Novel antibacterial azetidine lincosamides

Article abstract of DOI:10.1016/j.bmcl.2008.03.032

The synthesis and evaluation of novel azetidine lincosamides 1 are described. Eleven new (3-trans-alkyl)azetidine-2-carboxylic acids were synthesized via alkylation of N-TBS-4-oxo-azetidine-2-carboxylic acid and subsequent elaboration then coupled to 7-ch

Full text of DOI:10.1016/j.bmcl.2008.03.032

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Bioorganic & Medicinal Chemistry Letters 18 (2008) 2645–2648
Novel antibacterial azetidine lincosamidesq
Hardwin O’Dowd,* Jason G. Lewis, Joaquim Trias, Rumi Asano, Johanne Blais,
Sara L. Lopez, Craig K. Park, Charlotte Wu, Wen Wang and Mikhail F. Gordeev
Pfizer Global Research & Development, Fremont, Formerly Vicuron Pharmaceuticals, Inc., CA 94555, USA
Received 23 January 2008; accepted 7 March 2008
Available online 14 March 2008
Abstract—The synthesis and evaluation of novel azetidine lincosamides 1 are described. Eleven new (3-trans-alkyl)azetidine-2-car-
boxylic acids were synthesized via alkylation of N-TBS-4-oxo-azetidine-2-carboxylic acid and subsequent elaboration then coupled
to 7-chloro-1-methylthio-lincosamine. The resulting lincosamides differ from the drug clindamycin in both the size of the ring and
the position/structure of the alkyl side-chain. SAR within the series was explored with attention to alkyl variants in positions 1 and 3
of the azetidine ring.
Ó 2008 Elsevier Ltd. All rights reserved.
Of the lincosamide class of antibacterial protein synthe-
sis inhibitors, clindamycin (CLI) is the most widely used
in human medicine. The clinical importance of clinda-
mycin has increased with the emergence of commu-
nity-associated methicillin-resistant Staphylococcus
aureus (CA-MRSA) infections, where it offers an effec-
tive therapeutic alternative to b-lactam antibiotics.1,2
We have chosen to explore this class further, with the
goal of discovering novel antibacterial agents with im-
proved potency, spectrum and ADME/toxicity profile.
In terms of chemical structure clindamycin consists of
two subunits: an amino acid moiety (N-methyl-40-(R)-
propyl-L-proline, or 4-n-propylhygric acid) and an
amino sugar moiety (7-chloro-1-methylthio-lincos-
mediocre activity (MIC vs S. aureus = 25 lg/mL).3 We
amine, 7-Cl-MTL). Increasing the amino acid ring-size
from pyrrolidine (five-membered) to piperidine (six-
membered) has resulted in potent synthetic lincosamide
derivatives (e.g., the veterinary antibiotic pirlimycin3
and the recently disclosed VIC-105555).4 This report de-
scribes our investigation of azetidine (four-membered
ring) lincosamides.
have reasoned that it would be possible to improve the
activity of azetidine lincosamides by incorporating an al-
kyl side-chain on the ring. Molecular modeling of the
conformation of substituted azetidine, pyrrolidine and
piperidine amino acids indicated that the 3-trans-alkyl
azetidine amino acid structure would be a close mimic
for both the clindamycin and pirlimycin amino acid
moieties. Twelve new (30-trans-alkyl)azetidine lincosa-
mides (1) were synthesized with variation at the 30 and
10 positions; 7-chloro-1-methylthio-lincosamine was
used as the sugar moiety in all cases.
The unsubstituted azetidine-2-carboxamide derivative of
7-Cl-MTL has previously been reported to display only
Keywords: Antibacterial; Antibiotic; Protein synthesis inhibitor; Clin-
damycin; Lincosamide; Azetidine; Unnatural amino acid.
q
The synthesis of 3-trans-alkyl-L-azetidine carboxylic
acids 2 is depicted in Scheme 1. N-TBS-4-oxo-azeti-
dine-2S-carboxylic acid was doubly deprotonated with
LDA then alkylated with either an alkyl or allyl electro-
phile with high stereoselectivity.5,6 Conversion of the
This work was presented in part at the 45th Interscience Conference
on Antimicrobial Agents and Chemotherapy, 2005, Washington,
DC, Poster F-2037.
Corresponding author. E-mail: hardwin@hotmail.com
*
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.03.032

