Novel peptidomimetics of the antifungal cyclic peptide rhodopeptin: Synthesis of mimetics and their antifungal activity

Article abstract of DOI:10.1021/ol016394w

(matrix presented) Novel peptidomimetics of the antifungal cyclic peptide Rhodopeptin were synthesized. As with the cyclic peptides, the presence of all three Rhodopeptin side chains was found to be indispensable for peptidomimetic activity. We discovered

Full text of DOI:10.1021/ol016394w

ORGANIC
LETTERS
2001
Vol. 3, No. 22
3451-3454
Novel Peptidomimetics of the Antifungal
Cyclic Peptide Rhodopeptin: Synthesis
of Mimetics and Their Antifungal
Activity
Haruko C. Kawato,* Kiyoshi Nakayama, Hiroaki Inagaki, and Toshiharu Ohta*
Medicinal Chemistry Research Laboratory, Daiichi Pharmaceutical Co., Ltd.,
1-16-13 Kitakasai, Edogawa-ku, Tokyo 134-8630, Japan
kawatlm8@daiichipharm.co.jp
Received July 5, 2001
ABSTRACT
Novel peptidomimetics of the antifungal cyclic peptide Rhodopeptin were synthesized. As with the cyclic peptides, the presence of all three
Rhodopeptin side chains was found to be indispensable for peptidomimetic activity. We discovered new compounds exhibiting greater antifungal
activity and improved physiochemical properties in comparison to the parent compounds.
In the preceding paper,1 we described our concept and the
design of peptidomimetics of Rhodopeptin derivatives utiliz-
ing scaffolding methodology. Here, we report the synthesis,
the antifungal activity, and the water solubility of those
mimetics.
naldehyde provided compound 6 as a mixture of olefin
isomers. Subsequent introduction of a protected aminoalkyl
moiety at the 3-position and hydrogenolysis furnished
trisubstituted hydantoin 7 as a racemate.
The glucosamine derivatives 11 and 14 were synthesized
from bromo sugar 8 (Scheme 2). Incorporation of the
lipophilic straight chain was accomplished by zinc triflate
promoted glycosilation3 and was followed by deprotection
of the 2,2,2-trichloroethoxycarbonyl (Troc) group, introduc-
tion of the tert-butylacetyl group, and acetate removal to
provide 10. Protection of the 4,6-diol as the corresponding
acetonide allowed the 3-hydroxyl group to be acylated and
was followed by deprotection of the acetonide and removal
of the Cbz group to afford 11. Alternatively, benzylidene
formation on the 4,6-diol of 10, protection of the 3-hydoxyl
group with the Troc group, and NaBH3CN reduction4 under
Synthetic preparation of the hydantoin derivatives was
accomplished as shown in Scheme 1. Acylation of urea 3
with chloroacetyl chloride, followed by cyclization with
neopentylamine, gave compound 4. Introduction of a pro-
tected aminoalkyl moiety at the 3-position and hydrogeno-
lysis furnished compound 5, which contains two of the
three critical side chains. Alternatively, bromination of 4,
Michaelis-Arbuzov reaction with triethyl phosphite, and a
Horner-Wadsworth-Emmons-type reaction2 with n-capri-
(1) (a) Kawato, H. C.; Inagaki, H.; Ohta, T.; Nakayama, K. Org. Lett.
2001, 3, 3447.
(2) Meanwell, N. A.; Roth, H. R.; Smith, C. R.; Wedding, D. L.; Wright,
J. J. K. J. Org. Chem. 1991, 56, 6897.
(3) Nakayama, K.; Higashi, K.; Soga, T.; Uoto, K.; Kusama, T. Chem.
Pharm. Bull. 1992, 40, 2909.
10.1021/ol016394w CCC: $20.00 © 2001 American Chemical Society
Published on Web 10/04/2001

