Design, synthesis and biological evaluation of azithromycin glycosyl derivatives as potential antibacterial agents

Article abstract of DOI:10.1016/j.bmcl.2013.07.042

A series of 11,12-cyclic carbonate azithromycin-4″-O-carbamoyl glycosyl derivatives were designed, synthesized, and evaluated as antibacterial agents to search for target compounds with excellent activity. The results of preliminary antibacterial tests against eight strains in vitro revealed that all of the title compounds exhibited improved activities with broad spectrum compared with the parent compound. The glycosylated side chains may be the pharmacophores responsible for the improved activity.

Full text of DOI:10.1016/j.bmcl.2013.07.042

Bioorganic & Medicinal Chemistry Letters 23 (2013) 5057–5060
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design, synthesis and biological evaluation of azithromycin glycosyl
derivatives as potential antibacterial agents
Lei Zhang a,b, , Xiaoyun Chai a, , Baogang Wang a, Shichong Yu a, Honggang Hu a, Yan Zou a, Qingjie Zhao a,
Qingguo Meng b, , Qiuye Wu a,
⇑
⇑
a Department of Organic Chemistry, School of Pharmacy, Second Military Medical University, 325 Guohe Road, Shanghai 200433, China
b Department of Medicinal Chemistry, School of Pharmacy, Yantai University, 30 Qingquan Road, Yantai 264005, China
a r t i c l e i n f o
a b s t r a c t
A series of 11,12-cyclic carbonate azithromycin-400-O-carbamoyl glycosyl derivatives were designed, syn-
thesized, and evaluated as antibacterial agents to search for target compounds with excellent activity.
The results of preliminary antibacterial tests against eight strains in vitro revealed that all of the title
compounds exhibited improved activities with broad spectrum compared with the parent compound.
The glycosylated side chains may be the pharmacophores responsible for the improved activity.
Ó 2013 Elsevier Ltd. All rights reserved.
Article history:
Received 13 June 2013
Revised 15 July 2013
Accepted 17 July 2013
Available online 25 July 2013
Keywords:
Azithromycin
Antibacterial activity
Glycosyl
Synthesis
Over the past decade, macrolide antibiotics have played an
important role in the clinical treatment of upper and lower respi-
ratory tract infections. These antibiotics act by binding to the ribo-
somal RNA of bacteria. The previous studies provided strong,
detailed evidence of this aforementioned interaction at the molec-
ular level via high-resolution X-ray cocrystal structures.1–4 These
X-ray studies demonstrated that macrolides inhibit bacterial pro-
tein synthesis by sterically blocking the passage of nascent poly-
peptides through the exit tunnel of the ribosome.5
Macrolide antibiotics, such as erythromycin A (ERY), clarithro-
mycin (CLA), and azithromycin (AZI) (Fig. 1), are widely prescribed
clinically. However, these antibiotics have various disadvantages.
ERY, a first-generation macrolide, is readily degraded under acidic
conditions, thus losing its antibacterial activity;6 these degraded
products are responsible for undesirable gastrointestinal side
effects.7,8 Second-generation macrolides, such as CLA and AZI, have
been widely used for respiratory tract infection due to their supe-
rior antibacterial activity, pharmacokinetic properties and fewer
gastrointestinal side effects compared to ERY.9 However, their clin-
ical uses have been limited by the emergence of drug resistance.10
The increasing incidence of bacterial resistance is becoming a major
threat to the successful treatment of infectious diseases. The most
serious threat is the increasing resistance of community-acquired
respiratory tract infections to various antimicrobials, which is a
pandemic phenomenon.11 Third-generation macrolide ketolides,
such as telithromycin, can effectively address bacterial resistance
and other issues associated with current macrolide regimens.12,13
However, the use of telithromycin suffers from limitations such
as hepatotoxicity.14
With the widespread emergence of antibacterial resistance
against macrolides, there is still a need to develop and extend safe
chemotherapeutic agents with potent antibacterial activity. Re-
cently, considerable efforts have been focused on discovering novel
macrolides to combat the resistance. Research has indicated that
compounds containing particular groups in the 400-position of the
cladinose sugar were effective against macrolide resistant strains.15
Additionally, the C-11,12 carbamate side chain of macrolides may
aid in interactions with the target enzyme.16 So, we have chosen
the 400-position of the cladinose sugar as the modification position.
Glycosyl moieties exist extensively in organisms and exhibit a
variety of biological functions, such as diagnostic and therapeutic
potential. D-desosamine and L-cladinose, the two monosaccha-
rides in macrolide structures, are important for the binding of mac-
rolides to ribosomes. X-ray analyses of macrolides have revealed
that the hydroxy group of a glycosyl moiety was important for
hydrogen bond formation.2 However, there have been few reports
about glycosylation modification of macrolide antibiotics.17,18 As
we believe that the glycosylation may help for the hydrogen bond
formation, we therefore introduced glycosyl moieties to AZI
through a carbamoyl group and examined the antibacterial activi-
ties of the resultant products.
⇑
Corresponding authors. Tel./fax: +86 21 81871225. (Q.W.).
E-mail addresses: qinggmeng@163.com (Q. Meng), wuqy640319@163.com
(Q. Wu).
These authors contributed equally to this work.
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.07.042

