Design and synthesis of novel imidazole-substituted dipeptide amides as potent and selective inhibitos of Candida albicans myristoylCoA:protein N- myristoyltransferase and identification of related tripeptide inhibitors with mechanism-based antifungal activity

Article abstract of DOI:10.1021/jm970094w

A new class of antifungal agents has been discovered which exert their activity by blockade of myristoylCoA:protein N-myristoyltransferase (NMT; EC 2.1.3.97). Genetic experiments have established that NMT is needed to maintain the viability of Candida albicans and Cryptococcus neoformans, the two principal causes of systemic fungal infections in immunocompromised humans. Beginning with a weak octapeptide inhibitor ALYASKLS-NH₂ (2, k(i) = 15.3 ± 6.4 μM), a series of imidazole-substituted Ser-Lys dipeptide amides have been designed and synthesized as potent and selective inhibitors of Candida albicans NMT. The strategy that led to these inhibitors evolved from the identification of those functional groups in the high-affinity octapeptide substrate GLYASKLS-NH₃ 1a necessary for tight binding truncation of the C-terminus, replacement of the four amino acids at the N-terminus by a spacer group, and substitution of the glycine amino group with an N-linked 2- methylimidazole moiety. Initial structure-activity studies led to the identification of 31 as a potent and selective peptidomimetic inhibitor with an IC₅₀ of 56 nM and 250-fold selectivity versus human NMT. 2- Methylimidazole as the N-terminal amine replacement in combination with a 4- substituted phenacetyl moiety imparts remarkable potency and selectivity to this novel class of inhibitors. The (S:S) stereochemistry of serine and lysine residues is critical for the inhibitory activity, since the (R,R) enantiomer 40 is 10²-fold less active than the (S,S) isomer 31. The inhibitory profile exhibited by this new class of NMT ligands is a function of the pK(o) of the imidazole substituent as illustrated by the benzimidazole analog 35 which is about 10-fold less potent than 31. The measured pK(o) (7.1 ± 0.5) of 2-methylimidazole in 31 is comparable with the estimated pK(o) (~8.0) of the glycyl residue in the high-affinity substrate 1a. Groups bulkier than methyl, such as ethyl, isopropyl, or iodo, at the imidazole 2- position have a detrimental effect on potency. Further refinement of 31 by grafting an α-methyl group at the benzytic position adjacent to the serine residue led to 61 with an IC₅₀ of 40 nM. Subsequent chiral chromatography of 61 culminated in the discovery of the most potent Candida NMT inhibitor 61a reported to date with an IC₅₀ of 20 nM and 400-fold selectivity versus the human enzyme. Both 31 and 61a are competitive inhibitors of Candida NMT with respect to the octapeptide substrate GNAASARE-NH₂ with K(i)(app) = 30 and 27 nM, respectively. The potency and selectivity displayed by these inhibitors are dependent upon the size and orientation of the o-substituent. An α-methyl group with the R configuration corresponding to the (S)-methyl- 4-alanine in 2 confers maximum potency and selectivity. Structural modification of 31 and 61 by appending an (S)-carboyl group β to the cyclohexyl moiety provided the less potent tripeptice inhibitors 73a and 73b with an IC₅₀ of 1.45 ± 0.08 and 0.38 ± 0.03 μM, respectively. However, these tripeptides (73a and 73b) exhibited a pronounced selectivity of 560- and 2200-fold versus the human NMT. More importantly 73a displayed fungistatic activity against C. albicans with an EC₅₀ ± 17 μM in cell culture. Compound 73b also exhibited a similar antifungal activity. An Arf protein gel mobility shift assay for monitoring intracellular myristoylation revealed that a single dose of 200 μM of 73a or 73b produced <50% reduction in Arf N-myristoylation, after 24 and 48 h, consistent with their fungistatic rather than fungicidal activity. In contrast, the enantiomer 73d which had an IC₅₀ >1000 μM against C. albicans NMT did not exhibit antifungal activity and produced no detectable reduction in Arf N-myristoylation in cultures of C. albicans. These studies confirm that the observed antifungal activity of 73a adn 73b is due to the attenuation of NMT activity and that NMT represents an attractive target for the development of novel antifungal agents.

