Benzoxaborole antimalarial agents. Part 3: Design and syntheses of (carboxy-13C-3,3-2H2)-labeled and (3- 14C)-labeled 7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1- benzoxaboroles

Article abstract of DOI:10.1002/jlcr.2926

Two new isotopically labeled compounds, (carboxy-¹³C-3,3- ²H₂)-7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1- benzoxaborole (2) and (3-¹⁴C)-7-(2-carboxyethyl)-1,3-dihydro-1- hydroxy-2,1-benzoxaborole (3), were designed and synthesized to support the preclinical development studies of a potential new antimalarial agent, 7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole (1). Copyright

Full text of DOI:10.1002/jlcr.2926

Research Article
Received 03 February 2012,
Revised 01 April 2012,
Accepted 05 April 2012
Published online in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/jlcr.2926
Benzoxaborole antimalarial agents. Part 3:
design and syntheses of (carboxy-13C-3,3-2H2)-
labeled and (3-14C)-labeled 7-(2-carboxyethyl)-
1,3-dihydro-1-hydroxy-2,1-benzoxaboroles
a
a
a
a
*
Yong-Kang Zhang, Jacob J. Plattner, Eric E. Easom, Liang Liu,
Daniel M. Retz,b Min Ge,c and Hui-Hong Zhouc
Two new isotopically labeled compounds, (carboxy-13C-3,3-2H2)-7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole
(2) and (3-14C)-7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole (3), were designed and synthesized to support
the preclinical development studies of a potential new antimalarial agent, 7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,
1-benzoxaborole (1).
Keywords: isotopically labeled synthesis; carbon-14; deuterium; benzoxaborole; antimalarial agent; Plasmodium falciparum
including the trypanosome, Trypanosoma brucei and the
Introduction
filarial worm, Brugia malayi. As an extension of this
Malaria is a disease that is caused by parasites of the
work, we have screened our boron-containing compound
plasmodium genera that are transmitted from person to
person through the bites of infected mosquitoes and can
collection in a whole cell assay against the malaria parasite
P. falciparum. This screening campaign identified a number
be transmitted to people of all ages. About half of the world’s
of potent hits under 1 mM, including 7-(2-carboxyethyl)-1,
population is at risk of malaria causing about 216 million
clinical cases and 655,000 deaths every year, 85% of
which occur in children younger than 5 years. Malaria traps
families and communities in a downward spiral of poverty,
3-dihydro-1-hydroxy-2,1-benzoxaborole (1 in Figure 1) with
an IC50 of 26 nM.6 With its excellent antimalarial activity,
drug-like property, and in vivo pharmacological results tested,
this compound has been selected for preclinical develop-
disproportionately affecting marginalized and poor people
ment studies. During the preclinical development process,
(carboxy-13C-3,3-2H2)-7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,
in the poorest countries who cannot afford treatment or
have limited access to health care. Over the past decade,
artemisinin-based combination therapies have brought new
hope to the fight against malaria, although resistance of
malaria parasites to old drugs such as chloroquine is already
1-benzoxaborole (2 in Figure 1), a stable isotope-labeled
compound of 1, was needed for use as an internal standard
in LC/MS/MS (liquid chromatography/mass/mass) bioanalysis
to support the pharmacokinetic study of 1. And (3-14C)-7-(2-
widespread. However, it has been reported that Plasmodium
carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole (3 in
falciparum parasites in the border region of western
Figure 1), a radioactive isotope-labeled compound of 1, was
Cambodia and eastern Thailand have shown evidence of
resistance to artemisinins. This region has historically given
rise to other drug-resistant strains of P. falciparum that
also required for an in vivo mass balance study of 1. The pre-
sent article describes the design and chemical syntheses of
these two compounds.