Scheme 1
Scheme 2
acidic conditions provided 3,6-O-protected derivative 12.
Acylation of 12, followed by stepwise removal of all
protecting groups, furnished compound 14, which bears the
basic side chain on the 4-hydroxyl group. It was crystallized
as a citric acid salt.
The benzimidazole derivatives were prepared as depicted
in Scheme 3. Acylation with isobutyryl chloride on both
amino and hydroxyl groups of 2-amino-3-nitrophenol 15,
followed by hydrolysis of the resultant ester, afforded amide
16. Alkylation with n-nonyl bromide, hydrogenolysis, and
cyclization under acidic conditions then provided disubsti-
tuted benzimidazole 17. Attempted introduction of the
aminoalkyl moiety onto nitrogen gave a mixture of regio-
isomers 18 and 19, whose structures were determined by
NOE experiments. Isolation of the desired isomer 18 from
this unoptimized transformation was followed by hydro-
genolysis to furnish 20.
Synthesis of the quinolone derivative 24,5 as depicted in
Scheme 4, began with a thermal condensation and ring
closure sequence from m-nitrophenol 21 to afford 22.
Subsequent alkylation and hydrolysis furnished the N-
alkylated carboxylic acid 23. Amide formation, which thus
far has only been successfully achieved by a mixed anhydride
protocol, followed by deprotection of the Boc group,
completed the construction of 24.
yields. At this stage, acylation of compound 28 met with
some difficulties. The extremely weak nucleophilicity of the
amine moiety resulted in initial acylation exclusively at the
alcohol residue. Although subsequent acylation of the amine
group was unsuccessful under the typical DCC/DMAP
conditions, we discovered that dipyridyl carbonate (DPC),9
along with DMAP, provided a superior coupling protocol
to furnish 30 in high yields. We utilized DL-leucine deriva-
tives in these coupling reactions, not only to allow simul-
taneous biological testing of all stereoisomers but also to
avoid any issues associated with possible racemization. From
30, removal of the Fmoc group and cyclization with acetic
acid provided benzodiazepine 31. Hydrolysis and a Mit-
sunobu reaction were utilized to introduce the basic side
chain, and final alkylation on the nitrogen of the benzodi-
azepine ring and treatment with hydrazine completed the
synthesis of 32. The other benzodiazepine, 36, was prepared
The synthesis of benzodiazepine derivatives 32 and 366
(Scheme 5) commenced with the palladium-mediated cross-
coupling reaction7 between aryl iodide 25 and acetylene
derivatives. Selective hydrolysis at the benzylic position with
a Na2S/HCl protocol8 afforded compounds 28 and 29 in high
(4) Wischnat, R.; Martin, R.; Wong, C.-H. J. Org. Chem. 1998, 63, 8361.
(5) Mozek, I.; Sket, B. J. Heterocycl. Chem. 1994, 31, 1293.
(6) Wu, C.; Biediger, R. J.; Kogan, T. P. Synth. Commun. 1999, 29,
3509.
(8) Chapdelaine, M. J.; Warwick, P. J.; Shaw, A. J. Org. Chem. 1989,
54, 1218.
(7) Arcadi, A.; Cacchi, S.; Carnicelli, V.; Marinelli, F. Tetrahedron 1994,
50, 437.
(9) Uoto, K.; Ohsuki, S.; Takenoshita, H.; Ishiyama, T.; Iimura, S.; Hirota,
Y.; Mitsui, I.; Terasawa, H.; Soga, T. Chem. Pharm. Bull. 1997, 45, 1793.
3452
Org. Lett., Vol. 3, No. 22, 2001

Scheme 3
Scheme 5
in seven steps in an analogous fashion from compound 29.
It was crystallized as a citric acid salt.
The minimum inhibitory concentration values (MIC, µg/
mL) against Candida albicans ATCC24433, Cryptococcus
neoformans IAM12253, and Aspergillus fumigatus ATCC-
36607 are presented in Table 1,10 with the activities of AMPH
and FLCZ11 as references. Solubilities in water12 of selected
compounds are also listed.
Compounds 4, 5, and 17, which do not contain all three
side-chain moieties, showed no potency or lower activity than
the parent peptides 1 and 2.
Scheme 4
According to our molecular modeling studies in the
preceding paper, each of the designed trisubstituted pepti-
domimetics displayed reasonably good three-dimensional
overlap with the cyclic peptide. However, the location of
the bulky hydrophobic side chain of the glucosamine
(10) The in vitro susceptibility testing was done by a microdilution
method. Minimum inhibitory concentration (MIC) was defined as the lowest
concentration of a compound which gave no visibly detectable fungal growth
after incubation at 30 °C for 72 h. Medium: synthetic amino acid medium
fungal (SAAMF). Tested concentration of compounds: 2, 10, 50 µg/mL.
(11) Amphotericin B (AMPH; Fungizone, Bristol Myers Squibb, New
Brunswick, NJ) and fluconazole (FLCZ; Diflucan, Pfizer, NY).
Org. Lett., Vol. 3, No. 22, 2001
3453