Full text of DOI:10.1021/jm970094w

J . Med. Chem. 1997, 40, 2609-2625
2609
Design a n d Syn th esis of Novel Im id a zole-Su bstitu ted Dip ep tid e Am id es a s
P oten t a n d Selective In h ibitor s of Ca n d id a a lbica n s Myr istoylCoA:P r otein
N-Myr istoyltr a n sfer a se a n d Id en tifica tion of Rela ted Tr ip ep tid e In h ibitor s
w ith Mech a n ism -Ba sed An tifu n ga l Activity†
Balekudru Devadas,* Sandra K. Freeman, Mark E. Zupec, Hwang-Fun Lu, Srinivasan R. Nagarajan,
Nandini S. Kishore, J ennifer K. Lodge, David W. Kuneman,‡ Charles A. McWherter, Dutt V. Vinjamoori,‡
Daniel P. Getman, J effrey I. Gordon,§ and J ames A. Sikorski
Department of Medicinal and Structural Chemistry, G. D. Searle and Company, 700 Chesterfield Parkway North,
St. Louis, Missouri 63198, Department of Analytical Chemistry, Monsanto Corporate Research, Monsanto Company,
800 North Lindbergh Boulevard, St. Louis, Missouri 63167, and Department of Molecular Biology and Pharmacology,
Washington University School of Medicine, St. Louis, Missouri 63110
Received February 12, 1997X
A new class of antifungal agents has been discovered which exert their activity by blockade of
myristoylCoA:protein N-myristoyltransferase (NMT; EC 2.1.3.97). Genetic experiments have
established that NMT is needed to maintain the viability of Candida albicans and Cryptococcus
neoformans, the two principal causes of systemic fungal infections in immunocompromised
humans. Beginning with a weak octapeptide inhibitor ALYASKLS-NH2 (2, Ki ) 15.3 ( 6.4
µM), a series of imidazole-substituted Ser-Lys dipeptide amides have been designed and
synthesized as potent and selective inhibitors of Candida albicans NMT. The strategy that
led to these inhibitors evolved from the identification of those functional groups in the high-
affinity octapeptide substrate GLYASKLS-NH2 1a necessary for tight binding, truncation of
the C-terminus, replacement of the four amino acids at the N-terminus by a spacer group, and
substitution of the glycine amino group with an N-linked 2-methylimidazole moiety. Initial
structure-activity studies led to the identification of 31 as a potent and selective peptidomimetic
inhibitor with an IC50 of 56 nM and 250-fold selectivity versus human NMT. 2-Methylimidazole
as the N-terminal amine replacement in combination with a 4-substituted phenacetyl moiety
imparts remarkable potency and selectivity to this novel class of inhibitors. The (S,S)
stereochemistry of serine and lysine residues is critical for the inhibitory activity, since the
(R,R) enantiomer 40 is 103-fold less active than the (S,S) isomer 31. The inhibitory profile
exhibited by this new class of NMT ligands is a function of the pKa of the imidazole substituent
as illustrated by the benzimidazole analog 35 which is about 10-fold less potent than 31. The
measured pKa (7.1 ( 0.5) of 2-methylimidazole in 31 is comparable with the estimated pKa
(∼8.0) of the glycyl residue in the high-affinity substrate 1a . Groups bulkier than methyl,
such as ethyl, isopropyl, or iodo, at the imidazole 2-position have a detrimental effect on potency.
Further refinement of 31 by grafting an R-methyl group at the benzylic position adjacent to
the serine residue led to 61 with an IC50 of 40 nM. Subsequent chiral chromatography of 61
culminated in the discovery of the most potent Candida NMT inhibitor 61a reported to date
with an IC50 of 20 nM and 400-fold selectivity versus the human enzyme. Both 31 and 61a
are competitive inhibitors of Candida NMT with respect to the octapeptide substrate
GNAASARR-NH2 with Ki(app) ) 30 and 27 nM, respectively. The potency and selectivity
displayed by these inhibitors are dependent upon the size and orientation of the R-substituent.
An R-methyl group with the R configuration corresponding to the (S)-methyl-4-alanine in 2
confers maximum potency and selectivity. Structural modification of 31 and 61 by appending
an (S)-carboxyl group â to the cyclohexyl moiety provided the less potent tripeptide inhibitors
73a and 73b with an IC50 of 1.45 ( 0.08 and 0.38 ( 0.03 µM, respectively. However, these
tripeptides (73a and 73b) exhibited a pronounced selectivity of 560- and 2200-fold versus the
human NMT. More importantly 73a displayed fungistatic activity against C. albicans with
an EC50 of 51 ( 17 µM in cell culture. Compound 73b also exhibited a similar antifungal
activity. An Arf protein gel mobility shift assay for monitoring intracellular myristoylation
revealed that a single dose of 200 µM of 73a or 73b produced <50% reduction in Arf
N-myristoylation, after 24 and 48 h, consistent with their fungistatic rather than fungicidal
activity. In contrast, the enantiomer 73d which had an IC50 >1000 µM against C. albicans
NMT did not exhibit antifungal activity and produced no detectable reduction in Arf
N-myristoylation in cultures of C. albicans. These studies confirm that the observed antifungal
activity of 73a and 73b is due to the attenuation of NMT activity and that NMT represents an
attractive target for the development of novel antifungal agents.
S0022-2623(97)00094-0 CCC: $14.00 © 1997 American Chemical Society

2610 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16
Devadas et al.
In tr od u ction
MyristoylCoA:protein N-myristoyltransferase (NMT;
EC 2.1.3.97) catalyzes the cotranslational transfer of the
rare cellular fatty acid myristate (C14:0) from myris-
toylCoA to the N-terminal glycine residue of a number
of diverse eukaryotic cellular protein substrates. These
include serine/threonine and tyrosine kinases, protein
phosphatases such as calcineurin, the R subunits of
heterotrimeric G proteins, retroviral gag polyprotein
precursors such as Pr55gag of human immunodeficiency
virus I, and the capsid proteins of a number of picor-
naviruses and papovaviruses. Different proteins utilize
their covalently bound myristoyl moiety for different
purposes including regulation of protein-protein and
protein-lipid interactions.1-3
F igu r e 1. Octapeptide substrates (1a ,b) and inhibitor (2).
