Exploring the optimal site for modifications of pyranmycins with the extended arm approach.

Article abstract of DOI:10.1021/ol027288c

[reaction: see text] Continuing from the syntheses and the antibacterial studies of a library of pyranmycins, we further probed the proximity around ring III of pyranmycin by introducing an "extended arm" that has hydroxyethyl or aminoethyl groups at the

Full text of DOI:10.1021/ol027288c

ORGANIC
LETTERS
2003
Vol. 5, No. 4
431-434
Exploring the Optimal Site for
Modifications of Pyranmycins with the
Extended Arm Approach
Jie Li, Jinhua Wang, Yu Hui, and Cheng-Wei Tom Chang*
Department of Chemistry and Biochemistry, Utah State UniVersity,
0300 Old Main Hill, Logan, Utah 84322-0300
chang@cc.usu.edu
Received November 15, 2002
ABSTRACT
Continuing from the syntheses and the antibacterial studies of a library of pyranmycins, we further probed the proximity around ring III of
pyranmycin by introducing an “extended arm” that has hydroxyethyl or aminoethyl groups at the O-2′′, O-3′′, or O-4′′ positions. The results
from the antibacterial studies reveal the optimal structural motif is the attachment of an extended arm with a terminal hydroxyl group at the
O-3′′ position.
Aminoglycoside antibiotics, such as neomycin, have been
administered clinically for over fifty years.1,2 Overuse and
misuse of antibiotics have incurred the emergence of
antibiotic resistant bacteria,3,4 which significantly limits the
usefulness of aminoglycoside antibiotics. To counteract this
problem, our group has recently developed an expedited
method for the preparation of a novel class of aminoglycoside
antibiotics, pyranmycins,5 based on our aminosugar library6
and the reported X-ray and NMR structural studies (Figure
1).7-10 Herein, we wish to report the results from further
Figure 1. Neomycin-based aminoglycosides and pyranmycin.
(1) Hooper, I. R. Aminoglycoside Antibiotics; Springer-Verlag: New
York, 1982.
modifications of pyranmycins using the extended arm
approach.
Most of the aminoglycoside-resistant bacteria inactivate
the aminoglycosides via aminoglycoside-modifying enzymes
that introduce modifications on the neamine (rings I and
II).11,12 Chemical derivatization on neamine often results in
(2) Haddad, J.; Kotra, L. P.; Mobashery, S. In Glycochemistry Principles,
Synthesis, and Applications; Wang, P. G., Bertozzi, C. R., Eds.; Marcel
Dekker: New York, 2001; p 307.
(3) Cohen, M. L. Science 2002, 257, 1050-1055.
(4) Neu, H. C. Science 2002, 257, 1064-1072.
(5) (a) Wang, J.; Li, J.; Tuttle, D.; Takemoto, J.; Chang, C.-W. T. Org.
Lett. 2002, 4, 3997-4000, (b) Chang, C.-W. T.; Hui, Y.; Elchert, B.; Wang,
J.; Li, J.; Rai, R. Org. Lett. 2002, 4, 4603-4606.
(6) Elchert, B.; Yu, H.; Chang, C.-W. T. Submitted for publication.
(7) Fourmy, D.; Recht, M. I.; Puglisi, J. D. J. Mol. Biol. 1998, 277, 347-
362.
(9) Fourmy, D.; Yoshizawa, S.; Puglisi, J. D. J. Mol. Biol. 1998, 277,
333-345.
(10) Ma, C.; Baker, N. A.; Joseph, S.; McCammon, J. A. J. Am. Chem.
Soc. 2002, 124, 1438-1442.
(8) Fourmy, D.; Recht, M. I.; Blanchard, S. C.; Puglisi, J. D. Science
1996, 274, 1367.
10.1021/ol027288c CCC: $25.00 © 2003 American Chemical Society
Published on Web 01/24/2003

