Synthesis and evaluation of novel purple acid phosphatase inhibitors

Article abstract of DOI:10.1039/C8MD00491A

Transgenic studies in animals have demonstrated a direct association between the level of expression of purple acid phosphatase (PAP; also known as tartrate-resistant acid phosphatase) and the progression of osteoporosis. Consequently, PAP has emerged as

Full text of DOI:10.1039/C8MD00491A

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RESEARCH ARTICLE
Synthesis and evaluation of novel purple acid
phosphatase inhibitors†
Cite this: DOI: 10.1039/c8md00491a
ac
Waleed M. Hussein,ab Daniel Feder,a Gerhard Schenk,
a
Luke W. Guddata and Ross P. McGeary
*
Transgenic studies in animals have demonstrated a direct association between the level of expression of
purple acid phosphatase (PAP; also known as tartrate-resistant acid phosphatase) and the progression of
osteoporosis. Consequently, PAP has emerged as a promising target for the development of novel thera-
peutic agents to treat this debilitating disorder. PAPs are binuclear hydrolases that catalyse the hydrolysis of
phosphorylated substrates under acidic to neutral conditions. A series of phenyltriazole carboxylic acids,
prepared by the reactions of azide derivatives with propiolic acid through copperIJI)-catalysed azide–alkyne
cycloaddition click reactions, has been assessed for their inhibitory effect on the catalytic activity of pig and
red kidney bean PAPs. The binding mode of most of these compounds is purely uncompetitive with Kiuc
values as low as ∼23 μM for the mammalian enzyme. Molecular modelling has been used to examine the
binding modes of these triazole compounds in the presence of a substrate in the active site of the enzyme
in order to rationalise their activities and to design more potent and specific derivatives.
Received 29th September 2018,
Accepted 9th November 2018
DOI: 10.1039/c8md00491a
rsc.li/medchemcomm
to as uteroferrin (Uf).2a,3b–d In humans, PAP is involved in the
inflammatory response of antigen-presenting cells6 and in
Introduction
Purple acid phosphatases (PAPs) are binuclear metallo-
enzymes found in animals, plants and fungi.1 The PAP active
site contains two metal ions that assist in the hydrolysis
of phosphate esters and anhydrides under acidic to neutral
conditions.2 PAPs are the only known metallohydrolases that
require a heterovalent dimetallic active site to promote hydro-
lysis.1 While animal PAPs generally employ a redox-active
FeIII–FeII/III combination, in plants FeIII–ZnII and FeIII–MnII
centres are found.1–3
PAPs can dephosphorylate small substrates such as adeno-
sine diphosphate (ADP), adenosine triphosphate (ATP) and
para-nitrophenyl phosphate (pNPP),1a–d,3a as well as larger
substrates such as the phosphoproteins, osteopontin and
sialoprotein.4 Several biological roles for PAPs have been
suggested. PAP from the allantoic fluid of pregnant sows is
implicated in the transport of iron from mother to foetus
during gestation.5 Consequently, this enzyme is also referred
the process of bone remodelling in the bone resorptive
space.4b,7
Osteoporosis is a disease that affects mainly the elderly and
in particular post-menopausal women. In the developed world,
mortality and morbidity rates are increasing together with life
expectancy.8 In Europe and the United States, 30% women are
osteoporotic; 40% of post-menopausal women and 30% of men
will experience osteoporotic fractures in their lifetime.9
In humans, elevated PAP levels in the blood are directly
correlated with the onset and/or progression of osteoporo-
sis.10 In mice that overexpress PAP, a decrease in bone den-
sity is observed, consistent with the onset of osteoporosis.4a
In mice where the gene encoding for PAP is selectively
disrupted, there is an increase in bone mineralisation, symp-
tomatic of osteopetrosis (the opposite phenotype of
osteoporosis).10a These observations suggest that PAP inhibi-
tors could be used to treat osteoporosis. There is also the
possibility that potent PAP inhibitors will find additional
clinical applications in the treatment of other conditions
which are associated with elevated levels of this enzyme in
the serum, e.g. AIDS encephalopathy,11 Gaucher's disease,12
hairy cell leukemia,13 Alzheimer's disease14 and bone
metastases.15
a The University of Queensland, School of Chemistry and Molecular Biosciences,
Brisbane, QLD 4072, Australia. E-mail: r.mcgeary@uq.edu.au;
Tel: +61 7 33653955
b Helwan University, Pharmaceutical Organic Chemistry Department, Faculty of
Pharmacy, Ein Helwan, Helwan, Egypt
c The University of Queensland, Australian Centre for Ecogenomics, Brisbane, QLD
4072, Australia
Known inhibitors of PAPs include fluoride and tetrahedral
oxoanions such as phosphate, vanadate, tungstate, arsenate
and molybdate, but they have not been further developed
into anti-osteoporotic drugs due to their lack of specificity
† Electronic supplementary information (ESI) available: Enzyme kinetic data for
inhibitors 1a–f against pig PAP; 1H and 13C NMR spectra for compounds 1a–f,
3a–f and 5b–f. See DOI: 10.1039/c8md00491a
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and their potential to be cytotoxic.1a–d,2c,d,3d,4b,16 Vanadate
(VO43−) is a strong inhibitor of pig and red kidney bean PAP
(rkbPAP), with Ki values of 0.040 mM and 0.030 mM, respec-
tively, which is approximately 50 times more potent than the
inhibitory effect of phosphate (Ki ∼ 2 mM).1,16b This observa-
tion is consistent with similar studies that indicate that the
orthovanadate anion is significantly more potent at
inhibiting protein phosphatases than orthophosphate.17 This
difference is likely to be due to vanadate being able to stabi-
lise a five-coordinate trigonal bipyramidal complex at the ac-
tive site, which resembles the transition state of phosphoryl
transfer reactions.17
a Ki value of 4 μM against rkbPAP and 17 μM against pig
PAP.21 We have also described the synthesis, PAP inhibition
and computational modelling of a series of structurally analo-
gous α-amino-substituted naphthylmethylphosphonic acids,
which display Ki values of 5 μM against rkbPAP and 8 μM
against pig PAP.22 Our group also synthesised a series of acyl
derivatives of 6-aminopenicillanic acid and these showed po-