eventually spread through Asia to Africa. A deterioration of
the susceptibility of P. falciparum to artemisinins and the
possibility that resistance will spread beyond the Greater
Mekong Subregion are real threats to global strategies for
aAnacor Pharmaceuticals, Inc., 1020 E. Meadow Circle, Palo Alto, CA 94303, USA
the control and elimination of malaria. Therefore, there is an
urgent need to enhance the pipeline of antimalarial medica-
tions with novel chemical structures that are counterresistant
bMoravek Biochemicals, Inc., 577 Mercury Lane, Brea, CA 92821, USA
cAcesys Pharmatech, Inc., Science and Technology Building 24B, 5 Xing Mo Fan
Road, Nanjing 21009, China
and that are safe and easy for use in the most vulnerable
populations.1
Previously, we have reported a class of boron-containing
*Correspondence to: Yong-Kang Zhang, Anacor Pharmaceuticals, Inc., 1020 E.
Meadow Circle, Palo Alto, CA 94303, USA.
E-mail: yzhang@anacor.com
compounds, called benzoxaboroles, with selective activity
against fungi,2 bacteria,3 inflammation4, and parasites,5
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Y.-K. Zhang et al.
reduce the double bond to afford the ester intermediate 8. This
compound was further hydrolyzed to give the final compound 2.
Design and synthesis of (3-14C)-7-(2-carboxyethyl)-1,3-
dihydro-1-hydroxy-2,1-benzoxaborole (3)
The planned mass balance study of 1 needs a radiolabeled
compound that is to be used to mix with the material of 1, known
as ‘cold material’, for orally dosing rodents to locate the where-
abouts of compound 1 and its metabolites in the testing animals.
The radioactive isotope serves as a ‘hot trace’ to be tracked by the
analytical instrument for the mass balance analysis, and therefore,
it may best serve the purpose if the ‘hot trace’ remains in the
parent compound and metabolites during the in vivo biological
processes of oral absorption, distribution in body, metabolism,
and excretion. A preliminary analysis with Meteor metabolism soft-
ware (Lhasa Limited, Leeds, UK) predicted that the carboxyethyl
side chain of (2-carboxyethyl)benzene (9) may partially breakdown
to give benzoic acid (10) as shown in Figure 2. Unfortunately, this
metabolic pathway eliminates a potential cost-saving synthesis of
radiolabeling the carboxylic carbon of compound 1. Subsequently,
we chose to radiolabel the three-position carbon of the compound
(3) as shown in Figure 1.
The chemistry for the synthesis of 3 is illustrated in Scheme 2.
Bromination of 11 generated the bromo compound 12 of which
the amino group was converted to iodo atom of 13. One of the
key steps of this radiolabel synthesis was the Grignard reaction
from 13 to introduce the radioisotope onto 14 with the use of
freshly generated low cost radioactive reagent 14CO2. After the
esterification of the acid 14 to the ester 15, the bromo group in
15 was catalytically boronylated to generate the pinancolatoboron
of 16. Reduction of the ester in 16 generated the alcohol group
that was spontaneously cyclized by connecting to the boron to
form the oxaborole ring of 17 under aqueous acidic condition. It
was critical to purify and isolate compound 17 because structure
identification was difficult with compounds 15 and 16 owing to
poor MS ionization and complex NMR spectra. Reduction of the
cyano group of 17 provided the aldehyde compound 18 that
underwent a Wittig reaction to generate 19 with an extended side
chain. Reduction of the double bond in 19 and subsequent hydro-
lysis of the ester 20 afforded the final 14C-labeled compound 3.
Figure 1. Structures of 7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole
(1), (carboxy-13C-3,3-2H2)-7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole
(2), and (3-14C)-7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole (3).
