Synthesis of sordaricin analogues as potent antifungal agents against Candida albicans

Article abstract of DOI:10.1016/S0960-894X(02)00020-3

Sordaricin derivatives possessing a cyclohexane ring appendage attached via an ether, thioether, amine, oxime, ester or amide linkage were synthesized and their antifungal activity was evaluated in vitro. Compounds containing a thioether bond or an oxime

Full text of DOI:10.1016/S0960-894X(02)00020-3

Bioorganic & Medicinal Chemistry Letters 12 (2002) 803–806
Synthesis of Sordaricin Analogues as Potent Antifungal
Agents against Candida albicans
Satoru Kaneko,a,* Takuya Uchida,a Satoshi Shibuya,a Takeshi Honda,a Isao Kawamoto,a
Tamako Harasaki,b Takashi Fukuokab and Toshiyuki Konosua,*
aMedicinal Chemistry Research Laboratories, Sankyo Co., Ltd., 2-58, Hiromachi 1-chome, Shinagawa-ku, Tokyo 140-8710, Japan
bBiological Research Laboratories, Sankyo Co., Ltd., 2-58, Hiromachi 1-chome, Shinagawa-ku, Tokyo 140-8710, Japan
Received 5 November 2001; accepted 28 December 2001
Abstract—Sordaricin derivatives possessing a cyclohexane ring appendage attached via an ether, thioether, amine, oxime, ester or
amide linkage were synthesized and their antifungal activity was evaluated in vitro. Compounds containing a thioether bond or an
oxime bond as a linkage exhibited potent MICs (ꢀ0.125 mg/mL) against four Candida albicans strains including azole-low-suscep-
tible strains. They were also active (MIC ꢀ0.125 mg/mL) against Candida glabrata. Their in vivo efficacy was confirmed in a murine
intravenous infection model with Candida albicans. # 2002 Elsevier Science Ltd. All rights reserved.
Zofimarin (1) is an antifungal natural product isolated
from Zopfielle marina SANK21274.1 Sordarin (2) con-
tains a 6-desoxyaltrose moiety that does not have any
acyl chains, and it is also a natural glycoside isolated
from Sordaria araneosa.2 Sordaricin (3) is a common
tetracyclic diterpene aglycon present in both 1 and 2.
While 1 exhibits moderate inhibitory activity in the
growth of pathogenic fungi, 2 and 3 show almost no
inhibitory activity. Recently, some potent antifungal
derivatives among the sordarin family have been repor-
ted.3,4 When a hydrophobic aliphatic side chain was
attached to 3 instead of the sugar moiety, its activity
increased enough to inhibit the growth of fungi in an in
vitro assay. In particular, the isopentyl ether derivative
4 exhibited good potency as an antifungal agent.5 The
sordarin family inhibits fungal protein synthesis by
selectively interrupting the functions of fungal elonga-
tion factor 2, which plays important roles with ribosome
in the translation process.6ꢁ8 Since the disclosure of the
mode of action, chemical modification of 3 has been
considered as one of new attractive targets for the
development of novel antifungal agents.9
group. Herein, we describe the synthesis of the com-
pounds having a cyclohexyl ring linked by various
bonds and their in vitro antifungal activity against
Candida spp (Fig. 1).
Synthesis of Sordaricin Analogues
The synthetic routes of various derivatives are shown in
Scheme 1. Sordaricin benzyl ester 5a and sordaricin
benzhydryl ester 5b were prepared according to the
reported methods.5 For cyclohexyl ether derivative 7,
In the course of our modification, it was found that a
cyclohexyl group was one of the most effective aliphatic
components as a side chain. Therefore, our attention
was focused on the biological activity of derivatives with
various linkages between the aglycon and a cyclohexyl
*Corresponding author. Tel.: +81-3-3492-3131; fax: +81-3-5436-
8563; e-mail: skanek@shina.sankyo.co.jp
Figure 1. Chemical structures of zofimarin 1, sordarin 2, sordaricin 3
and isopentyl ether derivative 4.
0960-894X/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(02)00020-3