NMT is a cytosolic enzyme. It is encoded by a single
copy gene that appears to be represented in a broad
range of eukaryotes, e.g. Saccharomyces cerevisiae, a
variety of pathogenic fungi (Candida albicans, Crypto-
coccus neoformans, Histoplasma capsulatum), and Homo
sapiens.4-7 C. albicans NMT has been purified to
homogeneity. This monomeric protein of 451 amino acid
residues has no known co-factor requirements and
exhibits 46% primary sequence identity with the human
enzyme.7
F igu r e 2. NMT inhibitors with anti-Candida activity.
NMT has an ordered Bi-Bi mechanism.8 The apo-
enzyme first binds myristoylCoA, forming a binary
complex which results in conformational changes giving
rise to a functional peptide binding site and formation
of a myristoylCoA:NMT:peptide ternary complex. Ca-
talysis then occurs with the transfer of the myristoyl
moiety to the peptide substrate. This is then followed
by the sequential release of CoA and the acylpeptide
product.
Genetic studies have established that the NMT gene
is essential for C. neoformans,9 and C. albicans.10 To
do so, conditional lethal nmt alleles were incorporated
into their genomes by homologous recombination. These
mutant alleles encode NMTs with reduced affinity for
myristoylCoA due to substitution of a conserved glycine
near their C-terminus with aspartic acid. The mutant
nmt alleles produce temperature-sensitive growth arrest
and myristic acid auxotrophy. Removal of myristate
from the medium results in cell death, thus demonstrat-
ing that NMT is required for their viability. Moreover,
in the case of C. albicans, the myristic acid auxotrophy
was quite stable: the reversion rate was only 2.5 × 10-8
(ref 10).
These observations suggested that fungal NMTs may
serve as a target for the design and development of a
new class of fungicidal agents for treating systemic
fungal infections in the rapidly expanding population
of immunocompromised patients.11-13 Subsequent de-
sign of inhibitors was guided by the observation that,
although the acylCoA binding sites of orthologous NMTs
are highly conserved throughout the course of eukary-
otic evolution, their peptide substrate specificities have
diverged.5,9,12,14,15 This difference in substrate specificity
among NMTs was exploited to develop the first potent
and species-selective peptidomimetic inhibitors of the
acyltransferase. A high-affinity octapeptide substrate
GLYASKLS-NH2, 1a , was initially identified (Figure 1,
Km ) 0.07, 0.3, and 0.7 µM for S. cerevisiae,16 C.
albicans,17 and human15 NMT, respectively). The oc-
tapeptide sequence was derived from the N-terminal
fragment of Arf2p (ADP ribosylation factor 2), a yeast
protein that must be myristoylated by NMT for expres-
sion of its essential biological function.18 Next, a simple
substitution of alanine for glycine in 1a provided an
inhibitor 2 which was competitive for peptide (Ki ) 15.3
( 6.4 µM) and uncompetitive for myristoylCoA (Ki )
31.2 ( 0.7 µM).16,17 Further in vitro kinetic studies
using purified C. albicans NMT and a panel of peptides
having single alanine substitutions at each position of
1a revealed that the primary amino group of glycine,
the hydroxyl group of serine at position 5, and the
ꢀ-amino group of lysine at position 6 are the important
components involved in recognition by the NMT:myris-
toylCoA binary complex.17 Incorporation of these key
structural motifs into new scaffolds has generated a
series of potent and selective NMT inhibitors as exem-
plified by 31 and 61.12,13 This paper describes their
complete synthesis, structure-activity relationship (SAR)
studies, and further structural modifications which led
to the discovery of related tripeptide NMT inhibitors
73a and 73b (Figure 2) with a mechanism-based anti-
fungal activity.
Ch em istr y
N-Linked imidazole-substituted Ser-Lys dipeptide
amides with varying alkyl chain length between the N-1
of imidazole and Ser were synthesized according to the
routes shown in Scheme 2. The dipeptide intermediate
4 was conveniently synthesized as outlined in Scheme
1 from the commercially available lysine derivative 3b.
The ω-imidazol-1-ylalkanoic acids 5-719a were activated
as hydroxy benzotriazole esters and coupled with the
protected amine 4 in dimethylformamide. The resulting
products were deprotected by catalytic hydrogenation
to afford 8-10. The synthesis of analog 13 was achieved
† Presented in part at the 212th National Meeting of the American
Chemical Society, Orlando, FL, August 24-29, 1996.
* Author for correspondence.
‡ Monsanto Company.
§ Washington University School of Medicine.
X Abstract published in Advance ACS Abstracts, J uly 1, 1997.

Inhibitors of Candida albicans NMT
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16 2611
Sch em e 1. Synthesis of Ser-Lys Dipeptide Amine Precursora
a
Reagents: (a) EDC, HOBt, 2-cyclohexylethylamine, DMF, room temperature, 16 h; (b) 4 N HCl/dioxane, room temperature, 4 h; (c)
BOC-Ser(OBn)OSu, DMF, Et3N, room temperature, 16 h.
Sch em e 2. Synthesis of Imidazole-Linked NMT Inhibitors with Varying Chain Lengthsa
a
Reagents: (a) HOBt, EDC, Et3N, DMF, 30 min, room temperature; (b) H2, Pd/C (10%), MeOH, HCl, room temperature, 18 h, 40 psi;
(c) DCC, HOBt, CH2Cl2-DMF, 0 °C to room temperature; N-methylmorpholine; (d) trifluoroacetic acid, room temperature, 3 h.
according to Scheme 2 by condensing the acid 1119b with
the protected amine 1212 via formation of the benzo-
triazole active ester using DCC and hydroxybenzotria-
zole, followed by deprotection of the crude product with
trifluoroacetic acid, and purification by reverse-phase
(C18) HPLC.
ated from 12, using DCC/HOBt. The crude products
after reaction workup were treated with trifluoroacetic
acid and purified by reverse-phase HPLC to afford the
final compounds 30-35 as trifluoroacetate salts.
