26 ( 14
2 h (sc)
2 h (po)
13 ( 3
8 ( 3
16 ( 4
32 ( 19
24 ( 5
9 ( 3
4 h (sc)
4 h (po)
2 ( 0
5 ( 0
5 ( 3
2 ( 0
13 ( 2
2 ( 0
a The rodents (n ) 3/time point) were given a single 300 µmol/kg dose
of the chelators sc or po.
4 h (p < 0.005) postdrug. In the kidney, while the 1 and 2 h sc
versus po chelator concentrations were similar, the level of the
drug in the 4 h po dosed animals was significantly higher than
the sc dosed rats, p < 0.001. In the heart, the chelator content
was significantly higher in the sc versus po dosed animals at
1 h (p < 0.007); the levels were within error of each other at
2 h. However, by 4 h postdrug, the ligand was only detectable
in the heart of the po dosed rats. Route of administration did
not have any impact on pancreatic ligand content; all levels,
where detectable, were within error of each other (Table 4). In
the plasma, the chelator concentrations were significantly higher
in the po versus sc dosed animals only in the 4 h time point, p
< 0.001.

3920 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 13
Bergeron et al.
In the case of ligand 2, the liver and kidney concentrations
were significantly higher in the sc versus po dosed animals 1
and 4 h postdrug. The p-values for the liver (sc versus po) were
p < 0.001 (1 h) and p < 0.03 (4 h); the p-values for the kidney
were <0.001 (1 h) and <0.04 (4 h). However, the sc versus po
levels were within error of each other for the liver and kidney
at 2 h. Route of administration did not have any impact on
pancreatic ligand content; all levels were within error of each
other, where detectable. In the heart and plasma, the chelator
concentrations, where detectable, were significantly higher in
the sc versus po dosed animals only in the 1 h time point, p <
0.04 (heart) and p < 0.02 (plasma).
In the case of analogue 3, the concentration of the drug in
the liver was not affected by route of administration at the 1
and 2 h time points. However, the chelator content was
significantly higher in the liver of the sc versus po dosed animals
4 h postdrug, p < 0.003. In the kidney, the chelator content
was significantly greater for the sc versus po dosed animals at
all time points, p < 0.04, p < 0.01, and p < 0.001 for 1, 2, and
4 h, respectively. In the heart, the ligand content, where
detectable, was also significantly greater for the sc versus po
dosed animals at the 1 and 2 h time points, p < 0.05 and p <
0.009, respectively. Route of administration did not have any
impact on pancreatic ligand content; all levels were within error
of each other (Table 4). The chelator content in the plasma of
the sc versus po dosed animals was higher at the 2 and 4 h
time points, p < 0.007 and p > 0.006, respectively.
though, are hardly profound, in keeping with the high oral
bioavailability of these ligands.
Finally, and most important, are the data surrounding the
toxicity studies. Both the impact of the polyether side chain
and the dosing schedule (s.i.d. vs b.i.d.) were shown to have a
profound effect on nephrotoxicity. In the first study, non-iron-
loaded rodents were given 384 µmol/kg/day po of either 1, 2,
or 3 for 10 days and then sacrificed (Table 3). All histopatholo-
gies were normal except for the kidneys of rats treated with 1,
which showed mild nephrotoxicity.52 In a second trial, iron-
loaded rodents were given 119 µmol/kg po of 1 for 30 days
and sacrificed. No drug-related abnormalities were noted. When
unloaded rodents were given 63.3 µmol/kg po of 2, a dose which
clears the same amount of iron in the primates as 119 µmol/kg
of 1, for 30 days and sacrificed, all histopathologies were
normal.45 In a final experiment with 3, two sets of animals were
studied, one iron-loaded and one non-iron-loaded. In each
instance, groups of six animals were given either 75, 150, or
300 µmol/kg po of the ligand once daily for 28 days and then
sacrificed. Note that the higher doses were intentionally chosen
in an attempt to induce renal toxicity. Because of the number
of tissue samples and the results from the 10-day, 384 µmol/
kg/day experiments, we elected only to evaluate the histo-
pathologies of the kidneys and bone marrow; all were normal.
Certainly, the most astounding finding relates to dosing
schedules in the renal perfusion studies. When non-iron-
overloaded rodents were given a dose of 474 µmol/kg of 1 po
s.i.d. for 7 days, anesthetized, and their kidneys perfused and
fixed, light microscopy of the samples revealed moderate
damage to the proximal tubules, most notable, vacuolization.
However, when the same dose, 474 µmol/kg/day, was split in
half to 237 µmol/kg/day and given b.i.d. for 7 days, the
difference in proximal tubule damage was profoundly worse45
(Figure 5). Thus, b.i.d. dosing of this drug would not be a viable
plan. This strongly suggests that careful consideration must be
given as to how often chelators are dosed relative to nephro-
toxicity. However, when either 4′-polyether analogue 2 or 3′-
polyether 3 was given at 237 µmol/kg/day po b.i.d. (474 µmol/
kg/day) for 7 days, there were only minor architectural changes
in the proximal tubules (occasional pale, refractile inclusions
in the S1 segments), which were more prevalent in the 2 treated
rats, further underscoring the likely large therapeutic window
for these ligands.
Conclusion
The synthetic schemes for assembling 2 and 3 offer significant
improvements over the initial methodologies. Although prior
methods generated enough ligand to determine proof of
principle, because of low yields and separation problems, they
did not lend themselves to scale-up for the level of systematic
and inclusive biological studies required to assess the clinical
potential of the ligands.
