Development of quinone analogues as dynamin GTPase inhibitors

Article abstract of DOI:10.1016/j.ejmech.2014.06.070

Virtual screening of the ChemDiversity and ChemBridge compound databases against dynamin I (dynI) GTPase activity identified 2,5-bis-(benzylamino)-1,4- benzoquinone 1 as a 273 ± 106 μM inhibitor. In silico lead optimization and focused library-led synthesis resulted in the development of four discrete benzoquinone/naphthoquinone based compound libraries comprising 54 compounds in total. Sixteen analogues were more potent than lead 1, with 2,5-bis-(4-hydroxyanilino)-1,4-benzoquinone (45) and 2,5-bis(4-carboxyanilino)- 1,4-benzoquinone (49) the most active with IC₅₀ values of 11.1 ± 3.6 and 10.6 ± 1.6 μM respectively. Molecular modelling suggested a number of hydrogen bonding and hydrophobic interactions were involved in stabilization of 49 within the dynI GTP binding site. Six of the most active inhibitors were evaluated for potential inhibition of clathrin-mediated endocytosis (CME). Quinone 45 was the most effective CME inhibitor with an IC_50(CME) of 36 ± 16 μM.

Full text of DOI:10.1016/j.ejmech.2014.06.070

European Journal of Medicinal Chemistry 85 (2014) 191e206
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Development of quinone analogues as dynamin GTPase inhibitors
,
,
c
,
,
Kylie A. MacGregor a 2, Mohammed K. Abdel-Hamid a 2, Luke R. Odell a 1, Ngoc Chau b,
Ainslie Whiting b, Phillip J. Robinson b, Adam McCluskey a
,
*
a Centre for Chemical Biology, Chemistry, School of Environmental and Life Sciences, The University of Newcastle, Callaghan, NSW 2308, Australia
b Children's Medical Research Institute, University of Sydney, Westmead, NSW 2145, Australia
c Department of Medicinal Chemistry, Faculty of Pharmacy, Assiut University, Assiut 71526, Egypt
a r t i c l e i n f o
a b s t r a c t
Article history:
Virtual screening of the ChemDiversity and ChemBridge compound databases against dynamin I (dynI)
Received 6 March 2014
Received in revised form
27 June 2014
Accepted 28 June 2014
Available online 5 July 2014
GTPase activity identified 2,5-bis-(benzylamino)-1,4-benzoquinone 1 as a 273
106 mM inhibitor. In
silico lead optimization and focused library-led synthesis resulted in the development of four discrete
benzoquinone/naphthoquinone based compound libraries comprising 54 compounds in total. Sixteen
analogues were more potent than lead 1, with 2,5-bis-(4-hydroxyanilino)-1,4-benzoquinone (45) and
2,5-bis(4-carboxyanilino)-1,4-benzoquinone (49) the most active with IC50 values of 11.1
3.6 and
10.6 1.6 M respectively. Molecular modelling suggested a number of hydrogen bonding and hydro-
phobic interactions were involved in stabilization of 49 within the dynI GTP binding site. Six of the most
active inhibitors were evaluated for potential inhibition of clathrin-mediated endocytosis (CME).
Quinone 45 was the most effective CME inhibitor with an IC50(CME) of 36 16 mM.
m
Keywords:
Dynamin
Endocytosis
Quinones
Crown Copyright © 2014 Published by Elsevier Masson SAS. All rights reserved.
Dynamin inhibitors
Modelling
1. Introduction
particular, two crystal structures for the near full length mamma-
lian dynI (lacking the PRD domain) have been solved, providing
valuable information regarding the complex dynamics involved in
dynamins' GTPase cycle and its role as a fission protein in cells
[10,11].
Dynamin GTPase belongs to the dynamin super-family of large
GTPases which play crucial roles in the catalysis of membrane
fission during clathrin mediated endocytosis (CME), regulation of
cellular actin dynamics and are important for cytokinesis [1]. There
are three classical dynamins (dynI, dynII and dynIII) which each
CME is a highly regulated cellular process in which growth
factors and membrane-bound receptors are concentrated in
have low affinity for guanine nucleotides (10e100
m
M), high basal
invaginating clathrin-coated pits (CCPs). These CCPs pinch off to
form vesicles which carry the cargo into the cell [10]. In the final
stages of CME, dynamin GTPase can assemble into collars at the
necks of the deeply invaginated CCPs to catalyse membrane fission
apparently by neck constriction [3,11,12]. Synaptic vesicle endocy-
tosis (SVE) in neural cells is based on a similar mechanism to CME
and plays a role in the internalization of membrane to allow syn-
aptic vesicle recycling thus sustaining synaptic transmission [11].
SVE and CME differ mainly in the dynamin genes that are utilized in
each case and unique regulation of SVE by protein dephosphory-
lation. While dynII is found throughout the body and is the major
scission protein for CME, dynI is mainly found in neuronal cells
where it coexists with dynII and dynIII. In neural cells dynI protein
is expressed at about 50-fold higher levels than the two other
classical dynamins and is critical for neural cell functions [1].
We have previously reported the identification and develop-
ment of the MiTMAB [13], Bis-T [14,15], RTIL [16], Iminiodyn™ [17],
Pthaladyn™ [18], Dyngo™ [19,20], Rhodadyn™ [21] series and two
generations of Dynole™ [22e24] compounds as dynamin
GTP turnover (0.4e1 minꢀ1) and the propensity for oligomerization
into helical arrays around a template [2]. Each dynamin comprises
five structural domains including an amino-terminal G domain that
binds and hydrolyses GTP, a middle domain involved in self-
assembly and oligomerization,
a pleckstrin homology (PH)
domain responsible for interactions with the plasma membrane
and inducing the hemifission state, a GTPase effector domain (GED)
which is also involved in self-assembly, and a proline- and arginine-
rich (PRD) domain that interacts with SH3 domains in accessory
proteins [3e5]. With the exception of the PRD, tertiary structures of
all dynamin's individual domains have been reported [6e9]. In
* Corresponding author.
E-mail address: Adam.McCluskey@newcastle.edu.au (A. McCluskey).
Current address: Organic Pharmaceutical Chemistry, Department of Medicinal
1
Chemistry, Uppsala Biomedical Centre, Uppsala University, Box 574, SE-751 23
Uppsala, Sweden.
2
These authors contributed equally to this work.
http://dx.doi.org/10.1016/j.ejmech.2014.06.070
0223-5234/Crown Copyright © 2014 Published by Elsevier Masson SAS. All rights reserved.

192
K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
inhibitors [25]. We have also described the virtual screening of the
ChemBridge and the ChemDiversity chemical databases against a
dynI GTPase domain homology model (hDynI) using ICM flexible
ligand-grid receptor algorithm to target the GTP binding site which
led to the identification of the Pthaladyn™ compound series [17].
Herein we report on the development of a second series of com-
pounds from our initial virtual screening hits were we identified
and with a range of substituted amines we assembled a discrete
library of amine substituted 1,4-naphthoquinones, which were
screened for inhibition of dynI GTPase activity (Scheme 1 and
Table 1) [35].
Of the 13 analogues synthesized, only 9, 10 and 12 showed
significant improvement in activity compared to the lead 1
(IC50 ¼ 273 106
mM), with IC50 values of 30.3, 29.5 and 22.4 mM
2,5-bis-(benzylamino)cyclohexa-2,5-diene-1,4-dione (1) as
273 106 M potent dynI inhibitor (Fig. 1).
a
respectively. However, the activity profile for other derivatives was
consistent, excepting 6e8, with the contact preference maps and
suggested the symmetry inherent in the parent compound was not
absolutely required, but may have bestowed additional features
advantageous for dynI inhibition.
m
Quinones have been previously explored in a wide range of
different drug development pathways with multiple analogues
reported to be active in a myriad of different biological screens
[26,27]. In our virtual screening approaches, quinones were
retained due to their ability to link potential binding elements that
would not otherwise be accessed [28,29]. A number of quinones
have progressed to clinical trials, e.g. ubiquinone 10, diaziquone,
mitoxantrone hydrochloride and monobenzone [29]; and in a
related area (to this study) the natural product bolinaquinone has
been shown to display high levels of specificity towards clathrin
[30]. Herein we anticipated that our medicinal chemistry efforts
with lead 1, would add to the SAR of dynamin inhibition already
established [13e25,31e33].
The predicted binding poses for the 9, 10 and 12 showed a high
degree of similarity and revealed a number of hydrophobic and
hydrogen bonding interactions with the binding site (Fig. 3). One of
the quinone carbonyl groups was predicted to accept two hydrogen
bonds from P-loop residues, specifically from the backbone and
side chain protons of Ser45 and Lys44. The other carbonyl group
formed a hydrogen bond with Ser41. The hydroxyl moiety donated
a hydrogen bond to either the backbone or the side chain of Arg237
and accepted a Ser46 side chain hydrogen bond. The phenyl sub-
stituent made key hydrophobic interactions with the Ala42 and
Ile242 side chains, whilst the naphthaquinone core participated in
an areneecation interaction with Lys44 in addition to making hy-
drophobic contacts with the Ile63, Leu137 and Pro138 side chains.
The amine linker donated a moderate strength hydrogen bond to
the backbone carbonyl of Gly60.
Given our findings with the naphthoquinone analogues (3e16),
we next synthesized a focused library of symmetrically substituted
p-benzoquinone analogues (1, 17e33) (Scheme 2). Treatment of p-
benzoquinone (16) with two equivalents of 2-benzylamine in
ethanol at reflux for 18 h [36]. These analogues were screened for
their ability to inhibit dynI and these data are presented in Table 2.
Of the p-benzoquinone focused library analogues only 18, 21, 23
and 33 returned improved activity with dynI IC50 values of 29, 170,
2. Results and discussion
Structural optimization of 1 commenced with molecular dock-
ing and investigation of its predicted orientation within dynI's GTP-
binding site. Docking studies were conducted using AutoDock 4.2
and the dynI-GMPPCP co-crystal structure (PDB: 3ZYC) [3,34]. Ex-
amination of the GMMPCP binding pose revealed a number of
strong hydrogen bonding interactions between the active site res-
idues and GMPPCP (ESI). The hydrogen bonding interactions were
similar to that observed with 1 (ESI). These included a strong
hydrogen bond (2.4 Å) between one of the quinone carbonyl
moieties and the side chain of the catalytically crucial Ser41 [2].
Other hydrogen bond interactions ranged between moderate and
weak (2.8e3.2 Å) and involved the backbones and/or side chains of
Ser45, Ser46 and Lys206. One of the terminal phenyl rings showed
an areneecation interaction with the side chain of Lys44, similar to
that observed between Lys206 and the GMPPCP guanine core
within the published co-crystal. However, compound 1 did not
show any of the contacts with amino acids at the “dynamin-specific
loop” (residues 236e246) which were present with GMPPCP [3].
The lipophilic/hydrophilic contact surface preference in the GTP
active site showed a requirement to balance the hydrophilic/hy-
drophobic characters in subsequent modified quinone (Fig. 2).
While the dynamin-specific loop favoured the presence of hydro-
phobic contacts (Fig. 2A) the P- and G4-loop region favoured hy-
drophilic interactions (Fig. 2B).
112 and 58
analogues within this focused library returning IC50 values in the
29e170 M range. The enhanced activity of 18 was attributed to
mM, respectively. Only the oxygen bearing benzylamine
m
interaction with most key residues at the active site as shown from
its “Protein Ligand Interaction Fingerprints” (PLIF, Supplementary
data). Although 23 bears a hydroxyl group suggesting a good
hydrogen bonding network it showed a lower level of interactions
compared to 18 (missing the interaction with Lys44, Gly139 and
Asn208) suggesting that the C]O moiety in 18 plays an important
role in favourable interactions within the GTP biding site. Methyl-
ation of the hydroxyl group giving the 4-methoxy substituted 23
further reduced potency with the loss of the interactions with Gly62
and Val64 in switch I region of the binding site (21, IC50 ¼ 170
mM).
This highlighted the potential importance of hydrogen bonding
within the GTP-binding site. Other interesting results regarding the
activities of the benzylamine derivatives were shown by the loss of
the inhibitory activity in the case of 25, directing attention to the
importance of optimizing the length of the linker between the
quinone and terminal aromatic moieties. In addition, the PLIF
highlighted the binding site residues that were commonly involved
in the interactions which include Ser41, Ser46 and Lys206.
Our initial modelling studies suggested that 1,4-
naphthoquinone analogues would retain the required interactions
within the active site and consequently fuller engagement with the
key active site residues. Thus, treatment of 1,4-naphthoquinone (2)
Most of the alkyl or alkenyl substituted quinones (26e33) dis-
played no dynI inhibition, except 33 which was more active
(IC50 ¼ 58 23
mM) than the lead 1. This promoted us to further
investigate the activity of aliphatic side chain derivatives through
the introduction of heteroatom bearing substituents to target more
hydrogen bond interactions at the binding site. Compounds 34e43
were synthesized essentially as described in Scheme 1, using a
selected library of primary amines, and subsequently screened for
dynI inhibition (Table 3) [35].
Fig. 1. Chemical structure of virtual screening lead 2,5-bis-(benzylamino)cyclohexa-
2,5-diene-1,4-dione (1).