Synthesis of 36 was accomplished using a procedure
similar to that described above, by condensing 29 with
the amine 12 employing DCC/HOBt. The crude product
was treated with trifluoroacetic acid and purified by
reverse-phase HPLC to obtain 36.
The (R,R) enantiomer 40, which is the optical isomer
of 31, was synthesized by solid-phase methodology22
according to Scheme 4, starting from the commercially
available protected R-lysine derivative 37 linked to
Merrifield’s resin. The resin-bound lysine derivative 37
was treated with 50% trifluoroacetic acid in dichlo-
romethane. It was then washed with diisopropylethy-
lamine (DIEA) to generate the free amine and coupled
to BOC-D-Ser(OBn)-OH with the aid of DCC/HOBt in
dimethylformamide at room temperature to obtain 38.
Exposure of 38 to 50% trifluoroacetic acid in dichlo-
romethane for 30 min, followed by formation of the free
amine using DIEA, and coupling with the activated
2-methylimidazole-substituted phenylacetic acid 24 us-
ing DCC/HOBt, gave the resin-bound 2-methylimidazole
derivative 39. The final compound 40 was released from
the resin by heating 39 with 2-cyclohexylethylamine at
Scheme 3 outlines a general methodology for prepar-
ing Ser-Lys dipeptides with imidazole, 2-substituted
imidazole, and 2-methylbenzimidazole bearing a 4-sub-
stituted phenacetyl ring. The commercially available
4-iodophenylacetic acid was esterified in the presence
of methanolic HCl to obtain the corresponding methyl
ester 14. Reaction of the ester 14 with butyn-1-ol in
the presence of triethylamine, PdCl2(PPh3)2, and CuI20
yielded the acetylenic product 15 which, after catalytic
reduction of the triple bond, provided the 4-hydroxybu-
tyl-substituted phenylacetate ester 17. The hydroxy
ester 17 was then converted to the iodo derivative 19
using (PhO)3PMeI and subjected to alkaline hydrolysis
to obtain the iodo acid 21 after acidification. Condensa-
tion of the acid 21 with imidazole, 2-methylimidazole,
2-iodoimidazole,21 2-ethylimidazole, 2-isopropylimida-
zole, or 2-methylbenzimidazole in the presence of NaH
in DMF or THF yielded the corresponding imidazole-
linked phenylacetic acids (23-28), respectively. These
acids were individually coupled with the amine gener-

2612 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16
Devadas et al.
Sch em e 3a
a
Reagents: (a) Alkyn-1-ol, PdCl2P(Ph3)2, CuI, Et3N, CH3CN, room temperature, 3 h; (b) H2, Pd/C (5%), 40 psi, room temperature, 4 h;
(c) (PhO)3PMeI, CH3CN, room temperature, 4 h; (d) 1 M LiOH, MeOH-H2O, room temperature, 2.5 h; (e) RNa, DMF, 18-crown-6, 60 °C,
2.5 h; (f) HCl; (g) DCC/HOBt, DMF + 12, N-methylmorpholine; (h) trifluoroacetic acid, room temperature, 3 h.
Sch em e 4. Solid-Phase Synthesis of the (R,R) Isomera
a
Reagents: (a) 50% TFA/CH2Cl2, 30 min, room temperature; (b) DIEA; (c) BOC-D-Ser(OBn)-OH, DCC, 1 h; (d) 24, DCC, HOBt, DMF,
18 h; (e) 2-cyclohexylethylamine, 50 °C, 5 h; (f) 90% HF/anisole, 0 °C 1 h.
50 °C for 5 h, treating the resin-free material with 90%
HF-anisole at 0 °C for 1 h, and purification by reverse-
phase HPLC.
The benzylic R-substituted imidazole compounds 61-
65 were prepared as illustrated in Scheme 5 starting
from methyl 4-iodophenylacetate 14. Synthesis of 61
began with alkylation of 14 with methyl iodide in the
presence of NaH in tetrahydrofuran to afford the
racemic propionate ester 41. The ester was then
coupled with 3-butyn-1-ol using PdCl2(PPh3)2, CuI, and
triethylamine. The resulting acetylenic product 44 was
purified and subjected to catalytic reduction to obtain

Inhibitors of Candida albicans NMT
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16 2613
Sch em e 5a
a
Reagents: (a) NaH, THF, room temperature, 16 h; (b) 3-butyn-1-ol, PdCl2(Ph3)2, CuI, Et3N, CH3CN, room temperature, 3 h; (c) H2,
Pd/C (5%), 40 psi, room temperature, 4 h; (d) (PhO)3PMeI, CH3CN, room temperature, 4 h; (e) 1 M LiOH, MeOH-H2O, room temperature,
2.5 h; (f) RNa, DMF, 18-crown-6, 60 °C, 2.5 h; (g) DCC/HOBt, DMF + 12, N-methylmorpholine; (h) trifluoroacetic acid, room temperature,
3 h.