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L. Zhang et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5057–5060
By substitution of the 400-position with various carbamoyl-
Committee of Clinical Laboratory Standard).19 The MIC80 was de-
fined as the first well with an approximate 80% reduction in growth
compared to the growth of the drug-free well. For the assays, the ti-
tle compounds to be tested were dissolved in dimethyl sulfoxide
(DMSO), serially diluted in growth medium, inoculated and incu-
bated at 35 °C.
groups, a series of novel 400-substituted azithromycin derivatives
were obtained. Further modification at 11,12-cyclic carbonate
azithromycin was performed. We hypothesized that the 11,12-cyc-
lic carbonate azithromycin-400-O-carbamoyl glycosyl derivatives
might enhance the antibacterial activity against resistant strains.
Eight glycosylated intermediates 1f–8f were prepared from the
corresponding commercially available saccharides by following a
series of established transformations. We have chosen 1f as an
example (Scheme 1). First, all the hydroxyl groups of glucose (1a)
were protected with acetyl groups to give the acetylated saccha-
ride (1b) in an excellent yield. The treatment of 1b with 2-azido-
ethanol in the presence of BF3ꢀEt2O afforded a series of protected
azidoethyl-glycosides (1c) in the b-configuration with a very good
yield. After deprotection of 1c, we obtained the azidoethyl-glyco-
sides (1d). Compound 1d was protected with benzyl groups to give
the benzylated saccharide (1e) in an excellent yield. Lastly, 1f was
reduced by Pd/CaCO3 under a hydrogen atmosphere. The other se-
ven glycosylated intermediates (2f–8f) were synthesized in the
same manner.
The selected strains evaluated were methicillin-susceptible
Staphylococcus aureus MSSA-1 (S. aureus MSSA-1), methicillin-resis-
tant Staphylococcus aureus MRSA-1 (S. aureus MRSA-1), Staphylococ-
cus aureus ATCC25923 (S. aureus ATCC25923), Streptococcus
pneumoniae 943 (S. pneumoniae943), Staphylococcus pneumonia
746 (S. pneumoniae 746), Streptococcus pyogenes 447 (S. pyogenes
447), Escherichia coli 236 (E. coli 236), and Escherichia coli
ATCC25922 (E. coli ATCC25922). ERY and AZI served as the positive
control and were obtained from their respective manufacturers.
The results of the assays are summarized in Table 1. The data points
express the mean of replicate experiments. All of our susceptibility
tests were performed three times using each antibacterial agent.
The results indicated that nearly all of the 11,12-cyclic car-
bonate
azithromycin-400-O-carbamoyl
glycosyl
derivatives
The general synthetic methodology for preparing the com-
pounds of interest (F1–F8) is outlined in Scheme 2. Protection of
the 20-hydroxyl group of azithromycin with acetic anhydride
provided 20-acetyl azithromycin (B) in 82% yield. 11,12-Cyclic car-
bonate azithromycin 400-O-acylimidazolide (C) was obtained in 62%
yield by treatment of compound B with NaH and N,N0-carbonyldi-
imidazole (CDI) in DMF. Then, the intermediates (D1–D8) were
prepared by coupling compound C, followed by benzyl protection
of the glycosylated group in the presence of 1,8-diazabicyclo
[5.4.0] undec-7-ene (DBU). Lastly, intermediates D1–D8 were re-
duced by 10% Pd-C under a hydrogen atmosphere. After methanol-
ysis, we obtained compounds F1–F8. The yields were within the
range of 70–80%.
showed moderate activity against all the strains, and someof
the derivatives exhibited improved activity compared with AZI
and ERY. Notably, the MIC80 values indicate that compounds
F1 and F2 showed improved activity against all of the bacterial
strains relative to the other compounds, expressing the same
or higher antibacterial activities as AZI. Compounds F7 and F8
with the disaccharide side chain showed the least activity; this
observation could be due to increasing side chain length. Among
the compounds tested, Compounds F1 and F6 showed the same
activity against the S. aureus MSSA-1 as AZI, and they exhibited
eightfold higher activity than ERY. Particularly, compounds F1
and F2 showed fourfold and eightfold higher activity against
the S. pneumoniae 943 than AZI and ERY, respectively. In addi-
The in vitro antibacterial activities were reported as minimum
inhibitory concentrations (MICs), which were determined using
astandard dilution assay as recommended by the NCCLS (National
tion, the activity of compoundsF1 and F2 (MIC80 2 lg/mL) in-
creased significantly against S. pneumoniae 746, showing
eightfold higher activity than ERY. The activity of compounds
Figure 1. The structures of macrolide antibiotics.
Scheme 1. The synthesis of the saccharide intermediates. Reactions and conditions: (a) Ac2O, CH3COONa, reflux, 5 h, in 90% yield; (b) HOCH2CH2N3, BF3ꢀEt2O, DCM, Ar, 0 °C,
18 h, in 80% yield; (c) CH3ONa, CH3OH, 4 h, in 93% yield; (d) BnBr, NaH, TBAI, DMF, in 52% yield; and (e) Pd/CaCO3, H2, CH3OH, in 98% yield.

