Substituted 1,6-diphenylnaphthalenes as FtsZ-targeting antibacterial agents

Article abstract of DOI:10.1016/j.bmcl.2013.02.016

Bacterial cell division occurs in conjunction with the formation of a cytokinetic Z-ring structure comprised of FtsZ subunits. Agents that disrupt Z-ring formation have the potential, through this unique mechanism, to be effective against several of the newly emerging multidrug-resistant strains of infectious bacteria. Several 1-phenylbenzo[c]phenanthridines exhibit notable antibacterial activity. Based upon their structural similarity to these compounds, a distinct series of substituted 1,6-diphenylnaphthalenes were synthesized and evaluated for antibacterial activity against Staphylococcus aureus and Enterococcus faecalis. In addition, the effect of select 1,6-diphenylnaphthalenes on the polymerization dynamics of S. aureus FtsZ and mammalian β-tubulin was also assessed. The presence of a basic functional group or a quaternary ammonium substituent on the 6-phenylnaphthalene was required for significant antibacterial activity. Diphenylnaphthalene derivatives that were active as antibiotics, did exert a pronounced effect on bacterial FtsZ polymerization and do not appear to cross-react with mammalian tubulin to any significant degree.

Full text of DOI:10.1016/j.bmcl.2013.02.016

Bioorganic & Medicinal Chemistry Letters 23 (2013) 2001–2006
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Substituted 1,6-diphenylnaphthalenes as FtsZ-targeting antibacterial
agents
Yongzheng Zhang a, Daniel Giurleo b, Ajit Parhi a, Malvika Kaul c, Daniel S. Pilch c, Edmond J. LaVoie b,
⇑
a TAXIS Pharmaceuticals Inc., North Brunswick, NJ 08902, USA
b Department of Medicinal Chemistry, Rutgers, The State University of New Jersey, Piscataway, NJ 08854-8020, USA
c Department of Pharmacology, The University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, Piscataway, NJ 08854, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Bacterial cell division occurs in conjunction with the formation of a cytokinetic Z-ring structure com-
prised of FtsZ subunits. Agents that disrupt Z-ring formation have the potential, through this unique
mechanism, to be effective against several of the newly emerging multidrug-resistant strains of infec-
tious bacteria. Several 1-phenylbenzo[c]phenanthridines exhibit notable antibacterial activity. Based
upon their structural similarity to these compounds, a distinct series of substituted 1,6-diphenylnaphtha-
lenes were synthesized and evaluated for antibacterial activity against Staphylococcus aureus and Entero-
coccus faecalis. In addition, the effect of select 1,6-diphenylnaphthalenes on the polymerization dynamics
of S. aureus FtsZ and mammalian b-tubulin was also assessed. The presence of a basic functional group or
a quaternary ammonium substituent on the 6-phenylnaphthalene was required for significant antibacte-
rial activity. Diphenylnaphthalene derivatives that were active as antibiotics, did exert a pronounced
effect on bacterial FtsZ polymerization and do not appear to cross-react with mammalian tubulin to
any significant degree.
Received 11 November 2012
Accepted 4 February 2013
Available online 13 February 2013
Keywords:
Antibacterial
FtsZ-targeting
Staphylococcus aureus
Enterococcus faecalis
Diphenylnaphthalenes
FtsZ polymerization
Ó 2013 Elsevier Ltd. All rights reserved.
Bacterial infections associated with drug-resistant pathogens
such as methicillin-resistant Staphylococcus aureus (MRSA) and
vancomycin-resistant enterococci (VRE) are recognized world-
wide as a major health concern.1,2 Bacterial resistance to conven-
tional antibacterial drugs has created a critical need for the identi-
fication of novel therapeutic antibacterial targets. In this
connection, FtsZ has been identified as an appealing new target
for antibacterial agents.10–17 FtsZ is an essential and highly
conserved bacterial cytokinesis protein.3–8 Cell division in bacteria
occurs at the site of formation of a cytokinetic Z-ring polymeric
structure, which is comprised of FtsZ subunits.9 The potential for
developing agents that target FtsZ and the recent advances in the
development of small molecules that target FtsZ have been the
subject of several recent reports.10–24
Sanguinarine 1 (Fig. 1) is a benzo[c]phenanthridine plant alka-
loid that has been identified as small molecule that alters FtsZ Z-
ring formation and has modest antibacterial activity.11,25 Studies
in our laboratory have also shown that chelerythrine 2 (Fig. 1)
has similar effects on S. aureus and Bacillus subtilis.26 In addition,
the presence of a phenyl functionality at the 1-position of 2 (to
yield 3) was shown to significantly enhance antibacterial activity
relative to either 1 or 2.25,26 Within the cyclic core structure of 3,
there is a scaffold consisting of a 1,6-diphenylnaphthalene. In light
of these results, we examined the relative structure–activity rela-
tionships associated with the antistaphylococcal and antientero-
coccal activity of several 1,6-diphenylnaphthalene derivatives,
including those with a constitutive cationic charge, those with ba-
sic functionalities that could be protonated to varying degrees at
physiological pH, and those without a cationic charge.
