Riccardin C derivatives as anti-MRSA agents: Structure-activity relationship of a series of hydroxylated bis(bibenzyl)s

Article abstract of DOI:10.1016/j.bmcl.2012.10.052

Members of a series of macrocyclic bis(bibenzyl) riccardin-class derivatives were found to exhibit antibacterial activity towards methicillin-resistant Staphylococcus aureus (anti-MRSA activity). Structure-activity relationship (SAR) studies were conducted, focusing on the number and position of the hydroxyl groups. The minimum essential structure for anti-MRSA activity was also investigated.

Full text of DOI:10.1016/j.bmcl.2012.10.052

Bioorganic & Medicinal Chemistry Letters 22 (2012) 7444–7447
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Riccardin C derivatives as anti-MRSA agents: Structure–activity relationship
of a series of hydroxylated bis(bibenzyl)s
Hiromi Sawada a, Miki Okazaki a, Daichi Morita a, Teruo Kuroda a, Kenji Matsuno a, Yuichi Hashimoto b,
Hiroyuki Miyachi a,
⇑
a Division of Pharmaceutical Sciences, Okayama University, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 1-1-1, Tsushima-Naka, Kita-ku, Okayama
700-8530, Japan
b Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Members of a series of macrocyclic bis(bibenzyl) riccardin-class derivatives were found to exhibit anti-
bacterial activity towards methicillin-resistant Staphylococcus aureus (anti-MRSA activity). Structure–
activity relationship (SAR) studies were conducted, focusing on the number and position of the hydroxyl
groups. The minimum essential structure for anti-MRSA activity was also investigated.
Ó 2012 Elsevier Ltd. All rights reserved.
Received 14 September 2012
Revised 6 October 2012
Accepted 10 October 2012
Available online 17 October 2012
Keywords:
Bis(bibenzyl)
Riccardin
Anti-MRSA
SAR
The emergence of multi-drug-resistant Gram-positive bacterial
pathogens, such as methicillin-resistant Staphylococcus aureus
(MRSA), vancomycin-resistant enterococci (VRE) and recently rec-
ognized vancomycin-intermediate-resistant Staphylococcus aureus
(VISA), has become a serious clinical problem worldwide, in terms
of both nosocomial and community-acquired infections.1 Conse-
quently, there has been considerable research aimed at the discov-
ery of novel anti-MRSA, anti-VRE and anti-VISA agents in recent
years.2 Some compounds have been developed, but they have
various limitations, as documented in the literature,3 especially
for high-risk patients. Therefore, new and potent anti-MRSA,
anti-VRE and anti-VISA agents are urgently required.
Recently, a series of bioactive polyphenolic compounds (flavo-
noids) was reported to show anti-MRSA and anti-VRE activities.4,5
Based on the reported structure–activity relationships (SAR), we
considered that phenolic structure might important for anti-bacte-
rial activity against multi-drug-resistant Gram-positive bacterial
pathogens. Therefore, we decided to screen the phenol-containing
compounds in our natural product-based synthetic library for anti-
MRSA activity, using the Methicillin-resistant strains of S. aureus,
OM481 and OM584, which were clinical isolates from Okayama
University hospital Japan.6 The MICs (minimum inhibitory concen-
trations) were determined by the liquid microdilution method. Se-
rial twofold dilutions of the drugs were used. The MIC was
determined as the lowest concentration of antimicrobial agent in
which cells were not able to grow.
To our delight, we found that a macrocyclic bis(bibenzyl) deriv-
ative, riccardin C (RC), exhibited anti-MRSA activity with a mini-
mum inhibitory concentration (MIC) of 3.2 lg/mL, which is
comparable to that of clinically used anti-MRSA agent, linezolid
(ZYVOXÒ) (Table 1). RC is a natural product characteristic of
liverworts.7 It has pleiotropic pharmacological effects, including
COX-inhibitory activity,7 HIV-1 reverse transcriptase-inhibitory
activity,8 antifungal activity,8 LXR-modulating activity,9,10
anti-cancer activity,11 and NOS-inhibiting activity.12 However,
anti-MRSA activity of RC has not been reported. Encouraged by
our finding, we further investigated the anti-MRSA activity of a
series of riccardin-class macrocyclic bis(bibenzyl) derivatives.
We have previously reported a series of macrocyclic bis(biben-
zyl) derivatives with 1–3 hydroxyl groups on the benzene moie-
ties.13 In addition, we newly prepared macrocyclic bis(bibenzyl)
derivatives with four hydroxyl groups on the benzene rings
(compounds 2 and 3).14,15 In addition, we prepared three frag-
ments of RC, as depicted in Figure 1. These riccardin derivatives
were synthesized as described previously, with slight modifica-
tions, using intramolecular Wittig reaction to construct the macro-
cycle as the key step (Scheme 1). The three fragments of RC were
prepared according to reported methods.16–18 The assay results
are summarized in Table 1.
These results indicate that the number of phenolic hydroxyl
groups, rather than their position, is critical for anti-MRSA activity
⇑
Corresponding author.
E-mail address: miyachi@pharm.okayama-u.ac.jp (H. Miyachi).
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.10.052