tent inhibition against PAP, with Ki values in a low micromo-
lar range.23
Human PAP is the clinically relevant target for the devel-
opment of novel PAP inhibitors, but to date this enzyme can
only be obtained in small quantities using a baculoviral ex-
pression system,2b and since large amounts of enzyme are re-
quired for kinetic testing and crystallisation studies, pig and
rkbPAPs have been used as models for human PAP to test the
in vitro effects of potential inhibitors. The validity of using
pig and rkbPAPs as models of the human enzyme is
supported by the observations that: (i) most of the amino
acids in the active site that coordinate the metal ions are con-
served across all PAPs,1,2c–g,24 (ii) pig and human PAPs share
88% amino acid sequence identity and the amino acid resi-
dues within the active site are highly conserved in human
and pig PAPs,25 and (iii) previous studies have shown that
pig, human and rkbPAPs catalyse the hydrolysis of similar
substrates with similar catalytic efficiencies.1a–d,2c–g,3a,d It
should be noted that in the human genome two genes
encoding PAPs have been identified, one that is very similar
to the pig enzyme (a monomeric enzyme of ∼35 kDa), and
one that resembles rkbPAP (a putatively homodimeric form
Myers and coworkers synthesised and tested the inhibitory
activity against rkbPAP for a group of alkylphosphonic acids
that contain metal-binding groups such as thiol, carboxylate
or phosphonate.18 The results of their study did not yield
strong drug leads due to the modest potencies of the inhibi-
tors (IC50 = 80–3000 μM), however, they did demonstrate that
metal-binding moieties are important for the enzyme-binding
affinities of these inhibitors.18
A subsequent study by
Valizadeh and colleagues described the design of a suite of
modified phosphotyrosine-containing tripeptides, and
showed that these compounds exhibit inhibitory activity
against several mammalian and plant PAPs in the mid-
micromolar range.19
Based on the work of Schwender et al.,20 our group
designed and developed a series of α-alkoxy-substituted
naphthylmethylphosphonic acids, some of which showed low
micromolar inhibitory activity, with the best inhibitor having
Scheme 1 Reagents and conditions: (a) NaN3, (CH3)2CO/H2O (4 : 1), r.t., 24 h. 3a (70%); (b) NaBH4, 2 M, NaOH, MeOH, r.t., 4 h. 5b (51%); 5c (44%);
5d (52%); 5e (61%); 5f (57%); (c) (PhO)2PON3, DBU, dry toluene, 0 °C, 2 h. Then r.t., 16 h. 3b (81%); 3c (89%); 3d (87%); 3e (84%); 3f (92%); (d)
propiolic acid, sodium ascorbate, CuIJAcO)2, H2O/tBuOH (1 : 1), r.t., 14 h. 1a (90%); 1b (83%); 1c (87%); 1d (61%); 1e (75%); 1f (70%).
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Table 1 Kinetic data for inhibitors 1a–f against fully reduced pig PAP and rkbPAP at pH 4.9 and docking parameters for the predicted human PAP-
inhibitor complexes
Compound
PAP
Kic (μM)
Kiuc (μM)
MolDock score/(kcal mol−1
)
Ligand efficiencya
1a
1a
1b
1b
1c
1c
1d
1d
1e
1e
1f
Pig
rkb
Pig
rkb
Pig
rkb
Pig
rkb
Pig
rkb
Pig
rkb
—
—
—
951
—
—
—
1073
—
438
—
23
963
76
—
48
—
103
—
276
—
184
—
−81.52
−5.60
−82.74
−81.41
−76.30
−68.14
−88.70
−5.18
−4.77
−4.94
−4.72
−4.67
1f
2276
a
Ligand efficiency is calculated as the binding free energy (in kcal mol−1) divided by the number of heavy atoms.
with ∼55 kDa subunits).26 The function of this larger form is
currently unknown but based on the arguments above, PAP
inhibitors tested against pig and rkbPAP are expected to af-
fect both human PAP isoforms similarly.
(pNPP) at 5 mM. The assay procedure was adapted from sim-
ilar studies with PAPs.22,23,27
Mode and magnitude of inhibitor binding
Subsequently, more detailed catalytic studies were performed
to determine the magnitude of the competitive (Kic) and/or
Results and discussion
uncompetitive (Kiuc
)
inhibitor dissociation constants,
PAP has been identified as an attractive target for the
development of chemotherapeutics to treat osteoporosis.1–4
Through the screening of a Maybridge™ compound library,
we previously identified a promising inhibitor, p-tolyl thiazole
carboxylic acid, with Ki values of 340 μM for rkbPAP and 42
μM for pig PAP.27 Here, a set of analogues of this thiazole in-
hibitor was synthesised in which the thiazole ring was re-
placed by 1,2,3-triazole ring to afford compounds 1a–f
(Scheme 1).
representing the equilibrium constants for the enzyme-
inhibitor and enzyme-substrate-inhibitor complexes, respec-
tively (Table 1). Details of the data collection and numeric
analyses are described in the Experimental section and
follow a similar procedure we employed for other systems
(e.g. the antibiotic-degrading metallo-β-lactamases).28 Com-
pounds 1a–f showed either no or only weak competitive
binding to either of the two enzymes. However, in particular,
compounds 1a, 1b and 1c displayed uncompetitive inhibition
against the mammalian PAP, with Kiuc values of 23, 76 and
48 μM, respectively. Of particular importance is that inhibi-
tors 1a–f exclusively bind in an uncompetitive mode to pig
PAP, whereas they bind largely in a competitive mode (albeit
very weakly) to the plant enzyme. This difference in binding
mode to active sites that share considerable similarity (see
above) suggests that the inhibitors engage in different bind-
ing interactions. Furthermore, since there is a second human
PAP isoform that is homologous to rkbPAP26 it may be possi-
ble to selectively inhibit only the PAP isoform associated with
bone-related ailments.
Chemistry
To synthesise benzyl azide (3a), benzyl bromide (2) was
treated with sodium azide in acetone : water (4 : 1) to give 3a
in 70% yield. The rest of the benzyl azide derivatives 3b–f
were synthesised over two steps. First, reduction of benzalde-
hyde derivatives 4b–f to their corresponding benzyl alcohols
5b–f (44–61% yield) using sodium borohydride, followed by
conversion the hydroxyl group to the azide moiety using
diphenylphosphoryl azide in dry toluene to give 3b–f in 81–
92% yield (Scheme 1). The final products 1a–f were obtained
in 61–90% yield via the copperIJI)-catalysed azide alkyne cyclo-
addition click reaction between azide derivatives 3a–f and
propiolic acid in the presence of sodium ascorbate and
copperIJII) acetate in t-butanol : water mixture (1 : 1).
