Hybrid glycopeptide antibiotics

Full text of DOI:10.1021/ja0166693

12722
J. Am. Chem. Soc. 2001, 123, 12722-12723
Table 1. MICs of Vancomycin Derivativesa
Hybrid Glycopeptide Antibiotics
Binyuan Sun, Zhong Chen, Ulrike S. Eggert, Simon J. Shaw,
John V. LaTour, and Daniel Kahne*
Department of Chemistry, Princeton UniVersity
Princeton, New Jersey 08544
ReceiVed July 20, 2001
The emergence of resistance to vancomycin (1a, Table 1) in
enterococcal strains has aroused considerable concern.1 Efforts
to overcome resistance have led to a new class of vancomycin
derivatives containing hydrophobic substituents on the van-
cosamine sugar (e.g., 1b, Table 1).2 These glycolipid derivatives
are more active than vancomycin against both sensitive and
resistant enterococcal strains (1b vs 1a, Table 1). Based on several
lines of evidence, we have previously proposed that these
glycolipid derivatives of vancomycin are bifunctional molecules,
consisting of two biologically active components that interact with
different cellular targets.3,4 The aglycone binds to the D-Ala-D-
Ala dipeptide terminus of peptidoglycan precursors; the func-
tionalized disaccharide interacts with proteins involved in the
transglycosylation step of peptidoglycan synthesis. We have
suggested that this latter mechanism explains how the compounds
overcome resistance.5 If this bifunctional model is correct, then
it should be possible to improve the activity of vancomycin
derivatives by optimizing the glycolipid moiety for inhibition of
transglycosylation.
E. faecium
resistantc
E. faecalis
resistante
(VanB)
compd sensitiveb
(VanA)
sensitived
S. aureusf
1a
1b
2
2
2048
12.5
63
16
2048
12.5
32
4
<0.025
<0.01
<0.1
0.1
0.05
0.25
<0.025
0.2
2
3
125
250
a MIC values (µg/mL) were obtained by using a standard microdi-
lution assay. The MIC is defined as the lowest antibiotics concentration
that resulted in no visible growth after incubation at 35 °C for 22 h.
b Bacterial strain 49624. c Bacterial strain CL4931. d Bacterial strain
29212. e Bacterial strain CL4877. f Bacterial strain 29213.
Scheme 1a
Substituent changes to the disaccharide of vancomycin have
been extensively explored, but there have been only limited efforts
to change the sugars attached to the vancomycin aglycone.
Glycosylation of the vancomycin aglycone is not a trival
operation.6 It would be useful to have an efficient, general strategy
to attach a wide variety of different sugars to the vancomycin
aglycone. We reasoned that if glycolipid derivatives of vanco-
mycin are bifunctional, then the glycosidic linkage to the phenol
might not be critical. If it is not, then we can substitute a simpler
linker, which would enable us to explore a wide range of different
carbohydrate moieties rapidly.
To evaluate the importance of the glycosidic linkage in the
activity of glycolipid derivatives of vancomycin, we prepared
compound 2 by the route shown in Scheme 1. The compound
was found to have excellent activity against sensitive strains
(Table 1). However, compared with compound 1b, which contains
a Conditions: (a) Tf2O, DTBMP, -78 to -20 °C, Et2O/CH2Cl2, 84%.
(b) i: NaI, acetone, 99%; ii: NaOMe, MeOH, 82%. (c) 9, Cs2CO3, DMF,
83%. (d) PdCl2(PPh3)2, Bu3SnH, DMF/AcOH, 79%. (e) 4,4′-Chlorobi-
phenyl aldehyde, DIEA, NaBH3CN, DMF, 46%.
the natural glycosidic linkage, 2 shows a modest decrease in
activity (2-5-fold) against resistant strains. Therefore, while linker
structure influences activity, it is possible to dispense with the
glycosidic linkage itself.7
The next issue to address was how to improve on the activity
of the linked compound 2. We have developed solid-phase
methods to make substituted disaccharide libraries containing
hundreds to thousands of members, but the effort involved is not
insignificant.8 Prior to undertaking the synthesis of a carbohydrate
library, we wanted to be sure that changing the carbohydrate
structure could lead to significant improvements in activity. We
explored a few conservative changes to the natural disaccharide,
but all of them led to a decrease in activity. For example,
(1) Walsh, C. T. Nature 2000, 406, 775.
(2) Nagarajan, R. J. Antibiot. 1993, 46, 118.