a lack or a decrease in the activity.13 To avoid the decrease
in activity, we have developed a general method for the
synthesis of pyranmycins, which consists of a neamine core
and variable pyranoses as the ring III component. We have
also identified the lead structure at ring III of pyranmycins.5b
We wish to further explore the possible site(s) for the
attachment of additional functionalities at the ring III
pyranose of pyranmycin. To achieve this goal, we synthe-
sized seven derivatives of pyranmycins by extending the
amino group or hydroxyl group away from the ring III
pyranose of pyranmycin (Figure 2). The identification of the
optimal site(s) will allow us to introduce more complex
structural components in the future.
extended arms at both O-2′′ and O-3′′ positions, such as
TC025 and TC031. By comparing to the single extended
arm constructs (O-3′′) like TC026, TC028, and TC032, we
can evaluate the effect of the extended arm at O-2′′.
The syntheses of single extended arm pyranmycins require
the preparation of several glycosyl donors 5, 13, 17, 22, and
25 (Schemes 1, 2, and 3). The synthesis of 5 begins from
methyl 6-deoxy-2,3-di-O-benzyl-R-D-glucopyranoside.6 Al-
lylation of 4-OH followed by ozonolysis and reductive
workup provides 3 with a hydroxyethyl group at O-4
(Scheme 1). The benzyl and methyl groups were converted
Scheme 1. Synthesis of Glycosyl Donora
a Conditions: (a) Allyl bromide, NaH, TBAI, THF. (b) (i) O3,
CH2Cl2; (ii) NaBH4, MeOH. (c) Ac2O, cat. H2SO4. (d) (i) NH2NH2-
HOAc, DMF; (ii) CCl3CN, DBU, CH2Cl2.
into acetyl groups by using Ac2O and a catalytic amount of
H2SO4 concomitant with the acetylation of the hydroxyl
group on the extended arm. Selective deprotection of the
anomeric acetyl group followed by reacting with trichloro-
acetonitrile and DBU gave the desired glycosyl donor, 5.
The synthesis of 13 begins with 615 (Scheme 2). Allylation
of 3-OH followed by the deprotection of benzylidene group
generated a diol, 7. Selective tosylation of 6-OH followed
by LiAlH4 reduction gave the 6-deoxygenated compound,
Figure 2. Structures of the designed pyranmycins.
As more diverse structural scaffolds are incorporated, the
overall contour of the modified pyranmycins will deviate
from their predecessors that resemble neomycin. Thus, these
modified pyranmycins are expected to become poor sub-
strates for the aminoglycoside-modifying enzymes. There-
fore, if the high potency of these extended arm-modified
pyranmycins can be maintained, we can generate effective
new aminoglycoside antibiotics against resistant strains of
bacteria.
Scheme 2. Synthesis of Glycosyl Donora
Our previous work elucidates the importance of having a
6′′-CH3 group.5 Therefore, all of the constructs will follow
this trait except TC024. We began our approach by extending
the 4′′-OH by two carbons creating TC023. It is difficult to
install a single extended arm at the O-2′′ position while
maintaining the crucial â-glycosidic bond during the glyco-
sylation.14 Therefore, we synthesized compounds with double
a Conditions: (a) (i) Allyl bromide, NaH, TBAI, THF; (ii)
TsOH-H2O, MeOH. (b) (i) TsCl, py.; (ii) LiAlH4, THF. (c) (i)
(COCl)2, DMSO; DIPEA; (ii) NaBH4, MeOH. (d) (i) Tf2O, py.,
CH2Cl2; (ii) NaN3, DMF. (e) (i) O3, CH2Cl2; (ii) NaBH4, MeOH.
(f) Ac2O, cat. H2SO4. (g) (i) NH2NH2-HOAc, DMF; (ii) CCl3CN,
DBU, CH2Cl2.
(11) Kotra, L. P.; Haddad, J.; Mobashery, S. Antimicrob. Agents
Chemother. 2000, 44, 3249-3256..
(12) Mingeot-Leclercq, M.-P.; Glupczynski, Y.; Tulkens, P. M. Antimi-
crob. Agents Chemother. 1997, 43, 727-737.
(13) Greenberg, W. A,; Priestley, E. S.; Sears, P. S.; Alper, P. B.;
Rosenbohm, C.; Hendrix, M.; Hung, S.-C.; Wong, C.-H. J. Am. Chem. Soc.
1999, 121, 6527-6541.
432
Org. Lett., Vol. 5, No. 4, 2003