K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
193
Fig. 2. Maps for contact preferences at the GTP-binding site of dynI with the docked compound 1 (sticks) showing hydrophobic favouring contacts (A, green) mostly near the
dynamin specific loop (residues 236e246) and hydrophilic favouring interactions (B, purple) closer to the PeG4-loops side. (For interpretation of the references to colour in this
figure legend, the reader is referred to the web version of this article.)
As was predicted from our modelling study, the introduction of
a terminal eOH moiety gave moderate level of dynI inhibition with
the ethyl (34) containing derivative returning an IC50 value of
116 49 (Table 3). However, further chain elongation to propyl (35)
from the data obtained with the alkylamine derivatives that a
hydrogen bond donating substituent with a two carbon spacer to
the amino group was important for retaining dynI inhibition
activity.
or butyl (36) resulted in
a
complete loss of activity (dynI
With only moderate enhancement in the activity observed by
the omission of the peripheral phenyl moiety, we decided to retain
this moiety during further structural optimization. It was also clear
from the inhibitory data that hydrogen bonding interactions were
crucial for inhibitor activity. To further explore this, the optimum
length for the linker between the quinone core and the peripheral
aromatic moiety was examined. Two molecules were designed by
elongating the aminomethyl spacer in 23 to aminoethyl (44) and
shortening it to amino (45) and these were subjected to modelling
analysis. Both docked poses were inspected prior to the post-
docking energy minimization in order to investigate possible ste-
ric clashes or unfavourable ligand orientations. The docking pose
for 44 revealed an unfavourable orientation within the active site
with one of the aromatic side chains clashing with GTP binding
pocket residues (Fig. 4A).
IC50 > 300 M). Methylation of the terminal OH group (37 and 38)
m
or its replacement with other heteroatoms (39 and 40) was
generally detrimental to activity. Replacement of the bromo sub-
stituent in 33 with chloro (41 and 43) lead to a compete loss of
activity. The same result was observed on shortening of the linker
unit between the bromo and the quinone moieties (42). It was clear
O
O
O
H
N
i
R
O
Analysis of the docking pose of 45 revealed a better hydrogen
bonding network with no steric clashes at the active site. The en-
ergy minimized pose of the docked 45 showed similar interactions
to those observed with the parent 1, with an essentially identical
2
3-15
Scheme 1. Reagents and conditions: (i).

194
K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
Table 1
internalization of Tfn-A594 into U2OS cells as well as dynII GTPase
activity (Table 5).
The results revealed that five compounds possessed some de-
gree of CME inhibitory activity using transferrin endocytosis, with
only 50 failing to exert any effect (50, IC50(CME): Not Active). Com-
pound 45 was the most potent CME inhibitor, returning an IC50(CME)
Inhibition of dynI PS-stimulated GTPase activity by amine substituted 1,4-
naphthoquinone analogues 3e15.
of 36
modest (1, IC50(CME): 120 27
levels of CME inhibitory activity.
16
m
M, whilst the remaining active analogues exhibited
m
M) to weak (48, IC50(CME): >300 M)
m
Whilst these results could indicate differences in dynI/II inhib-
itory activity and specificity for one isoform, results obtained from
dynII GTPase inhibition by selected compounds showed no isoform
selectivity (Fig. 5). Accordingly, the differences in inhibitory activ-
ities against full length dynI versus CME were likely due to differ-
ences in the membrane permeability of each compound. We
therefore calculated the polar accessible surface area (PASA) [37,38]
for 1, 45, and 47e50 which showed values that were consistent
with the apparent CME activities (Table 5). Of the six examined
analogues, 1 had the lowest PASA value (107 Å2) and the smallest
difference between the dynI IC50 value and the CME IC50 value.
Conversely, 48 and 49 both possessed higher PASA values (298 and
302 Å2 respectively), resulting in lower membrane permeability
Compound
R
DynI
IC50 (mM)
3
4
5
6
7
8
9
10
11
12
13
14
15
e(CH2)2OH
e(CH2)3OH
Ph
2-COOH-Ph
3-COOH-Ph
4-COOH-Ph
3-CH2OH-Ph
4-CH2OH-Ph
3-OH-Ph
4-OH-Ph
eCH2Ph
eCH2-4-OCH3-Ph
eCH2-4-OH-Ph
e
e
e
314
e
e
30.3 5.7a
29.5 5.7a
109
22.4 1.5a
e
e
150
and up to 20-fold decrease in the CME IC50 relative to the dynI IC50
.
(ꢀ) not active up to 300
mM.
This was also true when comparing 49 (PASA value of 302 Å2) and
45 (PASA value of 204 Å2), where a COOH to OH substitution caused
an equipotent dynI inhibitory activity, but a 5-fold improvement in
CME inhibitory activity.
a
n ¼ 3.
orientation at the active site (Fig. 5B). Both hydroxyl moieties were
involved in strong backbone and side chain hydrogen bonding with
different loops of the binding site. In addition, both amino groups
were able to act as hydrogen bond donors generating two strong
backbone interactions with Gly60 and Arg237.
Differences in membrane permeability were also likely to ac-
count for the differences in CME block noted for the isomeric de-
rivatives 47e49. The isomer 47 (o-COOH) was found to exhibit the
highest CME inhibitory activity of the three analogues, returning an
Based on the predicted more favourable binding for 45, a series
of substituted aniline analogues was synthesized for screening
against dynI GTPase activity (Table 4).
The data presented in Table 4 showed that the parent aniline
(46) was devoid of activity. However, the introduction of a 2-CO2H
(47), 3-CO2H (48) or 4-CO2H (49) saw a significant increase in dynI
inhibition in line with that predicted by our modelling analysis,
with these compounds returning dynI IC50 values of 50.2, 25.4 and
IC50(CME) of 162 41 mM. Whilst this represents more than 3-fold
reduction in potency compared to the dynI inhibitory activity, it
was a significant improvement compared to the greater than 20-
fold reduction in potency observed for 48 and 49. Further evi-
dence for the membrane permeability problem amongst the
carboxyl containing analogues was provided through the calcula-
tion of the hydrogen bond donating capacity (HBD_cap) for each
derivative (Table 5). The ortho carboxyl isomer (47) showed a
significantly lower HBD_cap (304) compared to the other two iso-
mers (31 and 32) which show HBD_cap of 399 and 479 respectively.
This low HBD_cap for 47 was due to the positioning of the COOH
moiety proximal to the NH group allowing for an intramolecular
hydrogen bond formation which contributes in shielding the po-
larity of 47, facilitating membrane permeability [39,40].
10.6 mM respectively. Polar substituents were generally best toler-
ated, with compounds containing hydroxyl and carboxylate sub-
stituents exhibiting the greatest inhibitory activity. The position of
the aromatic substituent was found to affect the inhibition activity
(47 vs. 49, 45 vs. 50 and 56 vs. 58). As anticipated from modelling
observations, the methyl (51), methoxy (52e54) and bromo (55)
substituents were poorly tolerated, with IC50 > 300 mM.
Despite the differences in the rank order between the CME and
dynI inhibitory activities of the examined quinones, the results
were consistent with the inhibition of dynI by aminoquinone
compounds resulting in inhibition of CME, provided the inhibitor
was both sufficiently potent against the protein target, and was cell
permeable.
Molecular docking simulations revealed similarities between
the binding pose of 49 and that of 45, with 49 forming strong
network of hydrogen bonding interactions with residues at the
dynamin-specific loop (Supplementary data). These new in-
teractions involved extensive hydrogen bonding between the
carboxyl groups and the backbones/side chains of residues Asp208,
Val234, Val235 and Asn236.
3. Conclusion
Finally, in order to confirm that these quinone based compounds
target the GTP active site of dynI, a series of Michaelis Menten ki-
netic experiments were conducted with quinone-58. The Line-
weavereBurke plots revealed clear competitive inhibition with
respect to Mg2þ.GTP for dynI (Fig. 5), consistent with the modelling
predictions.
Herein we have reported the identification 2,5-bis(benzyla-
mino)cyclohexa-2,5-diene-1,4-dione (1) as a 273 106 mM potent
inhibitor of dynI. This lead was identified through a virtual
screening approach. Detailed docking analysis suggested a number
of favourable hydrogen bonding, hydrophobic and electrostatic
interactions with the binding site. Using a combination of molec-
ular modelling led design and focused library synthesis we devel-
oped four discrete naphthoquinone (3e15) and p-benzoquinone
(17e33; 34e43; and 45e58) based compound libraries.
2.1. Inhibition of clathrin-mediated endocytosis
The parent compound 1 as well as five quinone analogues 45
and 47e50 were quantified for their ability to inhibit the
Biological evaluation highlighted a preference for polar sub-
stituents capable of forming hydrogen bonds and/or participating

K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
195
Fig. 3. A. Predicted binding pose of 10 in the dynI GTP-binding site. Hydrogen bonding interactions (purple dash lines) are observed with Ser41, Lys44, Ser45, Ser46, Gly60 and
Arg237, whilst hydrophobic contacts are observed between the ligand and Ala42, Ile63, Val64, Leu137, Pro138 and Ile242. B. 2D representation for the same pose shown in A
showing in addition the areneecation interaction between the naphthoquinone core and the side chain of Lys44. (For interpretation of the references to colour in this figure legend,
the reader is referred to the web version of this article.)
in electrostatic interactions with residues within the binding site.
This was consistent with our modelling analysis. Analogues con-
taining short-chain alcohol substituents (2 carbons) were the most
potent dynI inhibitors.
quinone moiety in the form of a substituted aniline was preferred
(c.f. substituted benzylamines). Among these analogues, the p-hy-
droxy and p-carboxy aniline derivatives 45 and 49 with IC50 values
of 11.1 3.6 and 10.6 1.6 mM, respectively were the most potent.
A positional preference was also noted on the aromatic rings,
with para-substituted analogues consistently exhibiting a higher
potency than their positional isomers. Direct coupling to the
This represented a >20-fold activity enhancement relative to the
lead, 1. Docking studies revealed that the binding conformation of
45 and 49 in the dynI GTP-binding site is stabilized primarily by
hydrogen bonding and hydrophobic interactions. The close prox-
imity of these compounds to the G2 consensus region of the GTP-
binding site leads to the suggestion that its activity may result
from the compound interfering with the co-ordination of Mg2þ
with the binding site. This was confirmed by enzyme kinetic ex-
periments which showed quinone-58 acted as a competitive in-
hibitor with respect to Mg2þ.GTP for dynI.
H
R
O
O
N
i
n
O
N
H
R
n
O
Six active analogues (1, 45 and 47e50) were evaluated for their
CME inhibition among which five were found to possess some
degree of CME inhibitory activity. The p-OH substituted 45 was the
16
1, 17-43, 45-58
Scheme 2. Reagents and conditions: (i) RNH2 (0.5 equivalents), ethanol, reflux, 18 h.
most active against CME with an IC50(CME) of 36 16 mM consistent

