Synthesis and structure-activity relationship of dicationic diaryl ethers as novel potent anti-MRSA and anti-VRE agents

Article abstract of DOI:10.1016/j.bmcl.2009.06.077

A series of dicationic diaryl ethers have been synthesized and evaluated for in vitro antibacterial activities, including drug resistant bacterial strains. Most of these compounds have shown potent antibacterial activities. Several compounds, such as pipe

Full text of DOI:10.1016/j.bmcl.2009.06.077

Bioorganic & Medicinal Chemistry Letters 19 (2009) 4626–4629
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and structure–activity relationship of dicationic diaryl ethers
as novel potent anti-MRSA and anti-VRE agents
Laixing Hu a, Maureen L. Kully b, David W. Boykin a, Norman Abood c,
*
a Department of Chemistry, Georgia State University, Atlanta, GA 30303-3083, USA
b Naeja Pharmaceutical Inc., Edmonton, AB, Canada T6E 5V2
c Immtech Pharmaceuticals, Inc., 150 Fairway Dr., Suite 150, Vernon Hills, IL 60061-1860, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of dicationic diaryl ethers have been synthesized and evaluated for in vitro antibacterial activi-
ties, including drug resistant bacterial strains. Most of these compounds have shown potent antibacterial
activities. Several compounds, such as piperidinyl and thiomorpholinyl compounds 9e and 9l, improved
the antimicrobial selectivity and kept potent anti-MRSA and anti-VRE activity. The most potent bis-indole
Received 1 June 2009
Revised 18 June 2009
Accepted 22 June 2009
Available online 25 June 2009
diphenyl ether 19 exhibited anti-MRSA MIC value of 60.06
l
g/mL and enhanced antimicrobial selectivity.
Ó 2009 Elsevier Ltd. All rights reserved.
Keywords:
Antibacterials
Diamidines
Benzimidazoles
Indoles
Interest in novel antimicrobial agents has been stimulated by
the emergence of multi-drug resistant Gram-positive bacteria,
including methicillin-resistant Staphylococcus aureus (MRSA),
methicillin-resistant Staphylococcus epidermidis (MRSE) and vanco-
mycin-resistant Enterococcus faecium (VRE).1 Severe nosocomial
and community-acquired infections caused by these pathogens
has become a significant challenge in the clinic.2
tory activity.7,8 In order to investigate the structure–activity rela-
tionship (SAR) of this series of novel dicationic bis-
benzimidazolyl diphenyl ether compounds, we designed and syn-
During the course of our efforts to develop novel antimicrobial
agents, we have discovered a new class of dicationic bis-benzimid-
azole derivatives which displayed potent anti-MRSA and anti-VRE
activities.3 The lead compound 1 (Fig. 1) has shown significant
antibacterial activity including MRSA and VRE (MIC 6 0.5 lg/mL).
Optimization of the central linker of lead compound 1 resulted in
the discovery of 4,40-bis-[2-(5-N-isopropylamidino)benzimidazol-
yl] diphenyl ether 2, which displayed more potent anti-MRSA
and anti-VRE activity than lead compound 1.4 Interestingly, the
mono-amidino benzimidazolyl diphenyl ether derivative 3 has
been previously reported as an inhibitor of bacterial two-compo-
nent system (TCS), which showed anti-MRSA and anti-VRE activity
(MIC = 16 l
g/mL).5 Another diamidino benzimidazolyl diphenyl
ether derivative 4, a DNA minor groove binder, displayed inhibi-
tory activity of arginine-specific esteroproteases and antifungal
activity.6 A 6-amidino indole analogue 5 of the diamidino benz-
imidazole compound 4 exhibited antimicrobial activity and human
mitogen-activated protein kinase phosphatase-3 (MKP-3) inhibi-
* Corresponding author.
E-mail address: nabood@immtechpharma.com (N. Abood).
Figure 1. Benzimidazole and indole amidine compounds.