Docking studies
Computational docking studies were undertaken with
Molegro Virtual Docker (MVD) to predict the binding mode
of 1a–f with human PAP (Fig. 1). Firstly, the substrate p-NPP
was docked into the active site of human PAP. In its optimal
conformation, p-NPP is predicted to bind through its phos-
phate group in a μ-1,3 bidentate mode to the metal centre,
with the phenyl ring and nitro group positioned in a groove
formed by the metal-coordinating residue H221 and the
sidechains of residues F54 and F242. Compounds 1a–f were
Initial assessment of inhibitory potential of novel
compounds
Initial percentage inhibition assays were performed at com-
pound concentrations of 100 μM to test the abilities of 1a–f
to inhibit rkbPAP and fully reduced pig PAP in the presence
of the chromophoric substrate para-nitrophenyl phosphate
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Fig. 1 Surface and stick representation of the MVD docking results for compounds 1a–f (green surface) (in respective order from left to right) to
the docked hPAP–p-NPP complex (grey and yellow surfaces, respectively). The inhibitors are predicted to bind in a groove between the
mammalian PAP repression loop and residues R248 and K252. Metal ions are shown in green and are marked with an asterisk.
then each docked to this hypothetical human PAP–p-NPP
complex. The lowest energy poses predicted that all com-
pounds 1a–f would bind in a positively charged pocket under
the mammalian repression loop between the side-chains of
R248 and H249 and the side-chains of S148 and Q149. The
similarity of the predicted binding modes (Fig. 2) for these
compounds is consistent with their inhibition constants
(Table 1). Docking scores were within the range of −68.14 and
−88.70 kcal mol−1 for all ligands and ligand efficiencies were
highest for 1a–1d (Table 1). This finding is consistent with
their improved inhibitory activity when compared to 1e–1f.
Compound 1a showed the highest ligand efficiency and a pre-
dicted binding mode that is different from the other inhibi-
tors in this series, likely due to its smaller size, allowing it to
bind more complementarily to the enzyme (Fig. 1).
Superimposition of the crystal structure of pig PAP onto
the crystal structure of human PAP shows that the only nota-
ble difference in the surface predicted to bind 1a–f and
p-NPP is a conservative substitution of R248 in human PAP
to K250 in pig PAP (Fig. 2). Therefore, the mode and
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Fig. 2 Superimposition of the crystal structure of human PAP (PDB ID: 1WAR, grey) and pig PAP (PDB ID 5UQ6, cyan) showing that the binding
site for p-NPP (pink carbons) and the groove formed under the repression loop are conserved between the two enzymes. The only difference rele-
vant to inhibitor binding being R248/K250. The docked poses for inhibitors 1a–f (carbons coloured by ID) and for p-NPP are superimposed on the
crystal structures. The active site is marked with an asterisk. Labels show both human PAP (first) and pig PAP (second) numbering.
magnitude of inhibition are likely to be conserved between
pig and human PAPs.
cellulose column followed by gel filtration on a Sephadex
S-300 column. The resulting preparation was concentrated to
23.8 mg mL−1 using a Millipore Amicon centrifugal concen-
trator and stored at 4 °C in 0.5 M sodium chloride. Pig PAP
was extracted from the uterine fluid of a pregnant sow and
purified by ion-exchange chromatography using CM-cellulose
followed by gel filtration on a Sephadex G-75.30 Purified pig
PAP was concentrated to 8.1 mg mL−1 and stored at −20 °C in
100 mM acetate buffer at pH 4.9. Protein concentrations were
determined by measuring the absorbance at 280 nm using ex-
tinction coefficients of 1.41 for a 1 mg mL−1 solution (28.6
μM) of pig PAP and 2.1 for a 1 mg mL−1 solution (9.1 μM) of
rkbPAP. SDS-Page analysis showed that the enzymes were
>95% pure.
Conclusions
Six novel inhibitors have been discovered that inhibit
mammalian PAP in an uncompetitive manner. Docking
and kinetic studies were used to identify the most proba-
ble binding site for these inhibitors. The inhibitors are all
predicted to bind in a pocket under the mammalian re-
pression loop, a structural feature that is unique to mam-
malian PAPs and absent in plant PAPs. This prediction is
supported by the fact that these compounds inhibit
rkbPAP, by and large, competitively. These findings thus
provide the basis for the design of improved human PAP
inhibitors that could exploit the structural features of the
mammalian repression loop as well as the immediate en-
vironment surrounding the metal centre.
Enzyme kinetics
Inhibition assays for both pig PAP and rkbPAP were
performed in 96-well 400 μL multi-titre plates using a UV/
vis multiplate spectrophotometer. Prior to its use in ki-
netic assays pig PAP was fully reduced with 0.77 mM
β-mercaptoethanol for ten minutes at 37 °C. Kinetic mea-
surements were carried out at pH 4.9 (0.1 M sodium ace-
tate buffer with 25% DMSO) at 25 °C using para-
nitrophenyl phosphate (pNPP) as substrate at concentra-
tions of 1, 3, 5, 7.5, 10 and 12.5 mM. The rate of product
formation was measured at λ = 405 nm (ε = 343 M−1
cm−1).3a The concentration of enzyme used was 20.9 nM
for pig PAP and 7.4 nM for rkbPAP, while the
Experimental
Enzyme preparation and purification
PAP from red kidney bean (rkbPAP) was purified following a
previously published protocol.29 Briefly, red kidney beans
(Phaseolus vulgaris) were ground in a Waring blender and
suspended in 0.5 M sodium chloride. The suspension was fil-
tered through a muslin cloth, followed by ethanol fraction-
ation and ammonium sulfate precipitation. Purification was
achieved by ion-exchange chromatography using
a CM-
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concentration of the inhibitors ranged from 50 μM to 300
μM. The data were analysed by non-linear regression
using the general inhibition equation (eqn (1)) and the
program WinCurveFit (Kevin Raner software).