(3) (a) Ge, M.; Chen, Z.; Onishi, H. R.; Kohler, J.; Silver, L. L.; Kerns,
R.; Fukuzawa, S.; Thompson, C.; Kahne, D. Science 1999, 284, 507. (b) Kerns,
R.; Dong, S. D.; Fukuzawa, S.; Carbeck, J.; Kohler, J.; Silver, L.; Kahne, D.
J. Am. Chem. Soc. 2000, 122, 12608.
(4) Other bifunctional antibiotics, see: (a) Sucheck, S. J.; Wong, A. L.;
Koeller, K. M.; Boehr, D. D.; Draker, K.; Sears, P.; Wright, G. D.; Wong,
C.-H. J. Am. Chem. Soc. 2000, 122, 5230. (b) Wiedemann, I.; Breukink, E.;
Kraaij, C. v.; Kuipers, O. P.; Bierbaum, G.; de Kruijff, B.; Sahl, H.-G. J.
Biol. Chem. 2001, 276, 1772. (c) McCafferty, D. G.; Cudic, P.; Yu, M. K.;
Behenna, D. C.; Kruger, R. Curr. Opin. Chem. Biol. 1999, 3, 672.
(5) Other explanations have also been proposed and tested. See: (a)
Williams, D. H.; Bardsley, B. Angew. Chem., Int. Ed. Engl. 1999, 38, 1172.
(b) Rao, J.; Lahiri, J.; Isaacs, L.; Weis, R. M.; Whitesides, G. M. Science
1998, 280, 708. (c) Sundram, U. N.; Griffin, J. H. J. Am. Chem. Soc. 1996,
118, 13107. (d) Nicolaou, K. C.; Hughes, R.; Cho, S. Y.; Winssinger, N.;
Smethurst, C.; Labischinski, H.; Endermann, R. Angew. Chem., Int. Ed. 2000,
39, 3823.
(6) Both chemical and enzymatic methods have been used to make a limited
number of derivatives. see: (a) Ge, M.; Thompson, D.; Kahne, D. J. Am.
Chem. Soc. 1998, 120, 11014. (b) Thompson, C.; Ge, M.; Kahne, D. J. Am.
Chem. Soc. 1999, 121, 1237. (c) Nicolaou, K. C.; Mitchell, H. J.; Jain, N. F.;
Winssinger, N.; Hughes, R.; Bando, T. Angew. Chem., Int. Ed. Engl. 1999,
38, 240. (d) Solenberg, P. J.; Matsushima, P.; Stack, D. R.; Wilkie, S. C.;
Thompson, R. C.; Baltz, R. H. Chem. Biol. 1997, 4, 195. (e) Losey, H. C.;
Peczuh, M. W.; Chen, Z.; Eggert, U. S.; Dong, S. D.; Pelczer, I.; Kahne, D.;
Walsh, C. T. Biochemistry 2001, 40, 4745.
(7) As a control, we have also coupled the natural, unsubstituted disac-
charide to the vancomycin aglycone through an ethylene glycol linker. Like
vancomycin itself, this compound (2a) is active against sensitive strains but
not active against resistant strains. We have also attached a chlorobiphenyl-
substituted disaccharide to the carboxy terminus of the vancomycin aglycone.
The activity of this compound (2b) against resistant strains is comparable to
the activity of 1b. Compound 2b supports the conclusion that the natural
linkage is not essential. See Supporting Information for the structures and
MIC values of compound 2a and 2b.
(8) Liang, R.; Yan, L.; Loebach, J.; Ge, M.; Uozumi, Y.; Sekanina, K.;
Horan, N.; Gildersleeve, J.; Thompson, C.; Smith, A.; Biswas, K.; Still, W.
C.; Kahne, D. Science 1996, 274, 1520.
10.1021/ja0166693 CCC: $20.00 © 2001 American Chemical Society
Published on Web 11/17/2001

Communications to the Editor
J. Am. Chem. Soc., Vol. 123, No. 50, 2001 12723
Table 2. MICs of Moenomycin Disaccharide Derivativesa
Scheme 2a
a Conditions: (a) (CF3CO2)2Hg, (C2H5)2O‚BF3, 2-chloroethanol, 83%.
(b) i: (Me)3P, H2O/THF/EtOH; ii: 4-chloro-3-(trifluoromethyl)phenyl
isocyanate, CH2Cl2/DMF, 72% for two steps. (c) i: NaI, acetone; ii:
NaOMe, MeOH, 86% for two steps. (d) 10, Cs2CO3, DMF, 84%. (e)
PdCl2(PPh3)2, Bu3SnH, DMF/AcOH, 66%.