Results and discussion
Design and synthesis of (carboxy-13C-3,3-2H2)-
7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole (2)
Because the purpose of synthesizing a stable isotope-labeled com-
pound is to provide an internal reference standard for LC/MS/MS
analysis to qualify 1 in biological samples, it is needed to have
3 Da plus of the standard as compared with 1 for sufficient mass
resolution, although 2 Da mass difference provided reasonable
resolution.7 As defined by the structure of 1, installation of three
deuteriums or three 13C atoms or combination of the two isotopes
in benzene ring would not be a good synthetic strategy because
this requires an expensive starting material and also involves big
loss of labeled materials during the synthesis from the starting
material to the final compound. On the basis of the previous
chemistry established for the synthesis of 1,6,8 we choose to intro-
duce two deuteriums to the three positions of 1 and a 13C atom to
the carboxylic acid carbon in the later stage of the synthesis
starting from a boron-containing compound 4 as illustrated in
Scheme 1. The ester group of compound 4, prepared by the
method described previously,6,8 was reduced with NaBD4 to
provide the deuterated alcohol 5, which was further hydrolyzed
under acidic condition to give the deuterated benzoxaborole 6.
The aldehyde group in 6 went through a Wittig reaction to gener-
ate the 13C-labeled side chain of 7, which was hydrogenated to
Experimental
Materials and methods
Starting materials and solvents were purchased from chemical
companies and were used without further purification except
where noted otherwise. 1H and 13C NMR spectra were recorded at
room temperature on Bruker 300 or 400 instruments (Bruker Cor-
poration, Billerica, Massachusetts, USA). Electrospray ionization–mass
Scheme 1. Reagents and conditions: (a) NaBD4, MeOD, 0ꢀC to rt, 52 h; (b) 2N DCl,
MeOD, rt, 2 h, yield 53% over two steps; (c) Ph3PCH123COOEt bromide, NaH,
THF, À5ꢀC to rt, 24 h, then 2N DCl, yield 93%; (d) H2, 40 psi, PtO2Á3H2O, MeOH,
rt, 16 h, yield 60%; (e) LiOHÁH2O, MeOH, 0ꢀC, 16 h, then 2N HCl, yield 80%.
Figure 2. One of the side chain metabolite structures predicted by Meteor
software program.
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Y.-K. Zhang et al.
vacuum, and the residue was dissolved in D2O (50 mL) and
extracted with ethyl acetate (5Â 50 mL). The combined organic
layer was dried over Na2SO4, filtered, and concentrated to give
the residue (7.0g), which was purified by silica gel column
chromatography eluted with CH2Cl2 to afford compound 6 (2.2g,
1
yield 53%) as white solid. H NMR (300 MHz, DMSO-d6): d 10.37
(s, 1H), 9.21 (s, 1H), 7.85 (dd, 1H), 7.74 (dd, 1H), 7.67 (t, 1H) ppm.
7-(2-Ethoxy(13C)carbonyl-trans-vinyl)-(3,3-2H2)-1,3-dihydro-
1-hydroxy-2,1-benzoxaborole (7)
To a solution of triphenylphosphine (3.9 g, 14.9 mmol) in EtOAc
(9.0 mL) was added dropwise a solution of ethyl bromo(1-13C)
acetate (2.45 g) in EtOAc (7.0 mL) during 3 min. White precipitates
were formed during the addition, and the mixture was stirred for
3.5 h at room temperature. The solid was collected by filtration,
rinsed with Et2O, and dried overnight under high vacuum at
45ꢀC to give (ethoxy(13C)carbonylmethyl)triphenylphosphonium
bromide (4.7 g, yield 73%) as white solid. 1H NMR (300 MHz,
CDCl3): d 7.96–7.89 (m, 6H), 7.83–7.78 (m, 3H), 7.72–7.66 (m, 6H),
5.62 (dd, 2H), 4.05 (qd, 2H), 1.09 (t, 3H) ppm. To a solution of
compound 6 (1.5 g, 9.15 mmol) and (ethoxy(13C)carbonylmethyl)
triphenylphosphonium bromide (4.2 g, 9.78 mmol, 1.1 eq) in THF
(100.0 mL) was added NaH (60% in mineral oil, 0.5 g, 12.5 mmol,
1.4 eq) under stirring at À5ꢀC. The reaction mixture was then
stirred at room temperature overnight. Another portion of NaH
(0.5 g) was added at 0ꢀC and stirred at room temperature for
8 h. The reaction was quenched with D2O (10 mL), acidified to
pH 2–3 with 2N DCl, extracted with ethyl acetate (5 Â 50 mL),
dried over anhydrous Na2SO4, and filtered. The residue after
evaporation was purified by silica gel column chromatography
eluted with petroleum ether/ethyl acetate (3:1) to provide
compound 7 (2.0 g, yield 93%). 1H NMR (300 MHz, DMSO-d6):
d 9.31 (s, 1H), 8.09 (d, 1H), 7.83 (dd, 1H), 7.50 (t, 1H), 7.43 (d, 1H),
6.80 (d, 1H), 4.18 (q, 2H), 1.24 (t, 3H) ppm.