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.06.077

L. Hu et al. / Bioorg. Med. Chem. Lett. 19 (2009) 4626–4629
4627
thesized analogues of the new lead compound 2. In this article, we
report the synthesis, in vitro antibacterial activity and SAR of these
compounds as novel potent anti-MRSA and anti-VRE agents.
We have reported the synthesis of lead compound 2 by the
condensation of 4-(N-isopropylamidino)-1,2-phenylenediamine
hydrochloride with commercial available 4-(4-formylphen-
oxy)benzaldehyde in presence of benzoquinone as oxidative re-
agent.4 The analogs 9a–l of the lead compound 2 was prepared
by following this procedure as shown in Scheme 1. The cyano
group of starting material 3,4-diaminobenzonitrile 6 was con-
verted into the imidate ester 7 by using the Pinner method.9 Then,
the imidate ester was used directly to react with suitable commer-
cially available amines to yield the 4-(N-substituted amidino)-1,2-
phenylenediamines 8a–l. Condensation of these derivatives with
4-(4-formylphenoxy)benzaldehyde in presence of benzoquinone
as oxidative reagent afforded the corresponding dicationic bis-
benzimidazolyl diphenyl ethers 9a–l. Various derivatives 13a–e
substituted on the phenyl ring of the lead compound 2 were pre-
pared from bis-benzaldehydes 12a–e by condensation with 4-(N-
isopropylamidino)-1,2-phenylenediamine hydrochloride using
the same approach as described above (Scheme 2). Nucleophilic
aromatic substitution reactions of 4-hydroxybenzaldehydes 10a–
c with 4-fluorobenzaldehydes 11a–c generated bis-benzaldehydes
12a–e in reasonable yields.10 The preparation of pyridinyl bis-
benzimidazole amidines 15a–b was achieved by using the previ-
ously described procedure in two steps from 4-hydroxybenzalde-
hyde 10a, as shown in Scheme 3. 4,40-Bis-[2-(6-cyanoindolyl)]
diphenyl ether 18 was prepared from 4-methyl-3-nitrobenzonitri-
le 16 by condensation with 4-(4-formylphenoxy)benzaldehyde and
followed by heating with neat triethylphosphite (Scheme 4).7,8 The
dicyano compound 18 was converted to the desirable 6-(N-isopro-
pylamidino)indole compound 19 by using the Pinner method as
previously reported.7,8
Scheme 2. Reagents and conditions: (a) K2CO3, DMAC, 150 °C, 60–76%; (b) 4-(N-
isopropylamidino)-1,2-phenylenediamine, 1,4-benzoquinonE, EtOH. reflux, 45–
78%.
All the dicationic diaryl ether hydrochloride compounds pre-
pared herein were screened for their potential antibacterial activi-
ties in vitro against ten selected Gram-positive bacterial strains
Scheme 3. Reagents and conditions: (a) 2-fluoro-5-formylpyridine or 5-fluoro-2-
formylpyridine, K2CO2, DMAC, 150 °C, 73% or 68%; (b) 4-(N-isopropylamidino)-1,2-
phenylenediamine hydrochloride, 1,4-benzoquinone, EtOH, reflux, 58% or 67%.
Scheme 1. Reagents and conditions: (a) HCl (gas), EtOH; (b) R–NH2, EtOH, reflux,
45–90% in two steps; (c) 4-(4-formylphenoxy)benzaldehyde, 1,4-benzoquinone,
EtOH, reflux, 36–85%.
Scheme 4. Reagents and conditions: (a) 4-(4-formylphenoxy)benzaldehyde, piper-
idine, 100 °c, 50%; (b) P(OEt)3, 160 °C, 39%; (c) HCl (gas), EtOH; (d) I–PrNH2, EtOH,
reflux, 70% in two steps.