E. faecium
resistant
sensitive (VanA) sensitive
E. faecalis
resistant
(VanB)
compd
S. aureus
4
5
6
4
0.1
4
16
4
<0.1
125
8
1
1
2
0.2
125
2
compound 3 (Table 1), an isomer of compound 2, is less active
against both sensitive and resistant strains than 1b or 2. Although
the activity of compound 3 did change, it was not for the better.
We needed a more rational approach for compound selection.
We have suggested that the chlorobiphenyl disaccharide moiety
on vancomycin analogues 1b and 2 overcomes resistance by
interacting with proteins involved in the transglycosylation step
of cell wall biosynthesis.3,9 If this hypothesis explains the activity
of 1b and 2 against resistant bacterial strains, then one might
predict that replacing the disaccharide on 2 with a known
transglycosylase inhibitor would produce a still more active
compound. The best transglycosylase inhibitor known,10 moeno-
mycin, is a glycophospholipid containing five hexoses. Sofia and
co-workers recently made a solid-phase library of disaccharides
based on a fragment of moenomycin and identified compound 4
as having both good antibacterial activity and an ability to inhibit
transglycosylation.11 This disaccharide seemed like a much better
starting point than the vancomycin disaccharide because it has
better transglycosylase inhibitory activity. Therefore, we prepared
the disaccharide without the anomeric phospholipid (Scheme 2)
and linked it to the vancomycin aglycone to make 5. The activity
of compound 5 is shown in Table 2 along with the activities of
4 and a related analogue 6 that lacks the phospholipid (Table 2).
5 is significantly more active than compound 2 against resistant
strains. In fact, it is comparable to or better against these strains
than compound 1b (Table 1), the glycosidically linked prototype,
while maintaining excellent activity against sensitive strains. 5
is also more active than 4 even though it lacks the phospholipid
anchor, which was previously suggested to be critical for
biological activity (6, which does not contain either a phospholipid
anchor or the vancomycin aglycone, has minimal activity).11a
The preceding results support the hypothesis that better
vancomycin analogues can be made by attaching carbohydrates
with good transglycosylase inhibitory activity to the vancomycin
aglycone. The functionalized carbohydrate in 5 is based on a
disaccharide analogue (4) of moenomycin, a known transglyco-
250
2
1
250
>500
250
250
>500
250
VAb
6 + VAc
4
2
a Same bacterial strains were used as in Table 1. b Vancomycin
aglycone. c Add mixture of 6 and vancomycin aglycone.
sylase inhibitor.12,13 The phospholipid anchor in 4 was replaced
with the vancomycin aglycone to produce a hybrid compound
with activity that far exceeds the activity of the individual
components (compare 5 to the mixture of 6 with the vancomycin
aglycone, Table 2). The synthesis of a large collection of
vancomycin analogues will be greatly facilitated by the replace-
ment of the glycosidic linkage with a simple ethylene glycol
linker. Vancomycin has inspired the design of many synthetic
peptide binders.14 Attaching transglycosylase inhibitors to those
compounds could be a fruitful approach to the design of new
antibiotics. Hence, this work suggests significant opportunities
for the design of a large class of new antibiotics comprised of
hosts that bind to cell surface peptides attached to specific
inhibitors for peptidoglycan-processing enzymes.
Acknowledgment. This work was supported by Advanced Medicine,
Inc.
Supporting Information Available: Experimental details, including
spectral information, for the synthesis of compounds 11, 12, and 5, as
well as MIC data of related compounds 2a and 2b (see footnote 7) (PDF).
This material is available free of charge via the Internet at http://pubs.acs.org.
JA0166693
(12) We have confirmed that 5 inhibits transglycosylation in a permeabilized
E. coli model, like 4.3a,11b In addition, mutant bacterial strains described in
ref 13, which are resistant to transglycosylase inhibitors, are also resistant to
compounds 4 and 5.
(13) Eggert, U. S.; Ruiz, N.; Falcone, B. V.; Branstrom, A. A.; Goldman,
R. C.; Silhavy, T. J.; Kahne, D. Science 2001, 294, 361.
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(9) Goldman, R. C.; Baizman, E. R.; Longley, C. B.; Branstrom, A. A.
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(10) van Heijenoort, Y.; Derrien, M.; van Heijenoort, J. FEBS Lett. 1978,
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