Design, synthesis and antibacterial activity of 3-methylenepyrrolidine formyl hydroxyamino derivatives as novel peptide deformylase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2010.11.102

The synthesis and antibacterial activity of 3-methylenepyrrolidine formyl hydroxyamino derivatives are reported. The antibacterial activities of these derivatives were evaluated to discover SAR at P^1′ and P ^3′ positions, and most of

Full text of DOI:10.1016/j.bmcl.2010.11.102

Bioorganic & Medicinal Chemistry Letters 21 (2011) 1060–1063
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design, synthesis and antibacterial activity of 3-methylenepyrrolidine formyl
hydroxyamino derivatives as novel peptide deformylase inhibitors
Wei Shi a, Haikun Ma a, Yuejiao Duan a, Kelly Aubart b, Yuhong Fang b, Rimma Zonis b, Liping Yang a,
Wenhao Hu a,
⇑
a Institute of Drug Discovery and Development, and Department of Chemistry, East China Normal University, Shanghai 200062, China
b Antibacterial DPU, GlaxoSmithKline, Collegeville, PA 19426, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
The synthesis and antibacterial activity of 3-methylenepyrrolidine formyl hydroxyamino derivatives are
0
0
reported. The antibacterial activities of these derivatives were evaluated to discover SAR at P1 and P3
Received 25 September 2010
Accepted 22 November 2010
Available online 30 November 2010
positions, and most of these derivatives exhibit better in vitro antibacterial activity than existing drugs
against drug-resistant clinical isolates including MRSA, PRSP, and Haemophilus influenzae.
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
PDF inhibitor
Antibiotics
Drug-resistant bacteria
Pyrrolidine derivatives
The emergence of life threatening, multidrug-resistant (MDR)
bacterial infections such as methicillin-resistant Staphylococcus
aureus (MRSA) have been a serious concern both in the hospital
and community settings.1 MRSA infections are the leading cause
of nosocomial infections and cause increased number of deaths
every year.2 The latest antibiotic Linezolid (ZyvoxTM) was approved
by US-FDA in 2000, unfortunately, already a few cases of Linezolid
resistant pathogens in hospital isolates have been reported.3 While
vancomycin is an efficient therapeutic agent for most antibiotic-
resistant Gram-positive bacteria, vancomycin-resistant S. aureus
(VRSA) was reported in the U.S. in 2002.4 This crisis has resulted
in an intensive research effort to develop a new class of compounds
that exhibit novel mechanisms of antibacterial activity. Antibacte-
rial drugs with a new mode of action are expected to have no pre-
existing resistance.
H
O
HO
N
F
O
O
H
N
N
HO
N
HO
N
N
O
H
O
N
O
O
H
Actinonin
LBM415
OH
O
O
H
O
H
N
N
N
O
O
N
Peptide deformylase (PDF) catalyzes the removal of the
N-formyl group from the N-terminal methionine during bacterial
protein maturation, and mammalian cytosolic protein synthesis
does not produce N-formylated polypeptides,5 making PDF an
attractive target for developing antibiotics with novel mechanisms
of action. Actinonin (Fig. 1), a naturally occurring antibiotic iso-
lated in 1962 from an actinomycete,6 was the first PDF inhibitor re-
ported by the researchers from Vicuron. It showed moderate
antibacterial activity against several Gram-positive and Gram-neg-
ative bacteria,7 however, actinonin did not show good in vivo anti-
bacterial activity due to poor pharmacokinetic properties, either
BB83698
Figure 1. Structures of actinonin, LBM415 and BB83698.
poor absorption or quick clearance.8 Many pharmaceutical compa-
nies and academic institutions have focused on the development of
novel PDF inhibitors.9 Thus far, two PDF inhibitors underwent hu-
man clinical trials, BB83698 (Fig. 1, discovered by British Biotech,
in collaboration with Genesoft) and LBM415 (Fig. 1, discovered
by Vicuron pharmaceuticals, in collaboration with Novartis), but
there are no currently marketed PDF inhibitors. We have reported
the synthesis of a series of PDF inhibitors with a 2,5-dihydropyrrole
motif and evaluated their antibacterial activity.10 The motif of
⇑
Corresponding author.