Synthesis of benzyl azide (3a)33
Vmax
S
 
 
(1)

I 

I 
 
 
S 1
 K 1
 




m 


A solution of sodium azide (1.95 g, 30 mmol) in water (10
mL) was added to a stirred solution of benzyl bromide (2)
(3.42 g, 2.38 mL, 20 mmol) in acetone (40 mL). The reaction
mixture was stirred for 24 h at room temperature. Acetone
was evaporated in vacuo and the aqueous layer was extracted
with EtOAc (3 × 10 mL). The combined organic layer was
dried over Na2SO4, filtered and evaporated in vacuo to afford
the benzyl azide (3a) as a yellow oil (1.86 g, 70%) Rf: 0.60 (2%
Kiuc
Kic

In this equation, Kic and Kiuc represent the equilibrium
dissociation constants for competitive and uncompetitive
inhibitor binding, respectively, while v, Vmax, Km, [S] and
[I] represent the rate of product formation, the maximum
rate of product formation, the Michaelis constant, the sub-
strate concentration and the inhibitor concentration,
respectively.
1
EtOAc in LP, UV and KMnO4 dip). H NMR (300 MHz, CDCl3)
δ 4.34 (2H, s), 7.32–7.44 (5H, m); 13C NMR (75 MHz, CDCl3) δ
54.7, 128.1, 128.2, 128.8, 135.3 in agreement with that de-
scribed by Campbell-Verduyn.33
Docking studies
General method for preparation of benzyl alcohol derivatives
(5b–f)
Docking studies were undertaken with Molegro Virtual
Docker (MVD)31 using the MolDock SE algorithm. The li-
gand search space was confined to an 8 Å sphere originat-
ing from the metal centre of PAP. For human PAP, the
coordinates used were those of recombinant human PAP
in complex with phosphate (PDB code 1WAR)25a with the
phosphate anion omitted from the active site. Water mole-
cules were removed from all coordinate files prior to
docking.
These compounds 5b–f were prepared using the general
method described by Zartler and Shapiro.34 A solution of
sodium borohydride (0.37 eq., 173 mg, 4.58 mmol) in 2
M NaOH (247 μL) diluted with water (2.22 mL) was added
dropwise to a solution of aromatic aldehydes (4b–f) (12.38
mmol) in methanol (12.5 mL) at 18–25 °C. The reaction
mixture was stirred for 4 h at room temperature. The re-
action mixture was evaporated in vacuo; methanol (2 × 10
mL) was added and evaporated in vacuo. HCl (5%, 50
mL) was added to the residue and extracted with diethyl
ether (2 × 100 mL). The combined organic layer was dried
over anhydrous Na2SO4, and evaporated in vacuo to afford
5b–f.
Chemistry
For TLC staining, KMnO4 was used. KMnO4 dye ingredients:
3 g potassium permanganate; 20 g potassium carbonate; 5
mL 5% sodium hydroxide and 300 mL water.
Light petroleum (LP, b.p. 40–60 °C) was distilled before
use. Flash chromatography was carried out with Merck
Kieselgel 60 as described by Still.32 NMR spectra were
recorded on 300, 400 and 500 MHz spectrometers (Bruker,
Rheinstetten, Germany). Chemical shifts are reported in
parts per million (ppm) on a δ scale, relative to the sol-
vent peak (CDCl3 δH 7.24, δC 77.0; (CD3)2SO δH 2.49, δC
39.5). Coupling constants (J) are reported in hertz and
peak multiplicities described as singlet (s), doublet (d),
triplet (t), quartet (q), septet (sept), multiplet (m), or
broad (br). High resolution electrospray ionisation accurate
mass measurements were recorded in positive and nega-
tive mode on a quadrupole – time of flight instrument
(Bruker) with an ESI† source. Accurate mass measure-
ments were carried out with external calibration using so-
dium formate as reference calibrant and/or Agilent tune
mix (mw > 500). Low- and high-resolution electron impact
ionisation mass measurements were recorded using
perfluorokerosene-H as reference calibrant.
p-Tolylmethanol (5b)
The crude product was purified by silica flash column
chromatography (0–10% EtOAc in LP) to afford 5b as
white solid (770 mg, 51%) Rf: 0.14 (10% EtOAc in LP, UV
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and KMnO4 dip). 1H NMR (400 MHz, CDCl3) δ 2.40 (3H,
s), 2.61 (1H, s, OH), 4.61 (2H, s), 7.20 (2H, d, J 7.6 Hz),
7.26 (2H, d, J 7.9 Hz) in agreement with that described
by Shaikh.35 13C NMR (100 MHz, CDCl3) δ 21.0, 64.9,
127.0, 129.1, 137.1, 137.8.
13C NMR (100 MHz, CDCl3) δ 62.7, 127.0, 128.6, 128.7, 129.3,
132.6, 138.1 in agreement with that described by Shaikh.35
Naphthalen-1-ylmethanol (5f)
(4-Methoxyphenyl)methanol (5c)
The crude product was purified by silica flash column
chromatography (0–10% EtOAc in LP) to afford 5f as yel-
low solid (1.12 g, 57%) Rf: 0.15 (10% EtOAc in LP, UV
and KMnO4 dip). 1H NMR (300 MHz, CDCl3) δ 2.05 (1H,
s, OH), 5.09 (2H, s), 7.40–7.56 (4H, m), 7.80 (1H, d, J 7.9
Hz), 7.85–7.89 (1H, m), 8.07–8.10 (1H, m) in agreement
with that described by Ragnarsson.36 13C NMR (75 MHz,
CDCl3) δ 63.5, 123.6, 125.2, 125.3, 125.8, 126.3, 128.5,
128.6, 131.2, 133.7, 136.2.
The crude product was purified by silica flash column chro-
matography (0–60% EtOAc in LP) to afford 5c as yellow oil
(755 mg, 44%) Rf: 0.09 (10% EtOAc in LP, UV and KMnO4
1
dip). H NMR (300 MHz, CDCl3) δ 2.68 (1H, s, OH), 3.79 (3H,
s), 4.56 (2H, s), 6.87 (2H, d, J 8.7 Hz), 7.26 (2H, d, J 8.8 Hz);
13C NMR (75 MHz, CDCl3) δ 55.2, 64.7, 113.8, 128.5, 133.0,
159.0 in agreement with that described by Shaikh.35
General method for preparation of azide derivatives (3b–f)
(4-Chlorophenyl)methanol (5d)
These compounds 3b–f were prepared using the general
method described by Thompson.37
Diphenyl phosphoryl azide (2.16 g, 7.80 mmol, 1.21 eq.)
was added to a solution of alcohol 5b–f (790 mg, 6.50 mmol
of 5b, 1 eq.) in dry toluene (11 mL) at 0 °C under argon atmo-
sphere. DBU (1.19 g, 7.80 mmol, 1.21 eq.) was added to the
reaction mixture. The reaction mixture was stirred for 2 h at
0 °C then for 16 h at 20 °C. The resulting two-phase mixture
was washed with water (2 × 10 mL) and 5% HCl (10 mL). The
organic layer was evaporated in vacuo and purified by flash
column chromatography.