The critical intermediate for the preparation of the various 1,6-
diphenylnaphthalenes that were synthesized was 5-bromo-2-
naphthol, which was prepared from 6-hydroxy-1-naphthylamine
as described in the literature (19).27 Suzuki coupling of 4-tert-buty-
phenylboronic acid to this naphthylbromide provided 5-(4-t-butyl-
phenyl)-2-naphthol, which was then converted to its triflate
derivative with Tf2O. This triflate derivative could then be used
in a second Suzuki coupling reaction for the preparation of 5, 6,
and 18 (see Scheme 1).
The triflate of 1-[(4-t-butyl)phenyl]-6-hydroxynaphthalene was
also used in a Suzuki coupling reaction with 2-formylphenylboron-
ic acid. The resulting aldehyde was reduced with NaBH4 and the
hydroxymethyl derivative converted to its benzyl chloride. Treat-
ment of this chloride with either methylamine or dimethylamine
provided 7 and 8, respectively. Treatment of 8 with methyl iodide
provided the quaternary ammonium derivative, 9. The triflate of
1-[(4-t-butyl)phenyl]-6-hydroxynaphthalene was also coupled
with 4-carboxymethyl-2-methylphenylboronic acid. NBS bromina-
tion of 5 and this 6-(2-methyl-4-carboxymethyl)-1-(4-t-butylphe-
nylphenyl)naphthalene provided the bromomethyl intermediates
⇑
Corresponding author. Tel.: +1 848 445 2674; fax: +1 732 445 6312.
E-mail address: elavoie@pharmacy.rutgers.edu (E.J. LaVoie).
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.02.016

2002
Y. Zhang et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2001–2006
O
O
ammonium derivative, 15. Treatment of 1-(4-t-butyl)phenyl-6-
OCH3
OCH3
(2-bromomethyl-4-carboxymethylphenyl)naphthalene with NaN3
followed by reduction with triphenylphosphine provide the meth-
ylamino derivative, 12. The methyl ester of 12 was hydrolyzed
using LiOH in aqueous THF to yield its carboxy derivative, 17.
Using the appropriate triflate of 6-hydroxynaphthalene, various
1,6-diphenylnaphthalenes could be synthesized (Scheme 3). Treat-
ment of the requisite N,N-dimethylamino and N,N-dimethylanilino
derivatives with methyl iodide provided the quaternary ammo-
nium salts, 19–27. The triflate of 1-(3-[1,10]-biphenyl)-6-hydroxy-
naphthalene served as an intermediate for Suzuki coupling with
3-aminophenylboronic acid to give 18. Alternatively, this triflate
intermediate was treated with 3-cyanophenylboronic acid, which
was reduced with LAH to give 28, which in the presence of formal-
dehyde and formic acid provided the N,N-dimethyl derivative, 29.
Treatment of 29 with methyl iodide resulted in the formation of
the quaternary ammonium derivative, 30.
N
N
O
H3CO
CH3
CH3
O
OCH3
1
2
OCH3
OCH3
R1
R2
N
N
R3
H3CO
CH3
OCH3
4
3
Figure 1. The structures of sanguinarine 1, chelerythrine 2 and 1-phenyl-5-methyl-
2,3,7,8-tetramethoxybenzo[c]phenanthridine 3 relative to a generalized structure
of a 1,6-diphenylnaphthalene 4.
Several 1,6-diphenylnaphthalenes (DPNs) were evaluated for
antibiotic activity against methicillin-sensitive and methicillin-
resistant S. aureus (MSSA and MRSA, respectively) and vancomy-
cin-sensitive and vancomycin-resistant Enterococcus faecalis (VSE
and VRE, respectively), as summarized in Table 1.
A and B that were employed for the preparation of 10–17, as out-
lined in Scheme 2.