H. Sawada et al. / Bioorg. Med. Chem. Lett. 22 (2012) 7444–7447
7445
Table 1
Anti-MRSA activities of the present series of compounds
R1
C
R4
3
2
O
A
D
B
R3
R2
No.
R1
R2
R3
R4
MIC (l
g/mL)
OM481
OM584
Linezolid
1.0
1.0
24
25
26
27
28
29
30
31
32
1 (RC)
2
OCH3
H
H
OH
H
H
OH
OH
H
OH
OH
OH
H
OCH3
H
H
OH
H
OH
H
H
OH
OH
OH
H
H
OCH3
H
H
H
H
H
H
H
H
H
H
H
H
2-OH
3-OH
>100
>100
>100
>100
1.6
3.2
3.2
3.2
3.2
>100
>100
>100
>100
>100
>100
3.2
3.2
3.2
3.2
16
OH
H
OH
OH
OH
OH
OH
3.2
16
8
3
8
4
6.25
6.25
5
6
>100
>100
>100
>100
Linezolid ((S)-N-[[3-(3-fluoro-4-morpholinophenyl)-2-oxooxazolidin-5-yl]methyl]acetamide) was used as a positive control.
OH
OH
2
OH
OH
OH
O
4
O
3
O
HO
O
OH
HO
5
OH
OH
OH
OH
HO
HO
HO
HO
2
3
1
6
Figure 1. Chemical structure of riccardin C, its derivatives and its synthesized fragments.
against both OM481 and OM584 strains. That is, bis(bibenzyl)
derivatives with a single methoxy group did not show anti-MRSA
activity towards either strain. Bis(bibenzyl) derivatives with one
hydroxyl group also did not show apparent anti-MRSA activity,
except for compounds 28 and 29, which showed activity only to-
wards OM481. However, bis(bibenzyl)s with two and three pheno-
lic hydroxyl groups (30–32, 1 (RC)) showed apparent anti-MRSA
activity, because all of the compounds bearing two hydroxyl
groups exhibited anti-MRSA activity.
Next, we prepared riccardins 2 and 3 hydroxylated on various
benzene rings. However, the introduction of an additional hydroxyl
group at the 2- or 3-position of the A-benzene ring considerably
decreased the anti-MRSA activity. It remains to be examined
whether an additional hydroxyl group on rings B to D would influ-
ence the activity, and we are now preparing four appropriately
substituted bis(bibenzyl)s to investigate this issue.
activity towards both strains, with MIC’s of 3.2
lg/mL each. Thus,
the positions of the hydroxyl groups may not be relevant for the