E-mail address: huwenhao@yahoo.com (W. Hu).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.11.102

W. Shi et al. / Bioorg. Med. Chem. Lett. 21 (2011) 1060–1063
1061
the results are summarized in Table 1. Moderate antibacterial
H
O
R1
activity (MIC 8–16 lg/ml) was observed for compound 1a that
0
bears an aliphatic amino morpholine group at P3 position. This is
interesting because no antibacterial activity was observed from
corresponding 2,5-dihydropyrrole derivatives in our previous
N
N
HO
R2
O
N
O
R3
1a-1n
30
study.10 Compounds 1b–1d bearing aromatic amines at P position
typically gave moderate to good antibacterial activity (MIC 0.5–
Figure 2. Generic structure of 1a–1n.
8
4
l
g/ml). Good to excellent antibacterial activities (MIC 0.125–
lg/ml) were observed when heterocyclic aromatic amines were
0
introduced at the P3 position in compounds 1e–1g. Preliminary
3-methylenepyrrolidine is found in many biologically active prod-
ucts.11 In continuation of our efforts to find new PDF inhibitors
through structural modification of LBM415, we continued research
0
SAR study at P1 position was also investigated. Because cyclopen-
0
tylmethyl group at P1 position in BB83698 showed excellent anti-
bacterial activity, we decided to screen antibacterial activity of
on the exploring the structure–activity relationship (SAR) by
0
replacing the pyrrolidine functionality at the P2 position with
compounds 1h–1n that bear a cyclopentyl methyl group at the
0
P1 position. We were gratified to find that compounds 1h–1n
3-methylenepyrrolidine.
A number of 3-methylenepyrrolidine
exhibited better antibacterial activity than corresponding analogs
formyl hydroxyamino derivatives 1a–1n (Fig. 2) were synthesized
as novel PDF inhibitors, and their in vitro antibacterial activities
were evaluated.
1a–1g that bear an n-butyl group (1a vs 1h, 1b vs 1j, 1c vs 1k, 1f
0
vs 1m and 1g vs 1n). Similar SAR at P3 position was observed for
1h–1n. Among the compounds screened, 1n gave the best antibac-
terial activity (0.0625–0.5 lg/ml) against S. aureus, MSSA, MRSA
and S. epidermidis.
As PDF inhibitors were originally pursued with the belief to
generate a respiratory drug, we tested the antibacterial activity
of 1g, 1i, 1k–1n against MRSA, MRSE, Streptococcus pneumoniae,
PRSP, Haemophilus influenzae clinical isolates. The screening results
are summarized in Table 2. While moderate antibacterial activity
A general synthesis of 3-methylenepyrrolidine formyl hydrox-
yamino derivatives 1a–1n is illustrated in Scheme 1, and the
synthetic route is similar to that we reported previously.10 In the
synthesis shown in Scheme 1, we started from the methyl ester
of N-Boc-trans-4-hydroxy-L-proline (2). A Swern oxidation of 2
afforded corresponding ketone 3 in good yield according to a liter-
ature procedure.12 Treatment of 3 with a Wittig reagent (prepared
by methyl triphenyl phosphonium bromide and potassium tert-
butoxide) gave olefin 4,13 followed by removal of the Boc group
to afford building block 5 in a quantitative yield. The coupling
reaction between 5 and 6 (R1 is n-butyl or cyclopentyl methyl)
was carried out using 1-hydroxy-benzotriazole monohydrate
(1–32
lg/ml) against Gram-negative strain H. influenzae was ob-
served, excellent antibacterial activity (0.0625–4
l
g/ml) against
the other Gram-positive tested strains was observed. Here, com-
pound 1m showed better antibacterial activity than LBM415 and
other existing drugs such as Penicillin, Ciprofloxacin, Linezolid
and Vancomycin.
At last, two representative compounds 1e and 1g, along with
the control compound amoxicillin, were screened against a panel
of 14 isolates representing a broad-spectrum of activity. The re-
sults are summarized in Table 3. Compound 1g shows good activity
against all isolates tested except for E. coli.
