Synthesis and antimicrobial activity of novel globomycin analogues

Article abstract of DOI:10.1016/S0960-894X(03)00432-3

Globomycin, a signal peptidase II inhibitor, and its derivatives show potent antibacterial activity against Gram-negative bacteria. The synthesis and antimicrobial activity of novel globomycin analogues are reported. One of the analogues showed a more pot

Full text of DOI:10.1016/S0960-894X(03)00432-3

Bioorganic & Medicinal Chemistry Letters 13 (2003) 2315–2318
Synthesis and Antimicrobial Activity of Novel
Globomycin Analogues
Toshihiro Kiho,a Mizuka Nakayama,a Kayo Yasuda,b Shunichi Miyakoshi,b
Masatoshi Inukaib and Hiroshi Kogena,*
aExploratory Chemistry Research Laboratories, Sankyo Co., Ltd., Tokyo 140–8710, Japan
bLead Discovery Research Laboratories, Sankyo Co., Ltd., Tokyo 140–8710, Japan
Received 11 March 2003; revised 22 April 2003; accepted 22 April 2003
Abstract—Globomycin, a signal peptidase II inhibitor, and its derivatives show potent antibacterial activity against Gram-negative
bacteria. The synthesis and antimicrobial activity of novel globomycin analogues are reported. One of the analogues showed a more
potent activity against Gram-negative bacteria than globomycin and also exhibited antibacterial activity against methicillin-resis-
tant Staphylococcus aureus (MRSA).
# 2003 Elsevier Science Ltd. All rights reserved.
Globomycin (1a)1 and its congeners, SF-1902A –A5,2
longer alkyl side chain in the fatty acid unit than that of
1a. It was reported that the antibacterial activity is quite
sensitive to the length of the alkyl side chain, either in a
fatty acid or in an amino acid.2 The congeners which
have a longer side chain, 1b, SF-1902A and A4b, are
4a
more potent than 1a (MIC: 1a, 6.25 mg/mL; 1b, 1.56
mg/mL against E. coli NIHJ JC-2). However, SF-1902
2
isolated by different researchers as antibiotics against
only Gram-negative bacteria are 19-membered cyclic
depsipeptides.1,2 The major component, 1a, has only
been proven to be a specific inhibitor of signal peptidase
II, a prolipoprotein-processing enzyme,3 that processes
the acylated precursor form of lipoprotein into
apolipoprotein and a signal peptide in Escherichia coli.4
Inhibition of signal peptidase II leads to the accumula-
tion of the lipoprotein precursor in the cytoplasmic
membrane and consequently to the death of the cell.5
Signal peptidase II represents an attractive target
because the mechanism is different from currently
available drugs. Previously, we reported the absolute
structure of 1a obtained by X-ray analysis and the first
asymmetric total synthesis of 1a and SF-1902A5 (1b).1d,e
Now, we wish to report the structure–activity relation-
ships (SARs) of synthetic new globomycin analogues
that have potent inhibitory activity against Gram-nega-
tive bacteria. Structurally, these congeners were con-
structed from four natural amino acids, one N-methyl
amino acid and a b-hydroxy-a-methyl carboxylic acid.
Naturally occurring globomycin congeners are as shown
in Figure 1. The minor congeners, SF-1902A 3 and A4b,
have an l-Val in place of l-allo-Ile, and the other con-
geners, SF-1902A 2, A4a and A4b, have a shorter or
*Corresponding author. Tel.: +81-3-3492-3131; fax: +81-3-5436-
8570; e-mail: hkogen@shina.sankyo.co.jp
Figure 1. Structure of naturally occurring globomycin (1a) and its
congeners.
0960-894X/03/$ - see front matter # 2003 Elsevier Science Ltd. All rights reserved.
doi:10.1016/S0960-894X(03)00432-3