804
S. Kaneko et al. / Bioorg. Med. Chem. Lett. 12 (2002) 803–806
alkylation of the hydroxyl group of 5a was carried out
with 2-cyclohexenyl bromide and silver oxide at room
temperature in dichloromethane, to afford 6 (36%).
Subsequent hydrogenolysis brought about both satura-
tion of the cyclohexene ring and removal of the benzyl
ester, to yield 7 (70%). For cyclohexyl thioether deriva-
tive 10, compound 5a was first converted to unstable
triflate 8a (95%). Thioether formation was performed
by treatment with mercaptocyclohexane and sodium
hydride at 0 ꢂC in N,N-dimethylformamide to give 9
(59%). Then, hydrogenolysis of 9 yielded 10 (56%). For
N-methyl cyclohexylamine derivative 12, substitution of
8a was carried out with lithium N-methyl cyclohexyl-
amide at ꢁ50 ꢂC in tetrahydrofuran, to give 11 (21%).
Subsequent hydrogenolysis yielded 12 (61%). For
cyclohexanone oxime derivative 13, compound 5b was
first converted to triflate 8b. A displacement reaction of 8b
was carried out with potassium hydride and cyclohexanone
oxime at 0 ꢂC in N,N-dimethylformamide. Finally, de-
protection was performed with trifluoroacetic acid at 0 ꢂC
in dichloromethane to give 13 (57% in two steps).
Next, we set out for the oxidation of the hydroxyl-
methyl group of 5a as shown in Scheme 2. The aldehyde
of 5a was initially protected as an ethylene acetal.
Treatment of 5a with ethylene glycol catalyzed by
toluenesulfonic acid in methanol afforded 14 (78%).
Swern oxidation of 14 gave aldehyde 15 (87%). For O-
cyclohexyl oxime derivative 17, formation of oxime 1610
was performed with hydroxylamine hydrochloride in a
Scheme 1. (a) BnBr, NaHCO3/DMF; (b) Ph2CN2/MeOH; (c) 2-cyclohexenyl bromide, Ag2O, MS4A/CH2Cl2; (d) H2, Pd(OH)2-C/MeOH; (e) Tf2O,
Py/CH2Cl2; (f) mercaptocyclohexane, NaH/DMF; (g) N-methyl cyclohexylamine, BuLi/THF; (h) cyclohexanone oxime, KH/DMF; (i) TFA/
CH2Cl2.
.
Scheme 2. (a) CH(OMe)3, (CH2OH)2, TsOH/MeOH; (b) (COCl)2, Me2SO/CH2Cl2; Et3N; (c) NH2OH HCl/EtOH-Py; (d) cyclohexyl mesylate,
NaH/DMF; (e) (i) 1N-HCl/MeOH; (ii) H2, Pd(OH)2-C/MeOH; (f) NaClO2, NaH2PO4, 2-methyl-2-butene/tBuOH-H2O; (g) cyclohexanol, Ph3P,
DEAD/DMF; (h) 2,4,6-triisopropylbenzenesulfonyl chloride, Et3N, DMAP/CH2Cl2; cyclohexylamine.