(HOBt)
hydrochloride (EDCI) as coupling reagents in the presence of
N-methylmorpholine (NMM) to produce coupling product
and
1-(3-dimethylaminopropyl)-3-ethylcarbodiimide
7
smoothly, which gave corresponding acid 8 after hydrolysis. Penul-
timate compounds 9a–9n were obtained by coupling of 8 with
various amines R2R3NH. Finally, deprotection of the PMB group
with trifluoroacetic acid successfully provided the desired 3-meth-
ylenepyrrolidine derivatives 1a–1n. All the compounds were
characterized by 1H NMR, 13C NMR and HR-MS (ESI), and HPLC
purity of the compounds ranges from 90% to 97% AP.
In summary, we have synthesized a number of 3-methylene-
pyrrolidine formyl hydroxyamino derivatives 1a–1n, and screened
their antibacterial activity against a wide range of drug-resistant
0
bacteria including some clinical isolates. Preliminary SAR on P1
0
The new compounds 1a–1n were screened against Gram-
positive bacterial strains such as S. aureus and S. epidermidis, and
and P3 positions of the compounds was studied. Compounds 1g,
O
HO
O
O
O
O
a
c
b
N
N
N
H
N
O
O
O
O
Boc
Boc
Boc
2
5
3
4
+
H
O
H
O
H
O
R1
R1
R1
e
d
N
N
N
N
N
N
OH
PMBO
PMBO
PMBO
O
O
O
OH
O
8
O
6
7
O
O
f
H
O
H
O
R1
R1
g
N
N
N
PMBO
HO
R2
R2
O
N
R3
N
R3
O
O
1a-1n
9a-9n
Scheme 1. Reagents and conditions: (a) (i) DMSO, oxalyl chloride, CH2Cl2, ꢀ78 °C, 10 min; (ii) Et3N, ꢀ78 °C, 80%; (b) (i) Ph3P+CH3ꢁBrꢀ, t-BuOK, THF, 0 °C–25 °C, 1 h; (ii) 3, 1 h,
60%; (c) TFA/CH2Cl2, Et3N, 30 min, quantitative yield; (d) HOBT, EDCI, NMM, CH2Cl2, 25 °C, 16 h, 84%; (e) LiOH, dioxane/H2O, 25 °C, 5 h, 90%; (f) Et3N, ClCO2Et, R1R2NH, THF,
0 °C–25 °C, 16 h, 60–81%; (g) TFA/CH2Cl2, column chromatography purification, 35–45%.

1062
W. Shi et al. / Bioorg. Med. Chem. Lett. 21 (2011) 1060–1063
Table 1
In vitro minimum inhibitory concentration (MIC,
l
g/ml) values of 1a–1r against bacterial strainsa
Compound
R1
–NR2R3
S. aureus
MSSA
MRSA
S. epidermidis
ATCC25923
ATCC29213
ATCC43300
ATCC12228
1a
8
2
1
>64
8
8
16
2
N
O
HN
HN
1b
1c
1
4
0.5
1
F
1d
1e
HN
HN
OH
F
8
1
32
2
4
8
0.125
0.5
N
F
HN
O
1f
2
4
1
1
2
N
N
1g
0.5
0.125
0.125
HN
S
1h
1i
4
8
8
1
2
4
N
O
H
N
4
1
4
1j
0.5
0.25
0.5
HN
HN
F
1k
1
2
0.5
1
HN
F
F
1l
0.5
1
2
0.0625
0.125
0.125
0.5
N
HN
O
1m
0.125
N
N
1n
0.0625
0.5
0.0625
0.125
HN
S
LBM415
Penicillin
Ciprofloxacin
Linezolid
Vancomycin
0.5
0.0625
0.5
2
1
2
1
0.5
2
1
0.5
64
1
4
1
1
2
0.25
0.5
2
a
MIC were determined by broth micro dilution technique. MSSA, methicillin-susceptible Staphylococcus aureus; MRSA, methicillin-resistant Staphylococcus aureus.