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T. Kiho et al. / Bioorg. Med. Chem. Lett. 13 (2003) 2315–2318
A2 with the shortest side chain showed the weakest
activity.2b
formed with DIPC8 and Fragment B under Keck’s
condition11 to afford a fully protected seco-acid 3.
Sequential deprotections of TBS, allyl and Boc group
in 3 provided macrocyclization precursors. The mac-
rolactamization was performed by HATU8 or TBTU8
to give O-Bn derivatives under highly diluted condi-
tions. Finally, the removal of the benzyl group by
hydrogenolysis yielded novel globomycin analogues
(1c–1i). N-Demethyl derivative, 1i, only exists as a sin-
gle isomer in CD3OD, which is different from other
analogues.1d,e,13
Furthermore, the congeners containing an l-Val, SF-
1902A and A4b, are less active compared with 1a and
3
1b, respectively. In this paper, we focused on the length
of the alkyl side chain, the stereochemistry of allo-type
amino acids, the hydroxyl group in l-allo-Thr and the
N-methyl group.
Synthesis
The novel synthetic analogues (1c–1i) were prepared by
the convergent macrolactamization method using three
fragments6 (Fragment A, B and C) as shown in Scheme
1. Fragment A was prepared by an anti-selective boron
aldol reaction followed by hydrolysis.7 Fragments B and
C were synthesized from commercially available pro-
tected amino acids with PyBOP.8 Fragment C was con-
densed with Fragment A mediated by DEPC8 to give the
acylated tripeptide 2. The esterification of 2 was per-
Antibacterial Activity
Antibacterial activities of the synthetic globomycin
analogues (1a–1i) against Gram-negative bacteria are
summarized in Table 1. As a result, 1c14 possessing the
longest alkyl side chain shows the most potent activity
among the analogues (MIC: 1a, 12.5 mg/mL; 1b, 3.13
mg/mL; 1c, 1.56 mg/mL against E. coli SANK 70569).
The length of the alkyl side chain greatly affects the
Scheme 1.a Synthesis of globomycin (1a) and its analogues.