the hydroxy ester 47. Treatment of 47 with (PhO)3PMeI,
followed by hydrolysis of the ester 50 and acidification,
yielded the acid 53. In the following step, 53 was
condensed with sodium 2-methylimidazolide in dimeth-
ylformamide to obtain the corresponding imidazole-
substituted carboxylic acid 56. Subsequent activation
of 56 using DCC/HOBt and coupling with the free amine
generated from 12, followed by deprotection and puri-
fication by reverse-phase HPLC, provided 61 as a
diastereomeric mixture. The (R,S)-ethyl-62, (R,S)-pro-
pyl-63, and (R,S)-2-chloroimidazole 64 and (R,S)-2-
methylbenzimidazole 65 analogs were synthesized
using a similar reaction sequence as that outlined in
Scheme 5.
and (S)-phenylpropionic acids 66 and 67 were individu-
ally coupled with the free amine generated from 12,
using DCC/HOBt. The resulting products were treated
with trifluoroacetic acid, and the dipeptides 68 and 69
were isolated by reverse-phase chromatography.
Scheme 7 illustrates the synthesis of tripeptides 73a -
73c starting from the differentially protected lysine
derivative 70. Coupling of 70 with â-cyclohexylalanine
methyl ester using DCC/HOBt provided the dipeptide
ester 71. The carbobenzoxy group in 71 was removed
by catalytic hydrogenation, and the resulting amine was
condensed with the 2-methylimidazole derivative 24
using DCC/HOBt to afford the tripeptide 72. The ester
group in 72 was converted to the corresponding acid
with 1 M LiOH followed by acidification, and the
resulting crude product was treated with trifluoroacetic
acid. The final compound 73a was isolated by reverse-
phase HPLC purification. The racemic R-methyl analog
73b was prepared and purified in a similar manner as
described for 73a , using the imidazole acid 56 instead
of 24. The tripeptide ester 73c was prepared by
coupling the free amine generated from 72 with the acid
24 via activation using DCC/HOBt, followed by depro-
tection of the crude product with trifluoroacetic acid and
HPLC purification.
Chiral separation of 61 was achieved by HPLC
(Figure 3) using a chirobiotic-V column packed with the
chiral selector vancomycin linked to silica gel23 as the
stationary phase and eluting with 23% acetonitrile in
ammonium phosphate buffer at pH 4.2 to afford the
corresponding diastereomers 61a and 61b having >95%
1
purity as indicated by their H-NMR spectra.
Reference dipeptides 68 and 69, which enabled as-
signment of absolute configuration at the new chiral
center in diastereomers 61a and 61b, were prepared
according to Scheme 6. The commercially available (R)-

2614 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16
Devadas et al.
F igu r e 3. Chiral separation of 61.
Sch em e 6. Synthesis of Reference Peptidesa
a
Reagents: (a) DCC, HOBt, 0 °C 1 h, N-methylmorpholine, room temperature, 16 h; (b) trifluoroacetic acid, room temperature, 3 h.
Sch em e 7. Synthesis of Imidazole-Substituted Tripeptide NMT Inhibitorsa
a
Reagents: (a) DCC, HOBt, dimethylacetamide, CH2Cl2, 0 °C, 1 h; (b) L-cyclohexylalanine methyl ester hydrochloride, N-
methylmorpholine, room temperature, 16 h; (c) H2, 5% Pd/C, room temperature, 1 h; (d) Z-Ser(OtBu)-OH, DCC, HOBt, CH2Cl2,
dimethylacetamide, 0 °C, 1.5 h, room temperature, 48 h; (e) 24, DCC, HOBt, 0 °C, 1.5 h, room temperature, 24 h, (f) trifluoroacetic acid,
room temperature, 3 h; (g) 56, DCC, HOBt, 0 °C, 1.5 h, room temperature, 24 h; (h) 2 M LiOH, dioxane, H2O, room temperature, 2 h,
HOAc.
The enantiomer 73d was synthesized using solid-
phase methods, according to Scheme 8. The first step
involved the attachment of BOC-protected D-cyclohexyl-
alanine to the polystyrene resin using DCC to afford
74. Cleavage of the BOC group in 74 with 50%
trifluoroacetic acid, followed by generation of the free
amine and condensation with the BOC-protected D-
lysine derivative via activation using DCC, gave the
dipeptide 75 linked to the resin. This reaction sequence
was repeated on 75 to attach the BOC-protected D-
serine to the lysine residue on the resin to obtain 76.
In a similar fashion using 76, the 2-methylimidazole-

Inhibitors of Candida albicans NMT
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16 2615
Sch em e 8. Solid-Phase Synthesis of the (R,R,R) Isomera
a
Reagents: (a) Boc-â-cyclohexyl-D-Ala, DCC, 1 h; (b) 50% TFA/CH2Cl2, 30 min, room temperature; (c) DIEA, CH2Cl2; (d) Boc-D-Lys(2-
Cl-Cbz), DCC, 1 h; (e) BOC-D-Ser(OBn)-OH, DCC, 1 h; (f) 24, DCC, HOBt, DMF, 18 h; (g) 90% HF/anisole, 0 °C, 1 h; (h) reverse-phase
(C18) HPLC, 5-55%.
carboxylic acid 24 was coupled to the serine residue on
the resin. The peptide product was simultaneously
deprotected and cleaved from the resin and purified by
reverse-phase HPLC to yield the tripeptide 73d .