Table 2
In vitro minimum inhibitory concentration (MIC,
l
g/ml) values of 1g, 1i, 1k–1n against various clinical isolatesa
Compound
MSSA 1 strain
MRSA 3 strains
MSSE 1 strain
MRSE 3 strains
PSSP 1 strain
PRSP 3 strains
H. F 4 strains
1g
1i
1k
1l
1m
1n
LBM415
Penicillin
Ciprofloxacin
Linezolid
Vancomycin
1
2
1
0.5
0.5
1
0.5
0.5
1
0.5–1
0.5–2
1
1
2
4
0.25
1
0.5–32
1–2
0.25–4
0.25–0.5
0.0625–0.25
0.25
0.5–1
64–>64
0.5–8
4–32
4–32
4–8
2–8
2–8
1–8
2–8
16–>64
0.125–4
ND
0.25–4
0.25–1
0.125–2
0.0625–1
0.125–1
64–>64
1, 32–64
0.5–2
0.5
0.5
0.125
1
0.5
4
0.25
1
1
0.25–2
0.5–4
0.25–0.5
0.5–1
0.125–1
64–>64
1, 64–>64
1
0.25
0.25
0.25
0.25
0.125
2
2
0.5
1
1
1
1, 8
1, 16
1–2
1–4
ND
a
MIC were determined by agar dilution method. MSSE, methicillin-susceptible Staphylococcus epidermidis; MRSE, methicillin-resistant Staphylococcus epidermidis; PSSP,
penicillin-susceptible Streptococcus pneumoniae; PRSP, penicillin-resistant Streptococcus pneumoniae; H. F, Haemophilus influenzae.

W. Shi et al. / Bioorg. Med. Chem. Lett. 21 (2011) 1060–1063
1063
Table 3
637; (g) Huigens, R. W., III; Reyes, S.; Reed, C. S.; Bunders, C.; Rogers, S. A.;
Steinhauer, A. T.; Melander, C. Bioorg. Med. Chem. 2010, 18, 663.
2. (a) Appelbaum, P. C. Clin. Microbiol. Infect. 2006, 12, 16; (b) Yarwood, J. M.;
Bartels, D. J.; Volper, E. M.; Greenberg, E. P. J. Bacteriol. 2004, 186, 1838.
3. (a) Wilson, P.; Andrews, J. A.; Charlesworth, R.; Walseby, R.; Singer, M.; Farrell,
D. J.; Robbins, M. J. Antimicrob. Chemother. 2003, 51, 186; (b) Prystowsky, J.;
Siddiqui, F.; Chosay, J.; Shinabarger, D. L.; Millichap, J.; Peterson, L. R.; Noskin,
G. A. Antimicrob. Agents Chemother. 2001, 45, 2154; (c) Gonzales, R. D.;
Schreckenberger, P. C.; Graham, M. B.; Kelkar, S.; DenBesten, K.; Quinn, J. P.
Lancet 2001, 357, 1179; (d) Tsiodras, S.; Gold, H. S.; Sakoulas, G.; Eliopoulos, G.
M.; Wennersten, C.; Venkataraman, L.; Mocllering, R. C., Jr.; Ferraro, M. J. Lancet
2001, 358, 207; (e) Lohray, B. B.; Lohray, V. B.; Srivastava, B. K.; Gupta, S.;
Solanki, M.; Pandya, P.; Kapadnis, P. Bioorg. Med. Chem. Lett. 2006, 16, 1557.
4. Centers for Disease Control and Prevention. Morb. Mortal. Wkly. Rep. 2004, 53,
322.
In vitro minimum inhibitory concentration (MIC,
lg/ml) values of 1e, 1g against 14
bacterial strainsa
1e
1g
Amoxicillin
Staphylococcus aureus OXFORD
Staphylococcus aureus WCUH29
Enterococcus faecalis I
Enterococcus faecium X7501
Haemophilus influenzae Q1
Haemophilus influenzae H128
Haemophilus influenzae H128 Acr A-
Moraxella catarrhalis 1502
Streptococcus pneumoniae 1629
Streptococcus pneumoniae N1387
Streptococcus pneumoniae ERY2
Escherichia coli 3
4
0.5
0.125
0.125
64
0.5
32
4
16
16
0.5
4
0.5
2
16
4
4
0.25
32
60.06
64
0.25
16
4
60.06
60.06
60.06
2
2
1
5. (a) Adams, J. M. J. Mol. Biol. 1968, 33, 571; (b) Nguyen, K. T.; Hu, X.; Colton, C.;
Chakrabarti, R.; Zhu, M. X.; Pei, D. Biochemistry 2003, 42, 9952.
6. Gordon, J. J.; Kelly, B. K.; Miller, G. A. Nature 1962, 195, 701.
7. Chen, D. Z.; Patel, D. V.; Hackbarth, C. J.; Wang, W.; Yuan, Z., et al Biochemistry
2000, 39, 1256.