T. Kiho et al. / Bioorg. Med. Chem. Lett. 13 (2003) 2315–2318
2317
Table 1. Antibacterial spectrum of globomycin (1a), SF-1902A 5 (1b) and the synthetic analogues (1c–1i)
Organisms
MIC (mg/mL)
1a
1b
3.13
3.13
6.25
3.13
1.56
1c
1.56
1.56
3.13
1.56
1.56
3.13
3.13
1d
1e
1f
1g
25
25
100
50
1h
25
25
50
50
12.5
100
>100
>100
>100
>100
>100
>100
1i
Escherichia coli SANK 70569 (NIHJ JC-2)
Escherichia coli SANK 72290
Salmonella enteritidis SANK 72390
Klebsiella pneumoniae SANK 72490
Enterobacter cloacae 846
Enterobacter cloacae SANK 72690
Serratia marcescens SANK 72790
Proteus vulgaris SANK 72890
Morganella morganii SANK 72990
Pseudomonas aeruginosa SANK 73090
Pseudomonas aeruginosa SANK 73190
Pseudomonas aeruginosa SANK 3719
12.5
12.5
25
25
6.25
12.5
12.5
50
25
6.25
>100
>100
>100
>100
100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
100
>100
>100
>100
>100
>100
>100
>100
12.5
50
100
12.5
12.5
>100
>100
>100
>100
>100
50
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
antibacterial activity. Four-carbon increase in the fatty
acid side chain enhanced the activity by 4- to 8-fold
compared with 1a. Therefore, it may be possible to
produce a more potent inhibitor.
only Gram-negative bacteria but also Gram-positive
bacteria. Now, further investigations on SARs are cur-
rently underway.
With regard to stereoisomers, the activity of 1d dimin-
ished and the activity of 1e and 1f were completely lost.
In particular, the stereochemistry of the hydroxyl group
in l-Thr is quite important for the activity. Compound
1e was inactive although the deoxy derivative 1g and
methyl ether derivative 1h retained their activity.
Therefore, the hydroxyl group in l-allo-Thr is not
essential for the activity.15 Finally, N-demethyl deriva-
tive 1i also lost its activity.
Measurement of Antibacterial Activity
Bacteria were inoculated on Nutrient Agar (Eiken
Chemical Co., Ltd.) and the MIC was determined by
the agar dilution method.16
References and Notes
1. (a) Taxonomy of producing organism and fermentation:
Inukai, M.; Enokita, R.; Torikata, A.; Nakahara, M.; Iwado,
S.; Arai, M. J. Antibiot. 1978, 31, 410. (b) Isolation and phys-
ico-chemical and biological characterization: Inukai, M.;
Nakajima, M.; Osawa, M.; Haneishi, T.; Arai, M. J. Antibiot.
1978, 31, 421. (c) Structural determination of 1a: Nakajima,
M.; Inukai, M.; Haneishi, T.; Terahara, A.; Arai, M. J. Anti-
biot. 1978, 31, 426. (d) Kogen, H.; Kiho, T.; Nakayama, M.;
Furukawa, Y.; Kinoshita, T.; Inukai, M. J. Am. Chem. Soc.
2000, 122, 10214. (e) Kiho, T.; Nakayama, M.; Kogen, H.
Tetrahedron 2003, 59, 1685.
Surprisingly, 1c showed moderate activity against all
Gram-positive bacteria tested such as Staphylococcus
aureus (MRSA) (MIC=12.5 mg/mL) even though 1a
and 1b were almost inactive as shown in Table 2. This is
the first example that the antibacterial spectrum of glo-
bomycin analogues was expanded to also include Gram-
positive bacteria. These results suggest that lipoproteins
are essential for not only Gram-negative bacteria but
also Gram-positive bacteria and signal peptidase II
inhibitors would probably be effective against most
bacteria. Finding such an inhibitor would lead to
development of a new class of antibiotics. Finally, the
antifungal activity of these analogues was tested. How-
ever, no activity was observed against Candida albicans,
Candida glabrata and Aspergillus clavatus.
2. (a) Omoto, S.; Suzuki, H.; Inouye, S. J. Antibiot. 1979, 32,
83. (b) Omoto, S.; Ogino, H.; Inouye, S. J. Antibiot. 1981, 34,
1416.
3. Pugsley, A. P. Microbiol. Rev. 1993, 57, 50.
4. (a) Inukai, M.; Takeuchi, M.; Shimizu, K.; Arai, M. J.
Antibiot. 1978, 31, 1203. (b) Hussain, M.; Ichihara, S.;
Mizushima, S. J. Biol. Chem. 1980, 255, 3707. (c) Dev, I. K.;
Harvey, R. J.; Ray, P. H. J. Biol. Chem. 1985, 260, 5891. (d)
Ichihara, S.; Hussain, M.; Mizushima, S. J. Biol. Chem. 1981,
256, 3125. (e) Witke, C.; Gotz, F. FEMS Microl. Lett. 1995,
126, 233. (f) Braun, B.; Wu, H. C. In Bacterial Cell Wall;
Ghuysen, J. M., Hakenbeck, R., Eds.; Elsevier: Amsterdam,
1994; Chapter 14.
In summary, we disclosed the SAR of synthetic new
globomycin analogues and succeeded in producing a
promising antibiotic, which shows activity against not
Table 2. Antibacterial spectrum of globomycin (1a–1c) against
Gram-positive bacteria
5. Cavard, D. Arch. Microbiol. 1998, 171, 50.
6. Synthesis of three fragments, Fragment A, B and C, were
performed by the method described in refs 1d and 1e.
7. Abiko, A.; Liu, J.-F.; Masamune, S. J. Am. Chem. Soc.
1997, 119, 2586.
Organisms
MIC (mg/mL)
1a
1b
1c
8. PyBOP:9 (benzotriazolyloxy) tris(pyrrolydino)phosphonium
hexafluorophosphate, DEPC:10 diethylcyanophosphate, DIPC;
diisopropylcarbodiimide, HATU:12 O-(7-azabenzotriazole-1-
yl)-1,1,3,3-tetramethyluronium hexafluorophosphate, TBTU:12
2-(1H-Benzotriazole-1-yl)-1,1,3,3-tetramethyluronium tetra-
fluoroborate.
Staphylococcus aureus SANK 70668
Staphylococcus aureus SANK 71790
Staphylococcus aureus SANK 71890a
Enterococcus faecalis SANK 71990
>100
>100
>100
>100
50
50
50
100
6.25
6.25
12.5
12.5
aMRSA.