S. Kaneko et al. / Bioorg. Med. Chem. Lett. 12 (2002) 803–806
Table 1. In vitro antifungal activity of novel sordaricin derivatives
805
Organism
MIC (mg/mL)
10 12
FCZa
ITCZb
1
7
13
17
19
20
Candida albicans ATCC24433
Candida albicans SANK51486
Candida albicans TIMM3164
Candida albicans ATCC64550
Candida glabrata ATCC90030
Candida tropicalis ATCC750
0.5
0.125
0.031
0.25
1
1
0.5
0.5
0.25
0.5
0.5
>4
0.5
0.125
ꢀ0.063
0.25
0.031
8
8
16
16
8
0.031
0.031
0.063
0.063
0.063
1
0.031
0.031
0.063
0.063
0.125
1
ꢀ0.125
ꢀ0.125
0.25
0.5
0.5
4
>4
>4
>4
>4
0.5
>4
0.25
>4
>4
>4
0.5
ꢀ0.016
0.063
0.125
0.031
2
0.25
ꢀ0.063
2
>16
aFluconazole.
bItraconazole.
mixture of pyridine and ethanol (1:1) (99%). O-alkyla-
tion of 16 was carried out with cyclohexyl mesylate and
sodium hydride at 0 ꢂC in N,N-dimethylformamide
(60%). Finally, cleavage of ethylene acetal and benzyl
ester gave 1710 (92%).
against most tested strains. From these results, it is
speculated that lipophilicity of the linkage is very
important to exhibit the biological activity, and that the
linkage directly affects the binding interaction with EF-
2. The two representative compounds (10 and 17) were
evaluated for their in vivo activity using a murine model
of systemic candidasis.13 The results are illustrated in
Figure 2. Remarkably, both compounds showed good
efficacy against systemic candidasis after both oral and
subcutaneous administration.
Carboxylic acid 18 was obtained by NaClO2 oxidation
of 15 (70%). For cyclohexyl ester derivative 19, ester-
ification of 18 was achieved by Mitsunobu reaction.
Treatment of 18 with cyclohexanol, triphenylphosphine
and diethylazodicarboxylate in N,N-dimethylformamide
gave the cyclohexyl ester (42%). Subsequent hydrolysis
and hydrogenolysis yielded 19 (75%). For cyclohexyl
amide derivative 20, amidation of 18 was carried out with
2,4,6-triisopropylbenzenesulfonyl chloride, triethylamine
and cyclohexylamine to afford the desired amide.
Hydrolysis and hydrogenolysis of this amide produced
20 (39% in three steps).11
In conclusion, several novel sordaricin derivatives were
synthesized. It was found that thioether 10 and oxime
13 and 17 had excellent activity as antifungal agents.
These compounds will serve to investigate the mode of
action of this class of compounds. They are considered
to be promising candidates for the therapeutic treat-
ment of fungal infection.
Acknowledgements
Biological Activity
Minimum inhibitory concentrations (MICs) of the ana-
logues are summarized in Table 1.12
The authors thank Dr. T. Ogita and our colleagues in
New Lead Discovery Research Laboratories (Sankyo
Co., Ltd.) for preparing the natural product and pro-
viding the raw materials.
Compounds 10, 13, and 17 showed excellent activity
against Candida albicans. In particular, the MIC value
of thioether 10 was ꢀ 0:016 mg/mL against C. albicans
SANK51486. Moreover, these compounds were active
against azole-low-susceptible strains as well as azole
susceptible ones. In particular, the inhibitory potency of
13 and 17 was 16-fold stronger than that of itraconzole
against C. albicans ATCC64550. In addition, these
compounds showed potent activity against C. glabrata
with MICs of 0.125 mg/mL or lower. However, com-
pounds 7 and 19 only showed moderate activity and
compounds 12 and 20 had extremely weak activity
References and Notes
1. Ogita, T.; Hayashi, T.; Sato, A.; Furutani, W. JP Patent
62,040,292, 1987.
2. Hauser, D.; Sigg, H. P. Helv. Chim. Acta 1971, 54, 1178.
3. Gargallo-Viola, D. Curr. Opin. Antiinfect. Invest. Drugs
1999, 1, 297.
4. Odds, F. C. Exp. Opin. Ther. Pat. 2001, 11, 283 and refer-
ences therein.
5. Tse, B.; Balkovec, J. M.; Blazey, C. M.; Hsu, M. J.;
Nielsen, J.; Schmatz, D. Bioorg. Med. Chem. Lett. 1998, 8,
2269.
6. Domınguez, J. M.; Kelly, V. A.; Kinsman, O. S.; Marriott,
M. S.; Gomez, H. F.; Martin, J. J. Antimicrob. Agents Che-
mother. 1998, 42, 2274.
7. Domınguez, J. M.; Martin, J. J. Antimicrob. Agents Che-
mother. 1998, 42, 2279.
8. Capa, L.; Mendoza, A.; Lavandera, J. L.; Gomez, H. F.;
Garcıa-Bustos, J. F. Antimicrob. Agents Chemother. 1998, 42,
2694.
9. Justice, M. C.; Hsu, M. J.; Tse, B.; Ku, T.; Balkovec, D.;
Schmatz, D.; Nielsen, J. J. Biol. Chem. 1998, 273, 3148.
Figure 2. In vivo efficacy of sordaricin derivatives against murine
intravenous infection with Candida albicans SANK51486. *P<0.05.