Notably, the location of the basic side chain of this compound
was slightly altered relative to the parent compounds and
the other mimetics due to hydrogen bonding between the
amide proton and the 4-oxo group. This result suggests a
correlation between this conformational restriction and a
more active conformation. The mechanism of the action of
these peptidomimetics and Rhodopeptin itself is now being
investigated in our laboratory.
Table 1. Antifungal Activity and Solubility in Water of
Peptidomimetics
MIC (µg/mL)
solubility
C. albicans C. neoformans A. fumigatus in water
compd ATCC24433
IAM12253
ATCC36607 (mg/mL)
4
5
7
11
14
17
20
>50
>50
50
>50
>50
>50
10
>50
>50
2
50
50
2
>50
>50
10
>50
>50
>50
50
Of equal importance, the peptidomimetics possessed
greatly improved solubility in water. Whereas Rhodopeptin
analogues typically showed a solubility of only 1-2 mg/
mL in water, the peptidomimetics all exhibited solubilities
>50 mg/mL. This result demonstrates that the scaffolding
methodology is very effective for improving physiochemical
properties without losing the desired biological activity.
>100
>100
>100
2
>100
>100
>100
>50
24
32
36
2
10
10
2
10
2
10
50
10
1
2
10
10
2
>50
10
10
2
>50
10
10
0.9
2.6
In conclusion, novel peptidomimetics of the antifungal
cyclic peptide Rhodopeptin were designed and synthesized,
utilizing a scaffolding methodology. It was observed that the
three side chains are indispensable for antifungal activity of
the peptidomimetics, which is a similar requirement for the
parent cyclic peptides. We have successfully produced potent
mimics of Rhodopeptin. Interestingly, the similarity of the
three-dimensional structures of the new peptidomimetics
relative to Rhodopeptin correlated extremely well with the
magnitude of antifungal activity. In addition, we discovered
new compounds exhibiting greater antifungal activity and
improved physiochemical properties. Further investigations
to identify additional mimetics through this scaffolding
methodology are ongoing in our laboratory and will be
reported in due course.
AMPH
FLCZ
10
>50
derivatives 11 and 14 deviated slightly from the parent cyclic
peptide. Those mimics showed only weak activity against
Cryptococcus neoformans, suggesting that the position of
the bulky side chain is critical. Additionally, it appears that
the position of the bulky neutral moiety of mimetics using
monocyclic subunits, 7, 11, and 14, contain significant
conformation flexibility. In contrast, compound 20, whose
three critical moieties are more constrained in the proper
positions, seem to fit the parent peptide better, which might
explain its better antifungal activity.
The other peptidomimetics (20, 24, 32, and 36) exhibited
antifungal activity with equal to or greater magnitude than
that of the parent cyclic peptides. Particularly encouraging
was the extremely high potency of quinolone analogue 24.
Acknowledgment. We thank Dr. Ryohei Nakajima, Mr.
Akihiro Kitamura, and Mr. Kazuhiko Someya of Daiichi
Pharmaceutical Co., Ltd. for the evaluation of antifungal
activity. We also thank Prof. William Moser of IUPUI for
critiquing the manuscript.
(12) For peptidomimetics. About 1 mg of each compound was accurately
weighed and transferred to a glass tube. About 10 µL of distilled water
(the concentration: 100 mg/mL) was added to the tube. The dissolution of
the compound was checked by visual observation after the sample tube
was shaken vigorously. When the sample was not completely dissolved, a
2-fold dilution (the concentration: 50 mg/mL) was then checked. For
Rhodopeptin analogues. Since Rhodopeptin analogues showed a low
solubility, another protocol was employed. Thus, the powder (ca. 1 mg)
was combined with 200 µL of distilled water (the concentration: ca. 5 mg/
mL). After mixing the sample, followed by centrifugation, the concentration
of top clear layer was measured by HPLC and the concentration was
calculated using a calibration curve.
Supporting Information Available: Spectroscopic and
analytical data for compounds 4-7, 9-14, 16-20, 22-24,
26-29, and 31-36. This material is available free of charge
via the Internet at http://pubs.acs.org.
OL016394W
3454
Org. Lett., Vol. 3, No. 22, 2001