4628
L. Hu et al. / Bioorg. Med. Chem. Lett. 19 (2009) 4626–4629
and two anaerobic bacterial strains, including MRSA, multi-drug
resistant S. aureus (MDRSA), MRSE and VRE strains.3 We also
screened their antifungal activity against Candida albicans in order
to evaluate the specificity of their antimicrobial activity. Penicillin
G (Pen G), ciprofloxacin (CPLX) and vancomycin (VCM)) were used
as reference standards. The minimum inhibitory concentration
(MIC) results for the test compounds are shown in Table 1. Of
the twenty compounds tested against the Gram-positive aerobic
strains, almost all of these compounds showed potent antibacterial
pounds were comparable to that of the lead compound 1, however,
the c-pentyl compound gave decreased anti-anaerobic bacterial
activity. N-butyl and c-propyl substituents led to some loss of anti-
bacterial potency. The pyrrolidinyl derivative showed similar anti-
S. aureus and anti-S. epidermidis activities but was less active against
theother bacterialstrains compared to thatof lead compound1. Fur-
ther SAR study of thislead compound 1 by replacementof the central
ethylene linker with various two-atom or one-atom groups has
shown that the antibacterial activity of the dicationic bis-benzimid-
azole compounds is related to the relative electronegativity of the
central linkers, however, the central linkers of one-atom and two-
atoms showed opposite effects on activity. This result maybe related
to the difference in geometry of the two series since the single-atom
linker produces a much more bent molecule than the two-atom lin-
ker.4 Based on the above SAR information, we further designed and
prepared the N-substituted analogs 9a–l of lead compound 2 for
probing the SAR of this series of novel dicationic bis-benzimidazolyl
diphenyl ether compounds. sec-Butyl, sec-pentyl and c-pentyl com-
pounds 9a–c showed similar effects on antibacterial activity to that
activities, including MRSA and VRE strains (MIC value 61 lg/mL);
only one compound 9h was found to have only moderate antibac-
terial activity. Nine compounds 9b, 9d–g, 9i, 9l, 13a and 19 also
showed good activity against two anaerobic bacterial strains
(MIC value 64 lg/mL).
The SAR study of the lead compound 1 has shown that bis-5-N-
substituted amidine of 1H-benzimidazole is very important for
achieving potent antibacterial activity.3 The parent diamidine com-
pound of the lead compound 1 exhibited almost no antibacterial
activity. Antibacterial activity of the sec-butyl and c-pentyl com-
Table 1