While the ICE of the parent drug 1 in rodents was uniformly
poor at both evaluated 300 and 150 µmol/kg doses (e1.5%),
the polyethers 2 and 3 were significantly more efficient at all
doses (Table 1). However, there were rather profound differ-
ences in the ICE versus dose response curves for the two
polyether analogues in rodents (Figure 1). In both instances,
their deferration properties were shown to be saturable. The ICE
for 2 increased from 5.5 ( 1.9% to 21.7 ( 3.5%, a nearly 4-fold
difference, as the dose was decreased from 300 to 50 µmol/kg.
The ICE for 3 increased from 10.6 ( 4.4% to 20.7 ( 4.4%,
again on moving from 300 to 50 µmol/kg. However, the change
was not nearly as great with the 3′-polyether analogue 3, only
a 2-fold difference, suggesting greater flexibility in dosing
ranges. In addition, 3 was significantly more effective than the
4′-polyether analogue 2 when given po at 150 or 300 µmol/kg
(Table 1). The other notable difference between the two
polyether analogues can be found in their ferrokinetic profiles
(Figures 3 and 4). At all concentrations, ligand-induced defer-
ration is over more quickly with 2 (Figure 3) than for 3 (Figure
4). This data will serve as a useful guide for dose scheduling.
In both the rats and the primates, the po versus sc data for
all three ligands strongly suggest good oral bioavailability.
Although complete pharmacokinetic studies remain to be carried
out, the ICE values are essentially the same whether the drug
is given po or sc. The tissue distribution relative chelator levels
follow the order liver > kidney > plasma > pancreas > heart.
The sc versus po dosing data suggest generally higher concen-
trations early on with sc administered drug. The differences,
Experimental Section
Cebus apella monkeys were obtained from World Wide Primates
(Miami, FL). Male Sprague-Dawley rats were procured from
Harlan Sprague-Dawley (Indianapolis, IN). Ultrapure salts were
obtained from Johnson Matthey Electronics (Royston, UK). All
hematological and biochemical studies57 were performed by Antech
Diagnostics (Tampa, FL). Atomic absorption (AA) measurements
were made on a Perkin-Elmer model 5100 PC (Norwalk, CT).
Histopathological analysis was carried out by Florida Vet Path
(Bushnell, FL).
Cannulation of Bile Duct in Non-Iron-Overloaded Rats. The
cannulation has been described previously.57,58 Bile samples were
collected from male Sprague-Dawley rats (400-450 g) at 3 h
intervals for 24 h. The urine sample was taken at 24 h. Sample
collection and handling are as previously described.57,58
Iron Loading of Rats. Male Sprague-Dawley rats initially
weighing approximately 200-225 g were iron overloaded by the
intraperitoneal administration of iron dextran (Sigma, 100 mg of
iron/mL). The rats were given four doses of the iron (105 mg/kg/
dose) over a two week period on a Monday, Friday, Monday, Friday
schedule. After a two week equilibration period, the animals were
weighed again and their total iron burden was determined (total

Non-Nephrotoxic Desazadesferrithiocin Polyether Analogues
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 13 3921
mg of Fe/final body weight). The rats were then assigned to groups
so that the body weight and iron burden between groups were within
error of each other.
Iron Loading of C. apella Monkeys. The monkeys (3.5-4 kg)
were iron overloaded with iv iron dextran as specified in earlier
publications to provide about 500 mg of iron per kg of body
weight;59 the serum transferrin iron saturation rose to between 70
and 80%. At least 20 half-lives, 60 days,60 elapsed before any of
the animals were used in experiments evaluating iron-chelating
agents.
Primate Fecal and Urine Samples. Fecal and urine samples
were collected at 24 h intervals and processed as described
previously.57,58,61 Briefly, the collections began 4 days prior to the
administration of the test drug and continued for an additional 5
days after the drug was given. Iron concentrations were determined
by flame atomic absorption spectroscopy as presented in other
publications.58,62
Drug Preparation and Administration. In the iron clearing
experiments the rats were given a single 50, 150, or 300 µmol/kg
dose of the drugs po and/or sc. The compounds were administered
as (1) a solution in water, 2 300 µmol/kg dose only or (2) as the
monosodium salt of the compound of interest (prepared by addition
of the free acid to 1 equivalent of NaOH). The chelators were given
to the monkeys po and sc at a dose of 150 µmol/kg. The drugs
were prepared as for the rats; 2 was given po and sc as a solution
in water.
Calculation of Iron Chelator Efficiency. The theoretical iron
outputs of the chelators were generated on the basis of a 2:1 ligand:
iron complex. The efficiencies in the rats and monkeys were
calculated as set forth elsewhere.52 Data are presented as the mean
( the standard error of the mean; p-values were generated via a
one-tailed Student’s t-test in which the inequality of variances was
assumed; and a p-value of <0.05 was considered significant.
Toxicity Evaluation in Rodents. Male Sprague-Dawley rats
(250-300 g) were fasted overnight and were given 1, 2, or 3 po
by gavage once daily for 10 days at a dose of 384 µmol/kg/day.
This dose is equivalent to 100 mg/kg/day of the DFT sodium salt.
The animals were fed ∼3 h postdrug and had access to food for
5 h before being fasted overnight. The rats were euthanized 24 h
postdrug and extensive tissues were collected for histopathological
analysis.