It is evident from the lack of antimicrobial activity for 5 and 6
that the presence of a sufficiently basic functional group, which
would be significantly charged at physiological pH, is critical for
activity. The first series of DPNs that was studied consisted of
1-[4-(t-butyl)phenyl]naphthalene derivatives that had an o-ami-
nomethyl substituent on the 6-phenyl moiety. Among the analogs
related to 6-(o-aminomethyl)phenyl, the presence of a single
N-methyl group as in 7 is associated with significant activity
against all S. aureus and E. faecalis strains. However, the N,N-
dimethyl derivative, 8, is substantially less active. Conversion of
Treatment of these bromomethyl derivatives with 1,3-bis(t-
butoxycarbonyl)guanidine followed by removal of the N-Boc pro-
tecting groups provided the guanidinomethyl derivatives 10 and
16. Treatment of 1-(4-t-butyl)phenyl-6-(2-bromomethylphe-
nyl)naphthalene with potassium cyanide followed by reduction
of the resulting nitrile with LAH yielded the 2-aminoethyl deriva-
tive 11. 1-(4-t-Butyl)phenyl-6-(2-bromomethyl-4-carboxymethyl-
phenyl)naphthalene when treated with either methylamine or
dimethylamine provided 13 and 14, respectively. Compound 14
was methylated using methyl iodide to provide the quaternary
Br
c
,d
a
b
HO
HO
TfO
6
NH2
f
g
e
h
18
NH2
R"
i,j
CH2Br
R"
CH3
5, R" = H
CHO
k
IntermediateA, R" = H
IntermediateB, R'' = CO2Me
R'' = CO2Me
Cl
CH2NRCH3
7, R = H
8
, R = CH3
l
9
Scheme 1. Methods used for the preparation of 1,6-diphenylnaphthalenes. Reagents and conditions: (a) 4-(t-butyl)phenylboronic acid, Pd(PPh3)4, K2CO3, dioxane/H2O,
100 °C ; (b) Tf2O, Et3N, DCM, ꢀ78 °C; (c) 2-NO2-phenylboronic acid, Pd(OAc)2, XPhos, K2CO3, CH3CN/H2O, 100 °C; (d)H2, Pd/C (10%), EtOH; (e) 2-methylphenylboronic acid-
B(OH)2 or 2-methyl-4-carboxymethylphenylboronic acid, Pd(OAc)2, XPhos, K2CO3, CH3CN/H2O, 100 °C; (f) 2-formylphenylboronic acid, Pd(OAc)2, XPhos, K2CO3, CH3CN/H2O,
100 °C; (g) 3-aminophenylboronic acid, Pd(OAc)2, XPhos, K2CO3, CH3CH/H2O (1:1), 100 °C; (h) NBS, AIBN, CCl4, 85 °C; (i) NaBH4, EtOH; (j) MsCl, Et3N, CH2Cl2, rt; see Ref. 28; (k)
7, CH3NH2 in THF or 8, (CH3)2NH in THF, 60 °C, sealed tube; (l) MeI, sealed tube 100 °C.

Y. Zhang et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2001–2006
2003
R4"
CH2Br
4" = H
Intermediate A,
R
CH2Br
H3CO2C
CH2Br
Intermediate B,R4" = CO2CH3
g, h
e
c,d
a,b
RO2C
CH2NH2
H3CO2C
CH2NRCH3
CH2CH2NH2
R4"
CH2NC(NH2)2
13
, R = H
12
, R = CH3
17
, R = H
11
14, R = CH3
i
10,
R4" = H
16, R4" = CO2CH3
f
15
Scheme 2. Methods used for the preparation of 1,6-diphenylnaphthalenes. Reagents and conditions: (a) 1,3-bis(tert-butoxycarbonyl)guanidine, K2CO3, DMF, 50 °C,
acetonitrile; (b) TFA, DCM, 50 °C, sealed vial, (c) KCN, DMF, rt; (d) LAH, toluene/THF(1:1), 100 °C; (e) 13, CH3NH2 in THFor14, (CH3)2NH in THF, 60 °C, sealed tube; (f) MeI,
sealed tube 100 °C; (g) NaN3, DMF, rt; (h) PPh3 polymer bound, THF/H2O, rt; (i) LiOH, THF-H2O.