8. Epimerization of the equatorial 4-OH using Swern
oxidation and NaBH4 reduction allowed an SN2 azide-
substitution via treatments of Tf2O then NaN3. The resulting
product, 10, which decomposed gradually at room temper-
ature, was subjected to ozonolysis and reductive workup
providing 11. Standard procedures, described in Scheme 1,
were used to convert 11 into the corresponding glycosyl
donor, 13.
the 5,6-O-isopropylidene group of 14 allowed the deoxy-
genation of 6-OH, using TsCl and LiAlH4 treatments. The
deoxygenated product, 19, underwent ozonolysis and reduc-
tive workup generating 20, which led to two different routes.
One route employed similar hydrolysis, acetylation, then the
protocols for the synthesis of glycosyl trichloroacetimidate
offering 22 with a hydroxyethyl group at O-3 and 6-deoxy-
genation. The other route proceeded via an azide substitution
at the primary hydroxyl group on the extended arm. The
product, 23, gave rise to the glycosyl trichloroacetimidate,
25, with the terminal azido (amino) group at the extended
arm and 6-deoxygenation.
The syntheses of 17, 22, and 25 started from diacetone
D-glucose through a divergent approach (Scheme 3). After
Standard procedures modified from the literature for
glycosylation16 and final syntheses17 were used for the
preparation of final products ready for antibacterial assay
(Scheme 4).4
Scheme 3. Syntheses of Glycosyl Donorsa
Scheme 4. Synthesis of the Designed Pyranmycinsa
a Conditions: (a) 5, 13, 17, 22, or 25 and BF3-OEt2, CH2Cl2, 4
A MS. (b) (i) K2CO3, MeOH; (ii) PMe3, THF/H2O; (iii) H2,
Pd(OH)2/C; (iv) Dowex 1X8 (Cl- form).
a Conditions: (a) Allyl bromide, NaH, nBu4NI, THF. (b) TsOH,
MeOH. (c) (i) TsCl, py.; (ii) LiAlH4, THF. (d) (i) HOAc, TFA,
H2O; (ii) Ac2O, cat. H2SO4. (e) (i) H2NNH2-HOAc, DMF; (ii)
CCl3CN, DBU, CH2Cl2. (f) (i) O3, CH2Cl2; (ii) NaBH4, MeOH.
(g) (i) TsCl, py.; (ii) NaN3, DMF.
The syntheses of double extended arm pyranmycins began
with the precursor we have prepared.5 Incorporation of allyl
groups, followed by ozonolysis and reductive workup,
provides one of the designed diols, 33. Further azide
substitution gave the pyranmycin precursor with a double
extended arm, 34. Both compounds underwent the same final
syntheses generating the desired final products (Scheme 5).
All of the final products were assayed against Escherichia
coli with neomycin and ribostamycin as the controls (Table
1).18 Our preliminary goal is to generate the comparable
results with the pyranmycins without an extended arm
allylation at the 3-OH, the product 14 was diverted into two
distinct routes. Ozonolysis and reductive workup of 14,
followed by acid hydrolysis of isoproplyidene groups and
acetylation, gave 16 with a hydroxyethyl group at O-3.
Standard procedures were used to convert 16 into the
corresponding glycosyl donor, 17. Selective deprotection of
(14) The â-glycosidic bond is essential for the formation of intramolecular
hydrogen bonding between 2′-NH2 and O-5′′ of the constructed pyranmy-
cins. Please refer to the NMR study for the binding of aminoglycoside
toward RNA in ref 8.
(16) Hannessian, S. PreparatiVe Carbohydrate Chemistry; Marcel De-
kker: New York, 1997; pp 283-312.
(17) Sucheck, S. J.; Greenberg, W. A.; Tolbert, T. J.; Wong, C.-H. Angew.
Chem., Int. Ed. 2000, 39, 1080-1084.
(15) Dasgupta, F.; Garegg, P. J. Synthesis 1994, 1121-1123.
Org. Lett., Vol. 5, No. 4, 2003
433