In vitro antibacterial activity of benzimidazole amidine compoundsa
Strain/compound
MICs (lg/mL)
2
9a
9b
9c
9d
9e
9f
9g
9h
S. aureus ATCC 29213
S. aureus BAA-39b
0.25
0.5
0.5
0.5
0.25
0.5
0.25
0.5
0.5
0.25
0.5
0.25
0.12
0.12
<0.06
0.25
<0.06
<0.06
0.12
2
0.25
0.5
0.25
0.25
0.25
0.12
0.12
0.12
0.5
0.25
0.12
<0.06
4
0.5
0.5
0.25
0.25
0.5
0.5
1
0.5
0.25
1
8
4
4
1
1
>32
8
2
4
S. aureus ATCC 33591c
S. epidermidis ATCC 12228
S. epidermidis ATCC 51625d
S. pneumoniae ATCC 6301
E. faecalis ATCC 51575e
E. faecium ATCC 700221e
B. subtils ATCC 23857
B. cereus ATCC 11778
B. fragilis ATCC 23745
C. perfringens ATCC 10388
C. albicans ATCC 90028
0.12
<0.06
<0.06
<0.06
0.12
<0.06
<0.06
<0.06
2
0.25
0.12
0.12
<0.06
0.25
<0.06
<0.06
<0.06
16
0.25
0.12
0.25
<0.06
0.5
<0.06
0.12
0.5
0.12
<0.06
0.12
<0.06
0.25
0.12
<0.06
0.25
2
0.12
0.12
<0.06
0.25
<0.06
<0.06
<0.06
32
1
4
0.5
4
4
2
8
>32
>32
16
0.5
1–2
0.5
2
0.25
2
0.5
4
0.25
8
1
4
9i
9j
9k
9l
13a
13b
13c
13d
13e
S. aureus ATCC 29213
S. aureus BAA-39b
0.25
0.25
0.25
0.12
0.25
0.12
0.5
0.5
0.25
0.5
4
1
1
1
0.5
0.5
0.5
1
1
0.5
0.5
0.25
0.12
0.25
0.25
0.5
0.5
0.25
0.5
4
0.25
0.5
0.5
0.5
0.5
0.5
0.25
0.5
0.25
0.25
0.25
60.06
60.06
60.06
0.25
60.06
0.12
0.12
16
0.5
0.5
0.25
0.5
0.5
0.5
0.5
0.5
0.12
8
S. aureus ATCC 33591c
S. epidermidis ATCC 12228
S. epidermidis ATCC 51625d
S. pneumoniae ATCC 6301
E. faecalis ATCC 51575e
E. faecium ATCC 700221e
B. subtils ATCC 23857
B. cereus ATCC 11778
B. fragilis ATCC 23745
C. perfringens ATCC 10388
C. albicans ATCC 90028
0.12
60.06
60.06
60.06
0.12
60.06
60.06
0.12
4
0.25
60.06
0.12
60.06
0.5
60.06
60.06
0.12
>32
0.25
60.06
0.12
60.06
0.12
60.06
60.06
0.12
8
0.25
60.06
60.06
60.06
0.25
60.06
60.06
0.12
>32
1
0.5
0.5
16
16
32
2
8
2
16
1
16
0.25
2
0.25
2
0.25
4
0.25
4
0.5
1
15a
15b
19
Pen-G
CPLX
VCM
S. aureus ATCC 29213
S. aureus BAA-39b
1
1
0.5
0.5
0.5
0.5
0.12
1
>32
>32
32
32
<0.06
4
0.5
8
60.12
60.12
60.12
0.5
0.5
1
1
1
1
1
>64
>64
0.12–0.5
1–60.12
4–8
0.12–0.25
>32
60.06
60.06
60.06
60.06
60.06
0.25
60.06
60.06
60.06
0.125
0.5
S. aureus ATCC 33591c
S. epidermidis ATCC 12228
S. epidermidis ATCC 51625d
S. pneumoniae ATCC 6301
E. faecalis ATCC 51575e
E. faecium ATCC 700221e
B. subtils ATCC 23857
B. cereus ATCC 11778
B. fragilis ATCC 23745
C. perfringens ATCC 10388
C. albicans ATCC 90028
0.12
0.12
0.12
0.25
60.06
0.25
0.12
>32
2
0.12
0.12
60.06
0.12
60.06
60.06
0.12
32
0.5
>32
>64
60.12
60.12
0.5
0.25
—
<0.06
4–>32
4–8
60.06–0.12
>32
0.5
4
2
0.5
a
NCCLS guidelines M11-A6 and M7-A6 followed.
b
c
MDRSA.
MRSA.
MRSE.
VRE.