En zym e Assa y
The final compounds were evaluated for the inhibition
F igu r e 4. Dipeptide inhibitor of Candida albicans NMT.
of recombinant human and Candida NMTs as described
previously.12 Both enzymes were expressed in E. coli
and were purified to apparent homogeneity according
wells. Controls included DMSO alone and amphotericin
B (final concentration ) 0.05, 1, and 5 µg mL-1; positive
to protocols described in ref 24. Standard solutions of
control). Plates were incubated at 30 °C, and growth
inhibitors were prepared at 22 mM in 0.5% DMSO. The
was scored by monitoring the OD490 using an ELISA
IC50 determinations (concentration required for 50%
plate reader (Dynatech). EC50, the concentration of the
compound inhibiting the growth by 50% compared to
untreated controls, was determined at 24 h. All experi-
ments were done in triplicate on at least three occasions.
Within a given experiment, the triplicate determina-
tions varied by <10% per sample.
inhibition) involved incubation of the compound at a
series of known concentrations with C. albicans NMT
in the presence of 0.11 nmol of [3H]myristoyl CoA (1 µCi,
9.09 Ci/mmol) and 2.2 nmol of GNAASARR-NH2 in a
total volume of 60 µL. After a 10 min incubation at 24
°C, the reaction was quenched by addition of ice-cold
methanol. The products were separated by HPLC on a
Vydac-C4 column and quantitated by in-line scintillation
Resu lts a n d Discu ssion
counting.
Our initial structural modification of 1a by replacing
the amino acid residues at positions 1-4 with an 11-
aminoundecanoyl moiety, and substituting the C-ter-
minal Leu-Ser-carboxamide with a cyclohexylethyl group,
led to the identification of the dipeptide amide lead 3a
(Figure 4) with good activity against C. albicans NMT
(IC50 ) 0.11 ( 0.03 µM) and little selectivity (7-fold)
versus the human enzyme.12,26 Unfortunately no anti-
fungal activity was detected for 3a in cell cultures even
at concentrations of 100 µM.26 This may be a conse-
quence of inadequate inhibitor potency or ineffective
cellular penetration due to the presence of two basic
(pKa ∼10) primary amine groups in 3a . To overcome
this problem, efforts were directed toward finding a less
basic replacement for the N-terminal amino group. An
N-linked imidazole in place of the N-terminal amino
group was one of the choices investigated since the pKa
The reported IC50s represent an average of two
independent determinations each done in duplicate. Ki’s
are reported as apparent inhibition constants and were
determined as described in ref 12.
An tifu n ga l Assa y
NMT inhibitors were tested for anti-Candida activity
according to the following protocol.25 Briefly, 20 mM
stock solutions of the inhibitors were prepared in DMSO
(1% v/v) and diluted into sterile deionized water to
obtain twice the final desired concentration for the
cultured-based assay. Portions (100 µL) of the stock
solutions were dispensed into triplicate wells of a 96-
well microtiter plate (Costar). C. albicans strain B311
[a 100 µL portion containing 105 cells in 2 × yeast
nitrogen broth (YNB)] was distributed into each of the

2616 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16
Devadas et al.
Ta ble 1. Potency of Imidazole-Substituted NMT Inhibitors
the potencies of 13 (IC50 ) 1.55 ( 0.07 µM) and 30
indicated that introduction of conformational restric-
tions in the flexible alkyl chain linker confers about
5-fold improvement in potency and a 10-fold increase
in selectivity versus the human NMT (IC50 ) 16 ( 2.97
µM). The analog 30 is the first example of a competitive
peptidomimetic NMT inhibitor (Ki ) 0.59 ( 0.07 µM)
having an amphoteric group at the N-terminus and a
potency below 1 µM against the C. albicans enzyme.
A further improvement in potency was realized by
replacing the imidazole with a more basic 2-methylimid-
azole as exemplified by 31. It is remarkable that a
seemingly minor structural modification of 30, i.e.
introduction of a methyl group into an imidazole ring,
provided both enhanced potency (IC50 ) 0.056 ( 0.01
µM, C. albicans NMT; and 0.035 ( 0.008 µM, S.
cerevisiae NMT) and an additional 10-fold improvement
in selectivity (250-fold versus human NMT, Table 2).
This may be attributable to the measured pKa of the
2-methylimidazole (7.1 ( 0.5), which is about 3 pH units
lower than the measured pKa (∼10.0) of the N-terminal
amine group in 3a and comparable to the estimated pKa
(7.7) of alanine in 2. Kinetic analysis indicated that 31
is a competitive inhibitor (Ki ) 0.031 ( 0.003 µM) with
respect to the peptide substrate GNAASARR-NH2.
with Varying Chain Lengthsa
IC50, µM
compd
n
C. albicans NMT
human NMT
8
9
10
13
8
5.5 ( 1.3
3.0
3.0
ND
ND
ND
11.35 ( 0.49
9
11
10
1.55 ( 0.07
a
Potency against the indicated NMT as assessed by IC50 using
the peptide GNAASARR-NH2 at its apparent Km (see text) and
myristoylCoA at 1 µM. ND ) not determined.