The crude product was purified by silica flash column chro-
matography (0–10% EtOAc in LP) to afford 5d as white solid
(911 mg, 52%) Rf: 0.14 (10% EtOAc in LP, UV and KMnO4
1
dip). H NMR (400 MHz, CDCl3) δ 2.45 (1H, s, OH), 4.62 (2H,
s), 7.26 (2H, d, J 8.5 Hz), 7.32 (2H, d, J 8.4 Hz); 13C NMR (100
MHz, CDCl3) δ 64.3, 128.2, 128.6, 133.2, 139.2 in agreement
with that described by Shaikh.35
1-(Azidomethyl)-4-methylbenzene (3b)
(2-Chlorophenyl)methanol (5e)
The crude product was purified by silica flash column chroma-
tography (0–10% EtOAc in LP) to afford 5e as white solid (1.08
g, 61%) Rf: 0.17 (10% EtOAc in LP, UV and KMnO4 dip). 1H
NMR (400 MHz, CDCl3) δ 2.37 (1H, s, OH), 4.78 (2H, s), 7.23–
7.31 (2H, m), 7.37 (1H, dd, J 1.6 Hz, J 7.6 Hz), 7.47–7.50 (1H, m);
The crude product was purified by silica flash column chro-
matography (5% EtOAc in LP) to afford 3b as colourless oil
(766 mg, 81%) Rf: 0.66 (10% EtOAc in LP, UV and KMnO4
1
dip). H NMR (300 MHz, CDCl3) δ 2.41 (3H, s), 4.32 (2H, s),
7.25 (4H, s); 13C NMR (75 MHz, CDCl3) δ 21.1, 54.5, 128.2,
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129.4, 132.2, 138.0 in agreement with that described by
Kim.38
1-(Azidomethyl)naphthalene (3f)
1-(Azidomethyl)-4-methoxybenzene (3c)
The crude product was purified by silica flash column
chromatography (5% EtOAc in LP) to afford 3f as
colourless oil (645 mg, 92%) Rf: 0.72 (10% EtOAc in LP,
UV and KMnO4 dip). 1H NMR (300 MHz, CDCl3) δ 4.76
(2H, s), 7.47–7.48 (2H, m), 7.52–7.67 (2H, m), 7.86–7.94
(2H, m), 8.04–8.07 (1H, m) in agreement with that de-
scribed by Amegadzie.42 13C NMR (75 MHz, CDCl3) δ 52.9,
123.4, 125.1, 126.1, 126.6, 127.2, 128.7, 129.3, 130.9, 131.3,
133.8.
The crude product was purified by silica flash column chro-
matography (5% EtOAc in LP) to afford 3c as colourless oil
(554 mg, 89%) Rf: 0.58 (10% EtOAc in LP, UV and KMnO4
1
dip). H NMR (400 MHz, CDCl3) δ 3.84 (3H, s), 4.29 (2H, s),
6.94 (2H, d, J 8.7 Hz), 7.27 (2H, d, J 8.7 Hz); 13C NMR (100
MHz, CDCl3) δ 54.3, 55.2, 114.1, 127.3, 129.7, 159.6 in agree-
ment with that described by Pietruszka.39
General method for preparation of 1,2,3-triazole carboxylic
acid derivatives (1a–f)
1-(Azidomethyl)-4-chlorobenzene (3d)
These compounds 1a–f were prepared using the general
method described by Mindt.43 To a stirred solution of azides
3a–f (3.00 mmol) in t-butanol (15 mL), propiolic acid (210
mg, 185 μL, 3.00 mmol) was added, followed by a solution of
sodium ascorbate (119 mg, 0.6 mmol) and copperIJII) acetate
monohydrate (60 mg, 0.30 mmol) in water (15 mL). The
greenish blue reaction mixture was stirred for 14 h. The reac-
tion mixture was evaporated in vacuo and the residue was fil-
tered through a plug of silica to afford the triazole com-
pounds 1a–f.
The crude product was purified by silica flash column chro-
matography (5% EtOAc in LP) to afford 3d as colourless oil
(556 mg, 87%) Rf: 0.64 (10% EtOAc in LP, UV and KMnO4
1
dip). H NMR (300 MHz, CDCl3) δ 4.33 (2H, s), 7.26 (2H, d, J
8.7 Hz), 7.37 (2H, d, J 8.6 Hz); 13C NMR (75 MHz, CDCl3) δ
54.0, 129.0, 129.5, 133.8, 134.2 in agreement with that de-
scribed by Elson.40
1-Benzyl-1H-1,2,3-triazole-4-carboxylic acid (1a)43
1-(Azidomethyl)-2-chlorobenzene (3e)
The crude product was purified by silica flash column chro-
matography (5% EtOAc in LP) to afford 3e as colourless oil
(539 mg, 84%) Rf: 0.69 (10% EtOAc in LP, UV and KMnO4
1
dip). H NMR (400 MHz, CDCl3) δ 4.48 (2H, s), 7.25–7.30 (2H,
m), 7.36–7.43 (2H, m); 13C NMR (100 MHz, CDCl3) δ 52.2,
127.1, 129.6, 129.7, 130.0, 133.3, 133.7 in agreement with that
described by Ikawa.41
The crude product was purified with 60% EtOAc in LP
followed by 50% methanol in CHCl3 to afford 1a as a green
solid (548 mg, 90%). Rf: 0.15 (60% EtOAc in LP and 3 drops
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acetic acid, UV and KMnO4 dip). ESI-MS, m/z: 226 [M + Na]+.
1-(4-Chlorobenzyl)-1H-1,2,3-triazole-4-carboxylic acid (1d)
+
HRMS calculated for C10H9N3NaO2
226.0592, found
1
226.0589. H NMR (300 MHz, DMSO-d6) δ 5.63 (2H, s), 7.31–
7.40 (5H, m), 8.74 (1H, s); 13C NMR (75 MHz, DMSO-d6) δ
53.0, 128.0, 128.3, 128.76, 128.84, 135.6, 206.6 in agreement
with that reported by Mindt.43
1-(4-Methylbenzyl)-1H-1,2,3-triazole-4-carboxylic acid (1b)
The crude product was purified with 60% EtOAc in LP followed
by 50% methanol in CHCl3 to afford 1d as a light brown solid
(433 mg, 61%). Rf: 0.16 (60% EtOAc in LP and 3 drops acetic
acid, UV and KMnO4 dip). ESI-MS, m/z: 260 [M + Na]+. HRMS
calculated for C10H8ClN3NaO2 260.0203, found 260.0195. 1H
+
NMR (300 MHz, DMSO-d6) δ 5.61 (2H, s), 7.34 (2H, d, J 8.5 Hz),
7.43 (2H, d, J 8.5 Hz), 8.56 (1H, s); 13C NMR (75 MHz, DMSO-
d6) δ 52.1, 125.0, 128.8, 129.9, 132.9, 133.6, 134.8, 162.5.