The rats given 1 or 2 in the 30 day experiments were given the
drugs po at a dose that, in the primates, results in the excretion of
450 µg of Fe/kg, 119 µmol/kg/day and 63.3 µmol/kg/day, respec-
tively. Rats treated with 1 had been previously iron overloaded to
a level of 350 mg Fe/kg; the rodents treated with 2 for 30 days
were not iron-loaded. Iron-overloaded and non-iron-loaded rats were
given 3 po at doses of 75, 150, and 300 µmol/kg/day for 28 days.
Note that the doses chosen for 3 are approximately 1, 2, and 4
times the dose necessary to cause the excretion of 450 µg Fe/kg in
the primates. Because of the protracted nature of the 28 or 30 days
experiments, the animals were not fasted. To minimize food content
in the stomach, the drug was not given until the afternoon.
Additional animals served as age-matched controls. The rats were
euthanized 24 h postdrug, and extensive tissues were collected for
histopathological analysis.
A Comparison of the Impact of 1, 2, and 3 on Rodent
Kidneys. Male Sprague-Dawley rats (200-250 g) were housed
two to a cage and were fasted overnight. The rats were given 1, 2,
or 3 po twice daily at a dose of 237 µmol/kg/dose (474 µmol/kg/
day) for 7 days. The rats were fed approximately 3 h postdrug and
had access to food for 5 h before being fasted again. An additional
group of rodents was given 1 once daily at a dose of 474 µmol/
kg/day for 7 days. The single dose was given in the mornings; the
rats were fed and fasted as described for the b.i.d. dosed animals.
Untreated rodents served as age-matched controls.
One day postdrug, the rats were anesthetized with sodium
pentobarbital. The abdominal aorta was isolated and cleansed of
extraneous tissue and cannulated. The kidneys were perfusion-fixed
with glutaraldehyde in Tyrode’s buffer containing 3% PVP,
removed, and immersed in a glass vial containing fresh fixative at
room temperature. Four hours later, the kidneys were again rinsed
with Tyrode’s buffer. The tissue samples were placed in sodium
cacodylate. After two rinses with cacodylate buffer, the samples
were dehydrated with graded concentrations of ethyl alcohol and
infiltrated with propylene oxide overnight. Finally, they were
embedded in TAAB with DMP-30. The TAAB was polymerized
at 60 °C for 48 h.
Thick sections (500 nm) were cut from TAAB embedded tissue,
mounted onto glass slides, and stained with Toluidine Blue. The
tissue slices were photographed with a Zeiss Axioskop II at 400×
magnification.
Collection of Tissue Samples from Rodents: Tissue Distribution.
Male Sprague-Dawley rats (250-350 g) were given 1, 2, or 3 po
and sc at a dose of 300 µmol/kg. The compounds were administered
as (1) a solution in water, 2 (sc) or (2) as the monosodium salt of
the compound of interest (prepared by addition of the free acid to
1 equivalent of NaOH) 1, 2 (po), and 3. At times 1, 2, and 4 h
after dosing (n ) 3 rats per time point), the animals were euthanized
by exposure to CO2 gas. Blood was obtained via cardiac puncture
into vacutainers containing sodium citrate. The blood was centri-
fuged and the plasma separated for analysis. The liver, heart,
kidneys, and pancreas were then removed from the animals.
Tissue Analytical Methods. Tissue samples from animals treated
with 1-3 were prepared for HPLC analysis as previously
described.45,46 Analytical separation was performed on a Discovery
RP Amide C16 HPLC system with UV detection at 310 nm as
discussed previously.63,64 Mobile phase and chromatographic
conditions were as follows: solvent A, 5% CH3CN/95% buffer;
solvent B, 60% CH3CN/40% buffer.
The concentrations were calculated from the peak area fitted to
calibration curves by nonweighted least-squares linear regression
with Rainin Dynamax HPLC Method Manager software (Rainin
Instrument Co.). The method had a detection limit of 0.25 µM and
was reproducible and linear over a range of 1-1000 µM.
Tissue distribution data are presented as the mean; p-values were
generated via a one-tailed student’s t-test, in which the inequality
of variances was assumed and a p-value of <0.05 was considered
significant.
Synthetic Methods. Reagents were purchased from Aldrich
Chemical Co. (Milwaukee, WI), and Fisher Optima-grade solvents
were routinely used. Reactions were run under a nitrogen atmo-
sphere, and organic extracts were dried with sodium sulfate. Silica
gel 40-63 from SiliCycle, Inc. (Quebec City, QC, Canada) was
used for column chromatography. Glassware that was presoaked
in 3 N HCl for 15 min, washed with distilled water and distilled
EtOH, and oven-dried was used in the isolation of 2 and 3. Optical
rotations were run at 589 nm (sodium D line) utilizing a Perkin-
Elmer 341 polarimeter, with c being concentration in grams of
compound per 100 mL of CHCl3 solution. Chemical shifts (δ) for
1H NMR spectra at 400 MHz are given in parts per million
downfield from tetramethylsilane for CDCl3 (not indicated) or
sodium 3-(trimethylsilyl)propionate-2,2,3,3-d4 for D2O. Chemical
shifts (δ) for 13C NMR spectra at 100 MHz are given in parts per
million referenced to 1,4-dioxane (δ 67.19) in D2O or to the residual
solvent resonance in CDCl3 (δ 77.16). Coupling constants (J) are
in hertz. Elemental analyses were performed by Atlantic Microlabs
(Norcross, GA).