2646
H. O’Dowd et al. / Bioorg. Med. Chem. Lett. 18 (2008) 2645–2648
Scheme 1. Reagents: (a) LDA, R00CH2Br, THF; (b) TMSCHN2,
MeOH; (c) H2, Pd/C, EtOAc; (d) Et3NÆ3HF, THF; (e) LiAlH4, THF;
(f) Boc2O, CH2Cl2; (g) RuCl3ÆH2O, NaIO4, aq acetone; (h) 7-Cl-MTL,
HBTU, DIPEA, DMF; (i) TFA, H2O, DCE.
Scheme 2. Reagents: (a) LDA, THF, Me2C@CHCH2Br; (b)
Et3NÆ3HF, THF; (c) LiAlH4, THF; (d) Boc2O, CH2Cl2; (e) TBSCl,
imidazole, DMF; (f) O3, PPh3, CH2Cl2; (g) XYC@PPh3, THF; (h) H2,
Pd/C, EtOAc; (i) KO2CN@NCO2K, AcOH, dioxane; (j) TBAF, THF;
(k) RuCl3H2O, NaIO4, aq acetone; (l) 7-Cl-MTL, HBTU, DIPEA,
DMF; (m) TFA, H2O, DCE.
acid to a methyl ester was followed by hydrogenation (if
required) to give the saturated N-TBS-3-trans-alkyl-4-
oxo-azetidine-2-methyl ester. The TBS-group was re-
moved, and then both the amide and ester were reduced
using LiAlH4. The resulting free amine was Boc-pro-
tected, then the primary alcohol was oxidized to provide
the desired Boc-protected 3-trans-alkyl-L-azetidine car-
boxylic acids 2.6,7 The Boc-protected azetidine acids 2
were coupled with 7-Cl-MTL, then deprotected to fur-
nish the 30-trans-alkyl azetidine lincosamides 1.8
ment of 1a with ethylene oxide (Scheme 4). Each final
compound was purified to >95% via HPLC.
Novel azetidine lincosamides 1a–l were assayed against
a panel of bacterial isolates, using standard broth micro-
dilution techniques,9,10 and the resulting minimum
inhibitory concentration (MIC) data are summarized
in Table 1. Select compounds were also screened for
inhibition of the bacterial protein synthesis pathway in
a cell-free Escherichia coli transcription/translation
(TT) inhibition assay11 (Table 1). Five of the new
azetidine analogs were tested for in vivo efficacy against
S. aureus (Smith strain) in a murine septicemia protec-
tion model,12,13 Table 2.
An alternative approach that allowed for diversification
of the amino acid side-chain at a later stage was also
developed, as shown in Scheme 2. N-TBS-4-oxo-azeti-
dine-2S-carboxylic acid was alkylated with 3,3-dimeth-
ylallyl bromide and elaborated in a similar manner as
before to the Boc-amino alcohol. The alcohol was
TBS-protected, then the side-chain alkene was oxida-
tively cleaved to provide an aldehyde ‘handle’ for fur-
ther modification. Olefination of aldehyde 8 followed
by hydrogenation gave access to a variety of cyclo-
alkyl-alkyl side-chains, in the case of cyclopropyl ana-
logs diimide reduction was used in order to achieve
selective saturation of the alkene. TBS-deprotection
was followed by oxidation, coupling and Boc-deprotec-
tion as before.
N-Methyl azetidine amino acid 2k was synthesized via a
combined one-pot deprotection/Eschweiler–Clark reac-
tion of 2a with formic acid and coupled directly with
7-Cl-MTL to give 1k (Scheme 3). 10-(2-Hydroxy-
ethyl)azetidine lincosamide, 1l, was obtained via treat-
Scheme 3. Reagents: (a) CH2O aq, CHO2H; (b) 7-Cl-MTL, HBTU,
DIPEA, DMF.