196
K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
Table 2
code: 3ZYC) [3]. The pdb file was imported into Accelrys Discovery
Studio package and was processed through deletion of the ligand,
the water of crystallization and the co-crystallized ions. Residues
at the enzyme active site that interact with the bound ligand were
set as flexible residues during docking simulations. The ligand 3D
structures were built using Molecular Operating Environment
(MOE) software, their partial charges were assigned and energy
minimized through MMFF94x force field to a gradient of 0.001,
then saved as pdb files. Docking simulations were performed us-
ing AutoDock 4.2 [34] and AutoDock tools (ADT) [41] packages
applying the selective receptor flexibility function. A grid map of
44 ꢁ 44 ꢁ 44 grid points, centred on the GMPPCP ligand of the
complex structures, was used to cover the binding pocket. A
spacing of 0.375 Å was set centred at X ¼ 12.685; Y ¼ 18.265;
Z ¼ ꢀ4.842 and flexible residues were set to Ser41, Lys44, Ser46
and Arg237. A Lamarckian genetic algorithm was used with the
following parameters: number of individuals in population, 100;
maximum number of energy evaluations, 2,500,000; maximum of
generations, 27,000. Docking results were clustered with a rms
tolerance of 2.0 Å. The lowest energy conformation from each
cluster was taken as possible binding mode and was typed with
CHARMm force field then subjected to a steepest descent energy
minimization to a maximum steps of 5000 or until converged to
rms gradient of 0.01. The minimized structures were investigated
using MOE software for possible interactions and contacts map
calculations.
Inhibition of dynI PS-stimulated GTPase activity by quinone analogues 1, 17e33.
Compound
n
R
DynI
IC50
(mM)
1
1
1
1
1
1
1
1
1
1
2
1
2
3
4
5
2
1
3
Ph
273 106a
e
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
3-pyridyl
4-COOH-Ph
4-CH3-Ph
4-F-Ph
4-OCH3Ph
4-Br-Ph
4-OH-Ph
4-Cl-Ph
Ph
CH3
CH3
CH3
CH3
29
e
e
170
e
112
e
e
e
e
e
e
CH3
>300
e
CH(CH3)2
CH]CH2
Br
e
58 23b
(ꢀ) not active up to 300
m
M.
a
n ¼ 3.
Polar accessible surface area (PASA) and hydrogen bonding ca-
pacity (HBD_cap) were calculated using the “Descriptors” function
implemented in MOE software. The 3D structures of the ligands
were built using MOE builder and were energy minimized to a
gradient of 0.001 using MMFF94x force field and were saved as an
mdb file for descriptors calculation.
b
n ¼ 2.
with its dynI inhibition. No isoform-selectivity was observed as the
4 compounds tested essentially equipotent against dynII. The low
levels of CME inhibition observed with 47e49 were attributed to
their low levels of predicted membrane permeability and thus
reduced cellular uptake.
4.2. Biology
4.2.1. Materials
4. Experimental
Phosphatidylserine (PS), phenylmethylsulfonylfluoride (PMSF),
Tween 80, fibronectin and DAPI were from Sigma Aldrich (St. Louis,
MO). GTP was from Roche Applied Science (Basel, Switzerland) and
leupeptin was from Bachem (Bubendorf, Switzerland). Para-
formaldehyde (PFA) was from BDH (AnalaR Merck Chemicals,
Darmstadt, Germany). Phosphate buffered salts, foetal bovine
serum (FBS) and Dulbecco's Minimal Essential Medium (DMEM)
were from Invitrogen (Paisley, Strathclyde, UK). Texas-red conju-
gated transferrin (Tfn-TxR), Alexa-488 conjugated EGF (EGF-A488),
Alexa-594 conjugated Transferrin (Tfn-A594), DAPI and Fluo3-AM
were from Molecular Probes (Eugene, OR, USA). All other reagents
were of analytical reagent grade or better.
4.1. Molecular modelling
Molecular modelling studies were performed using the crystal
structure of dynI GTPase domain bound to GMPPCP ligand (PDB
Table 3
Inhibition of dynI PS-stimulated GTPase activity by quinone analogues 34e43.
4.2.2. Compounds
Compounds were synthesized in-house and made up as 30 mM
stock solutions in 100% DMSO and diluted in 50% v/v DMSO/20 mM
Tris/HCl pH 7.4 or cell media prior to further dilutions in the assay.
The final DMSO concentrations in the GTPase or endocytosis assays
were at most 3.3% or 1% respectively. The GTPase assay for dynI was
unaffected by DMSO up to 3.3% or 2.5% respectively. Stock solutions
were stored at ꢀ20 ꢂC for several months. Dynole-34-2™ was used
as a positive control showing comparable activity to the previously
reported data [22].
Compound
n
R
DynI
IC50
(mM)
34
35
36
37
38
39
40
41
42
43
2
3
4
2
3
2
2
2
2
3
OH
OH
OH
OCH3
OCH3
SCH3
N(CH3)2
Cl
116 49b
ea
e
e
e
e
>300
ea
e
4.2.3. Protein production
Br
Cl
Native endogenous dynI was purified from sheep brain by
extraction from the peripheral membrane fraction of whole brain
[13], and affinity purification on GST-Amph2-SH3-sepharose as
previously described [18], yielding 8e10 mg of protein from 250 g
e
(ꢀ) not active up to 300
mM.
a
n ¼ 2.
b
n ¼ 3.

K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
197
Fig. 4. A. Predicted binding pose of 44 (sticks representation) in the GTP-binding site showing steric clashes between the hydroxyphenyl moiety and residues at the active site in red
line spheres. B. 2D predicted binding pose of 45 in the GTP-binding site. Hydrogen bonding interactions are observed between 45 and Ser41, Ser45, Ser46, Gly60, Thr65, Asp136,
Leu137, Lys206, Asn236 and Arg237 with potential hydrophobic contacts observed with Val64 and Leu137 and Val234 (see Fig. 2 for legend). (For interpretation of the references to
colour in this figure legend, the reader is referred to the web version of this article.)
of sheep brain. Recombinant rat dynamin II was expressed and
purified as previously described [13]. In brief, dynII (His-6-tagged)
DNA was inserted in to the pIEx-6 vector and the plasmid was used
to transfect Sf21 insect cells using transfection with poly-
ethylenimine using a DNA:polyethylenimine ratio of 1:5 for 48 h.
Following transfection the cells were harvested and 9 mg dynII was
purified to >98% purity from 1 L of insect cell suspension culture
(1 ꢁ 106 cells/mL) using GST-Amph2-SH3-sepharose as reported
for dynI [13].
4.2.5. CME assay
CME was examined by quantifying the uptake of Alexa-495-
conjugated transferrin (Tfn-A495) in serum-starved U2OS cells
using methods described previously [22]. Briefly, cells were plated
on fibronectin-coated 96-well glass-bottomed plate to 70e80%
confluency, then serum-starved overnight (16 h). Cells were pre-
incubated with test compounds or vehicle (1% DMSO) for 30 min
prior to the addition of 4
m
g/mL Tfn-A495 for 8 min at 37 ꢂC.
Surface-bound Tfn was removed with an ice-cold acid wash solu-
tion (0.2 M acetic acid, 0.5 M NaCl, pH 2.8) for 15 min followed by an
ice-cold PBS wash for 5 min. Cells were immediately fixed with 4%
PFA for 10 min at room temperature. Nuclei were stained using
DAPI. Quantitative analysis of Tfn-A594 uptake in U2OS cells was
performed by high content imaging. Nine images were collected
from each well, averaging 40e50 cells per image. The average in-
tegrated intensity of Tfn-A594 signal per cell was calculated for
each well using MetaXpress (Molecular Devices), and the data
expressed as a percentage of control cells (vehicle treated only).
4.2.4. Dynamin GTPase assay
The Malachite Green method was used for the sensitive colori-
metric detection of orthophosphate (Pi) release from GTP as pre-
viously described [17]. DynI or dynII (each at 50 nM in the reaction)
activity was stimulated by phosphatidylserine (PS) liposomes for
30 (dynI) or 90 (dynII) minutes. Data analysis and enzyme kinetics
using non-linear regression was performed using Prism 5 (Graph-
Pad Software Inc., San Diego, CA).

198
K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
Büchi or Heidolph rotary evaporators. Melting points were recor-
ded in open capillaries on a Stuart SMP11 Melting Point Apparatus.
Temperatures are expressed in degrees Celsius (ꢂC) and are un-
corrected. Where available, literature values are provided and
appropriately referenced. Electrospray mass spectra were recorded
using 10% DMSO/H2O or HPLC-grade methanol or acetonitrile as
carrier solvents on a Shimadzu LCeMS spectrometer.
Nuclear magnetic resonance (NMR) spectroscopy was per-
formed on a Bruker Avance 300 MHz spectrometer, where proton
NMR (1H NMR) spectra and carbon NMR (13C NMR) spectra were
acquired at 300 and 75 MHz respectively, or a Bruker Avance III
400 MHz spectrometer, where 1H NMR and 13C NMR were acquired
at 400 and 100 MHz respectively. All spectra were recorded in
deuterated dimethyl sulfoxide (DMSO-d6), obtained from Sigma
Aldrich or Cambridge Isotope Laboratories Inc., unless otherwise
stated, with the residual solvent peaks used as the internal refer-
ence (
d
2.49 (quintet) and
d
39.7 (septet) for 1H NMR and 13C NMR
) were measured in parts per
respectively). Chemical shifts (
d
Fig. 5. Competitive kinetics of quinone-58 with respect to Mg2þ.GTP. The data depicts
quinone-58 concentration dependent changes in double-reciprocal plot between
million (ppm) and referenced against the internal reference peaks.
Coupling constants (J) were measured in Hertz (Hz).
substrate (GTP at 60e200
mM) and reaction velocity (V). The data corresponds to
NMR assignments were determined through the interpretation
of one- and two-dimensional spectra, specifically gradient hetero-
nuclear single quantum correlation (gHSQC), gradient hetero-
nuclear multiple bond correlation (gHMBC) and distortionless
enhancement by polarization transfer quaternary (DEPTQ) spec-
troscopy. Multiplicities are denoted as singlet (s), broad singlet (br
s), doublet (d), doublet of doublets (dd), doublet of doublet of
doublets (ddd), triplet (t), triplet of doublets (td), doublet of triplets
(dt), quartet (q), quintet (quin) and multiplet (m). Peaks are listed in
increasing chemical shift in the following format: chemical shift
(integration (1H), multiplicity (1H), coupling constant (1H).
quinone-58 concentrations of 30 (C), 20 (
mean SEM of four independent experiments each conducted in triplicate.
◊
), 12 ( ), and 8 (▫) M. Error bars represent
D
m
IC50 values were calculated using Graphpad Prism 5 and data was
expressed as mean 95% confidence interval (CI) for 3 wells and
~1200 cells.
4.3. Chemistry
4.3.1. General methods
All reactions were performed using standard laboratory equip-
ment and glassware. Solvents and reagents were purchased from
Sigma Aldrich, Lancaster International or TCI and used as received.
Organic solvents were bulk quality, and were distilled from glass
prior to use. Organic solvent extracts were dried with magnesium
sulfate (MgSO4), and dried under reduced pressure with either
4.3.2. General procedure for the synthesis of 2,5-bis(benzylamino)-
1,4,benzoquinone (1)
To a stirred solution of 1,4-benzoquinone (0.403 g, 3.73 mmol)
in ethanol (5 mL), was added a solution of benzylamine (0.224 g,
2.41 mmol) in ethanol (5 mL). The resulting dark red solution was
stirred at reflux for 18 h. The obtained mixture was allowed to cool
to room temperature, then kept at 4 ꢂC for 24 h. The precipitate was
washed sequentially with ethanol and ether, and dried under
vacuum to afford the desired compound as an orange-red solid.
Yield 0.088 g (47%); mp 229e230 ꢂC (Lit. 252 ꢂC; 234 ꢂC; 259 ꢂC;
259 ꢂC; 253 ꢂC) [42].
Table 4
Inhibition of dynI PS-stimulated GTPase activity by substituted aniline analogues
45e58.
1H NMR (DMSO-d6)
(10H, m), 8.30 (2H, t, J ¼ 6.6 Hz, NH).
13C NMR (DMSO-d6)
45.3, 93.4, 127.3, 127.4, 128.6, 137.5, 151.1,
177.9.
d
4.35 (4H, d, J ¼ 6.6 Hz), 5.16 (2H, s), 7.27
d
MS (ESIþ) m/z: 319 (MþH), HRMS (ESIþ) m/z calculated for
C
20H18N2O2 (MþH) 319.1447; found 319.1441.
Compound
R1
R2
R3
DynI
IC50
(mM)
4.3.3. General procedure for the synthesis of 2-(2-
hydroxyethylamino)-1,4-naphthoquinone (3)
45
46
47
48
49
50
51
52
53
54
55
56
57
58
H
H
COOH
H
H
H
H
OCH3
H
H
H
COOH
H
OH
H
H
OCH3
H
H
H
CH2OH
H
OH
H
H
H
COOH
H
CH3
H
H
OCH3
H
11.1 3.6b
e
50.2 14.4b
25.4 11.3b
10.6 1.6a
57.9 17.2b
e
To
a stirred suspension of 1,4-naphthoquinone (0.328 g,
2.07 mmol) in methanol (8 mL) was added ethanolamine (0.104 g,
1.70 mmol). The resulting dark red solution was stirred at 25 ꢂC for
18 h. The obtained mixture was poured into cold water (40 mL), and
the resulting orange-brown suspension was stirred at 25 ꢂC for
30 min. The mixture was kept at 4 ꢂC overnight. The precipitate was
collected by filtration, washed with ice-cold water, and dried under
vacuum to afford the desired compound as a red-brown solid. Yield
0.125 g (37%); mp 146e147 ꢂC (Lit. 159e160 ꢂC) [43].
e
H
H
>300
>300
387
>>300
e
Br
CH2OH
H
H
H
H
1H NMR (DMSO-d6)
d
3.23 (2H, q, J ¼ 6.0 Hz), 3.59 (2H, t,
CH2OH
21.5 3.1a
J ¼ 6.0 Hz), 4.85 (1H, br s, OH), 5.72 (1H, s), 7.28 (1H, t, J ¼ 6.0 Hz,
NH), 7.71 (1H, td, J ¼ 1.5, 7.5 Hz), 7.81 (1H, td, J ¼ 1.5, 7.5 Hz), 7.93
(1H, dd, J ¼ 1.5, 7.5 Hz), 7.97 (1H, dd, J ¼ 1.5, 7.5 Hz).
(ꢀ) not active up to 300
mM.
a
n ¼ 2.
b
n ¼ 3.