Scheme 2. Reagents and conditions: (a) NBS, 1,4-dioxane, rt, 1 h, yield 41%; (b) (1)
NaNO2, H2SO4, H2O, 0ꢀC, 30 min, (2) NaI, 0ꢀC to rt, 1 h, yield 70%; (c) (1) i-PrMgCl,
14
THF, À40ꢀC, 20 min, (2) CO2, À10ꢀC, 2 h, then 3N HCl, yield 60%; (d) Me2SO4,
K2CO3, acetone, 35ꢀC, 16 h, yield 83%; (e) Pin2B2, Pd(PPh3)2Cl2, KOAc, 1,4-dioxane,
90ꢀC, 40 h; (f) NaBH4, EtOH, rt, 2 h, then 2N HCl, yield 56% over two steps; (g) Raney
Ni, NaH2PO2ÁH2O, pyridine/AcOH/H2O (2:1:1), 45ꢀC, 2 h, yield 68%;
(h) Ph3P═CHCO2Et, DCM, rt, 16 h, yield 74%; (i) H2, 10% Pd/C, EtOAc, rt, 1 h, yield
100%; (j) LiOHÁH2O, MeOH/THF/H2O (1:1:1), rt, 1 h, then 2N HCl, yield 100%. Asterisk
(*) is used to mark the position of 14C-isotope in this scheme.
spectrometry (ESI–MS) and LC–MS were carried out on an API2000
(AB Sciex, Framingham, Massachusetts, USA) or a Finnigan LCQ mass
spectrometer (Thermo Fisher Scientific, Waltham, Massachusetts,
USA). HPLC was conducted on an Agilent 1200 system (Agilent Tech-
nologies, Inc., Santa Clara, California, USA) using a BDS Hypersil C-18
column (150 Â 4.6mm, 5micron, 120nm pore size, Thermo Fisher
Scientific, Waltham, Massachusetts, USA). The mobile phase used
was composed of buffer A (H2O containing 0.1% phosphoric acid)
and buffer B (CH3CN). The column was eluted with a gradient of
95% buffer A and 5% buffer B to 40% buffer A and 60% buffer B over
10 min, followed by 1 min at 40% buffer A and 60% buffer B,
followed by 1min gradient to 95% buffer A and 5% buffer B that
was kept for three more minutes at a flow rate of 1.0 mL/min
with column temperature at 30ꢀC and UV detection at 220 or
230 nm, and 266 nm. Silica gel 60 nm (200–425 mesh, Thermo Fisher
Scientific, Waltham, Massachusetts, USA) was used for flash column
chromatography.
7-(2-Ethoxy(13C)carbonylethyl)-(3,3-2H2)-1,3-dihydro-1-
hydroxy-2,1-benzoxaborole (8)
To a solution of compound 7 (2.0g, 8.51 mmol) in CH3OH (20 mL)
was added PtO2Á3H2O (0.2 g). The flask of the reaction mixture
was evacuated and refilled with H2 twice. The reaction mixture
was stirred under 40 psi at room temperature overnight. TLC ana-
lysis confirmed that there was no starting material left. The reac-
tion mixture was filtered and evaporated. The residue was purified
by silica gel column chromatography eluted with petroleum ether/
ethyl acetate (3:1) to give compound 8 (1.2 g, yield 60%). 1H NMR
(300 MHz, CD3OD): d 7.37 (t, 1H), 7.22 (d, 1H), 7.15 (d, 1H), 4.10 (q,
2H), 3.37 (s, 1H), 3.09 (t, 2H), 2.65 (t, 2H), 1.24 (t, 3H) ppm.