(S)-4,5-Dihydro-2-[2-hydroxy-4-(3,6,9-trioxadecyloxy)phenyl]-4-
methyl-4-thiazolecarboxylic Acid (2). A solution of 50% (w/w)
NaOH (10.4 mL, 199 mmol) in CH3OH (90 mL) was added to 6
(6.54 g, 15.3 mmol) in CH3OH (200 mL) with ice bath cooling.
The reaction mixture was stirred at room temperature for 16 h,
and the bulk of the solvent was removed by rotary evaporation.
The residue was treated with dilute NaCl (150 mL) and was
extracted with ether (3 × 150 mL). The basic aqueous phase was
cooled in ice, acidified with 2 N HCl to a pH ≈ 2, and extracted
with EtOAc (4 × 100 mL). The EtOAc extracts were washed with
saturated NaCl (200 mL) and were concentrated in vacuo. Drying
under high vacuum furnished 5.67 g of 245 (93%) as an orange
1
oil: [R]25 +53.1° (c 0.98). H NMR (D2O) δ: 1.76 (s, 3 H), 3.35
(s, 3 H), 3.54-3.61 (m, 3 H), 3.64-3.72 (m, 4 H), 3.74-3.78 (m,

3922 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 13
Bergeron et al.
2 H), 3.90-3.94 (m, 2 H), 3.96 (d, 1 H, J ) 12.0), 4.25-4.29 (m,
2 H), 6.53 (d, 1 H, J ) 2.4), 6.64 (dd, 1 H, J ) 9.0, 2.2), 7.61 (d,
1 H, J ) 9.2). 13C NMR (D2O) δ: 23.65, 39.56, 58.65, 68.34, 69.33,
70.07, 70.18, 70.44, 71.62, 77.58, 102.11, 106.72, 109.66, 134.67,
161.27, 167.07, 176.86, 180.70. Anal. (C18H25NO7S) C, H, N.
(S)-4,5-Dihydro-2-[2-hydroxy-3-(3,6,9-trioxadecyloxy)phenyl]-4-
methyl-4-thiazolecarboxylic Acid (3). Compound 8 (7.63 g, 27.1
solely the responsibility of the authors and does not necessarily
represent the official views of the National Institute of Diabetes
and Digestive and Kidney Diseases or the National Institutes
of Health. We thank Jill Verlander Reed and Hua Yao for the
perfusion of the rodent kidneys, Elizabeth M. Nelson, Tanaya
Lindstrom, and Katie Ratliff-Thompson for their technical
assistance, and Carrie A. Blaustein for her editorial and
organizational support.
mmol), degassed 0.1 M pH 5.95 phosphate buffer (200 mL),65
9
(6.98 g, 40.7 mmol), and NaHCO3 (4.33 g, 51.5 mmol, in portions)
were successively added to distilled, degassed CH3OH (200 mL).
The reaction mixture, pH 6.2-6.6, was heated at 70 °C for 72 h.
After cooling to room temperature, the bulk of the solvent was
removed by rotary evaporation. The residue was dissolved in 8%
NaHCO3 (200 mL) and was extracted with CHCl3 (3 × 100 mL).
The aqueous portion was cooled in an ice-water bath, acidified to
pH ≈ 1 with 5 N HCl, and extracted with EtOAc (4 × 100 mL).
The EtOAc extracts were washed with saturated NaCl and were
concentrated in vacuo. Drying under high vacuum furnished 9.74 g
References
(1) Raymond, K. N.; Carrano, C. J. Coordination Chemistry and Microbial
Iron Transport. Acc. Chem. Res. 1979, 12, 183–190.
(2) Bernier, G.; Girijavallabhan, V.; Murray, A.; Niyaz, N.; Ding, P.;
Miller, M. J.; Malouin, F. Desketoneoenactin-Siderophore Conjugates
for Candida: Evidence of Iron Transport-Dependent Species Selectiv-
ity. Antimicrob. Agents Chemother. 2005, 49, 241–248.
(3) Walz, A. J.; Mollmann, U.; Miller, M. J. Synthesis and Studies of
Catechol-Containing Mycobactin S and T Analogs. Org. Biomol.
Chem. 2007, 5, 1621–1628.
1
of 346 (90%) as an orange oil: [R]20 +61.9° (c 1.55). H NMR
(D2O) δ: 1.77 (s, 3 H), 3.35 (s, 3 H), 3.56-3.62 (m, 3 H), 3.64-3.73
(m, 4 H), 3.75-3.79 (m, 2 H), 3.92-3.96 (m, 2 H), 3.99 (d, 1 H,
J ) 11.6), 4.25-4.31 (m, 2 H), 6.99 (t, 1 H, J ) 8.2), 7.26-7.33
(m, 2 H). 13C NMR δ 24.52, 39.93, 59.07, 69.04, 69.83, 70.49,
70.64, 70.86, 71.97, 83.21, 116.33, 117.94, 118.50, 122.80, 147.67,
150.24, 172.38, 176.10. HRMS m/z calcd for C18H26NO7S, 400.1429
(M + H); found, 400.1413.
(4) Yuan, W. M.; Gentil, G. D.; Budde, A. D.; Leong, S. A. Characteriza-
tion of the Ustilago Maydis Sid2 Gene, Encoding a Multidomain
Peptide Synthetase in the Ferrichrome Biosynthetic Gene Cluster. J.