K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
199
Table 5
CME inhibition data for quinone based dynamin inhibitors, compared with in vitro dynI GTPase activity inhibition.
Compound
Structure
DynI IC50
(
m
M)
DynII IC50
(m
M)
CME IC50
(
mM)
PASAa
HBD_capb
1
273
e
120 27
107
193
329
45
11.1 3.6f
23.6
36 16
204
47
48
50.2 14.4f
NDc
162 41
243
298
304
399
25.4 11.3e
10.6 1.6e
57.9 17.2f
14.7
>300
49
50
9.1
200 41
302
479
43.7
e
NCd
NCd
(ꢀ) Not active up to 300
mM.
a
Polar accessible surface area (Å2).
Hydrogen bond donor capacity.
Not determined.
b
c
d
e
f
Not calculated.
n ¼ 2.
n ¼ 3.
13C NMR (DMSO-d6)
133.3, 135.0, 148.9, 181.5, 181.7.
d
44.8, 58.6, 99.8, 125.5, 126.1, 130.5, 132.4,
MS (ESIþ) m/z: 232 (MþH), HRMS (ESIþ) m/z calculated for
13H13NO3 (MþH) 232.0974; found 232.0968.
C
MS (ESIþ) m/z: 218 (MþH), HRMS (ESIþ) m/z calculated for
C
12H11NO3 (MþH) 218.0817; found 218.0812.
4.3.5. 2-(Anilino)-1,4-naphthoquinone (5)
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and aniline
to afford the title compound as a dark red solid. Yield 0.209 g (49%);
mp 185 ꢂC (Lit. 191e192 ꢂC) [44].
4.3.4. 2-(3-Hydroxypropylamino)-1,4-naphthoquinone (4)
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and 3-
amino-1-propanol to afford the title compound as an orange-
brown solid. Yield 0.158 g (40%); mp 120e121 ꢂC.
1H NMR (DMSO-d6)
d 6.09 (1H, s), 7.21 (1H, m), 7.41 (4H, m),
7.77 (1H, td, J ¼ 1.5, 7.5 Hz), 7.85 (1H, td, J ¼ 1.5, 7.5 Hz), 7.94 (1H,
dd, J ¼ 1.5, 7.5 Hz), 8.05 (1H, dd, J ¼ 1.5, 7.5 Hz), 9.21 (1H, br s,
NH).
1H NMR (DMSO-d6)
d 1.73 (2H, m), 3.23 (2H, m), 3.49 (2H, m),
4.61 (1H, t, J ¼ 5.1 Hz, OH), 5.67 (1H, s), 7.53 (1H, t, J ¼ 6.0 Hz, NH),
7.70 (1H, td, J ¼ 1.5, 7.5 Hz), 7.81 (1H, td, J ¼ 1.5, 7.5 Hz), 7.93 (1H, dd,
J ¼ 1.5, 7.5 Hz), 7.96 (1H, dd, J ¼ 1.5, 7.5 Hz).
13C NMR (DMSO-d6)
d 102.1, 123.9 (2C), 125.5 (2C), 126.3, 129.5
(2C), 130.6, 132.8 (2C), 135.1, 138.3, 146.4, 181.8, 182.8.
13C NMR (DMSO-d6)
d
30.6, 39.8, 58.9, 99.4, 125.5, 126.0, 130.6,
MS (ESIþ) m/z: 250 (MþH), HRMS (ESIþ) m/z calculated for
132.3, 133.4, 135.0, 148.7, 181.4, 181.7.
C
16H11NO2 (MþH) 250.0868; found 250.0863.