R1
R2
R1
R2
a, b
Z
X
TfO
Y
19-27
c
Ph
Ph
b
Ph
Ph
e
d
I
CH2NH2
CH2N(CH3)2
CN
CH2N(CH3)3
28
29
30
Scheme 3. Reagents and conditions: (a) R–B(OH)2, Pd(OAc)2, XPhos, K2CO3, CH3CN/H2O, 100 °C; (b) MeI, sealed tube 100 °C; (c) 3-cyanophenylboronic acid, Pd(OAc)2, XPhos,
K2CO3, CH3CN/H2O, 100 °C; (d) LAH, THF/toluene, 100 °C; (e) formaldehyde, NaBH3CN, pH 6
this tertiary amine to its N,N,N,-trimethyl ammonium derivative, 9,
resulted in a significant enhancement of its antibacterial activity
against E. faecalis. In contrast, the 6-[(2-aminomethyl)-4-
(carboxy)]phenylnaphthalene 17 did not exhibit appreciable anti-
bacterial activity. Within this group of compounds, the most
remarkable result, as was observed for 8, is the loss of antibacterial
activity for the N,N-dimethyl derivative, 14.
against both the MSSA and MRSA strains, with MICs of 0.5
mL. With regard to E. faecalis, 9 is slightly less active than 7, with
MICs of 4.0 g/mL against both VSE and VRE. The guanidinomethyl
lg/
l
derivative, 10, and the 2-aminoethyl analog, 11, also have similar
antibacterial activity to 7.
Several DPNs were evaluated that incorporated into their struc-
ture a 1-[4-(t-butyl)phenyl]naphthalene moiety and either an m-
amino or m-aminomethyl substituent on the 6-phenyl group.
While the aniline derivative, 18, is inactive, its N,N,N-trimethyl
ammonium derivative 19 had potent antibacterial activity against
all the S. aureus and E. faecalis strains.
The structure–activity relationship among the 6-[(2-amino-
methyl)-4-(carboxymethyl)]phenylnaphthalene analogs, 12–16,
was similar to that associated with 6-(o-aminomethyl)phenyl-
naphthalene derivatives, 7–11. The 6-[(2-aminomethyl)-4-(car-
boxymethyl)]phenylnaphthalene 12 has good antibacterial
activity against both MSSA and MRSA, but was notably less active
Those DPNs that incorporated within their structure a 6-[3-
(trimethylammonium)phenyl]naphthalene and either
a 1-[4-

2004
Y. Zhang et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2001–2006
Table 1
Antistaphylococcal and antienterococcal activities of substituted 1,6-diphenylnaphthalenes
R1
R2
Z
X
Y
X
Y
Z
R2
R1
MIC (
lg/mL)
S. aureus
S. aureus
E. faecalis
E. faecalis
8325-4 (MSSA)
ATCC 33591 (MRSA)
ATCC 19433 (VSE)
ATCC 51575 (VRE)
5
6
7
8
CH3
NH2
CH2NH(CH3)
CH2N(CH3)2
CH2N(CH3)3
CH2NC(NH2)2
CH2CH2NH2
CH2NH2
CH2NHCH3
CH2N(CH3)2
CH2N(CH3)3
CH2NC(NH2)2
CH2NH2
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
NH2
N(CH3)3
N(CH3)3
N(CH3)3
N(CH3)3
N(CH3)3
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
t-Butyl
Phenyl
CF3
>64
>64
2
32
0.5
1
2
2
4
>64
1
>64
>64
2
>64
0.5
1
2
2
4
>64
2
>64
>64
2
>64
4
2
4
16
8
>64
4
>64
>64
2
>64
4
2
4
16
4
>64
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
1
CO2CH3
CO2CH3
CO2CH3
CO2CH3
CO2CH3
CO2H
H
H
H
H
H
1
1
2
4
>64
>64
0.5
0.5
1
1
1
1
1
4
1
2
4
0.5
2
4
0.063
1
0.13
0.063
0.031
>64
>64
1
1
4
4
1
2
1
4
1
4
8
1
2
4
>64
2
>64
>64
2
2
8
8
2
4
2
4
2
4
16
4
8
32
8
1
1
>64
>64
2
4
8
8
2
4
2
4
2
8
16
4
16
32
>64
>64
>64
>64
>64
H
H
F
H
H
phenyl
H
H
N(CH3)3
N(CH3)3
N(CH3)3
N(CH3)3
H
t-Butyl
Phenyl
F
H
H
H
phenyl
phenyl
phenyl
phenyl
CH2NH2
CH2N(CH3)2
CH2N(CH3)3
H
H
H
H
H
H
Sanguinarine
Chelerythrine
2
Oxacillin
Vancomycin
Erythromycin
Tetracycline
Clindamycin
>64
64
>64
0.5
2
aMinimum inhibitory concentration (MIC) assays were conducted in accordance with Clinical and Laboratory Standards Institute (CLSI) guidelines for broth microdilution.29
MIC is defined as the lowest compound concentration at which bacterial growth is P90% inhibited.