Bacteriol. 2001, 183, 4040–4051.
(5) Byers, B. R.; Arceneaux, J. E. Microbial Iron Transport: Iron
Acquisition by Pathogenic Microorganisms. Met. Ions Biol. Syst. 1998,
35, 37–66.
(6) Kalinowski, D. S.; Richardson, D. R. The Evolution of Iron Chelators
for the Treatment of Iron Overload Disease and Cancer. Pharmacol.
ReV. 2005, 57, 547–583.
(7) Bergeron, R. J. Iron: A Controlling Nutrient in Proliferative Processes.
Trends Biochem. Sci. 1986, 11, 133–136.
Ethyl (S)-4,5-Dihydro-2-[2-hydroxy-4-(3,6,9-trioxadecyloxy)phe-
nyl]-4-methyl-4-thiazolecarboxylate (6). Flame activated K2CO3
(5.05 g, 36.6 mmol) followed by 5 (11.11 g, 34.9 mmol) in acetone
(50 mL) were added to 448 (9.35 g, 33.2 mmol) in acetone (300
mL). The reaction mixture was heated at reflux for 3 days.
Additional K2CO3 (4.59 g, 33.2 mmol) and 5 (2.12 g, 6.65 mmol)
in acetone (5 mL) were added, and the reaction mixture was heated
at reflux for 1 day. After cooling to room temperature, solids were
filtered and the solvent was removed by rotary evaporation. The
residue was dissolved in 1:1 0.5 M citric acid/saturated NaCl (320
mL) and was extracted with EtOAc (3 × 150 mL). The combined
organic extracts were washed with distilled H2O (200 mL) and
saturated NaCl (200 mL) and were concentrated in vacuo. Purifica-
tion using flash column chromatography eluting with 50% EtOAc/
petroleum ether generated 12.0 g of 645 (84%) as an oil: [R]23 +40.2
(c 1.09). 1H NMR δ: 1.30 (t, 3 H, J ) 7.2) 1.66 (s, 3 H), 3.19 (d,
1 H, J ) 11.2), 3.38 (s, 3 H), 3.54-3.57 (m, 2 H), 3.64-3.70 (m,
4 H), 3.72-3.76 (m, 2 H), 3.81-3.88 (m, 3 H), 4.12-4.17 (m, 2
H), 4.20-4.28 (m, 2 H), 6.46 (dd, 1 H, J ) 8.8, 2.4), 6.49 (d, 1 H,
J ) 2.4), 7.28 (d, 1 H, J ) 8.4), 12.69 (s, 1 H). 13C NMR δ 14.21,
24.58, 39.94, 59.17, 62.01, 67.65, 69.60, 70.69, 70.76, 70.98, 72.03,
83.22, 101.51, 107.41, 109.98, 131.77, 161.27, 163.09, 170.90,
172.95. Anal. (C20H29NO7S) C, H, N.
(8) Theil, E. C.; Huynh, B. H. Ferritin Mineralization: Ferroxidation and
Beyond. J. Inorg. Biochem. 1997, 67, 30.
(9) Ponka, P.; Beaumont, C.; Richardson, D. R. Function and Regulation
of Transferrin and Ferritin. Semin. Hematol. 1998, 35, 35–54.
(10) Ponka, P. Physiology and Pathophysiology of Iron Metabolism:
Implications for Iron Chelation Therapy in Iron Overload. In The
DeVelopment of Iron Chelators for Clinical Use; Bergeron, R. J.,
Brittenham, G. M. Eds. CRC: Boca Raton, FL, 1994; pp 1-29.
(11) Olivieri, N. F.; Brittenham, G. M. Iron-Chelating Therapy and the
Treatment of Thalassemia. Blood 1997, 89, 739–761.
(12) Vichinsky, E. P. Current Issues with Blood Transfusions in Sickle
Cell Disease. Semin. Hematol. 2001, 38, 14–22.
(13) Kersten, M. J.; Lange, R.; Smeets, M. E.; Vreugdenhil, G.; Roozendaal,
K. J.; Lameijer, W.; Goudsmit, R. Long-Term Treatment of Trans-
fusional Iron Overload with the Oral Iron Chelator Deferiprone (L1):
A Dutch Multicenter Trial. Ann. Hematol. 1996, 73, 247–252.
(14) Conrad, M. E.; Umbreit, J. N.; Moore, E. G. Iron Absorption and
Transport. Am. J. Med. Sci. 1999, 318, 213–229.
(15) Lieu, P. T.; Heiskala, M.; Peterson, P. A.; Yang, Y. The Roles of
Iron in Health and Disease. Mol. Aspects Med. 2001, 22, 1–87.
(16) Angelucci, E.; Brittenham, G. M.; McLaren, C. E.; Ripalti, M.;
Baronciani, D.; Giardini, C.; Galimberti, M.; Polchi, P.; Lucarelli, G.
Hepatic Iron Concentration and Total Body Iron Stores in Thalassemia
Major. N. Engl. J. Med. 2000, 343, 327–331.
2-Hydroxy-3-(3,6,9-trioxadecyloxy)benzonitrile (8). Compound
749 (5.3 g, 39.2 mmol) was added to a suspension of 60% NaH
(3.13 g, 78.2 mmol) in DMSO (60 mL) using oven-dried glassware.