Ta ble 2. Potency and Selectivity of Imidazole-Substituted
NMT Inhibitorsa
IC50, µM
compd
n
R
C. albicans NMT human NMT selectivityb
36
30
31
33
34
32
40
5
4
4
4
4
4
4
H
H
1.2
ND
ND
51
Replacement of the 2-methyl group in 31 by elec-
tronegative groups which would lower the pKa by 2-3
units,29 such as iodo, to obtain 32 (IC50 ) 1.9 ( 0.03
µM) markedly reduced potency by 36-fold. A similar
trend was observed when the 2-methyl group was
replaced by bulkier groups such as ethyl or isopropyl,
as illustrated by analogs 33 and 34. Substitution of
2-methylimidazole with less basic heterocycles such as
2-methylbenzimidazole to obtain 35 (IC50 ) 0.6 ( 0.16
µM) also led to diminished potency.
0.31 ( 0.06
0.056 ( 0.0
0.147 ( 0.01
0.2 ( 0.04
1.88 ( 0.03
62.4 ( 13
16 ( 2.97
14.1 ( 4.25
28.2
>100
ND
CH3
C2H5
i-C3H7
I
252
192
ND
ND
ND
CH3
ND
a
Potency against the indicated NMT as assessed by IC50 using
the peptide GNAASARR-NH2 at its apparent Km (see text) and
b
myristoylCoA at 1 µM. Selectivity is the ratio of the IC50 for
human NMT divided by the IC50 for C. albicans NMT. ND ) not
determined.
To further augment the inhibitory activity of NMT
inhibitors, we sought additional structural features that
might impart higher binding affinity. A comparision
of Km(apparent) values of the octapeptide substrates 1a and
1b revealed that substitution of alanine-4 in 1a by
glycine to give 1b resulted in a 10-fold increase in Km.17
This observation suggested that the side chain methyl
of alanine-4 in 1a also contributes to binding. Thus,
analogs of 31 with an R-alkyl substiutent R1 at the
benzylic position adjacent to serine might provide the
conformational bias present in 1a and consequently
exhibit higher potencies. This hypothesis set the stage
for synthesis of a series of benzylic R-substituted imid-
azole compounds 61-65.
The diastereomers 61a and 61b were isolated by
chiral chromatography as depicted in Figure 3. The
assignment of absolute configuration at the benzylic
chiral centers was established by comparing the 1H-
NMR spectra of 61a and 61b with those of reference
compounds 68 and 69. The methyl group in (S,S,S) 69
displayed a characteristic downfield shift when com-
pared with the corresponding (R,S,S) diastereomer 68.
A similar downfield shift of the methyl signal was also
observed in 61b when compared with its diastereomer
61a . Furthermore, the proton chemical shifts of the
R-methine protons for the serine and lysine residues in
61a and 61b, as confirmed by proton-COSY experi-
ments, were identical with the corresponding proton
chemical shift values observed for 68 and 69.
of imidazole (∼7.5) is comparable with that of alanine
or glycine.27 A brief structure-activity (SAR) study that
varied the connecting chain length between the N-1
nitrogen of imidazole and the serine residue (Table 1)
revealed that an 11-carbon atom linker, as exemplified
by 13, conferred maximum potency (IC50 ) 1.55 ( 0.07
µM) for C. albicans NMT. It is pertinent to note that
analogs of 3a containing a much less basic nitrogen
heterocycle such as 1,2,4-triazole, or more basic func-
tional groups such as guanidines or piperazines in place
of imidazole, were significantly less effective NMT
inhibitors.28
At this stage, further enhancements to this series to
attain potencies below 1 µM presented a synthetic
challenge. We reasoned that the activity of these
inhibitors might be improved by introducing conforma-
tional constraints in the flexible linear linker chain
using a 1,4-disubstituted phenyl ring. A recent study
had demonstrated that incorporation of a rigidifying
element such as 1,4-phenylene moiety â to the unde-
canoyl carbonyl group in 3a was well tolerated by the
enzyme.26 As the distance between the ω-amino group
and undecanoyl carbonyl functionality is critical for
molecular recognition, imidazole compounds 30-36
were synthesized with four or five methylene groups
connecting the imidazole nitrogen with the aromatic
ring. Comparision of the IC50s (Table 2) of 30 (IC50
)
0.31 ( 0.06 µM) and its homolog 36 (IC50 ) 1.2 µM)
suggested that placing the imidazole moiety four me-
thylene units from the phenyl ring resulted in a 4-fold
enhancement in potency. Furthermore, examination of
Analogs 61-65, 61a , and 61b were evaluated for
inhibition of C. albicans and human NMT (Table 3). The

Inhibitors of Candida albicans NMT
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16 2617
Ta ble 3. Potency and Selectivity of Benzylic-Substituted NMT
Substitution of the 2-methylimidazole in 61 with less
basic groups such as 2-chloroimidazole (64) or 2-meth-
ylbenzimidazole (65, pKa(est) ) 6.3) resulted in signifi-
cantly reduced potency (Table 3). In the case of the
2-chloro analog, although the size of the halo group is
comparable to that of the 2-methyl group, the inductive
effect is likely to attenuate the pKa of the imidazole ring
which appears detrimental to inhibitory activity.29
These results suggest that the protonation of the imi-
dazole ring is essential for tight binding of these
peptidomimetic ligands to the peptide binding site
provided by the myristoylCoA:NMT binary complex.