7-Formyl-(3,3-2H2)-1,3-dihydro-1-hydroxy-2,1-
benzoxaborole (6)
Compound 4 (8.4 g, 25.1 mmol) was mixed and evaporated with
CH3OD (20.0 mL) twice to minimize the possible presence of H2O
and was redissolved in freshly distilled CH3OD (50.0 mL).
7-(2-(13C)carboxyethyl)-(3,3-2H2)-1,3-dihydro-1-hydroxy-2,1-
benzoxaborole (2)
Under N2, NaBD4 (2.0 g) was added in portions at 0–5ꢀC, and To a solution of compound 8 (1.5 g, 6.3 mmol) in CH3OH
the reaction mixture was stirred at room temperature overnight. (16.0 mL) was added LiOHÁH2O (0.71 g, 16.9 mmol, 3.3 eq) under
Thin layer chromatography (TLC) analysis showed the remaining stirring at 0ꢀC. The reaction mixture was stirred overnight, and
of the starting material. Additional NaBD4 (0.5 g) was added, TLC analysis showed no starting material remaining. H2O
and the mixture was stirred for 36 h more. TLC analysis indicated (30 mL) was added dropwise into the reaction mixture at 0ꢀC.
the disappearance of the starting material, and nonisolated The mixture was adjusted to pH 1 with 2N HCl. The resulting
intermediate
3-(1,3-dioxolan-2-yl)-2-(4,4,5,5-tetramethyl-1,3,2- aqueous solution was extracted with ethyl acetate (5 Â 50 mL).
dioxaborolan-2-yl)-(a,a-2H2) benzyl alcohol (5) was presumably The combined organic layer was dried over Na2SO4, filtered,
generated. 2N DCl (26.0 mL) was added to adjust to pH 1, and and concentrated to dryness. The residue was purified by
the mixture was stirred for 2 h. Solvents were removed under silica gel column chromatography with hexane/ethyl acetate
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Y.-K. Zhang et al.
(3:1) and was further purified by recrystallization from methanol dried on the vacuum line to give 14 as a white solid (0.64 g, yield
1
1
to give the desired target compound 2 (1.06 g, yield 80%). H 60%). H NMR (400 MHz, CDCl3): d 8.03 (d, 1H), 7.77 (d, 1H), 7.52
NMR (300 MHz, DMSO-d6): d 7.36 (t, 1H), 7.21 (d, 1H), 7.14 (t, 1H) ppm; MS (ESI+): m/z = 226 and 228 (M-1).
(d, 1H), 3.03–2.96 (m, 2H), 2.57–2.50 (m, 2H) ppm; 13C NMR
Methyl 2-bromo-3-cyano-(carbonyl-14C)benzoate (15)
(75 MHz, DMSO-d6): d 173.87, high intensity), 154.13, 145.08,
130.83, 126.71, 119.12, 69.44–69.14 (m), 35.59 (d), 29.32 ppm.
HPLC purity: 98.3% at 220 nm; isotopic enrichment: 96.9%; MS:
m/z = 232 (M + 23, ESI+) and 208 (M À 1, ESIÀ).