After the reaction mixture was stirred at room temperature for 1 h,
5 (12.49 g, 39.22 mmol) in DMSO (25 mL) was introduced. After
24 h of stirring at room temperature, the reaction mixture was
poured with stirring into cold water (100 mL) and was extracted
with CHCl3 (3 × 100 mL). The aqueous phase was acidified to pH
≈ 1 with 6 N HCl and was extracted with CHCl3 (5 × 60 mL).
The latter CHCl3 extracts were concentrated in vacuo. Purification
using column chromatography by gravity eluting with 10% CH3OH/
CHCl3 gave 7.74 g of 8 (70%) as an oil. 1H NMR δ: 3.40 (s, 3 H),
3.58-3.62 (m, 2 H), 3.65-3.73 (m, 4 H), 3.75-3.78 (m, 2 H),
3.83-3.87 (m, 2 H), 4.14-4.18 (m, 2 H), 6.79-6.85 (m, 1 H),
(17) Bonkovsky, H. L.; Lambrecht, R. W. Iron-Induced Liver Injury. Clin.
LiVer Dis. 2000, 4, 409–429.
(18) Pietrangelo, A. Mechanism of Iron Toxicity. AdV. Exp. Med. Biol.
2002, 509, 19–43.
(19) Cario, H.; Holl, R. W.; Debatin, K. M.; Kohne, E. Insulin Sensitivity
and ꢀ-Cell Secretion in Thalassaemia Major with Secondary Haemo-
chromatosis: Assessment by Oral Glucose Tolerance Test. Eur.
J. Pediatr. 2003, 162, 139–146.
(20) Wojcik, J. P.; Speechley, M. R.; Kertesz, A. E.; Chakrabarti, S.; Adams,
P. C. Natural History of C282y Homozygotes for Hemochromatosis.
Can. J. Gastroenterol. 2002, 16, 297–302.
(21) Brittenham, G. M.; Griffith, P. M.; Nienhuis, A. W.; McLaren, C. E.;
Young, N. S.; Tucker, E. E.; Allen, C. J.; Farrell, D. E.; Harris, J. W.
Efficacy of Deferoxamine in Preventing Complications of Iron
Overload in Patients with Thalassemia Major. N. Engl. J. Med. 1994,
331, 567–573.
(22) Brittenham, G. M. Disorders of Iron Metabolism: Iron Deficiency and
Overload. In Hematology: Basic Principles and Practice, 3rd ed.;
Hoffman, R., Benz, E. J., Shattil, S. J., Furie, B., Cohen, H. J.,
Silberstein, L. E., McGave, P. Eds. Churchill Livingstone: New York,
2000; pp 397-428.
7.09 (dd, 1 H, J ) 7.8, 1.6), 7.15-7.18 (m, 1 H), 8.6 (s, 1 H). 13
C
NMR δ: 57.25, 67.76, 67.85, 68.79, 68.92, 69.06, 70.36, 98.38,
115.44, 116.55, 118.51, 123.13, 145.98, 149.46. HRMS m/z calcd
for C14H20NO5, 282.1341 (M + H); found, 282.1328.
(23) Zurlo, M. G.; De Stefano, P.; Borgna-Pignatti, C.; Di Palma, A.; Piga,
A.; Melevendi, C.; Di Gregorio, F.; Burattini, M. G.; Terzoli, S.
Survival and Causes of Death in Thalassaemia Major. Lancet 1989,
2, 27–30.
Acknowledgment. The project described was supported by
grant number R37DK049108 from the National Institute of
Diabetes and Digestive and Kidney Diseases. The content is

Non-Nephrotoxic Desazadesferrithiocin Polyether Analogues
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 13 3923
(24) Graf, E.; Mahoney, J. R.; Bryant, R. G.; Eaton, J. W. Iron-Catalyzed
Hydroxyl Radical Formation. Stringent Requirement for Free Iron
Coordination Site. J. Biol. Chem. 1984, 259, 3620–3624.
(25) Halliwell, B. Free Radicals and Antioxidants: A Personal View. Nutr.
ReV. 1994, 52, 253–265.
(46) Bergeron, R. J.; Wiegand, J.; Bharti, N.; Singh, S.; Rocca, J. R. Impact
of the 3,6,9-Trioxadecyloxy Group on Desazadesferrithiocin Analogue
Iron Clearance and Organ Distribution. J. Med. Chem. 2007, 50, 3302–
3313.
(47) Bergeron, R. J.; Wiegand, J.; McManis, J. S.; Bharti, N. The Design,
Synthesis, and Evaluation of Organ-Specific Iron Chelators. J. Med.
Chem. 2006, 49, 7032–7043.
(48) Bergeron, R. J.; Bharti, N.; Wiegand, J.; McManis, J. S.; Yao, H.;
Prokai, L. Polyamine-Vectored Iron Chelators: The Role of Charge.
J. Med. Chem. 2005, 48, 4120–4137.
(49) Bergeron, R. J.; Wiegand, J.; McManis, J. S.; Weimar, W. R.; Park,
J.-H.; Eiler-McManis, E.; Bergeron, J.; Brittenham, G. M. Partition-
Variant Desferrithiocin Analogues: Organ Targeting and Increased Iron
Clearance. J. Med. Chem. 2005, 48, 821–831.