8
2
64
2
60.06
1
60.06
60.06
>64
4
Streptococcus pyogenes 1307006P
Streptococcus pyogenes 1308007P
4
2
8. Jain, R.; Chen, D.; White, R. J.; Patel, D. V.; Yuan, Z. Curr. Med. Chem. 2005, 12,
1607.
a
MIC endpoints were determined by broth microdilution according to CLSI
9. (a) Jayasekera, M. M. K.; Kendall, A.; Shammas, R.; Dermyer, M.; Tomala, M.;
Shapiro, M. A.; Holler, T. P. Arch. Biochem. Biophys. 2000, 381, 313; (b) Hu, X.;
Nguyen, K. T.; Jiang, V. C.; Lofland, D.; Moser, H. E.; Pei, D. J. Med. Chem. 2004,
47, 4941; (c) Davies, S. J.; Ayscough, A. P.; Beckett, R. P.; Clements, J. M.; Doel, S.;
Pratt, L. M.; Spavold, Z. M.; Thomas, S. W.; Whittaker, M. Bioorg. Med. Chem. Lett.
2003, 13, 2715; (d) Molteni, V.; He, X.; Nabakka, J.; Yang, K.; Kreusch, A.;
Gordon, P.; Bursulaya, B.; Warner, I.; Shin, T.; Biorac, T.; Ryder, N. S.; Goldberg,
R.; Doughty, J.; He, Y. Bioorg. Med. Chem. Lett. 2004, 14, 1447; (e) Aubart, K. M.;
Benowitz, A. B.; Christensen, S. B.; Lee, J.; Silva, D. J. PCT WO Patent
2005017124, 2007.
guidelines
1m and 1n showed very promising antibacterial activity, and their
in vivo studies are currently in progress.
Acknowledgement
10. Shi, W.; Duan, Y. J.; Qian, Y.; Li, M.; Yang, L. P.; Hu, W. H. Bioorg. Med. Chem. Lett.
2010, 20, 3592.
This work was supported by the Science and Technology Com-
mission of Shanghai Municipality (07DZ19503-2, 10XD1401700),
NSFC (20932003), and MOST of China (2009ZX09501-017).
11. (a) Pendri, Y. R.; Chen, C.-P. H.; Patel, S. S.; Evans, J. M.; Liang, J.; Kronenthal, D.
R.; Powers, G. L.; Prasad, S. J., et al. WO Patent 2004052310, 2004; (b) Alegria, L.
A.; Dress, K. R.; Huang, B.; Kumpf, R. A.; Lewis, K. K.; Matthews, J. J.; Sakata, S.
K.; Wallace, M. B. WO Patent 2005026114, 2005; (c) Lange, U. E. W.; Baucke, D.;
Hornberger, W.; Mack, H.; Seitz, W.; Hoeffken, H. W. Bioorg. Med. Chem. Lett.
2006, 16, 2648; (d) Gauzy, L.; Zhao, R.; Deng, Y.H.; Li, W.; Bouchard, H.; Chari, R.
V. J.; Commercon, A. WO Patent 2007085930, 2007; (e) Cottrell, K. M.;
Maxwell, J.; Tang, Q.; Grillot, A.-L.; Le Tiran, A.; Perola, E. WO Patent
2008106139, 2008.
References and notes
1. (a) Wright, G. D. Nat. Rev. Microbiol. 2007, 5, 175; (b) Demain, A. L.; Sanchez, S. J.
Antibiot. 2009, 62, 5; (c) Gould, I. M. Int. J. Antimicrob. Agents 2008, 32S, 4; (d)
Lankeshwar, N.; Bagde, U. S. Asian J. Microbiol. Biotechnol. Environ. Sci. 2008, 10,
829; (e) Mahady, G. B.; Huang, Y.; Doyle, B. J.; Locklear, T. Stud. Nat. Prod. Chem.
2008, 35, 423; (f) Arias, C. A.; Murray, B. E. Expert Rev. Anti-Infect Ther. 2008, 6,
12. Qiu, X. L.; Qing, F. L. Synthesis 2004, 3, 334.
13. Dixon, S. M.; Milinkevich, K. A.; Fujii, J.; Ruiwu, L.; Nianhuan, Y.; Lam, K. S.;
Kurth, M. J. J. Comb. Chem. 2007, 9, 143.