To a round bottom flask (100 mL) were added 2-bromo-3-
cyano-(carboxy-14C)benzoic acid (0.64 g, 2.81 mmol) and acetone
(10 mL). To the mixture was added dimethylsulfate (0.42 g,
3.33 mmol, 1.2 eq) and potassium carbonate (0.46 g, 3.33 mmol,
3-Amino-2-bromobenzonitrile (12)
1.2 eq). The mixture was heated to 35ꢀC in an oil bath overnight,
To a solution of 3-aminobenzonitrile (11, 2.95 g, 25 mmol) in filtered, the solids washed with acetone, and the filtrate was
1,4-dioxane (40 mL) in a round bottom flask (200 mL) was added concentrated. The residue was purified by silica gel column
a solution of N-bromosuccinimide (NBS, 4.45 g, 25 mmol) in chromatography eluted with 10% ethyl acetate/hexane to give
1
1,4-dioxane (20 mL) at room temperature over 30 min with 15 as a white solid (0.56 g, yield 83%). H NMR (400 MHz, CDCl3):
magnetic stirring.9 The mixture was stirred for another 30 min d 7.95 (d, 1H), 7.76 (d, 1H), 7.50 (t, 1H), 3.97 (s, 3H) ppm; MS
and then concentrated onto silica gel. The crude product on (ESI+):m/z = 243 and 245 (M + 1).
silica gel was placed to the top of a silica gel column and eluted
7-Cyano-(3-14C)-1,3-dihydro-1-hydroxy-2,1-
with hexane/ethyl acetate/dichloromethane (8:1:1). The fractions
containing the desired product were concentrated to give a
solid residue that was recrystallized from toluene to provide
compound 12 (2.02 g, yield 41%). 1H NMR (400 MHz, CDCl3):
d 7.17 (t, 1H), 7.04 (d, 1H), 6.93 (d, 1H), 4.33 (s, 2H) ppm; MS
(ESI+): m/z = 197 and 199 (M + 1).
benzoxaborole (17)
To a round bottom flask (25 mL), fitted with a condenser under
argon, were added methyl 2-bromo-3-cyano-(carbonyl-14C)
benzoate (15, 0.23 g, 0.9 mmol) and 1,4-dioxane (6 mL, distilled
from LAH). To the mixture were added bis(pinacolato)diboron
(0.32 g, 1.4 eq) and potassium acetate (0.40 g, 4.05 mmol, 4.5 eq),
and argon was bubbled into the mixture for several minutes.8
2-Bromo-3-iodobenzonitrile (13)
A
mixture of 3-amino-2-bromobenzonitrile (12, 2.02 g, Dichlorobis(triphenylphosphine)palladium II (25 mg, 0.036 mmol,
10.25 mmol) in 2 M sulfuric acid (25 mL) in a 200 ml round 3 mol%) was added and the mixture heated to 90ꢀC in an oil bath
bottom flask (200 mL) was warmed to dissolve solids and then for 40 h. The cooled mixture was diluted with ethyl acetate (6 mL),
cooled in an ice bath.10 To the mixture was added sodium nitrite filtered, and the solid was washed with ethyl acetate. The filtrate
(0.74 g, 10.77 mmol, 1.05 eq) in water (5 mL), and the mixture was containing 16 was concentrated to syrup that was diluted with
stirred for 30 min in the ice bath, followed by addition of a ethanol (10 mL), and the reaction flask was placed in a water bath.
solution of sodium iodide (1.85 g, 12.3 mmol, 1.2 eq) in water To the mixture was added sodium borohydride (0.23 g, 6 mmol,
(5 mL) with additional stirring for 30 min at 0ꢀC and 30 min at 6.6 eq), and the mixture was stirred at room temperature (rt) for
room temperature. The mixture was extracted three times with 2 h. The mixture was added to 2N HCl (30 mL) and stirred for
ethyl acetate. The combined extracts were dried over magne- 20 min. The mixture was extracted three times with isopropa-
sium sulfate, filtered and concentrated. The residue was purified nol/chloroform (1:9). The combined extracts were dried over
by silica gel column chromatography eluted with 5% ethyl anhydrous magnesium sulfate, filtered, and concentrated. The
acetate/hexane to give the product 13 as a tan solid (2.16 g, yield crude product was dissolved in toluene/dichloromethane (1:1,
1
70%). H NMR (400 MHz, CDCl3): d 8.07 (d, 1H), 7.63 (d, 1H), 7.11 2 mL), placed to the top of a silica gel column, and eluted with
(t, 1H) ppm; MS (ESI+): m/z = 308 and 310 (M + 1).
ethyl acetate/hexane (3:7) then acetic acid/ethyl acetate/hexane
(1:40:60) to give 17 as a white solid (80 mg, yield 56%). H NMR
(400 MHz, DMSO-d6): 9.47 (s, 1H), 7.83 (d, 1H), 7.77 (d, 1H), 7.69
(t, 1H), 5.08 (s, 2H) ppm; MS (ESIÀ): m/z = 160 (M À 1).