200
K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
4.3.6. 2-(2-Carboxyanilino)-1,4-naphthoquinone (6)
1H NMR (DMSO-d6)
d
4.49 (2H, d, J ¼ 5.7 Hz), 5.19 (1H, t,
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and an-
thranilic acid to afford the title compound as an orange-red solid.
Yield 0.156 g (56%); mp > 250 ꢂC (Lit. 237e240 ꢂC (dec.)) [44].
J ¼ 5.7 Hz, OH), 6.06 (1H, s), 7.32 (2H, d, J ¼ 8.7 Hz), 7.38 (2H, d,
J ¼ 8.7 Hz), 7.77 (1H, td, J ¼ 1.5, 7.5 Hz), 7.85 (1H, td, J ¼ 1.5, 7.5 Hz),
7.94 (1H, dd, J ¼ 1.5, 7.5 Hz), 8.05 (1H, dd, J ¼ 1.5, 7.5 Hz), 9.20 (1H, br
s, NH).
1H NMR (DMSO-d6)
d
6.56 (1H, s), 7.22 (1H, m), 7.68 (2H, m), 7.81
13C NMR (100 MHz) (DMSO-d6)
d 62.7, 101.9, 123.7, 125.4, 126.3,
(1H, td, J ¼ 1.8, 7.5 Hz), 7.88 (1H, td, J ¼ 1.8, 7.5 Hz), 7.97 (1H, dd,
J ¼ 1.8, 7.5 Hz), 8.03 (1H, m), 8.08 (1H, dd, J ¼ 1.8, 7.5 Hz), 10.82 (1H,
br s, NH).
127.5, 130.6, 132.8, 135.1, 136.7, 139.9, 146.5, 181.8, 182.6.
MS (ESIþ) m/z: 280 (MþH), HRMS (ESIþ) m/z calculated for
C
17H13NO3 (MþH) 280.0974; found 280.0978.
13C NMR (DMSO-d6)
d 105.2, 120.0, 120.8, 123.5, 125.6, 126.6,
130.5, 132.2, 132.5, 133.2, 134.3, 135.2, 140.3, 143.9, 169.0, 181.6,
183.5.
4.3.11. 2-(3-Hydroxyanilino)-1,4-naphthoquinone (11)
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and 3-
aminophenol to afford the title compound as a dark brown solid.
Yield 0.250 g (53%); mp > 250 ꢂC (Lit. 242 ꢂC) [45].
MS (ESIꢀ) m/z: 292 (MꢀH), HRMS (ESIꢀ) m/z calculated for
C
17H11NO4 (MꢀH) 292.0610; found 292.0615.
1H NMR (DMSO-d6)
d 6.14 (1H, m), 6.61 (1H, m), 6.79 (1H, d,
4.3.7. 2-(3-Carboxyanilino)-1,4-naphthoquinone (7)
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and 3-
aminobenzoic acid to afford the title compound as a dark red
solid. Yield 0.239 g (64%); mp > 250 ꢂC (Lit. 264e265 ꢂC) [44].
J ¼ 1.8 Hz), 6.80 (1H, m), 7.21 (1H, t, J ¼ 8.4 Hz), 7.71 (1H, td, J ¼ 1.5,
7.5 Hz), 7.85 (1H, td, J ¼ 1.5, 7.5 Hz), 7.94 (1H, dd, J ¼ 1.5, 7.5 Hz), 8.05
(1H, dd, J ¼ 1.5, 7.5 Hz), 9.08 (1H, br s, NH), 9.62 (1H, br s, OH).
13C NMR (100 MHz) (DMSO-d6)
d 102.4, 110.5, 112.6, 114.4, 125.4,
1H NMR (DMSO-d6)
d
6.12 (1H, s), 7.55 (1H, t, J ¼ 7.8 Hz), 7.64
126.3, 130.2, 130.6, 132.8 (2C), 135.0, 139.3, 146.2, 158.2, 181.7, 182.7.
(1H, m), 7.76 (1H, m), 7.79 (1H, td, J ¼ 1.5, 7.5 Hz), 7.86 (1H, td,
J ¼ 1.5, 7.5 Hz), 7.92 (1H, d, J ¼ 1.8 Hz), 7.95 (1H, dd, J ¼ 1.5, 7.5 Hz),
8.07 (1H, dd, J ¼ 1.5, 7.5 Hz), 9.34 (1H, br s, NH), 13.15 (1H, br s,
COOH).
MS (ESIþ) m/z: 266 (MþH), HRMS (ESIþ) m/z calculated for
C
16H11NO3 (MþH) 266.0817; found 266.0812.
4.3.12. 2-(4-Hydroxyanilino)-1,4-naphthoquinone (12)
13C NMR (100 MHz) (DMSO-d6)
d 102.5, 124.2, 125.5, 126.0,
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and 4-
aminophenol to afford the title compound as a dark brown solid.
Yield 0.341 g (76%); mp > 250 ꢂC (Lit. 225 ꢂC) [45].
126.3, 128.1, 129.8, 130.6, 132.1, 132.7, 132.9, 135.1, 138.7, 146.3, 167.0,
181.6, 182.9.
MS (ESIꢀ) m/z: 292 (MꢀH), HRMS (ESIꢀ) m/z calculated for
C
17H11NO4 (MꢀH) 292.0610; found 292.0615.
1H NMR (DMSO-d6)
d
5.86 (1H, s), 6.82 (2H, d, J ¼ 8.7 Hz), 7.15
(2H, d, J ¼ 8.7 Hz), 7.75 (1H, td, J ¼ 1.5, 7.5 Hz), 7.83 (1H, td, J ¼ 1.5,
7.5 Hz), 7.93 (1H, dd, J ¼ 1.5, 7.5 Hz), 8.03 (1H, dd, J ¼ 1.5, 7.5 Hz),
9.04 (1H, br s, NH), 9.56 (1H, br s, OH).
4.3.8. 2-(4-Carboxyanilino)-1,4-naphthoquinone (8)
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and 4-
aminobenzoic acid to afford the title compound as a dark red
solid. Yield 0.148 g (40%); mp > 250 ꢂC (Lit. 319e321 ꢂC) [44].
13C NMR (DMSO-d6)
d 101.0, 116.0, 125.4, 126.0, 126.2, 129.1,
130.6, 132.6, 133.0, 135.0, 147.2, 155.5, 181.9, 182.3.
MS (ESIþ) m/z: 266 (MþH), HRMS (ESIþ) m/z calculated for
1H NMR (DMSO-d6)
d
6.35 (1H, s), 7.52 (2H, d, J ¼ 8.7 Hz), 7.80
C
16H11NO3 (MþH) 266.0817; found 266.0812.
(1H, td, J ¼ 1.5, 7.5 Hz), 7.87 (1H, td, J ¼ 1.5, 7.5 Hz), 7.96 (1H, dd,
J ¼ 1.5, 7.5 Hz), 7.97 (2H, d, J ¼ 8.7 Hz), 8.07 (1H, dd, J ¼ 1.5, 7.5 Hz),
9.38 (1H, br s, NH).
4.3.13. 2-(Benzylamino)-1,4-naphthoquinone (13)
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and ben-
zylamine to afford the title compound as an orange-brown solid.
Yield 0.235 g (56%); mp 154e155 ꢂC (Lit. 154.7e155.7 ꢂC; 155 ꢂC)
[44].
13C NMR (DMSO-d6)
d 104.3, 122.6, 125.7, 126.6, 126.8, 130.7,
131.0, 132.7, 133.3, 135.4, 142.9, 145.5, 167.2, 181.7, 183.5.
MS (ESIꢀ) m/z: 292 (MꢀH), HRMS (ESIꢀ) m/z calculated for
C
17H11NO4 (MꢀH) 292.0610; found 292.0616.
1H NMR (DMSO-d6)
d
4.44 (2H, d, J ¼ 6.6 Hz), 5.55 (1H, s), 7.27
4.3.9. 2-(3-(Hydroxymethyl)anilino)-1,4-naphthoquinone (9)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-naphthoquinone and 3-
aminobenzylalcohol to afford the title compound as a maroon
solid. Yield 0.243 g (52%); mp 186e187 ꢂC.
(5H, m), 7.71 (1H, td, J ¼ 1.5, 7.5 Hz), 7.80 (1H, td, J ¼ 1.5, 7.5 Hz), 7.89
(1H, dd, J ¼ 1.5, 7.5 Hz), 7.99 (1H, dd, J ¼ 1.5, 7.5 Hz), 8.15 (1H, t,
J ¼ 6.6 Hz, NH).
13C NMR (DMSO-d6)
d 45.3, 100.6, 125.5, 126.1, 127.3 (3C), 128.7
1H NMR (DMSO-d6)
d
4.52 (2H, d, J ¼ 5.7 Hz), 5.27 (1H, t,
(2C), 130.6, 132.4, 133.2, 135.0, 137.6, 148.7, 181.6, 181.8.
MS (ESIþ) m/z: 264 (MþH), HRMS (ESIþ) m/z calculated for
J ¼ 5.7 Hz, OH), 6.11 (1H, s), 7.15 (1H, d, J ¼ 7.8 Hz), 7.23 (1H, dd,
J ¼ 2.4, 7.8 Hz), 7.34 (1H, d, J ¼ 2.4 Hz), 7.38 (1H, t, J ¼ 7.8 Hz), 7.71
(1H, td, J ¼ 1.5, 7.5 Hz), 7.85 (1H, td, J ¼ 1.5, 7.5 Hz), 7.94 (1H, dd,
J ¼ 1.5, 7.5 Hz), 8.06 (1H, dd, J ¼ 1.5, 7.5 Hz), 9.19 (1H, br s, NH).
C
17H13NO2 (MþH) 264.1025; found 264.1019.
4.3.14. 2-(4-Methoxybenzylamino)-1,4-naphthoquinone (14)
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and 4-
methoxybenzylamine to afford the title compound as a red-
brown solid. Yield 0.250 g (45%); mp 159e160 ꢂC.
13C NMR (DMSO-d6)
d 62.7, 102.1, 121.5, 122.3, 123.5,125.4, 126.3,
129.2, 130.6, 132.7, 135.0, 138.1, 144.1, 146.4, 181.7, 182.7.
MS (ESIþ) m/z: 280 (MþH), HRMS (ESIþ) m/z calculated for
C
17H13NO3 (MþH) 280.0974; found 280.0968.
1H NMR (DMSO-d6)
d
3.71 (3H, s), 4.35 (2H, d, J ¼ 6.3 Hz), 5.57
4.3.10. 2-(4-(Hydroxymethyl)anilino)-1,4-naphthoquinone (10)
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and 4-
aminobenzylalcohol to afford the title compound as a maroon
solid. Yield 0.234 g (49%); mp 212 ꢂC.
(1H, s), 6.89 (2H, d, J ¼ 8.7 Hz), 7.27 (2H, d, J ¼ 8.7 Hz), 7.71 (1H, td,
J ¼ 1.5, 7.5 Hz), 7.80 (1H, td, J ¼ 1.5, 7.5 Hz), 7.89 (1H, dd, J ¼ 1.5,
7.5 Hz), 7.98 (1H, dd, J ¼ 1.5, 7.5 Hz), 8.09 (1H, t, J ¼ 6.3 Hz, NH).
13C NMR (DMSO-d6)
d 44.7, 55.2, 100.5, 114.1, 125.5, 126.0, 128.6,
129.3, 130.6, 132.4, 133.2, 135.0, 148.5, 158.6, 181.5, 181.8.

K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
201
MS (ESIþ) m/z: 294 (MþH), HRMS (ESIþ) m/z calculated for
18H15NO3 (MþH) 294.1130; found 294.1125.
(4JCF ¼ 2.6 Hz, C2’, C2”), 150.9 (2C2), 161.5 (1JCF ¼ 242.9 Hz, C5’, C5”),
178.0 (2C).
C
MS (ESIþ) m/z: 355 (MþH), HRMS (ESIþ) m/z calculated for
4.3.15. 2-(4-Hydroxybenzylamino)-1,4-naphthoquinone (15)
Synthesized using the same general procedure described for the
synthesis of 3, commencing with 1,4-naphthoquinone and 4-
hydroxybenzylamine to afford the title compound as a maroon
solid. Yield 0.219 g (46%); mp 222e223 ꢂC.
C
20H16F2N2O2 (MþH) 355.1258; found 355.1253.
4.3.20. 2,5-Bis(4-methoxybenzylamino)-1,4-benzoquinone (21)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 4-
methoxybenzylamine to afford the title compound as a dark
brown solid. Yield 0.485 g (67%); mp 222 ꢂC.
1H NMR (DMSO-d6)
d
4.30 (2H, d, J ¼ 6.6 Hz), 5.58 (1H, s). 6.71
(2H, d, J ¼ 8.7 Hz), 7.14 (2H, d, J ¼ 8.7 Hz), 7.70 (1H, td, J ¼ 1.5,
7.5 Hz), 7.80 (1H, td, J ¼ 1.5, 7.5 Hz), 7.89 (1H, dd, J ¼ 1.5, 7.5 Hz), 7.97
(1H, dd, J ¼ 1.5, 7.5 Hz), 8.03 (1H, t, J ¼ 6.6 Hz, NH), 9.30 (1H, br s,
OH).
1H NMR (DMSO-d6)
d
3.71 (6H, s), 4.27 (4H, d, J ¼ 6.6 Hz), 5.17
(2H, s), 6.87 (4H, d, J ¼ 8.7 Hz), 7.21 (4H, d, J ¼ 8.7 Hz), 8.19 (2H, t,
J ¼ 6.6 Hz, NH).
13C NMR (DMSO-d6)
d 44.9, 100.5, 115.4 (2C), 125.5, 126.0, 127.5,
13C NMR (DMSO-d6)
d 44.7 (2C), 55.2 (2C), 93.2 (2C), 114.1 (4C),
128.7 (1C), 130.6, 132.3, 133.2, 135.0, 148.5, 156.7, 181.5, 181.8.
128.5 (4C), 129.3 (2C), 151.0 (2C), 158.6 (2C), 177.9 (2C).
MS (ESIþ) m/z: 280 (MþH), HRMS (ESIþ) m/z calculated for
MS (ESIþ) m/z: 379 (MþH), HRMS (ESIþ) m/z calculated for
C
17H13NO3 (MþH) 280.0974; found 280.0968.
C
22H22N2O4 (MþH) 379.1658; found 379.1652.
4.3.16. 2,5-Bis(3-pyridylmethylamino)-1,4-benzoquinone (17)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 3-(ami-
nomethyl)pyridine to afford the title compound as an orange-
brown solid. Yield 0.367 g (67%); mp 222e223 ꢂC.
4.3.21. 2,5-Bis(4-bromobenzylamino)-1,4-benzoquinone (22)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 4-
bromobenzylamine to afford the title compound as a brown solid.
Yield 0.342 g (73%); mp 186e188 ꢂC.
1H NMR (DMSO-d6)
d
4.40 (4H, d, J ¼ 6.6 Hz), 5.25 (2H, s), 7.34
1H NMR (DMSO-d6)
d
4.33 (4H, d, J ¼ 6.6 Hz), 5.15 (2H, s), 7.24
(4H, d, J ¼ 8.4 Hz), 7.51 (4H, d, J ¼ 8.4 Hz), 8.29 (2H, t, J ¼ 6.6 Hz, NH).
13C NMR (DMSO-d6)
44.6 (2C), 93.6 (2C), 120.4 (2C), 129.6 (4C),
(2H, dd, J ¼ 4.8, 7.8 Hz), 7.68 (2H, dt, J ¼ 1.8, 7.8 Hz), 8.30 (2H, t,
J ¼ 6.6 Hz, NH), 8.45 (2H, dd, J ¼ 1.8, 4.8 Hz), 8.52 (2H, d, J ¼ 1.8 Hz).
d
13C NMR (100 MHz) (DMSO-d6)
d 42.8 (2C), 93.5 (2C), 123.8 (2C),
131.5 (4C), 137.1 (2C), 150.9 (2C), 178.0 (2C).
133.2 (2C), 135.2 (2C), 148.6 (2C), 149.0 (2C), 150.9 (2C), 178.1 (2C).
MS (ESIþ) m/z: 475 (MþH), HRMS (ESIþ) m/z calculated for
MS (ESIþ) m/z: 321 (MþH), HRMS (ESIþ) m/z calculated for
C
20H16Br2N2O2 (MþH) 474.9657; found 474.9651.
C
18H16N4O2 (MþH) 321.1352; found 321.1346.
4.3.22. 2,5-Bis(4-hydroxybenzylamino)-1,4-benzoquinone (23)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 4-
hydroxybenzylamine to afford the title compound as a dark pink
solid. Yield 0.170 g (54%); mp 242 ꢂC.
4.3.17. 2,5-Bis(4-carboxybenzylamino)-1,4-benzoquinone (18)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 4-(ami-
nomethyl)benzoic acid to afford the title compound as a maroon
solid. Yield 0.227 g (60%); mp > 250 ꢂC.
1H NMR (DMSO-d6)
d
4.22 (4H, d, J ¼ 6.6 Hz), 5.18 (2H, s), 6.69
1H NMR (DMSO-d6)
d
4.44 (4H, d, J ¼ 6.6 Hz), 5.15 (2H, s), 7.38
(4H, d, J ¼ 8.4 Hz), 7.89 (4H, d, J ¼ 8.4 Hz), 8.33 (2H, t, J ¼ 6.6 Hz, NH).
13C NMR (DMSO-d6)
45.1 (2C), 93.7 (2C), 127.4 (4C), 129.8 (4C),
(4H, d, J ¼ 8.7 Hz), 7.08 (4H, d, J ¼ 8.7 Hz), 8.11 (2H, t, J ¼ 6.6 Hz, NH),
9.33 (2H, br s, OH).
d
13C NMR (DMSO-d6)
d 45.0 (2C), 93.2 (2C), 115.4 (4C), 127.6 (2C),
130.0 (2C), 142.8 (2C), 151.1 (2C), 167.4 (2C), 178.1 (2C).
128.8 (4C), 151.1 (2C), 156.7 (2C), 177.9 (2C).
MS (ESIꢀ) m/z: 405 (MꢀH), HRMS (ESIꢀ) m/z calculated for
MS (ESIþ) m/z: 351 (MþH), HRMS (ESIþ) m/z calculated for
C
22H18N2O6 (MꢀH) 405.1087; found 405.1092.
C
20H18N2O4 (MþH) 351.1345; found 351.1339.
4.3.18. 2,5-Bis(4-methylbenzylamino)-1,4-benzoquinone (19)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 4-
methylbenzylamine to afford the title compound as a pink-brown
solid. Yield 0.262 g (64%); mp 238e240 ꢂC.
4.3.23. 2,5-Bis(4-chlorobenzylamino)-1,4-benzoquinone (24)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 4-
chlorobenzylamine to afford the title compound as an orange-
brown solid. Yield 0.277 g (58%); mp 222e224 ꢂC.
1H NMR (DMSO-d6)
(2H, s), 7.13 (8H, m), 8.21 (2H, t, J ¼ 6.6 Hz, NH).
13C NMR (DMSO-d6)
20.8 (2C), 45.1 (2C), 93.4 (2C), 127.2 (4C),
d
2.25 (6H, s), 4.30 (4H, d, J ¼ 6.6 Hz), 5.14
1H NMR (DMSO-d6)
d
4.34 (4H, d, J ¼ 6.6 Hz), 5.16 (2H, s), 7.30
(4H, d, J ¼ 8.7 Hz), 7.38 (4H, d, J ¼ 8.7 Hz), 8.29 (2H, t, J ¼ 6.6 Hz, NH).
13C NMR (DMSO-d6)
44.6 (2C), 93.6 (2C), 128.7 (4C), 129.3 (4C),
d
d
129.3 (4C), 134.4 (2C), 136.4 (2C), 151.1 (2C), 178.0 (2C).
131.9 (2C), 136.7 (2C), 151.0 (2C), 178.1 (2C).
MS (ESIþ) m/z: 347 (MþH), HRMS (ESIþ) m/z calculated for
MS (ESIþ) m/z: 387 (MþH), HRMS (ESIþ) m/z calculated for
C
22H22N2O2 (MþH) 347.1760; found 347.1754.
C
20H16Cl2N2O2 (MþH) 387.0667; found 387.0662.
4.3.19. 2,5-Bis(4-fluorobenzylamino)-1,4-benzoquinone (20)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 4-
fluorobenzylamine to afford the title compound as an orange-
brown solid. Yield 0.283 g (61%); mp 229e230 ꢂC.
4.3.24. 2,5-Bis(phenethylamino)-1,4-benzoquinone (25)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and phene-
thylamine to afford the title compound as a red-brown solid. Yield
0.375 g (61%); mp 218e220 ꢂC (Lit. 222 ꢂC) [46].
1H NMR (DMSO-d6)
d
4.34 (4H, d, J ¼ 6.6 Hz), 5.19 (2H, s), 7.14
(4H, t, J ¼ 8.7 Hz), 7.33 (4H, dd, J ¼ 5.4, 8.7 Hz), 8.26 (2H, t, J ¼ 6.6 Hz,
1H NMR (DMSO-d6)
7.23 (10H, m), 7.64 (2H, t, J ¼ 6.3 Hz, NH).
13C NMR (DMSO-d6)
33.6 (2C), 43.3 (2C), 92.4 (2C), 126.5 (2C),
128.5 (4C), 128.9 (4C), 138.9 (2C), 151.1 (2C), 177.5 (2C).
d 2.85 (4H, m), 3.37 (4H, m), 5.27 (2H, s),
NH).
13C NMR (DMSO-d6)
d
44.5 (2C), 93.4 (2C), 115.4 (2JCF ¼ 21.4 Hz,
d
C4’, C4”, C6’, C6”), 129.4 (3JCF ¼ 8.1 Hz, C3’, C3”, C7’, C7”), 133.7