806
S. Kaneko et al. / Bioorg. Med. Chem. Lett. 12 (2002) 803–806
1
10. The geometry of oximes 16 and 17 was determined by H
NMR chemical shifts. Syn-aldoximes show their –CH¼N– sig-
nals at a magnetic field lower than approximately d 7.0ppm,
whereas anti-isomers show their signals at a higher field. The
corresponding signals of 16 and 17 appeared at d 7.72 and d 7.35,
respectively. In addition, a-branched aldehydes tend to pre-
dominantly afford thermodynamically more stable syn-isomers.
See: (a) Karabatsos, G. J.; Hsi, N. Tetrahedron 1967, 23, 10 79. (b)
Karabatsos, G. J.; Taller, R. A. Tetrahedron 1968, 24, 3347.
11. All new compounds gave satisfactory spectroscopic and
analytical data. The representative data are shown as follows.
d, J=7.6 Hz), 0.81 (3H, d, J=6.6 Hz); FABMS (m/z): 428
[M+H]+. 17: 1H NMR (400 MHz, CDCl3) d 9.99 (1H, s), 7.35
(1H, s), 6.03 (1H, d, J=3.7 Hz), 4.02 (1H, m), 2.78 (1H, t,
J=4.4 Hz), 2.33–2.25 (2H, m), 2.11–1.28 (m), 1.00 (3H, d,
J=6.6 Hz), 0.98 (3H, d, J=6.6 Hz), 0.84 (3H, d, J=6.6 Hz);
FABMS (m/z): 428 [M+H]+.
12. In vitro antifungal activity was determined at 35 ꢂC in
RPMI1640at pH 7..0 Minimum inhibitory concentration
(MIC) was defined as the minimum concentration of the test
compound that inhibited the growth of the fungi by 80%.
13. In vivo activity was determined in mice (each group con-
sisted of 8–10male mice, ddY strain, 5 weeks old) infected
systemically by an intravenous administration of 6ꢃ105 CFU
of C. albicans SANK51486. Test compounds (60mg/kg/
administration) were administered orally (po) or sub-
cutaneously (sc) 2, 5 and 8 h post infection. An aqueous solu-
tion of carboxymethyl cellulose sodium salt (0.5%) containing
dimethyl sulfoxide (10%) was used as a vehicle for adminis-
tration. Viable cell numbers in the kidneys were determined 24
h post infection.
1
10: H NMR (500 MHz, CDCl3) d: 9.66 (1H, s), 6.11 (1H, dd,
J=3.4, 1.2 Hz), 3.07 (1H, d, J=12.0Hz), 2.80(1H, t, J=3.8),
2.65 (1H, d, J=12.0 Hz), 2.60–1.10 (m), 1.06 (3H, d,
J=6.8 Hz), 0.99 (3H, d, J=6.8 Hz), 0.79 (3H, d, J=6.8 Hz);
FABMS (m/z): 431 [M+H]+. 13: 1H NMR (400 MHz,
CDCl3) d 9.82 (1H, s), 6.07 (1H, d, J=2.8 Hz), 4.20(1H, d,
J=9.6 Hz), 4.13 (1H, d, J=9.6 Hz), 2.70(1H, t, J=3.6 Hz),
2.46 (2H, m), 2.36–2.24 (3H, m), 2.18–1.52 (m), 1.32 (1H, d,
J=12.4 Hz), 1.25 (2H, m), 1.00 (3H, d, J=6.4 Hz), 0.96 (3H,