biphenyl], 1-(4-fluorophenyl), 1-(4-trifluoromethylphenyl), or
1-[3-biphenyl] moiety (20–23) had significant antibacterial activ-
ity. Unexpectedly, similar activity was observed for 6-[4-(trime-
thylammonium)phenyl]naphthalenes that had incorporated in
their structure a 1-[4-(t-butyl)phenyl], 1-[4-biphenyl], 1-(4-fluoro-
phenyl), or 1-[3-biphenyl] moiety (24–27). The 6-[4-(trimethylam-
monium)phenyl]naphthalene analog, 24, is only slightly less active
than 19 against all the S. aureus and E. faecalis strains.
assay to monitor the impact, if any, of select diphenylnaphthalene
compounds on the dynamics of self-polymerization by S. aureus
FtsZ (SaFtsZ), which was expressed in Escherichia coli and purified
as previously described.32 In this assay, FtsZ polymerization is de-
tected in solution by a time-dependent increase in light scattering,
as reflected by a corresponding increase in solution absorbance at
340 nm (A340). As illustrative examples, Figure 2 shows the time-
dependent A340 profiles of SaFtsZ in the presence of vehicle (DMSO)
In a comparison of the 1-[3-biphenyl]-6-(3-aminomethylphe-
nyl)naphthalene derivatives, 28–30, the primary amine 28 was
significantly more active than the tertiary amine 29. The N,N,N-
trimethylammonium derivative, 30, was as or more potent than
28 against all the S. aureus and E. faecalis strains.
Stimulation of FtsZ self-polymerization and the concomitant
stabilization of FtsZ polymers has been implicated in the antibacte-
rial actions of several distinct classes of small molecules, including
substituted benzamides,10,13,14,20 dibenzoquinoliziniums,30 iso-
quinolines,31 and biaryls.32 We used a light scattering (turbidity)
onlyor compounds 9 and 11 at a concentration of 40 lg/mL. Vanco-
mycin was also included as a non-FtsZ-targeting control antibiotic.
Note that vancomycin exerts a negligible impact on SaFtsZ poly-
merization, an expected result given that the antibacterial target
of vancomycin is the bacterial cell wall and not FtsZ. By contrast,
compounds 9 and 11 increase both the kinetics and extent of SaF-
tsZ polymerization, with the relative magnitude of these increases
correlating with the relative antistaphylococcal potencies of the
compounds (see Table 1). Thus, as we have previously indicated
for the dibenzoquinoliziniums,30 isoquinolines,31 and biaryls,32

Y. Zhang et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2001–2006
2005
being a known stimulator of tubulin polymerization33,34 and the
latter drug being a known inhibitor of tubulin polymerization.35
Figure 3 shows the time-dependent A340 profiles of porcine b-tubu-
Vehicle
9
1.20
1.00
0.80
0.60
0.40
11
Vancomycin
lin in the presence of DMSO vehicle, 9 (at 40
lg/mL), paclitaxel (at
25 g/mL), or nocodazole (at 10 g/mL). Both paclitaxel and noco-
l
l
dazole produce their expected impacts on tubulin polymerization
dynamics. By contrast, 9 exerts little or no impact. Thus, a diphe-
nylnaphthalene compound that exerts a profound impact on bacte-
rial FtsZ polymerization does not appear to cross-react with
mammalian tubulin to any significant degree. This degree of target
specificity bodes well for the potential of desirable toxicological
properties on the part of the diphenylnaphthalenes.