d
e

L. Hu et al. / Bioorg. Med. Chem. Lett. 19 (2009) 4626–4629
4629
of the lead compound 1. However, the pyrrolidinyl substituent 9d
displayed only slightly less antibacterial potency compared to lead
compound 1, which is clearly different from the result in the ethyl-
ene series. It is also noteworthy that the compound 9d decreased the
antifungal activity by 2–4-folds. The differences in antimicrobial
activity are likely due to the difference in shapes of these two series
of compounds. Interestingly, the N-piperidinyl group in 9e further
enhanced the selectivity of antimicrobial activity and kept the po-
tent anti-MRSA and anti-VRE activities. 4-Methylpiperidnyl and
the 3,5-dimethylpiperidinyl derivatives 9f–g showed similar effects
on the activity in comparison to 9d. However, the 4-pyrrolidinyl-
piperidinyl group led to some loss of potency. This result may sug-
gest that the more bulky group is not beneficial to the
antibacterial activity. N-Hexamethyleneiminyl, ethylpiperazinyl,
morpholinyl, and thiomorpholinyl substituents 9i–l were slightly
less potent than lead compound 2. In particular, the thiomorpholinyl
compound 9l exhibited the best selectivity by a factor of 32 for anti-
the effect of the bis-benzimidazole rings of the lead compound 2
on antibacterial activity. Very interestingly, the bis-indole com-
pound 19 showed the most potent activity (MIC 6 0.5 lg/mL) com-
pared to that of lead compound 2 and the other analogues. In
particular, the anti-MDRSA activity of the compound 19,
MIC 6 0.06 lg/mL, was more active than that of the lead com-
pound 2 by 8 times and VCM by 16 times. The compound 19 was
also more potent than the lead compound 2 against the anaerobic
bacterial strain Bacillus subtilis and the fungal strain Candida albi-
cans. On the other hand, it is noteworthy that compound 19 exhib-
ited an antimicrobial selectivity factor of 8 for anti-MRSA activity
to antifungal activity, which is much better than that of lead com-
pound 2 by a factor of 2. This result suggests that replacement of
the bis-benzimidazole rings of the lead compound 2 can lead to
improved antibacterial activity and enhanced antimicrobial
selectivity.
In conclusion, we have synthesized and evaluated the antibac-
terial activities of the analogues of lead compound 2 for probing
the SAR of this system. Most of the compounds show significant
antibacterial activities against Gram-positive bacteria, including
drug resistant bacterial strains. Several compounds, such as pipe-
ridinyl and thiomorpholinyl compounds 9e, 9l, show improved
antimicrobial selectivity and at the same time keep potent anti-
MRSA and anti-VRE activity. The SAR study of two series of dicat-
ionic bis-benzimidazole compounds (1 and 2) containing ethylene
and oxygen central linker have shown that the central linker
causes different effect on the antibacterial activity. Replacement
of the benzimidazole ring of the lead compound 2 with an indole
ring resulted in improvement of antibacterial activity and en-
hanced antimicrobial selectivity. This series of dicationic diaryl
ethers merits further investigation as novel potent anti-MRSA
and anti-VRE agents.
MRSA and anti-VRE activity (MIC 6 0.5
(MIC = 16 g/mL).
lg/mL) to antifungal activity
l
The effect of the substituents on the central phenyl ring of the
lead compound 2 was examined. Both electron-donating groups,
such as methoxy or chloro group, and strong electron-withdrawing
group—trifluoromethyl group located at various positions on the
phenyl ring (13a–e) showed no apparent effect on the activity
against Gram-positive bacterial activity including MRSA and VRE,
however, decreased anti-anaerobic bacterial activity was noted.
Replacement of one of the 2-phenyl rings of 2 by 2-pyridin-5-yl
or 5-pyridin-2-yl ring yielded the compounds 15a–b. Both pyridi-
nyl compounds 15a–b showed a slight loss of activity against
Gram-positive bacterial strains and decreased potency against
anaerobic bacterial strains compared to 2. This result is in contrast
to that for the pyridinyl compounds of the lead compound 1 which
showed only moderate antibacterial activity.3 The SAR results for
these two series of dicationic bis-benzimidazole compounds con-
taining ethylene and oxygen central linkers suggested that the cen-
tral linker leads to different effects on the antibacterial activity.
Like imidazoles, benzimidazoles exhibit fast prototropic tau-
tomerism,11 which leads to an equilibrium mixture of symmetrical
tautomers 2a and 2b (Figure ure2). Since the 6-amidino indole ana-
logue 5 has been previously reported as an antimicrobial agent,7
we replaced of the bis-benzimidazole rings with bis-indole rings
to yield 4,40-bis-[2-(6-N-isopropylamidino)] diphenyl ether 19,
which is an isostere of one tautomer of 2, in order to investigate
References and notes
1. (a) Barrett, J. F. Expert Opin. Ther. Targets 2005, 9, 253; (b) Loffler, C. A.;
MacDougall, C. Exp. Rev. Anti Inf. Ther. 2007, 5, 961; (c) Bryskier, A. Exp. Rev. Anti
Inf. Ther. 2005, 3, 505.