202
K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
MS (ESIþ) m/z: 347 (MþH), HRMS (ESIþ) m/z calculated for
22H22N2O2 (MþH) 347.1760; found 347.1754.
4.3.30. 2,5-Bis(isopentylamino)-1,4-benzoquinone (31)
C
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and iso-pen-
tylamine to afford the title compound as an orange-pink solid. Yield
0.117 g (49%); mp 164e165 ꢂC (Lit. 167.3e169.7 ꢂC) [36].
4.3.25. 2,5-Bis(ethylamino)-1,4-benzoquinone (26)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and ethylamine
to afford the title compound as a pink-brown solid. Yield 0.071 g
(39%); mp 209e210 ꢂC (Lit. 218 ꢂC; 210 ꢂC) [44].
1H NMR (DMSO-d6)
d
0.88 (12H, d, J ¼ 6.6 Hz), 1.42 (4H, m), 1.56
(2H, m), 3.13 (4H, m), 5.20 (2H, s), 7.67 (2H, t, J ¼ 6.3 Hz, NH).
13C NMR (DMSO-d6)
22.4 (4C), 25.6 (2C), 36.3 (2C), 40.3 (2C),
d
92.0 (2C), 151.4 (2C), 177.4 (2C).
1H NMR (DMSO-d6)
(2H, s), 7.65 (2H, t, J ¼ 6.3 Hz, NH).
13C NMR (DMSO-d6)
13.3 (2C), 36.8 (2C), 92.0 (2C), 151.3 (2C),
177.5 (2C).
d
1.11 (6H, t, J ¼ 7.2 Hz), 3.15 (4H, m), 5.21
MS (ESIþ) m/z: 279 (MþH), HRMS (ESIþ) m/z calculated for
C
16H26N2O2 (MþH) 279.2073; found 279.2067.
d
4.3.31. 2,5-Bis(allylamino)-1,4-benzoquinone (32)
MS (ESIþ) m/z: 195 (MþH), HRMS (ESIþ) m/z calculated for
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and allylamine
to afford the title compound as a pink-red solid. Yield 0.172 g (79%);
mp 192e193 ꢂC (Lit. 170 ꢂC; 192e193 ꢂC) [48].
C
10H14N2O2 (MþH) 195.1134; found 195.1128.
4.3.26. 2,5-Bis(propylamino)-1,4-benzoquinone (27)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and propyl-
amine to afford the title compound as a pink solid. Yield 0.075 g
(37%); mp 142e143 ꢂC.
1H NMR (DMSO-d6)
d
3.79 (4H, m), 5.11 (2H, m), 5.16 (2H, dd,
J ¼ 1.8, 6.6 Hz), 5.19 (2H, s), 5.77 (2H, m), 7.84 (2H, t, J ¼ 6.3 Hz, NH).
13C NMR (DMSO-d6)
44.3 (2C), 93.1 (2C), 116.7 (2C), 133.1 (2C),
151.2 (2C), 177.8 (2C) ppm.
d
1H NMR (DMSO-d6)
d
0.85 (6H, t, J ¼ 7.5 Hz), 1.55 (4H, m), 3.08
MS (ESIþ) m/z: 219 (MþH), HRMS (ESIþ) m/z calculated for
C
12H14N2O2 (MþH) 219.1134; found 219.1128.
(4H, m), 5.23 (2H, s), 7.69 (2H, t, J ¼ 6.3 Hz, NH).
13C NMR (DMSO-d6)
d 11.4 (2C3), 21.0 (2C), 43.6 (2C), 92.1 (2C),
4.3.32. 2,5-Bis(3-bromopropylamino)-1,4-benzoquinone (33)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 3-
bromopropylamine hydrobromide. A stoichiometric quantity of
triethylamine was added to the reaction mixture to neutralize the
liberated HBr. The title compound was obtained as a brown solid.
Yield 0.230 g (67%); mp 180e181 ꢂC.
151.6 (2C), 177.4 (2C).
MS (ESIþ) m/z: 223 (MþH), HRMS (ESIþ) m/z calculated for
C
12H18N2O2 (MþH) 223.1447; found 223.1441.
4.3.27. 2,5-Bis(butylamino)-1,4-benzoquinone (28)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and butyl-
amine to afford the title compound as an orange-pink solid. Yield
0.070 g (30%); mp 149e150 ꢂC (Lit. 160 ꢂC; 154 ꢂC) [42].
1H NMR (DMSO-d6)
d 2.06 (4H, m), 3.24 (4H, m), 3.53 (4H, t,
J ¼ 6.6 Hz), 5.29 (2H, s), 7.78 (2H, t, J ¼ 6.3 Hz, NH).
13C NMR (DMSO-d6)
d 30.9 (2C), 32.3 (2C), 40.6 (2C), 92.5 (2C),
1H NMR (DMSO-d6)
d
0.87 (6H, t, J ¼ 7.2 Hz), 1.29 (4H, m), 1.50
(4H, m), 3.11 (4H, m), 5.22 (2H, s), 7.69 (2H, t, J ¼ 6.3 Hz, NH).
13C NMR (DMSO-d6)
13.8 (2C), 19.8 (2C), 29.7 (2C), 41.7 (2C),
151.4 (2C), 177.7 (2C).
MS (ESIþ) m/z: 379 (MþH), HRMS (ESIþ) m/z calculated for
d
C
12H16Br2N2O2 (MþH) 378.9657; found 378.9651.
92.0 (2C), 151.5 (2C), 177.4 (2C).
MS (ESIþ) m/z: 251 (MþH), HRMS (ESIþ) m/z calculated for
4.3.33. 2,5-Bis(2-hydroxyethylamino)-1,4-benzoquinone (34)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and ethanol-
amine to afford the title compound as a red-brown solid. Yield
0.224 g (60%); mp 244e246 ꢂC (Lit. > 350 ꢂC) [35].
C
14H22N2O2 (MþH) 251.1760; found 251.1754.
4.3.28. 2,5-Bis(pentylamino)-1,4-benzoquinone (29)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and pentyl-
amine to afford the title compound as a pink-red solid. Yield 0.171 g
(66%); mp 133e134 ꢂC (Lit. 134.6e137.1 ꢂC) [36].
1H NMR (DMSO-d6)
d
3.18 (4H, m), 3.54 (4H, q, J ¼ 5.4 Hz), 4.88
(2H, t, J ¼ 5.4 Hz, OH), 5.28 (2H, s), 7.51 (2H, t, J ¼ 5.1 Hz, NH).
13C NMR (DMSO-d6)
44.8 (2C), 58.6 (2C), 92.4 (2C), 151.7 (2C),
177.6 (2C).
d
1H NMR (DMSO-d6)
d
0.85 (6H, t, J ¼ 6.9 Hz), 1.26 (8H, m), 1.52
(4H, m), 3.11 (4H, m), 5.21 (2H, s), 7.69 (1H, t, J ¼ 6.3 Hz, NH).
13C NMR (DMSO-d6)
14.0 (2C), 21.9 (2C), 27.3 (2C), 28.8 (2C),
MS (ESIþ) m/z: 227 (MþH), HRMS (ESIþ) m/z calculated for
C
10H14N2O4 (MþH) 227.1032; found 227.1026.
d
41.9 (2C), 92.0 (2C), 151.5 (2C), 177.4 (2C).
4.3.34. 2,5-Bis(3-hydroxypropylamino)-1,4-benzoquinone (35)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 3-amino-
1-propanol to afford the title compound as a red-brown solid. Yield
0.227 g (55%); mp 160e162 ꢂC.
MS (ESIþ) m/z: 279 (MþH), HRMS (ESIþ) m/z calculated for
C
16H26N2O2 (MþH) 279.2073; found 279.2067.
4.3.29. 2,5-Bis(hexylamino)-1,4-benzoquinone (30)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and n-hexyl-
amine to afford the title compound as a bright orange-pink solid.
Yield 0.162 g (56%); mp 130e132 ꢂC (Lit. 114e116 ꢂC) [47].
1H NMR (DMSO-d6)
d 1.67 (4H, m), 3.18 (4H, m), 3.44 (4H, m),
4.62 (2H, t, J ¼ 4.8 Hz, OH), 5.23 (2H, s), 7.76 (2H, t, J ¼ 6.0 Hz, NH).
13C NMR (DMSO-d6)
d 30.7 (2C), 39.7 (2C), 58.8 (2C), 92.1 (2C),
151.6 (2C), 177.4 (2C).
1H NMR (DMSO-d6)
d
0.85 (6H, t, J ¼ 6.9 Hz), 1.25 (12H, m), 1.51
(4H, m), 3.11 (4H, m), 5.21 (2H, s), 7.69 (2H, t, J ¼ 6.0 Hz, NH).
13C NMR (DMSO-d6)
14.0 (2C), 22.2 (2C), 26.2 (2C), 27.6 (2C),
MS (ESIþ) m/z: 255 (MþH), HRMS (ESIþ) m/z calculated for
C
12H18N2O4 (MþH) 255.1345; found 255.1339.
d
31.0 (2C), 42.0 (2C), 92.0 (2C), 151.5 (2C), 177.4 (2C).
4.3.35. 2,5-Bis(4-hydroxybutylamino)-1,4-benzoquinone (36)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 4-amino-
MS (ESIþ) m/z: 307 (MþH), HRMS (ESIþ) m/z calculated for
C
18H30N2O2 (MþH) 307.2386; found 307.2380.