The structure-activity relationships associated with the anti-
staphylococcal and antienterococcal activity of several 1,6-diphe-
nylnaphthalenes, including those with a constitutive cationic
charge and those with basic functionalities that could be proton-
ated to varying degrees at physiological pH, reveal several deriva-
tives with significant antibacterial activity against S. aureus and E.
faecalis. The presence on the 6-phenyl moiety of a methylaminom-
ethyl-, 2-methylaminoethyl, guanidinomethyl, or (trimethylam-
monium)methyl at the o-position was associated with significant
antibacterial activity within a series of 6-phenyl-1-(4-t-butyl-
phenyl)naphthalenes. In addition, the presence of a trimethylam-
monium group at either the m- or p-position of the 6-phenyl
moiety was associated with antibacterial activity for 6-phenyl-1-
(4-t-butylphenyl)naphthalenes, 6-phenyl-1-(3-[1,10-phenyl)naph-
thalenes and 6-phenyl-1-(4-[1,10-biphenyl)naphthalenes. As was
observed with benzo[c]phenanthridinium derivatives 1 or 2, it ap-
pears that for the 1,6-diphenylnaphthalenes 9 and 11, that there
antibacterial activity is associated with their ability to effect FtsZ
polymerization. While having a pronounced impact on bacterial
FtsZ polymerization, they do not affect mammalian tubulin.
0
50
100
150
200
250
300
Time (minutes)
Figure 2. Impact of select diphenylnaphthalenes on the polymerization of S. aureus
FtsZ (SaFtsZ), as determined by monitoring time-dependent changes in absorbance
at 340 nm (A340). Polymerization profiles of SaFtsZ (10
DMSO vehicle (blue) or 40 g/mL of either 9 (red), 11 (black), or the comparator
antibiotic vancomycin (green) are depicted. Experiments were conducted at 25 °C
in solution containing 50 mM TrisÅHCl (pH 7.4), 50 mM KCl, 2 mM magnesium
acetate, 1 mM CaCl2, and 1 mM GTP. The reactions (100 lL total volume) were
assembled in half-volume, flat-bottom 96-well microtiter plates, and their A340
values were continuously monitored using a VersaMaxÒ (Molecular Devices, Inc.)
plate reader.
lM) in the presence of
l
0.40
Vehicle
9
0.30
0.20
0.10
0.00
paclitaxel
nocodazole
Acknowledgments
This study was supported by research agreements between
TAXIS Pharmaceuticals, Inc. and both Rutgers, The State University
of New Jersey (E.J.L.) and the University of Medicine and Dentistry
of New Jersey (D.S.P). We would like to thank Drs. Steve Tuske and
Eddy Arnold (Center for Advanced Biotechnology and Medicine,
Rutgers University) for their assistance with the expression and
purification of S. aureus FtsZ. S. aureus 8325-4 was the generous gift
of Dr. Glenn W. Kaatz (John D. Dingell VA Medical Center, Detroit,
MI). The Brucker Avance III 400 MHz NMR spectrometer used in
this study was purchased with funds from NCRR Grant No.
1S10RR23698-1A1. Mass spectrometry was provided by the Wash-
ington University Mass Spectrometry Resource with support from
the NIH National Center for Research Resources Grant No.
P41RR0954.
0
5
10
Time (minutes)
15
20
Figure 3. Impact of 9 on the polymerization of microtubule associated protein
(MAP)-rich porcine b-tubulin (70% tubulin, 30% MAPs), as determined by monitor-
ing time-dependent changes in A340. Polymerization profiles of tubulin (2 mg/mL)
in the presence of DMSO vehicle (blue), 40
(red), or 10 g/mL nocodazole (green) are depicted. Experiments were conducted at
37 °C in solution containing 80 mM PIPESÅNaOH (pH 7.0), 2 mM MgCl2, 1 mM EGTA,
and 1 mM GTP. The reactions (100 L total volume) were assembled in half-volume,
lg/mL 9 (black), 25 lg/mL paclitaxel
l
A. Supplementary data
l
flat-bottom 96-well microtiter plates, and their A340 values were continuously
monitored using a VersaMaxÒ plate reader.
Supplementary data associated with this article can be found,
in the online version, at http://dx.doi.org/10.1016/j.bmcl.2013.
02.016.
the antibacterial activities of the diphenylnaphthalenes appear to
be related to the impact the compounds have on the polymeriza-
tion dynamics of FtsZ.
References and notes
Tubulin is the closest mammalian functional homolog to bacte-
rial FtsZ. We therefore sought to determine whether 9, which is a
potent stimulator of FtsZ polymerization (Fig. 2), would exert a
similar effect on mammalian tubulin. To this end, we monitored
the impact of 9 on porcine b-tubulin polymerization using a light
scattering assay similar to that described above for FtsZ polymeri-
zation. We used the antineoplastic drugs paclitaxel (taxol) and
nocodazole as positive controls in these assays, the former drug
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