The crude product was purified with 60% EtOAc in LP
followed by 50% methanol in CHCl3 to afford 1b as a green
solid (543 mg, 83%). Rf: 0.19 (60% EtOAc in LP and 3 drops
1-(2-Chlorobenzyl)-1H-1,2,3-triazole-4-carboxylic acid (1e)
acetic acid, UV and KMnO4 dip). ESI-MS, m/z: 240 [M + Na]+
and 216 [M
−
H]−. HRMS calculated for C11H10N3O2
−
216.0773, found 216.0788. 1H NMR (400 MHz, DMSO-d6) δ
2.26 (s, 3H), 5.60 (2H, s), 7.16 (2H, d, J 6.6 Hz), 7.21 (2H, d, J
7.5 Hz), 8.84 (1H, br s); 13C NMR (100 MHz, DMSO-d6) δ 20.7,
52.5, 127.9, 128.1, 129.3, 129.4, 132.6, 137.7.
1-(4-Methoxybenzyl)-1H-1,2,3-triazole-4-carboxylic acid (1c)
The crude product was purified with 60% EtOAc in LP followed
by 50% methanol in CHCl3 to afford 1e as a brown solid (530
mg, 75%). Rf: 0.16 (60% EtOAc in LP and 3 drops acetic acid,
UV and KMnO4 dip). ESI-MS, m/z: 260 [M + Na]+. HRMS calcu-
+
1
lated for C10H8ClN3NaO2 260.0203, found 260.0199. H NMR
(300 MHz, DMSO-d6) δ 5.72 (2H, s), 7.23 (1H, dd, J 2.1 Hz, J 6.9
Hz), 7.33–7.42 (2H, m), 7.51 (1H, dd, J 1.8 Hz, J 7.5 Hz), 8.50
(1H, s); 13C NMR (75 MHz, DMSO-d6) δ 50.7, 127.8, 128.3,
129.6, 130.3, 130.5, 132.6, 133.1, 142.7, 162.6.
1-(Naphthalen-1-ylmethyl)-1H-1,2,3-triazole-4-carboxylic acid (1f)
The crude product was purified with 60% EtOAc in LP
followed by 50% methanol in CHCl3 to afford 1c as a
yellow solid (607 mg, 87%). Rf: 0.15 (60% EtOAc in LP
and 3 drops acetic acid, UV and KMnO4 dip). ESI-MS, m/
+
z: 256 [M + Na]+. HRMS calculated for C11H11N3NaO3
256.0698, found 256.0696. 1H NMR (300 MHz, DMSO-d6)
δ 3.72 (s, 3H), 5.57 (2H, s), 6.92 (2H, d, J 7.8 Hz), 7.31
(2H, d, J 8.0 Hz), 8.74 (1H, br s); 13C NMR (75 MHz,
DMSO-d6)
δ 52.1, 55.1, 114.09, 114.14, 127.4, 128.1,
129.5, 129.8, 159.3.
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The crude product was purified with 60% EtOAc in LP
followed by 50% methanol in CHCl3 to afford 1f as a brown
solid (529 mg, 70%). Rf: 0.16 (60% EtOAc in LP and 3 drops
acetic acid, UV and KMnO4 dip). ESI-MS, m/z: 276 [M + Na]+.
1979, 254, 8340–8342; (c) P. Ling and R. M. Roberts, J. Biol.
Chem., 1993, 268, 6896–6902; (d) M. B. Twitchett, G. Schenk,
M. A. S. Aquino, D. T. Y. Yiu, T. C. Lau and A. G. Sykes,
Inorg. Chem., 2002, 41, 5787–5794; (e) P. V. Bernhardt, G.
Schenk and G. J. Wilson, Biochemistry, 2004, 43,
10387–10392; ( f ) G. Schenk, Y. B. Ge, L. E. Carrington, C. J.
Wynne, I. R. Searle, B. J. Carroll, S. Hamilton and J. de
Jersey, Arch. Biochem. Biophys., 1999, 370, 183–189.
4 (a) K. Hollberg, K. Hultenby, A. R. Hayman, T. M. Cox and
G. Andersson, Exp. Cell Res., 2002, 279, 227–238; (b) G. W.
Oddie, G. Schenk, N. Z. Angel, N. Walsh, L. W. Guddat, J. De
Jersey, A. I. Cassady, S. E. Hamilton and D. A. Hume, Bone,
2000, 27, 575–584.
5 P. R. Nuttleman and R. M. Roberts, J. Biol. Chem., 1990, 265,
12192–12199.
6 A. J. Janckila, D. H. Neustadt, Y. R. Nakasato, J. M. Halleen,
T. Hentunen and L. T. Yam, Clin. Chim. Acta, 2002, 320,
49–58.
7 J. M. Halleen, S. Raisanen, J. J. Salo, S. V. Reddy, G. D.
Roodman, T. A. Hentunen, P. P. Lehenkari, H. Kaija, P. Vihko
and H. K. Vaananen, J. Biol. Chem., 1999, 274, 22907–22910.
8 F. Guillemin, L. Martinez, M. Calvert, C. Cooper, T. Ganiats,
M. Gitlin, R. Horne, A. Marciniak, J. Pfeilschifter, S.
Shepherd, A. Tosteson, S. Wade, D. Macarios and N.
Freemantle, Osteoporosis Int., 2013, 24, 3001–3010.
9 (a) T. Sozen, L. Ozisik and N. C. Basaran, Eur. J. Rheumatol.,
2017, 4, 46–56; (b) J. Y. Reginster and N. Burlet, Bone,
2006, 38, 4–9; (c) N. C. Wright, A. C. Looker, K. G. Saag, J. R.