Scheme 5. Synthesis of the Designed Pyranmycinsa
Figure 3. Pyranmycins without an extended arm.
activity (entries 4 vs 10, and entries 5 vs 9 and 12) in the
scaffolds with 6-deoxypyranose (6′′-CH3) at ring III. A
surprising decrease in the antibacterial activity against E. coli
(entries 9 and 12) is noticed, as a charged ammonium group,
rather than a neutral hydroxyl group, was introduced at the
terminal end of the extended arm at O-3′′. This finding
demonstrates the value of using real molecules for the SAR
studies, which is difficult to predict by computation simula-
tion that relies on charge-charge interaction.
In conclusion, we have developed a methodology to
introduce functionalities for probing the perimeter of the
binding sites of pyranmycins. This methodology can be
applied to study interactions in other systems. From the SAR
results, we are pleased to identify that the optimal site for
future modification is at O-3′′. As revealed by TC028, no
severe decrease in the antibacterial activity from its unmodi-
fied counterpart, TC005, is observed. We are currently using
solid-phase synthesis to explore more SAR information.
a Conditions: (a) (i) K2CO3, MeOH; (ii) allyl bromide, NaH,
TBAI, THF. (b) (i) O3, CH2Cl2; (ii) NaBH4, MeOH. (c) (i) K2CO3,
MeOH; (ii) PMe3, THF/H2O; (iii) H2, Pd(OH)2/C; (iv) Dowex 1X8
(Cl- form). (d) (i) Tf2O, py.; (ii) NaN3, DMF.
(Figure 3).5b The structure-activity relationship (SAR)
reveals that attaching an extended arm at O-2′′ will com-
pletely obliterate the antibacterial activity (entries 8 and 11),
while attaching the extended arm at O-4′′ dramatically
decreases the antibacterial activity (entries 5 vs 6). Attaching
an extended arm at O-3′′ also decreases the antibacterial
Acknowledgment. We acknowledge the financial support
from the Utah State University (CURI grant) and the support
from the Department of Chemistry and Biochemistry, Utah
State University. Mass spectrometry was provided by the
Washington University Mass Spectrometry Resource, and
the Mass Spectrometry Facilities, University of California,
Riverside.
Table 1. MIC against Escherichia coli
entry
compd
MICa (µM)
Supporting Information Available: Experimental pro-
cedures for the preparation of compounds 2-5, 7-27, 29,
30, TC023, TC024, TC025, TC026, TC028, TC031, and
1
2
3
4
5
6
7
8
9
neomycin
ribostamycin
TC001
TC005
TC020
TC023
TC024
TC025
TC026
2
7
42
9
22
38
40
inactive
27
1
TC032, and the corresponding H and 13C NMR spectral;
mass spectral data for compounds 2-4, 7-12, 14-16, 18-
21, 23, 24, 26, 27, 29, 30, TC023, TC024, TC025, TC026,
TC028, TC031, and TC032. This material is available free
of charge via the Internet at http://pubs.acs.org.
OL027288C
10
11
12
TC028
TC031
TC032
13
inactive
39
(18) Methods for Dilution Antimicrobial Susceptibility Testing for
Bacteria that Grow Aerobically, Approved standard M7-A5, and Perfor-
mance Standards for Antimicrobial Disk Susceptibility Tests, Approved
standard M2-A7, National Committee for Clinical Laboratory Standards,
Wayne, PA.
a MIC: minimum inhibitory concentration.
434
Org. Lett., Vol. 5, No. 4, 2003