(50) Kocian, O.; Chiu, K. W.; Demeure, R.; Gallez, B.; Jones, C. J.;
Thornback, J. R. Synthesis and Characterization of New Polyethyl-
eneoxy Substituted Salicylaldimine Schiff Bases and Some Corre-
sponding Reduced Tetra- and Penta-Aza Ligands and Their Gadolin-
ium(III) Complexes: New Potential Contrast Agents in Magnetic
Resonance Imaging. J. Chem. Soc., Perkin Trans. 1 1994, 527–534.
(51) Bergeron, R. J.; Wiegand, J.; McManis, J. S.; Bussenius, J.; Smith,
R. E.; Weimar, W. R. Methoxylation of Desazadesferrithiocin
Analogues: Enhanced Iron Clearing Efficiency. J. Med. Chem. 2003,
46, 1470–1477.
(52) Bergeron, R. J.; Wiegand, J.; McManis, J. S.; McCosar, B. H.; Weimar,
W. R.; Brittenham, G. M.; Smith, R. E. Effects of C-4 Stereochemistry
and C-4′ Hydroxylation on the Iron Clearing Efficiency and Toxicity
of Desferrithiocin Analogues. J. Med. Chem. 1999, 42, 2432–2440.
(53) Bergeron, R. J.; Wiegand, J.; McManis, J. S.; Weimar, W. R.; Huang,
G. Structure-Activity Relationships among Desazadesferrithiocin
Analogues. AdV. Exp. Med. Biol. 2002, 509, 167–184.
(54) Porter, J. B.; Morgan, J.; Hoyes, K. P.; Burke, L. C.; Huehns, E. R.;
Hider, R. C. Relative Oral Efficacy and Acute Toxicity of Hydroxy-
pyridin-4-one Iron Chelators in Mice. Blood 1990, 76, 2389–2396.
(55) Porter, J. B.; Hoyes, K. P.; Abeysinghe, R. D.; Brooks, P. N.; Huehns,
E. R.; Hider, R. C. Comparison of the Subacute Toxicity and Efficacy
of 3-Hydroxypyridin-4-one Iron Chelators in Overloaded and Non-
overloaded Mice. Blood 1991, 78, 2727–2734.
(56) Brittenham, G. M. Pyridoxal Isonicotinoyl Hydrazone (PIH): Effective
Iron Chelation after Oral Administration. Ann. N.Y. Acad. Sci. 1990,
612, 315–326.
(57) Bergeron, R. J.; Streiff, R. R.; Creary, E. A.; Daniels, R. D.; King,
W.; Luchetta, G.; Wiegand, J.; Moerker, T.; Peter, H. H. A Compara-
tive Study of the Iron-Clearing Properties of Desferrithiocin Analogues
with Desferrioxamine B in a Cebus Monkey Model. Blood 1993, 81,
2166–2173.
(58) Bergeron, R. J.; Streiff, R. R.; Wiegand, J.; Vinson, J. R. T.; Luchetta,
G.; Evans, K. M.; Peter, H.; Jenny, H.-B. A Comparative Evaluation
of Iron Clearance Models. Ann. N.Y. Acad. Sci. 1990, 612, 378–393.
(59) Bergeron, R. J.; Streiff, R. R.; Wiegand, J.; Luchetta, G.; Creary, E. A.;
Peter, H. H. A Comparison of the Iron-Clearing Properties of 1,2-
Dimethyl-3-Hydroxypyrid-4-one, 1,2-Diethyl-3-Hydroxypyrid-4-one,
and Deferoxamine. Blood 1992, 79, 1882–1890.
(60) Wood, J. K.; Milner, P. F.; Pathak, U. N. The Metabolism of Iron-
Dextran Given as a Total-Dose Infusion to Iron Deficient Jamaican
Subjects. Br. J. Hamaetol. 1968, 14, 119–129.
(61) Bergeron, R. J.; Wiegand, J.; Brittenham, G. M. HBED: A Potential
Alternative to Deferoxamine for Iron-Chelating Therapy. Blood 1998,
91, 1446–1452.
(26) Halliwell, B. Iron, Oxidative Damage, and Chelating Agents. In The
DeVelopment of Iron Chelators for Clinical Use; Bergeron, R. J.,
Brittenham, G. M. Eds.; CRC: Boca Raton, FL, 1994; pp 33-56.
(27) Koppenol, W. Kinetics and Mechanism of the Fenton Reaction:
Implications for Iron Toxicity. In Iron Chelators: New DeVelopment
Strategies; Badman, D. G., Bergeron, R. J., Brittenham, G. M. Eds.;
Saratoga: Ponte Vedra Beach, FL, 2000; pp 3-10.
(28) Babbs, C. F. Oxygen Radicals in Ulcerative Colitis. Free Radical Biol.
Med. 1992, 13, 169–181.
(29) Hazen, S. L.; d’Avignon, A.; Anderson, M. M.; Hsu, F. F.; Heinecke,
J. W. Human Neutrophils Employ the Myeloperoxidase-Hydrogen
Peroxide-Chloride System to Oxidize R-Amino Acids to a Family of
Reactive Aldehydes. Mechanistic Studies Identifying Labile Intermedi-
ates Along the Reaction Pathway. J. Biol. Chem. 1998, 273, 4997–
5005.
(30) Balla, J.; Vercellotti, G. M.; Jeney, V.; Yachie, A.; Varga, Z.; Jacob,
H. S.; Eaton, J. W.; Balla, G. Heme, Heme Oxygenase, and Ferritin:
How the Vascular Endothelium Survives (and Dies) in an Iron-Rich
Environment. Antioxid. Redox. Signal 2007, 9, 2119–2137.