K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
203
1-butanol to afford the title compound as an orange-brown solid.
4.3.40. 2,5-Bis(2-chloroethylamino)-1,4-benzoquinone (41)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 2-
Yield 0.139 g (54%); mp 122e124 ꢂC.
1H NMR (DMSO-d6)
d
1.41 (4H, m), 1.55 (4H, m), 3.13 (4H, m),
3.38 (4H, m), 4.42 (2H, t, J ¼ 5.1 Hz, OH), 5.23 (2H, s), 7.74 (2H, t,
chloroethylamine hydrochloride. A stoichiometric quantity of
J ¼ 6.0 Hz, NH).
triethylamine was added to the reaction mixture to neutralize the
liberated HCl. The title compound was obtained as a brown solid.
Yield 0.187 g (75%); mp 190e191 ꢂC.
13C NMR (DMSO-d6)
d 24.5 (2C), 30.0 (2C), 41.9 (2C), 60.5 (2C),
92.1 (2C), 151.6 (2C), 177.4 (2C).
MS (ESIþ) m/z: 283 (MþH), HRMS (ESIþ) m/z calculated for
1H NMR (DMSO-d6)
(2H, s), 7.70 (2H, t, J ¼ 6.3 Hz, NH).
13C NMR (DMSO-d6)
42.2 (2C), 43.7 (2C), 93.1 (2C), 151.0 (2C),
178.0 (2C).
d
3.50 (4H, m), 3.78 (4H, t, J ¼ 6.0 Hz), 5.37
C
14H22N2O4 (MþH) 283.1658; found 283.1652.
d
4.3.36. 2,5-Bis(2-methoxyethylamino)-1,4-benzoquinone (37)
To a stirred solution of 1,4-benzoquinone (0.542 g, 5.01 mmol)
in methanol (10 mL) was added 2-methoxyethylamine
(3.33 mmol). The resulting dark red solution was stirred at 25 ꢂC
for 18 h, during which time a red-brown solid began to precipitate.
The reaction mixture was kept at 4 ꢂC for 24 h. The precipitate was
collected by filtration, washed sequentially with methanol and
ether, and dried under vacuum to afford the title compound as
purple flakes. Yield 0.103 g (44%); mp 125e126 ꢂC.
MS (ESIþ) m/z: 263 (MþH), HRMS (ESIþ) m/z calculated for
C
10H12Cl2N2O2 (MþH) 263.0354; found 263.0349.
4.3.41. 2,5-Bis(2-bromoethylamino)-1,4-benzoquinone (42)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 2-
bromoethylamine hydrobromide. A stoichiometric quantity of
triethylamine was added to the reaction mixture to neutralize the
liberated HBr. The title compound was obtained as a brown solid.
Yield 0.202 g (67%); mp 160e161 ꢂC.
1H NMR (DMSO-d6)
d
3.24 (6H, s), 3.29 (4H, q, J ¼ 5.7 Hz), 3.49
(4H, t, J ¼ 5.7 Hz), 5.29 (2H, s), 7.52 (2H, t, J ¼ 5.7 Hz, NH).
1H NMR (DMSO-d6)
J ¼ 6.0 Hz, NH).
d
3.61 (8H, m), 5.37 (2H, s), 7.72 (2H, t,
13C NMR (DMSO-d6)
d 41.7 (2C), 58.2 (2C), 69.3 (2C), 92.4 (2C),
151.4 (2C), 177.6 (2C).
13C NMR (DMSO-d6)
178.1 (2C).
d 30.8 (2C), 43.6 (2C), 93.1 (2C), 150.8 (2C),
MS (ESIþ) m/z: 255 (MþH), HRMS (ESIþ) m/z calculated for
C
12H18N2O4 (MþH) 255.1345; found 255.1339.
MS (ESIþ) m/z: 351 (MþH), HRMS (ESIþ) m/z calculated for
C
10H12Br2N2O2 (MþH) 350.9344; found 350.9338.
4.3.37. 2,5-Bis(3-methoxypropylamino)-1,4-benzoquinone (38)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 3-
methoxypropylamine to afford the title compound as a light pink
solid. Yield 0.124 g (47%); mp 100e102 ꢂC.
4.3.42. 2,5-Bis(3-chloropropylamino)-1,4-benzoquinone (43)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 3-
chloropropylamine hydrochloride. A stoichiometric quantity of
triethylamine was added to the reaction mixture to neutralize the
liberated HCl. The title compound was obtained as a light brown
solid. Yield 0.220 g (61%); mp 178e180 ꢂC.
1H NMR (DMSO-d6)
(4H, t, J ¼ 6.0 Hz), 5.20 (2H, s), 7.70 (2H, t, J ¼ 6.0 Hz, NH).
13C NMR (DMSO-d6)
27.8 (2C), 39.6 (2C), 58.1 (2C), 69.8 (2C),
d 1.76 (4H, m), 3.17 (4H, m), 3.22 (6H, s), 3.34
d
92.1 (2C), 151.5 (2C), 177.5 (2C).
1H NMR (DMSO-d6)
d 1.98 (4H, m), 3.25 (4H, m), 3.65 (4H, t,
MS (ESIþ) m/z: 283 (MþH), HRMS (ESIþ) m/z calculated for
J ¼ 6.6 Hz), 5.28 (2H, s), 7.78 (2H, t, J ¼ 6.3 Hz, NH).
C
14H22N2O4 (MþH) 283.1658; found 283.1652.
13C NMR (DMSO-d6)
d 30.7 (2C), 39.5 (2C), 43.1 (2C), 92.5 (2C),
151.5 (2C), 177.8 (2C).
4.3.38. 2,5-Bis(2-(methylthio)ethylamino)-1,4-benzoquinone (39)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 2-(meth-
ylthio)ethylamine to afford the title compound as a brick-red solid.
Yield 0.321 g (70%); mp 157e158 ꢂC.
MS (ESIþ) m/z: 291 (MþH), HRMS (ESIþ) m/z calculated for
C
12H16Cl2N2O2 (MþH) 291.0667; found 291.0662.
4.3.43. 2,5-Bis(4-hydroxyanilino)-1,4-benzoquinone (45)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 4-
aminophenol to afford the title compound as a dark brown solid.
Yield 0.193 g (39%); mp > 250 ꢂC.
1H NMR (DMSO-d6)
d 2.08 (6H, s), 2.68 (4H, m), 3.34 (4H, m),
5.30 (2H, s), 7.68 (2H, t, J ¼ 6.0 Hz, NH).
13C NMR (DMSO-d6)
d 14.6 (2C), 31.3 (2C), 41.1 (2C), 92.5 (2C),
1H NMR (DMSO-d6)
(4H, d, J ¼ 8.7 Hz), 9.16 (2H, br s, NH), 9.58 (2H, br s, OH).
13C NMR (DMSO-d6)
94.2 (2C),115.9 (4C),125.7 (4C),129.0 (2C),
d
5.54 (2H, s), 6.80 (4H, d, J ¼ 8.7 Hz), 7.14
151.1 (2C), 177.7 (2C).
MS (ESIþ) m/z: 287 (MþH), HRMS (ESIþ) m/z calculated for
C
12H18N2O2S2 (MþH) 287.0888; found 287.0882.
d
148.6 (2C), 155.6 (2C), 179.2 (2C).
MS (ESIþ) m/z: 323 (MþH), HRMS (ESIþ) m/z calculated for
4.3.39. 2,5-Bis(2-(dimethylamino)ethylamino)-1,4-benzoquinone
(40)
C
18H14N2O4 (MþH) 323.1032; found 323.1026.
Synthesized using the same general procedure described for the
synthesis of 37, commencing with 1,4-benzoquinone and N,N-
dimethylethylenediamine to afford the title compound as a bright
pink solid. Yield 0.273 g (61%); mp 146e148 ꢂC (Lit. 155e159 ꢂC)
[49].
4.3.44. 2,5-Bis(phenylamino)-1,4-benzoquinone (46)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and aniline
hydrobromide to afford the title compound as a dark purple solid.
Yield 0.091 g (54%); mp > 250 ꢂC (Lit. > 350 ꢂC; 357e360 ꢂC; 345 ꢂC)
[42].
1H NMR (DMSO-d6)
d
2.16 (12H, s), 2.45 (4H, t, J ¼ 6.3 Hz), 3.19
(4H, m), 5.25 (2H, s), 7.39 (2H, t, J ¼ 5.7 Hz, NH).
13C NMR (DMSO-d6)
d
39.6 (2C), 45.1 (4C), 56.3 (2C), 92.3 (2C),
1H NMR (DMSO-d6) (50 ꢂC)
d 5.78 (2H, s), 7.22 (2H, m), 7.40 (8H,
151.2 (2C), 177.4 (2C).
m), 9.27 (2H, br s, NH).
MS (ESIþ) m/z: 281 (MþH), HRMS (ESIþ) m/z calculated for
13C NMR (DMSO-d6) (50 ꢂC)
d
95.7 (2C), 123.3 (2C), 125.3 (2C),
C
14H24N4O2 (MþH) 281.1978; found 281.1972.
129.1 (2C), 137.8 (2C), 147.2 (2C), 179.6 (2C).