2318
T. Kiho et al. / Bioorg. Med. Chem. Lett. 13 (2003) 2315–2318
9. Coste, J.; Le-Nguyen, D.; Castro, B. Tetrahedron Lett.
1990, 31, 205.
10. Shioiri, T.; Yokoyama, Y.; Kasai, Y.; Yamada, S. Tetra-
hedron 1976, 32, 2211.
4.90* (d, 1/6H, J=9.9 Hz), 5.06–5.10 (m, 5/6H), 6.92* (d, 1/
6H, J=9.1 Hz), 7.11 (d, 5/6H, J=7.4 Hz), 7.37* (br s, 1/6H),
7.41* (d, 1/6H, J=8.1 Hz), 7.53* (br t, 1/6H, J=3.1 Hz), 7.62
(d, 5/6H, J=4.4 Hz), 7.68 (br m, 10/6H); 13C NMR
(125 MHz, CDCl3, 33 mM, both rotamers) d ppm: 11.6, 12.3*,
14.1, 14.6, 14.8*, 15.0, 18.9, 19.1*, 21.9, 22.66, 22.72*, 23.0*,
24.3, 24.8*, 25.2, 25.9*, 26.9*, 27.1, 29.3, 29.42, 29.49, 29.56,
29.59, 29.7*, 31.3, 31.9, 36.6, 37.3*, 38.1, 38.4*, 39.2*, 40.1, 40.5,
41.1, 56.2*, 56.6, 57.7, 57.8*, 59.1, 59.2*, 60.7*, 61.5, 66.9, 67.2*,
68.0, 76.4, 77.2, 77.9*, 168.8, 170.3, 170.7, 170.9*, 171.0*, 173.3,
173.4*, 174.6*, 174.7, 177.0; IR (KBr) cmÀ1: 3326, 2959, 2927,
2856, 1758, 1656, 1545, 1466, 1377, 1196; HRMS m/z (M+H)+
calcd 712.4861, found 712.4849. Anal. calcd for
11. Boden, E. P.; Keck, G. E. J. Org. Chem. 1985, 50, 2394.
12. (a) Carpino, L. A.; El-Faham, A. J. Org. Chem. 1994, 59,
695. (b) Carpino, L. A. J. Am. Chem. Soc. 1993, 115, 4397. (c)
Carpino, L. A.; El-Faham, A. J. Org. Chem. 1995, 60, 3561.
(d) Humphrey, J. M.; Chamberlin, A. R. Chem. Rev. 1997, 97,
2243.
13. 1H NMR suggested that other analogues (1a–1h) exist as a
mixture of rotational isomers in solution.
14. The spectrum data for compound 1c. The minor con-
former is marked with an asterisk. 1H NMR [500 MHz,
CDCl3, 16 mM, two rotamers (major/minor=5.3/1)] d ppm:
0.85–0.97 (m, 15H), 1.10 (d, 15/6H, J=6.8 Hz), 1.15* (d, 3/
6H, J=7.1 Hz), 1.13–1.42 (m, 21H), 1.48–1.57 (m, 1H), 1.62–
1.71 (m, 3H), 1.76–1.88 (br, 2H), 2.02–2.10* (m, 1/6H), 2.10–
2.16 (m, 1H), 2.18–2.23 (m, 5/6H), 2.78* (s, 3/6H), 3.11–3.18
(m, 1H), 3.21 (s, 15/6H), 3.63 (br s, 5/6H), 3.75 (dd, 5/6H,
J=4.1, 17.2Hz), 3.85* (dd, 1/6H, J=4.0, 18.0 Hz), 3.94 (s, 10/
6H), 3.98* (s, 2/6H), 4.01–4.17 (m, 2H), 4.20–4.40 (m, 2H),
4.53 (dd, 1H, J=4.3, 7.4 Hz), 4.77* (dd, 1/6H, J=4.5, 9.4 Hz),
.
C36H65N5O9 H2O: C, 59.24; H, 9.25; N, 9.59. Found: C, 58.97;
H, 8.93; N, 9.48; [a]2D6=+19.0 (c 0.50, CH3OH).
15. Diacetylation of 1a in l-Ser and l-allo-Thr residue dimin-
ished the activity.2b O-Bn or O-Me globomycin derivatives in
l-Ser residue were inactive against E. coli SANK 70569 tested
only (MIC>50 mg/mL).
16. National Committee for Clinical Laboratory Standards.
Standard Methods for Dilution Antimicrobial Susceptibility
Tests for Bacteria that Grow Aerobically. Approved Standard
(M7-A5); NCCLS: Villanova, 2000; p 7.