7446
H. Sawada et al. / Bioorg. Med. Chem. Lett. 22 (2012) 7444–7447
OMe
MeO
OMe
O
MeO
O
O
OMe
O
MeO
3
2
O
MeO
a
b
OH
c
d
MeO
MeO
OHC
O
O
O
OHC
O
7
MeO
CO2Me
9a: 2-MeO
9b: 3-MeO
MeO
CO2Me
8a: 2-MeO
8b: 3-MeO
10a: 2-MeO
10b: 3-MeO
OMe
OMe
OMe
OMe
MeO
MeO
MeO
MeO
O
O
O
O
f
g
e
MeO
MeO
CHO
CHO
MeO
CHO
OMe
P+(Ph)3 Br-
OH
MeO
Br
MeO
14a: 2-MeO
MeO
MeO
14b: 3-MeO
11a: 2-MeO
11b: 3-MeO
12a: 2-MeO
12b: 3-MeO
13a: 2-MeO
13b: 3-MeO
OMe
OH
MeO
HO
O
O
h
i
OMe
OH
HO
MeO
15a: 2-MeO
2: 2-MeO
15b: 3-MeO
3: 3-MeO
CHO
CHO
OH
O
O
OH
O
k
l
O
j
18
17
20
22
4
16
19
Br
n
p
m
OMe
OH
MeO
HO
MeO
5
OH
OMe
PO(OEt)
2
Br
q
o
HO
OMe
MeO
OMe
21
23
6
Scheme 1. Synthesis of 2-OH and 3-OH riccardin C. Reagents and conditions: (a) 4-fluoro-3(or 2)-methoxybenzaldehyde, K2CO3, dry DMF, 120 °C, 18 h, 84%; (b) {5-methoxy-
2-[2-methoxy-4-(methoxycarbonyl)phenyl]benzyl}-triphenylphosphonium bromide, K2CO3, 18-crown-6, CH2Cl2, reflux, 15 h, quant.; (c) H2, 10% Pd/C, Et3N, AcOEt, rt, 5 h,
92%; (d) (1) LiAlH4, Et2O, 0 °C, 3 h; (2) H+, H2O, rt, 4 h, 63%; (e) CBr4, PPh3, benzene, reflux, 17 h, 72%; (f) PPh3, acetonitrile, reflux, 24 h, 98%; (g) NaOMe, CH2Cl2, rt, 24 h, 63%
(1.4 mM phosphonium salt solution was added dropwise at a rate of 14 mL/h); (h) H2 10% Pd/C, AcOEt, rt, 2 h, 66%; (i) BBr3, dry CH2Cl2, ꢀ78 to 0 °C, 7 h, 68%; (j) phenol, K2CO3,
dry DMF, 130 °C, 4 h, 84%; (k) mCPBA, dry CH2CL2, rt, 20 h, 89%; (l) 3 mol/KOH, EtOH, 50 °C, 3 h, 62%; (m) 4-bromoanisole, Pd(P(Ph)3)4, (nBu)4N+Brꢀ, K3PO4, DME, H2O, 70 °C,
3 h, 74%; (n) BBr3, dry CH2CL2, ꢀ78 °C, 22 h, 90%; (o) P(OEt)3, 80 °C, 22 h, 80%; (p) 4-methoxybenzaldehyde, MeONa, dry DMF, 120 °C, 4 h, 48%; (q) (1) H2 10% Pd/C, AcOEt, rt,
2 h, 82%; (2) BBr3, dry CH2CL2, ꢀ78 °C, 16 h, 61%.
We next wished to identify the minimum essential structure of
riccardin for anti-MRSA activity. For this purpose, we prepared
three fragments of RC, 4–6, according to reported methods
(Scheme 1). Compound 6 is identical with lunularin,16 which is a
precursor of the biosynthetic pathway of bis(bibenzyl)-type cyclic
phenolic natural products. Based on the brief SAR described above,
we speculated that all fragments with two phenolic hydroxyl
groups, that is, compounds 5 and 6, would exhibited substantial
activity, but to our surprise, the mono-hydroxy derivative, 4 exhib-
ited substantial anti-MRSA activity, with an MIC of 6.25 lg/mL. In
contrast, neither 5 nor 6 exhibited activity.
In conclusion, we found that riccardin-type macrocyclic bis(bib-
enzyl)s exhibit anti-MRSA activity against OM481 and OM584
strains. The SAR indicated that the number, but not the position,
of the phenolic hydroxyl groups is important for anti-MRSA activ-
ity in this series of compounds. However, the minimum essential

H. Sawada et al. / Bioorg. Med. Chem. Lett. 22 (2012) 7444–7447
7447
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14. Physicochemical properties of 2-OH-riccardin C: white powder, mp 123–125-
do, 1H NMR (CD3OD, 400 MHz): d = 6.88 (d, J = 8.4 Hz, 2H), 6.85 (d, J = 2.4 Hz,
2H), 6.79–6.65 (m, 5H), 6.35 (br, 2H), 6.19 (br, 1H), 5.97 (br, 1H), 5.42 (d,
J = 1.6 Hz, 1H), 4.61 (s, 2H), 3.00-2.48 (br, 8H). HRMS (FAB, [(M+H)+]):
structure for the activity appears to be the 2-phenoxyphenol part,
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anti-MRSA activity of these compounds.
Further structural development studies are under way, and we
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Acknowledgments
This work was supported in part by the Drug Discovery for
Intractable Infectious Diseases Project, Graduate School of
Medicine, Dentistry and Pharmaceutical Sciences, Okayama
University, the Uehara Memorial Foundation, the Tokyo Biochem-
ical Research Foundation (TBRF), and the Okayama Foundation
for Science and Technology (OFST).
References and notes
C28H25O5, calcd for 441.1702, found 441.1711.
15. Physicochemical properties of 3-OH-riccardin C: yellow amorphous solid, 1H
NMR (300 MHz, CDCl3) d 7.06–6.73 (m, 8H), 6.46–6.19 (m, 4H), 5.61 (s, 1H),
5.42–5.36 (m, 3H), 3.06–2.98 (m, 2H), 2.90–2.80 (m, 4H), 2.59–2.56 (m, 2H).
HRMS (FAB, [M+]): C28H24O5, calcd for 441.1702, found 441.1695.
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