H. O’Dowd et al. / Bioorg. Med. Chem. Lett. 18 (2008) 2645–2648
2647
Table 2. In vivo efficacy
CLI
1a
1d
1f
1h
1j
MIC (lg/mL)
ED50, IV (mg/kg)
0.125 0.25 0.06 0.03 0.25 0.125
2.8
1.1
10.8 28.0 NT
1.7
>10 3.0
NT
3.1
NT
ED50, PO (mg/kg)a 19.9
a NT, not tested.
cMLSB strain confirm that azetidine lincosamides share
the same protein synthesis inhibitory mode of action
and bacterial 50S RNA target with clindamycin.14,15
E. coli transcription/translation inhibition data for the
novel compounds were proportional to the observed
MIC in most cases, with linear R0 side-chains affording
superior activity. Increasing the length of the side-chain
Scheme 4. Reagents: (a) ethylene oxide, MeOH.
Results: The 3-trans-substituted-2S-azetidine moiety is
well-tolerated as a mimic of the 4-n-propylhygric acid
portion of clindamycin. Cell-free transcription/transla-
tion assay data and the lack of susceptibility of S. aureus
Table 1. In vitro antibacterial activity and translation/transcription inhibition activity
b
Compound
R
R0
MIC (lg/mL)
TT IC50
(lM)
Streptococcus S. aureus
pneumoniae
S. aureus Enterococcus
(cMLSB)a faecium (n = 2) faecalis (n = 2) (n = 2)
Enterococcus
Bacteroides fragilis
(n = 3)
(n = 1)
CLI
1a
1b
1c
1d
1e
1f
—
H
H
H
H
H
H
H
H
H
H
Me
—
0.06
0.125–0.25 >8
0.125–4
0.125–1
0.5–4
0.25–8
0.125–8
1–>8
0.06–1
0.125–2
0.5–4
2.70
0.36
11.06
12.85
1.13
15.53
2.44
NT
0.06
0.125–0.25 >8
0.125
0.125
0.03
0.5
0.5
>8
>8
0.5–4
0.5–8
0.5–4
0.06–0.125 >8
0.125–0.25
0.5–4
0.25–4
0.5–16
0.06–0.5
0.25–>4
0.25–>4
2–>4
0.125–0.5
0.5–4
0.125
0.016
0.125
0.125
0.125
0.06
0.5
>8
>4
>4
>4
>4
0.03
0.06–0.25
0.25–1
0.25
1g
1h
1i
0.25–0.5
0.25
0.25
0.125–1
0.5–>4
0.125–0.5
1–>4
0.25
7.80
NT
1–2
1–>4
1j
0.125–0.25 >4
0.25–2
2–>4
0.25–2
0.125–2
0.5–8
2.75
NT
1k
1l
0.5
1
>4
>8
0.125
0.5
0.25–4
0.5–>8
NT
a cMLSB: constitutive macrolide, lincosamide and streptogramin B resistance phenotype.
b NT, not tested.

2648
H. O’Dowd et al. / Bioorg. Med. Chem. Lett. 18 (2008) 2645–2648
4. Lewis, J. G.; Gu, S.; Kumar, S. A.; Chen, T.;
O’Dowd, H.; Patel, D. V.; Hackbarth, C. J.; Asano,
R.; Park, C. K.; Blais, J.; Wu, C.; Wang, W.; Yuan,
Z.; Trias, J.; White, R. J.; Gordeev, M. F. Novel
Antimicrobial 7-Methyl Lincosamides: Pipecolamide
Analogs. 44th Interscience Conference on Antimicro-
bial Agents and Chemotherapy. Washington, DC,
Poster F-1389, 2004.
was beneficial. Compound 1a displayed the best overall
activity, performing well both in vitro and in vivo, with
efficacy ca. 2-fold superior to that for clindamycin for
both IV and PO routes of administration. The high
in vitro potency of the cyclobutane-containing com-
pound 1f did not translate into in vivo efficacy, possibly
due to inferior PK. N-Alkylated azetidine lincosamides
1k, 1l showed lower activity than the parent azetidine
lincosamide 1a.
5. Baldwin, J. E.; Adlington, R. M.; Gollins, D. W.;
Schofield, C. J. Tetrahedron 1990, 46, 4733.
6. Kozikowski, A. P.; Liao, Y.; Tuckmantel, W.; Wang, S.;
Pshenichkin, S.; Surin, A.; Thomsen, C.; Wroblewski, J. T.
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Approved Standard-6th ed. NCCLS Document M7-A6,
Vol. 23, No. 2, 2003.
In summary, an SAR study of novel 30-trans-substituted
azetidine lincosamides was conducted. The azetidine is a
well-tolerated replacement for the pyrrolidine of clinda-
mycin. Several examples of azetidine lincosamides with
similar in vitro potency and spectrum to that of clinda-
mycin were identified. The efficacy of compound 1a
compared favorably to that of clindamycin in a murine
septicemia model, via both IV and PO routes of admin-
istration. Azetidine lincosamides are promising antimi-
crobial agents that warrant further study.
10. NCCLS (National Committee for Clinical Laboratory
Standards). Methods for Dilution Antimicrobial Suscep-
tibility Tests for Bacteria that Grow Anaerobically;
Approved Standard-6th ed. NCCLS Document M11-A6,
Vol. 24, No. 2, 2004.
11. Shinabarger, D. L.; Marotti, K. R.; Murray, R. W.; Lin,
A. H.; Melchior, E. P.; Swaney, S. M.; Dunyak, D. S.;
Demyan, W. F.; Buysse, J. M. Antimicrob. Agents
Chemother. 1997, 41, 2132.
Supplementary data
Copies of 1H NMR spectra for compounds 1a–l are
available. Supplementary data associated with this arti-
cle can be found, in the online version, at doi:10.1016/
j.bmcl.2008.03.032.
12. Goldstein, B. P.; Candiani, G.; Arain, T. M.; Romano, G.;
Ciciliato, I.; Berti, M.; Abbondi, M.; Scotti, R.; Mainini,
M.; Ripamonti, F.; Resconi, A.; Denaro, M. Antimicrob.
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