(31) Bergeron, R. J.; McManis, J. S.; Weimar, W. R.; Wiegand, J.; Eiler-
McManis, E. Iron Chelators and Therapeutic Uses. In Burger’s
Medicinal Chemistry, 6th ed.; Abraham, D. A. Ed.; Wiley: New York,
2003; Vol. III, pp 479-561.
(32) Brittenham, G. M. Iron Chelators and Iron Toxicity. Alcohol 2003,
30, 151–158.
(33) Hoffbrand, A. V.; Al-Refaie, F.; Davis, B.; Siritanakatkul, N.; Jackson,
B. F. A.; Cochrane, J.; Prescott, E.; Wonke, B. Long-Term Trial of
Deferiprone in 51 Transfusion-Dependent Iron Overloaded Patients.
Blood 1998, 91, 295–300.
(34) Olivieri, N. F. Long-Term Therapy with Deferiprone. Acta Haematol.
1996, 95, 37–48.
(35) Olivieri, N. F.; Brittenham, G. M.; McLaren, C. E.; Templeton, D. M.;
Cameron, R. G.; McClelland, R. A.; Burt, A. D.; Fleming, K. A. Long-
Term Safety and Effectiveness of Iron-Chelation Therapy with
Deferiprone for Thalassemia Major. N. Engl. J. Med. 1998, 339, 417–
423.
(36) Richardson, D. R. The Controversial Role of Deferiprone in the
Treatment of Thalassemia. J. Lab. Clin. Med. 2001, 137, 324–329.
(37) Nisbet-Brown, E.; Olivieri, N. F.; Giardina, P. J.; Grady, R. W.;
Neufeld, E. J.; Sechaud, R.; Krebs-Brown, A. J.; Anderson, J. R.;
Alberti, D.; Sizer, K. C.; Nathan, D. G. Effectiveness and Safety of
ICL670 in Iron-Loaded Patients with Thalassaemia: A Randomised,
Double-Blind, Placebo-Controlled, Dose-Escalation Trial. Lancet 2003,
361, 1597–1602.
(38) Galanello, R.; Piga, A.; Alberti, D.; Rouan, M.-C.; Bigler, H.; Se´chaud,
R. Safety, Tolerability, and Pharmacokinetics of ICL670, a New Orally
Active Iron-Chelating Agent in Patients with Transfusion-Dependent
Iron Overload Due to ꢀ-Thalassemia. J. Clin. Pharmacol. 2003, 43,
565–572.
(39) Cappellini, M. D. Iron-Chelating Therapy with the New Oral Agent
ICL670 (Exjade). Best Pract. Res. Clin. Haematol. 2005, 18, 289–
98.
(40) Exjade prescribing information: http://www.pharma.us.novartis.com/
product/pi/pdf/exjade.pdf; (accessed December 2007).
(41) Bickel, H.; Hall, G. E.; Keller-Schierlein, W.; Prelog, V.; Vischer,
E.; Wettstein, A. Metabolic Products of Actinomycetes. XXVII.
Constitutional Formula of Ferrioxamine B. HelV. Chim. Acta 1960,
43, 2129–2138.
(42) Pippard, M. J. Desferrioxamine-Induced Iron Excretion in Humans.
Bailliere’s Clin. Haematol. 1989, 2, 323–343.
(62) Bergeron, R. J.; Wiegand, J.; Wollenweber, M.; McManis, J. S.; Algee,
S. E.; Ratliff-Thompson, K. Synthesis and Biological Evaluation of
Naphthyldesferrithiocin Iron Chelators. J. Med. Chem. 1996, 39, 1575–
1581.
(43) Giardina, P. J.; Grady, R. W. Chelation Therapy in ꢀ-Thalassemia:
An Optimistic Update. Semin. Hematol. 2001, 38, 360–366.
(44) Galanello, R.; Forni, G.; Jones, A.; Kelly, A.; Willemsen, A.; He, X.;
Johnston, A.; Fuller, D.; Donovan, J.; Piga, A. A Dose Escalation
Study of the Pharmacokinetics, Safety & Efficacy of Deferitrin, an
Oral Iron Chelator in Beta Thalassaemia Patients. ASH Annu. Meet.
Abstr. 2007, 110, 2669.
(45) Bergeron, R. J.; Wiegand, J.; McManis, J. S.; Vinson, J. R. T.; Yao,
H.; Bharti, N.; Rocca, J. R. (S)-4,5-Dihydro-2-(2-Hydroxy-4-Hydrox-
yphenyl)-4-Methyl-4-Thiazolecarboxylic Acid Polyethers: A Solution
to Nephrotoxicity. J. Med. Chem. 2006, 49, 2772–2783.
(63) Bergeron, R. J.; Wiegand, J.; Weimar, W. R.; McManis, J. S.; Smith,
R. E.; Abboud, K. A. Iron Chelation Promoted by Desazadesferrithio-
cin Analogues: An Enantioselective Barrier. Chirality 2003, 15, 593–
599.
(64) Bergeron, R. J.; Wiegand, J.; Ratliff-Thompson, K.; Weimar, W. R.
The Origin of the Differences in (R)- and (S)-Desmethyldesferrithiocin:
Iron-Clearing Properties. Ann. N.Y. Acad. Sci. 1998, 850, 202–216.
(65) Gomori, G. Preparation of Buffers for Use in Enzyme Studies. Methods
Enzymol. 1955, 1, 138–146.
JM800154M