1
2-Bromo-3-cyano-(carboxy-14C)benzoic acid (14)
To a round bottom flask (100 mL) were added 2-bromo-3-
iodobenzonitrile (13, 2.23 g, 7.24 mmol, 1.55 eq) and anhydrous
THF (14 mL, distilled from Na). The flask was placed under argon,
capped with septum, and cooled to À40ꢀC in a dry ice acetone
7-Formyl-(3-14C)-1,3-dihydro-1-hydroxy-2,1-
benzoxaborole (18)
bath. To the mixture was added 2 M isopropyl magnesium To a round bottom flask (25 mL) were added sequentially
chloride (4 ml, 8 mmol, 1.7 eq), and the mixture was stirred for 7-cyano-(3-14C)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole (17,
20 min at À40ꢀC.11 The flask was attached to the vacuum line, 80 mg, 0.5 mmol), a mixed solvent of pyridine/acetic acid/water
frozen in liquid nitrogen, and evacuated. 14C-Carbon dioxide, (2:1:1, 4 mL), sodium hypophosphite hydrate (NaH2PO2ÁH2O,
which was previously generated from barium (14C)carbonate 0.44 g, 5 mmol, 10 eq), and Raney nickel (120 mg, Aldrich 2800,
(0.93 g, 4.66 mmol, 1 eq) and lactic acid, was condensed in the Sigma-Aldrich, St. Louis, MO, USA).12 The mixture was heated at
flask and valves closed. The reaction flask was placed in a dry 45ꢀC in an oil bath for 2 h. The catalyst was filtered off and
ice acetone bath at À10ꢀC and stirred for 2 h. The flask was washed with methanol. The filtrate was evaporated, and the resi-
removed from the vacuum line, and water (30 mL) was added due was diluted with 2N HCl and extracted four times with ethyl
to the flask. The mixture was washed with ether, acidified acetate. The combined extracts were dried over anhydrous mag-
with 3N HCl and extracted three times with ethyl acetate. The nesium sulfate, filtered, and concentrated. The crude product
combined ethyl acetate extracts were dried over magnesium dissolved in toluene/dichloromethane (1:1, 2 mL) was placed to
sulfate and concentrated to a white solid. The solid was washed the top of a silica gel column and eluted with acetic acid/ethyl
with 50% ether/hexane to remove 2-bromobenzonitrile and acetate/hexane (1:20:80) to give 18 as a white solid (55 mg, yield
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Y.-K. Zhang et al.
68%). 1H NMR (400 MHz, CDCl3): 10.03 (s, 1H), 8.08 (s, 1H), 7.87 (d, hydroxy-2,1-benzoxaborole (3), were successfully synthesized.
1H), 7.71 (t, 1H), 7.67 (d, 1H), 5.19 (s, 2H) ppm; MS (ESI+): m/ These isotopically labeled compounds were used for the preclinical
z = 165 (M + 1).
development studies of a potential new antimalarial agent,
7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole (1). The
results of these studies will be reported in due courses.
7-(2-Ethoxycarbonyl-trans-vinyl)-(3-14C)-1,3-dihydro-1-
hydroxy-2,1-benzoxaborole (19)
To a round bottom flask (25 mL) were added 7-formyl-(3-14C)-1,3-
dihydro-1-hydroxy-2,1-benzoxaborole (18, 55 mg, 0.33 mmol),
dichloromethane (4 mL), and (ethoxycarbonylmethylene)
triphenylphosphorane (230 mg, 0.67 mmol, 2 eq). The mixture
was stirred at room temperature overnight. The mixture was
placed directly to the top of a silica gel column and eluted with
acetic acid/ethyl acetate/hexane (1:20:80) to give 19 as a white
Acknowledgment
We thank the Medicines for Malaria Venture (MMV) for the
financial support to the project.