Curtis, E. S. Delzell, S. Randall and B. Dawson-Hughes,
J. Bone Miner. Res., 2014, 29, 2520–2526.
+
HRMS calculated for C14H11N3NaO2 276.0749, found
276.0734. 1H NMR (300 MHz, DMSO-d6) δ 6.07 (2H, s), 7.41
(1H, d, J 6.9 Hz), 7.48–7.61 (3H, m), 7.95 (2H, t, J 9.3 Hz),
8.18 (1H, d, J 7.7 Hz), 8.30 (1H, s); 13C NMR (75 MHz, DMSO-
d6) δ 50.7, 123.2, 125.6, 126.2, 126.8, 127.3, 128.7, 129.0,
130.6, 131.5, 133.3, 133.5, 163.1.
Conflicts of interest
There are no conflicts to declare.
Acknowledgements
We thank Dr. Tri Le for assistance with NMR studies and Mr.
Graham MacFarlane for MS measurements. This work was
funded by Australian Research Council Grants DP0986292
and DP150104358.
Notes and references
1 (a) N. Mitic, S. J. Smith, A. Neves, L. W. Guddat, L. R. Gahan
and G. Schenk, Chem. Rev., 2006, 106, 3338–3363; (b) G.
Schenk, N. Mitic, L. R. Gahan, D. L. Ollis, R. P. McGeary and
L. W. Guddat, Acc. Chem. Res., 2012, 45, 1593–1603; (c) G.
Schenk, N. Mitic, G. R. Hanson and P. Comba, Coord. Chem.
Rev., 2013, 257, 473–482; (d) N. Mitic, M. Miraula, C. Selleck,
K. S. Hadler, E. Uribe, M. M. Pedroso and G. Schenk, in
Advances in Protein Chemistry and Structural Biology: Metal-
containing Enzymes, ed. C. Z. Christov, Elsevier, Oxford,
2014, vol. 97, pp. 49–81; (e) A. Reithmeier, T. Lundbaeck, M.
Haraldsson, M. Frank, B. Ek-Rylander, P.-G. Nyholm, A.-L.
Gustavsson and G. Andersson, Chem. Biol. Drug Des.,
2018, 92, 1255–1271.
10 (a) J. M. Halleen, H. Kaija, J. J. Stepan, P. Vihko and H. K.
Vaananen, Arch. Biochem. Biophys., 1998, 352, 97–102; (b) H.
Yaziji, A. J. Janckila, S. C. Lear, A. W. Martin and L. T. Yam,
Am. J. Clin. Pathol., 1995, 104, 397–402.
11 J. Schindelmeiser, F. Gullotta and D. Munstermann, Pathol.,
Res. Pract., 1989, 185, 184–186.
2 (a) B. C. Antanaitis and P. Aisen, Adv. Inorg. Biochem.,
1983, 5, 111–136; (b) A. R. Hayman and T. M. Cox, J. Biol.
Chem., 1994, 269, 1294–1300; (c) S. J. Smith, A. Casellato,
K. S. Hadler, N. Mitic, M. J. Riley, A. J. Bortoluzzi, B.
Szpoganicz, G. Schenk, A. Neves and L. R. Gahan, J. Biol.
Inorg. Chem., 2007, 12, 1207–1220; (d) G. Schenk, R. A.
Peralta, S. C. Batista, A. J. Bortoluzzi, B. Szpoganicz, A. K.
Dick, P. Herrald, G. R. Hanson, R. K. Szilagyi, M. J. Riley,
L. R. Gahan and A. Neves, J. Biol. Inorg. Chem., 2008, 13,
139–155; (e) N. Mitic, C. J. Noble, L. R. Gahan, G. R. Hanson
and G. Schenk, J. Am. Chem. Soc., 2009, 131, 8173–8179; ( f )
N. Mitic, K. S. Hadler, L. R. Gahan, A. C. Hengge and G.
Schenk, J. Am. Chem. Soc., 2010, 132, 7049–7054; (g) G.
Schenk, C. L. Boutchard, L. E. Carrington, C. J. Noble, B.
Moubaraki, K. S. Murray, J. de Jersey, G. R. Hanson and S.
Hamilton, J. Biol. Chem., 2001, 276, 19084–19088.
12 J. Schindelmeiser, H. J. Radzun and D. Munstermann,
Pathol., Res. Pract., 1991, 187, 209–213.
13 J. D. Hoyer, C. Y. Li, L. T. Yam, C. A. Hanson and P. J.
Kurtin, Am. J. Clin. Pathol., 1997, 108, 308–315.
14 P. Lang, M. Schultzberg and G. Andersson, J. Histochem.
Cytochem., 2001, 49, 379–396.
15 (a) Y. C. Chung, C. H. Ku, T. Y. Chao, J. C. Yu, M. M. Chen
and S. H. Lee, Cancer Epidemiol., Biomarkers Prev., 2006, 15,
424–428; (b) L. B. Tanko, M. A. Karsdal, C. Christiansen and
D. J. Leeming, Cancer Metastasis Rev., 2006, 25, 659–668.
16 (a) D. C. Crans, C. M. Simone, R. C. Holz and L. Que,
Biochemistry, 1992, 31, 11731–11739; (b) T. W. Elliott, N.
Mitic, L. R. Gahan, L. W. Guddat and G. Schenk, J. Braz.
Chem. Soc., 2006, 17, 1558–1565.
17 (a) V. I. Kuznetsov, A. C. Hengge and S. J. Johnson,
Biochemistry, 2012, 51, 9869–9879; (b) D. C. Crans, J. J. Smee,
E. Gaidamauskas and L. Q. Yang, Chem. Rev., 2004, 104,
849–902.
3 (a) N. Mitic, M. Valizadeh, E. W. W. Leung, J. de Jersey, S.
Hamilton, D. A. Hume, A. I. Cassady and G. Schenk, Arch.
Biochem. Biophys., 2005, 439, 154–164; (b) B. P. Gaber, J. P.
Sheridan, F. W. Bazer and R. M. Roberts, J. Biol. Chem.,
18 J. K. Myers, S. M. Antonelli and T. S. Widlanski, J. Am. Chem.
Soc., 1997, 119, 3163–3164.
Med. Chem. Commun.
This journal is © The Royal Society of Chemistry 2018

View Article Online
MedChemComm
Research Article
19 M. Valizadeh, G. Schenk, K. Nash, G. W. Oddie, L. W.
Guddat, D. A. Hume, J. de Jersey, T. R. Burke and S.
Hamilton, Arch. Biochem. Biophys., 2004, 424, 154–162.