2. (a) Levy, S. B.; Marshell, B. Nat. Med. 2004, 10, S122; (b) Mitchell, M. O. Anti-Inf.
Agents Med. Chem. 2007, 6, 243.
3. Hu, L.; Kully, M. L.; Boykin, D. W.; Abood, N. Bioorg. Med. Chem. Lett. 2009, 19,
1291.
4. Hu, L.; Kully, M. L.; Boykin, D. W.; Abood, N. Bioorg. Med. Chem. Lett. 2009.
doi:10.1016/j.bmcl.2009.05.061.
5. Weidner-Wells, M. A.; Ohemeng, K. A.; Nguyen, V. N.; Fraga-Spano, S.;
Macielag, M. J.; Werblood, H. M.; Foleno, B. D.; Webb, G. C.; Barrett, J. F.;
Hlasta, D. J. Bioorg. Med. Chem. Lett. 2001, 11, 1545.
6. (a) Tidwell, R. R.; Geratz, J. D. J. Med. Chem. 1978, 21, 613; (b) Del Poeta, M.;
Schell, W. A.; Dykstra, C. C.; Jones, S. K.; Tidwell, R. R.; Kumar, A.; Boykin, D. W.;
Perfect, J. R. Antimicrob. Agents Chemother. 1998, 42, 2503; (c) Goodwin, K. D.;
Lewis, M. A.; Tanious, F. A.; Tidwell, R. R.; Wilson, W. D.; Georgiadis, M. M.;
Long, E. C. J. Am. Chem. Soc. 2006, 128, 7846; (d) Tanious, F. A.; Laine, W.;
Peixoto, P.; Bailly, C.; Goodwin, K. D.; Lewis, M. A.; Long, E. C.; Georgiadis, M.
M.; Tidwell, R. R.; Wilson, W. D. Biochemistry 2007, 46, 6944.
7. (a) Dann, O.; Fick, H.; Pietzner, B.; Walkenhorst, E.; Fernbach, R.; Zeh, D. Liebigs
Ann. Chem. 1975, 160; (b) Dann, O.; Ruff, J.; Wolff, H. P.; Griessmeier, H. Liebigs
Ann. Chem. 1984, 409.
8. Lazo, J. S.; Nunes, R.; Skoko, J. J.; Queiroz de Oliveira, P. E.; Vogt, A.; Wipf, P.
Bioorg. Med. Chem. Lett. 2006, 14, 5643.
9. Roger, R.; Neilson, D. G. Chem. Rev. 1961, 61, 179.
10. (a) Yeager, G. W.; Schissel, D. N. Synthesis 1991, 1991, 63; (b) Pfefferkorn, J. A.;
Greene, M. L.; Nugent, R. A.; Gross, R. J.; Mitchell, M. A.; Finzel, B. C.; Harris, M.
S.; Wells, P. A.; Shelly, J. A.; Anstadt, R. A.; Kilkuskie, R. E.; Kopta, L. A.;
Schwende, F. J. Bioorg. Med. Chem. Lett. 2005, 15, 2481.
11. (a) Grimmett, M. R.. In Comprehensive Heterocyclic Chemistry; Katritzky, A. R.,
Rees, C. W., Eds.; Pergamon: Oxford, 1984; Vol. 5, p 345; (b) Grimmett, M. R.. In
Comprehensive Heterocyclic Chemistry; Katritzky, A. R., Rees, C. W., Scriven, E. F.
V., Eds.; Elsevier: Oxford, 1996; Vol. 7, p 77; (c) Grimmett, M. R. Sci. Synth.
2002, 12, 529; (d) Alamgir, M.; Black, D. St. C.; Kumar, N. Top. Heterocycl. Chem.
2007, 9, 87.
Figure 2. The lead compound 2 tautomers.