Conflict of Interest
The authors did not report any conflict of interest.
1
solid (57 mg, yield 74%). H NMR (400 MHz, CDCl3): 8.16 (d, 1H),
7.62 (d, 1H), 7.50 (t, 1H), 7.34 (d, 1H), 6.65 (d, 1H), 5.09 (s, 2H),
4.29 (q, 2H), 1.35 (t, 3H) ppm; MS (ESI-): m/z = 233 (M-1).
References
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7-(2-Ethoxycarbonylethyl)-(3-14C)-1,3-dihydro-1-hydroxy-
2,1-benzoxaborole (20)
To a round bottom flask (25mL) were added 7-(2-ethoxycarbonyl-
trans-vinyl)-(3-14C)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole (19,
57 mg, 0.24 mmol), ethyl acetate (4mL), and 10% palladium on
carbon (10 mg). The mixture was stirred at room temperature
under a hydrogen balloon for 1 h. The resulting mixture was
filtered through a 0.2micron acrodisc and washed with ethyl
acetate. The solvent was concentrated to give 20 as a solid (57 mg,
1
yield 100%). H NMR (400 MHz, CDCl3): 7.39 (t, 1H), 7.17 (m, 2H),
5.60 (s, 1H), 5.05 (s, 2H), 4.14 (q, 2H), 3.15 (t, 2H), 2.70 (t, 2H), 1.23
(t, 3H) ppm. MS (ESIÀ): m/z=235 (MÀ 1).
7-(2-Carboxyethyl)-(3-14C)-1,3-dihydro-1-hydroxy-2,1-
benzoxaborole (3)
To a round bottom flask (25 mL) were added 7-(2-ethoxycarbo-
nylethyl)-(3-14C)-1,3-dihydro-1-hydroxy-2,1-benzoxaborole (20,
57 mg, 0.24 mmol), a mixed solvent of methanol/THF/water
(1:1:1, 3 mL) and lithium hydroxide hydrate (42 mg, 1 mmol,
4 eq). The mixture was stirred at rt for 1 h, acidified with 2N HCl
(1 mL), and concentrated. The solid was diluted with water and
extracted three times with ethyl acetate. The combined extracts
were dried over anhydrous magnesium sulfate and concentrated
to give the desired final compound 7-(2-carboxyethyl)-(3-14C)-
1,3-dihydro-1-hydroxy-2,1-benzoxaborole 3 as a white solid
1
(50 mg, yield 100%). H NMR (400 MHz, DMSO-d6): 12.06 (s, 1H),
8.99 (s, 1H), 7.35 (t, 1H), 7.21 (d, 1H), 7.14 (d, 1H), 4.95 (s, 2H),
2.99 (t, 2H), 2.54 (t, 2H) ppm; chemical purity by HPLC: 99.5% at
230 nm; radiochemical purity by HPLC: 99.7% with radiochemical
detection; MS (ESIÀ): m/z = 207 (M À 1). In addition, this radiola-
beled material had the same HPLC retention time as that of
the nonlabeled compound (1).
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Engl. 1998, 37, 1701.
Conclusion
In conclusion, two new isotopically labeled compounds,
(carboxy-13C-3,3-2H2)-7-(2-carboxyethyl)-1,3-dihydro-1-hydroxy-2,1-
benzoxaborole (2) and (3-14C)-7-(2-carboxyethyl)-1,3-dihydro-1-
[12] R. J. Heffner, J. Jiang, M. M. Joullie, J. Am. Chem. Soc. 1992, 114,
10181.
J. Label Compd. Radiopharm 2012
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