L. Zhang et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5057–5060
5059
Scheme 2. The synthesis of the compounds of interest. Reactions and conditions: (a) Ac2O, DCM, Et3N, rt, 24 h, in 82% yield; (b) CDI, NaH, DMF, rt, 24 h, in 62% yield; (c)
R’OCH2CH2NH2, DMF, DBU, rt, 12 h, in 21–38% yield; (d) Pd/C, H2, CH3OH, in 75–88% yield; and (e) CH3OH, 55 °C, 20 h, 70–80% yield.
Table 1
Antibacterial activities of the title compounds in vitro (MIC80, lg/mL)
Compd
S. aureus MSSA-1
S. aureus MRSA-1
S. aureus ATCC25923
S. pneumoniae 943
S. pneumoniae 746 S. pyogenes 447
E. coli 236
E. coli ATCC25922
F1
F2
F3
F4
F5
F6
F7
F8
AZI
ERY
2
4
8
16
8
2
16
32
2
16
32
8
16
8
8
32
8
8
4
16
32
16
16
16
1
1
4
2
4
4
8
8
4
8
2
2
4
4
8
16
64
32
16
64
64
32
64
32
32
64
128
64
64
64
128
128
128
256
256
128
128
128
256
128
128
256
256
256
256
8
8
16
32
16
16
16
16
8
16
16
F1 and F2 against E. coli 236 and E. coli ATCC25922 was either
maintained or slightly greater.
In conclusion, an efficient method has been developed for
the synthesis of 11,12-cyclic carbonate azithromycin-400-O-car-

5060
L. Zhang et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5057–5060
bamoyl glycosyl derivatives. The results of preliminary antibac-
terial tests against eight strains in vitro demonstrated that
these derivatives exhibited improved activities with a broad
spectrum compared with the parent compound, which could
be attributed to the glycosylated side chains. The monosaccha-
ride side chain may help the target compounds fit into the
binding pocket and interact with thebacterial ribosome. How-
ever, compounds F7 and F8 with disaccharide side chains ex-
pressed the lowest activity, indicating that the bulky
disaccharide side chain might hinder the interactions of the
derivative with the bacterial ribosome. This study has led to
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This work was supported by the Major Project of the Science
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Supplementary data
Supplementary dataassociated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2013.
07.042.