204
K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
MS (ESIþ) m/z: 291 (MþH), HRMS (ESIþ) m/z calculated for
18H14N2O2 (MþH) 291.1134; found 291.1128.
4.3.50. 2,5-Bis(2-methoxyanilino)-1,4-benzoquinone (52)
Synthesized using the same general procedure described for the
synthesis of 37, commencing with 1,4-benzoquinone and o-anisi-
dine to afford the title compound as a brown solid. Yield 0.286 g
(49%); mp 240e241 ꢂC (Lit. 250 ꢂC) [52].
C
4.3.45. 2,5-Bis(2-carboxyanilino)-1,4-benzoquinone (47)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and anthranilic
acid to afford the title compound as a dark red-brown solid. Yield
0.290 g (63%) mp > 250 ꢂC (Lit. 322e324 ꢂC; 335 ꢂC; 315 ꢂC) [50].
1H NMR (DMSO-d6)
d 3.84 (6H, s), 5.56 (2H, s), 7.02 (2H, td,
J ¼ 0.9, 7.5 Hz), 7.15 (2H, dd, J ¼ 0.9, 7.8 Hz), 7.26 (2H, td, J ¼ 1.2,
7.8 Hz), 7.36 (2H, dd, J ¼ 1.2, 7.8 Hz), 8.84 (2H, br s, NH).
1H NMR (DMSO-d6)
(2H, m, H), 10.73 (2H, br s, NH).
13C NMR (DMSO-d6)
98.4 (2C), 120.8 (2C), 121.4 (2C),124.1 (2C),
d
6.24 (2H, s), 7.26 (2H, m), 7.68 (4H, m), 8.02
13C NMR (100 MHz) (DMSO-d6)
d 56.0 (2C), 95.6 (2C), 112.3 (2C),
121.0 (2C), 123.8 (2C), 126.0 (2C), 127.2 (2C), 146.9 (2C), 152.3 (2C),
179.4 (2C).
d
132.1 (2C), 134.1 (2C), 139.5 (2C), 144.5 (2C), 168.4 (2C), 180.8 (2C).
MS (ESIþ) m/z: 351 (MþH), HRMS (ESIþ) m/z calculated for
MS (ESIꢀ) m/z: 377 (MꢀH), HRMS (ESIꢀ) m/z calculated for
C
20H18N2O4 (MþH) 351.1345; found 351.1339.
C
20H14N2O6 (MꢀH) 377.0774; found 377.0779.
4.3.51. 2,5-Bis(3-methoxyanilino)-1,4-benzoquinone (53)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and m-anisi-
dine to afford the title compound as a dark brown solid. Yield
0.096 g (51%); mp > 250 ꢂC.
4.3.46. 2,5-Bis(3-carboxyanilino)-1,4-benzoquinone (48)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 3-
aminobenzoic acid to afford the title compound as an olive-green
solid. Yield 0.139 g (60%). mp > 250 ꢂC.
1H NMR (DMSO-d6)
d
3.76 (6H, s), 5.82 (2H, s), 6.80 (2H, m), 6.94
(2H, m), 6.96 (2H, m) 7.33 (2H, t, J ¼ 8.4 Hz), 9.24 (2H, br s, NH).
13C NMR (100 MHz) (DMSO-d6)
55.4 (2C), 96.0 (2C), 109.7 (2C),
1H NMR (DMSO-d6)
d
5.81 (2H, s), 7.55 (2H, t, J ¼ 7.8 Hz), 7.62
(2H, m), 7.78 (2H, m), 7.90 (2H, t, J ¼ 1.8 Hz), 9.42 (2H, br s, NH),
d
13.12 (2H, br s, COOH).
111.3 (2C), 115.7 (2C), 130.2 (2C), 139.1 (2C), 147.2 (2C), 160.0 (2C),
180.0 (2C).
13C NMR (DMSO-d6)
d 96.0 (2C), 124.3 (2C), 126.4 (2C), 128.2
(2C), 129.8 (2C), 132.1 (2C), 138.3 (2C), 147.2 (2), 166.9 (2C), 180.1
(2C).
MS (ESIþ) m/z: 351 (MþH), HRMS (ESIþ) m/z calculated for
C
20H18N2O4 (MþH) 351.1345; found 351.1339.
MS (ESIꢀ) m/z: 377 (MꢀH), HRMS (ESIꢀ) m/z calculated for
C
20H14N2O6 (MꢀH) 377.0774; found 377.0779.
4.3.52. 2,5-Bis(4-methoxyanilino)-1,4-benzoquinone (54)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and p-anisi-
dine. The title compound was obtained as a dark brown solid. Yield
0.167 g (53%); mp > 250 ꢂC.
4.3.47. 2,5-Bis(4-carboxyanilino)-1,4-benzoquinone (49)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 4-
aminobenzoic acid to afford the title compound as a dark brown
solid. Yield 0.160 g (68%); mp > 250 ꢂC (Lit. > 260 ꢂC) [51].
1H NMR (DMSO-d6)
J ¼ 9.0 Hz), 7.27 (4H, d, J ¼ 9.0 Hz), 9.23 (2H, br s, NH).
13C NMR (DMSO-d6)
55.6 (2C), 95.8 (2C), 114.9 (4C), 125.8 (4C),
d 3.76 (6H, s), 5.60 (2H, s), 6.99 (4H, d,
1H NMR (DMSO-d6)
(4H, d, J ¼ 8.7 Hz), 9.44 (2H, br s, NH).
13C NMR (DMSO-d6)
97.8 (2C), 123.0 (4C), 127.2 (2C), 131.0 (4C),
d
6.06 (2H, s), 7.52 (4H, d, J ¼ 8.7 Hz), 7.96
d
130.7 (2C), 147.1 (2C), 157.2 (2C), 179.9 (2C).
d
MS (ESIþ) m/z: 351 (MþH), HRMS (ESIþ) m/z calculated for
142.4 (2C), 146.1 (2C), 167.2 (2C), 180.8 (2C).
C
20H18N2O4 (MþH) 351.1345; found 351.1339.
MS (ESIꢀ) m/z: 377 (MꢀH), HRMS (ESIꢀ) m/z calculated for
C
20H14N2O6 (MꢀH) 377.0774; found 377.0779.
4.3.53. 2,5-Bis(2-bromoanilino)-1,4-benzoquinone (55)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 2-
bromoaniline to afford the title compound as a brown solid. Yield
0.103 g (37%); mp 225e226 ꢂC.
4.3.48. 2,5-Bis(3-hydroxyanilino)-1,4-benzoquinone (50)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 3-
aminophenol to afford the title compound as a dark brown solid.
Yield 0.071 g (37%); mp > 250 ꢂC.
1H NMR (DMSO-d6)
(2H, m), 7.78 (2H, m), 9.15 (2H, br s, NH);
13C NMR (100 MHz) (DMSO-d6)
95.9 (2C), 120.3 (2C), 127.9
d 5.18 (2H, s), 7.29 (2H, m), 7.45 (2H, m), 7.49
1H NMR (DMSO-d6)
d
5.81 (2H, s), 6.61 (2H, dd, J ¼ 2.1, 7.8 Hz),
d
6.79 (4H, m), 7.20 (2H, dd, J ¼ 7.8, 8.1 Hz), 9.18 (2H, br s, NH), 9.63
(2C), 129.0 (2C), 129.1 (2C), 133.6 (2C), 136.2 (2C), 148.3 (2C), 179.3
(2C).
(2H, br s, OH).
13C NMR (DMSO-d6)
d
95.8 (2C), 110.5 (2C), 112.9 (2C), 114.4 (2C),
MS (ESIþ) m/z: 447 (MþH), HRMS (ESIþ) m/z calculated for
130.2 (2C), 138.9 (2C), 147.3 (2C), 158.2 (2C), 179.9 (2C).
C
18H12Br2N2O (MþH) 446.9344; found 446.9339.
MS (ESIþ) m/z: 323 (MþH), HRMS (ESIþ) m/z calculated for
C
18H14N2O4 (MþH) 323.1032; found 323.1026.
4.3.54. 2,5-Bis(2-(hydroxymethyl)anilino)-1,4-benzoquinone (56)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and 2-
aminobenzyl alcohol to afford the title compound as a yellow-
brown solid. Yield 0.090 g (45%); mp 206e207 ꢂC.
4.3.49. 2,5-Bis(2-methylanilino)-1,4-benzoquinone (51)
Synthesized using the same general procedure described for the
synthesis of 1, commencing with 1,4-benzoquinone and o-toluidine
to afford the title compound as dark purple flakes. Yield 0.096 g
(49%); mp > 250 ꢂC (253e254 ꢂC; 250e252 ꢂC) [52].
1H NMR (DMSO-d6)
d 4.47 (4H, s), 5.47 (2H, br s, OH), 5.50 (2H,
s), 7.26 (2H, m), 7.37 (4H, m), 7.46 (2H, d, J ¼ 7.5 Hz), 9.33 (2H, br s,
1H NMR (DMSO-d6)
(4H, m), 7.34 (2H, m), 9.11 (2H, br s, NH).
13C NMR (DMSO-d6)
17.5 (2C), 94.5 (2C), 126.6 (2C), 127.0 (2C),
d
2.17 (6H, s), 5.01 (2H, s), 7.21 (2H, m), 7.27
NH).
13C NMR (100 MHz) (DMSO-d6)
d 60.7 (2C), 95.2 (2C), 124.3 (2C),
d
126.3 (2C), 128.2 (2C), 128.8 (2C), 136.0 (2C), 136.4 (2C), 148.1 (2C),
179.5 (2C).
127.4 (2C), 131.2 (2C), 134.4 (2C), 136.2 (2C), 149.8 (2C), 179.0 (2C).
MS (ESIþ) m/z: 319 (MþH), HRMS (ESIþ) m/z calculated for
MS (ESIþ) m/z: 351 (MþH), HRMS (ESIþ) m/z calculated for
C
20H18N2O2 (MþH) 319.1447; found 319.1441.
C
20H18N2O4 (MþH) 351.1345; found 351.1339.

K.A. MacGregor et al. / European Journal of Medicinal Chemistry 85 (2014) 191e206
205
4.3.55. 2,5-Bis(3-(hydroxymethyl)phenylamino)-1,4-benzoquinone
(57)
Synthesized using the same general procedure described for the
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2014.06.070.
synthesis of 37, commencing with 1,4-benzoquinone and 3-
aminobenzylalcohol to afford the title compound as a brown
solid. Yield 0.294 g (51%); mp 243e244 ꢂC.
References
1H NMR (DMSO-d6)
d
4.51 (4H, d, J ¼ 5.7 Hz), 5.26 (2H, t,
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13C NMR (DMSO-d6)
d 62.7 (2C), 95.5 (2C), 121.5 (2C), 122.3 (2C),
123.8 (2C), 129.2 (2C), 137.8 (2C), 144.1 (2C), 147.5 (2C), 179.9 (2C).
MS (ESIþ) m/z: 351 (MþH), HRMS (ESIþ) m/z calculated for
C
20H18N2O4 (MþH) 351.1345; found 351.1339.
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4.3.56. 2,5-Bis(4-(hydroxymethyl)phenylamino)-1,4-benzoquinone
(58)
Synthesized using the same general procedure described for the
synthesis of 37, commencing with 1,4-benzoquinone and 4-
aminobenzylalcohol to afford the title compound as a brown
solid. Yield 0.197 g (47%); mp > 250 ꢂC.
1H NMR (DMSO-d6)
d
4.49 (4H, d, J ¼ 5.7 Hz), 5.19 (2H, t,
J ¼ 5.7 Hz, OH), 5.75 (2H, s), 7.31 (4H, d, J ¼ 8.7 Hz), 7.36 (4H, d,
J ¼ 8.7 Hz), 9.28 (2H, br s, NH).
13C NMR (DMSO-d6)
d 62.7 (2C), 95.3 (2C), 123.6 (4C), 127.5 (4C),
136.4 (2C), 140.2 (2C), 147.6 (2C), 179.8 (2C).
MS (ESIþ) m/z: 351 (MþH), HRMS (ESIþ) m/z calculated for
C
20H18N2O4 (MþH) 351.1345; found 351.1339.
Author statement
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The authors declare the following competing financial in-
terest(s): We have entered into a commercial agreement with
Abcam Biochemicals (Cambridge, UK) for the supply of our dyna-
min inhibitors. This includes some of the compounds listed in this
paper.
#Dynole™, Rhodadyn™, MiTMAB™, Bis-T™, Dyngo™, and Imi-
nodyn™ are trademarks of Children's Medical Research Institute
and Newcastle Innovation Ltd. Most of the lead dynamin inhibitors
described in this paper are available from Abcam Biochemicals
(Cambridge, UK).
Acknowledgements
This work was supported by grants from the National Health
and Medical Research Council (Australia) (Grant 571076 and
1011457), The Australia Research Council, The Australian Cancer
Research Foundation, The Ramaciotti Foundation, The Children's
Medical Research Institute, and Newcastle Innovation Ltd.
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MiTMAB myristoyl trimethyl ammonium bromide
OcTMAB Octyl trimethyl ammonium bromide
Bis-T
RTIL
PSA
Bis-tyrphostin
Room Temperature Ionic Liquid
Polar Surface Area
U2OS
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Dynamin I
Dynamin II
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