20 C. F. Schwender, S. A. Beers, E. Malloy, K. Demarest, L.
Minor and K. H. W. Lau, Bioorg. Med. Chem. Lett., 1995, 5,
1801–1806.
21 R. P. McGeary, P. Vella, J. Y. W. Mak, L. W. Guddat and G.
Schenk, Bioorg. Med. Chem. Lett., 2009, 19, 163–166.
22 S. H. Mohd-Pahmi, W. M. Hussein, G. Schenk and R. P.
McGeary, Bioorg. Med. Chem. Lett., 2011, 21, 3092–3094.
23 (a) W. Faridoon, M. Hussein, N. U. Islam, L. W. Guddat, G.
Schenk and R. P. McGeary, Bioorg. Med. Chem. Lett.,
2012, 22, 2555–2559; (b) W. M. Hussein, D. Feder, G.
Schenk, L. W. Guddat and R. P. McGeary, Eur. J. Med. Chem.,
2018, 157, 462–479.
24 (a) G. Schenk, L. T. Guddat, Y. Ge, L. E. Carrington, D. A.
Hume, S. Hamilton and J. de Jersey, Gene, 2000, 250,
117–125; (b) T. Klabunde, N. Strater, B. Krebs and H. Witzel,
FEBS Lett., 1995, 367, 56–60; (c) T. Klabunde, B. Stahl, H.
Suerbaum, S. Hahner, M. Karas, F. Hillenkamp, B. Krebs
and H. Witzel, Eur. J. Biochem., 1994, 226, 369–375.
25 (a) N. Strater, B. Jasper, M. Scholte, B. Krebs, A. P. Duff,
D. B. Langley, R. L. Han, B. A. Averill, H. C. Freeman and
J. M. Guss, J. Mol. Biol., 2005, 351, 233–246; (b) C. Selleck, D.
Clayton, L. R. Gahan, N. Mitic, R. P. McGeary, M. M.
Pedroso, L. W. Guddat and G. Schenk, Chem. – Eur. J.,
2017, 23, 4778–4781.
R. P. McGeary, P. Vella, G. Schenk and R. H. Abd El-hameed,
Eur. J. Med. Chem., 2011, 46, 6075–6082; ( f ) Y. Yusof,
D. T. C. Tan, O. K. Arjomandi, G. Schenk and R. P. McGeary,
Bioorg. Med. Chem. Lett., 2016, 26, 1589–1593.
29 J. L. Beck, L. A. Mcconachie, A. C. Summors, W. N. Arnold, J.
Dejersey and B. Zerner, Biochim. Biophys. Acta, 1986, 869,
61–68.
30 H. D. Campbell, D. A. Dionysius, D. T. Keough, B. E. Wilson,
J. D. Jersey and B. Zerner, Biochem. Biophys. Res. Commun.,
1978, 82, 615–620.
31 R. Thomsen and M. H. Christensen, J. Med. Chem., 2006, 49,
3315–3321.
32 W. C. Still, M. Kahn and A. Mitra, J. Org. Chem., 1978, 43,
2923–2925.
33 L. Campbell-Verduyn, P. H. Elsinga, L. Mirfeizi, R. A. Dierckx
and B. L. Feringa, Org. Biomol. Chem., 2008, 6, 3461–3463.
34 E. R. Zartler and M. J. Shapiro, Curr. Opin. Chem. Biol.,
2005, 9, 366–370.
35 N. S. Shaikh, K. Junge and M. Beller, Org. Lett., 2007, 9,
5429–5432.
36 U. Ragnarsson, L. Grehn, L. S. Monteiro and H. L. S. Maia,
Synlett, 2003, 2386–2388.
37 A. S. Thompson, G. R. Humphrey, A. M. Demarco, D. J.
Mathre and E. J. J. Grabowski, J. Org. Chem., 1993, 58,
5886–5888.
38 B. H. Kim, S. M. Park and Y. S. Lee, Postech Found (UYPO) Univ
Pushan Polytechnic (UYPU-Non-standard), US6884882-B1;
JP2005194257-A; KR2005072620-A; KR540509-B1; JP4071738-B2.
39 J. Pietruszka and G. Solduga, Eur. J. Org. Chem.,
2009, 5998–6008.
26 J. U. Flanagan, A. I. Cassady, G. Schenk, L. W. Guddat and
D. A. Hume, Gene, 2006, 377, 12–20.
27 D. Feder, W. M. Hussein, D. J. Clayton, M. W. Kan, G.
Schenk, R. P. McGeary and L. W. Guddat, Chem. Biol. Drug
Des., 2012, 80, 665–674.
40 K. E. Elson, I. D. Jenkins and W. A. Loughlin, Org. Biomol.
Chem., 2003, 1, 2958–2965.
28 (a) O. K. Arjomandi, W. M. Hussein, P. Vella, Y. Yusof, H. E.
Sidjabat, G. Schenk and R. P. McGeary, Eur. J. Med. Chem.,
2016, 114, 318–327; (b) W. M. Hussein Faridoon, P. Vella, N.
Ul Islam, D. L. Ollis, G. Schenk and R. P. McGeary, Bioorg.
Med. Chem. Lett., 2012, 22, 380–386; (c) W. M. Hussein, S. S.
Fatahala, Z. M. Mohamed, R. P. McGeary, G. Schenk, D. L.
Ollis and M. S. Mohamed, Chem. Biol. Drug Des., 2012, 80,
500–515; (d) R. P. McGeary, D. T. C. Tan, C. Selleck, M. M.
Pedroso, H. E. Sidjabat and G. Schenk, Eur. J. Med. Chem.,
2017, 137, 351–364; (e) M. S. Mohamed, W. M. Hussein,
41 T. Ikawa, H. Sajiki and K. Hirota, Tetrahedron, 2005, 61,
2217–2231.
42 A. K. Amegadzie, K. M. Gardinier, E. J. Hembre, P. A.
Hipskind, L. N. Jungheim, B. S. Muehl, K. A. Savin, K. J.
Thrasher, S. A. Boyd, A. Amegadzie, K. Gardinier, E.
Hembre, P. Hipskind, L. Jungheim, B. Muehl, K. Savin, K.
Thrasher and S. Boyd, Lilly & Co Eli (ELIL) WO2005000821-
A1; EP1638944-A1; US2006160794-A1.
43 T. L. Mindt and R. Schibli, J. Org. Chem., 2007, 72,
10247–10250.
This journal is © The Royal Society of Chemistry 2018
Med. Chem. Commun.