All of the NMT inhibitors described above were
evaluated for their antifungal activity in a cell culture
assay. Surprisingly none of the final compounds dis-
played any growth inhibitory activity (EC50 > 100 µM)
against C. albicans (or C. neoformans). Consequently,
our synthetic efforts were focused on improving the cell-
penetration properties of these NMT inhibitors by
installing a carboxyl group â to the cyclohexyl moiety.
This approach resulted in the synthesis of tripeptide
analogs 73a and 73b. Both these analogs were moder-
ate inhibitors of C. albicans NMT [IC50 ) 1.45 ( 0.08
and 0.38 ( 0.03 µM, respectively (Table 4)]. However,
both displayed a dramatically enhanced selectivity of
560- and 2200-fold, respectively, toward the C. albicans
NMT, an unexpected “beneficial” effect of introducing
the carboxyl group (Table 4). More importantly, com-
pound 73a exhibited growth inhibitory activity (Table
4) with an EC50 of 51 ( 17 µM, 24 h, after a single dose
administration to cultures of a well-characterized am-
photericin B-sensitive and fluconazole-sensitive clinical
isolate of C. albicans (strain B311, ref 25). The com-
pound had no growth inhibitory activity against C.
neoformans (EC50 >100 µM).
An assay has been developed recently to test the
effects of in vitro inhibitors of purified NMT on cellular
NMT activity in exponentially growing cultures of C.
albicans.25 The assay takes advantage of the fact that
one of the most prominent C. albicans N-myristoylpro-
teins is an Arf whose mobility during SDS-polyacryl-
amide gel electrophoresis is greater when it is N-
myristoylated than when it is not. The ratio of N-
myristoylated to nonmyristoylated Arf can be taken as
a measure of cellular NMT activity. This ratio is
defined by Western blot analysis of total cellular pro-
teins isolated prior to and after exposure to an enzyme
inhibitor. Studies of C. albicans strains with conditional
lethal NMT gene mutations indicate that a reduction
in the level of N-myristoylated Arf to e50% of total Arf
is associated with growth arrest and death.25 An Arf
protein gel mobility shift assay indicated that a single
200 µM dose of 73a or 73b produces a <50% reduction
in Arf N-myristoylation after 4 h, which is consistent
with their observed fungistatic but not fungicidal activ-
ity.25
Inhibitorsa
IC50, µM
C. albicans
human
NMT
compd
R1
NMT
selectivityb
61
CH3 (racemic)
CH3 (R)
CH3 (S)
C2H5 (racemic)
n-C3H7 (racemic) 0.130 ( 0.01
0.04 ( 0.003
0.02 ( 0.001
0.31
8 ( 0.42
200
410
160
37
4
69
61a
61b
62
63
64
8.2
49
0.042 ( 0.013
1.54 ( 0.01
0.49 ( 0.01
142.0
119.5 ( 31.8
CH3 (racemic)
CH3 (racemic)
2.05 ( 0.49
0.86 ( 0.48
65
139
a
Potency against the indicated NMT as assessed by IC50 using
the peptide GNAASARR-NH2 at its apparent Km and myristoylCoA
b
at 1 µM. Selectivity is the ratio of the IC50 for human NMT
divided by the IC50 for C. albicans NMT.
F igu r e 5. Double-reciprocal plot of 1/V vs 1/[GNAASARR-
NH2] for inhibitor 61a with C. albicans NMT. GNAASARR-
NH2 (Km ) 20 µM) concentrations of 5, 10, 20, 40, and 80 µM
were used with a fixed [3H]myristoylCoA concentration of 1
µM (9.09 Ci/mmol) and 15 ng of NMT. The secondary plot of
slopes vs 1/[61a ] yielded a Ki(app) of 27 ( 7 nM. The data are
plotted as the mean of the reciprocal velocity ( the standard
deviation of the triplicate measurements. Error bars that
would obscure data points are omitted for clarity.
SAR data suggest that the observed potency and selec-
tivity are a function of the size and orientation of the
R-substituent at the benzylic position. While the R-
methyl group in 61 delivers maximum potency (IC50
)
0.04 ( 0.003 µM, C. albicans NMT; and 0.017 µM, S.
cerevisiae NMT) and selectivity versus 31, the incorpo-
ration of a bulkier group such as n-propyl (63) results
in dramatically reduced potency against C. albicans
NMT and enhanced activity against human enzyme.
Inclusion of an R-ethyl group (62) retains potency
comparable to that of the R-methyl group against the
fungal NMT, but its selectivity is reduced significantly.
A detailed analysis of the inhibitory profile of indi-
vidual isomers revealed that the (R,S,S) diastereomer
61a is a highly potent inhibitor with an IC50 of 20 (
0.99 nM for C. albicans NMT and exhibits 400-fold
selectivity versus the human enzyme (Table 3). The
corresponding (S,S,S) isomer 61b is 15-fold less potent
(IC50 of 310 nM against the fungal enzyme) and “only”
150-fold selective versus human NMT. A more detailed
enzyme kinetic analysis of 61a revealed it to be a
competitive inhibitor (Ki ) 27 ( 7 nM, Figure 5) with
respect to the peptide substrate GNAASARR-NH2.
Importantly, the (R)-methyl group in 61a , which cor-
responds to the (S)-4-alanine methyl configuration in
1a , confers decreased potency against human NMT
while increasing it against the fungal enzyme.
The racemic R-methyl analog 73b is 4-fold more
potent and exhibits similar anti-Candida activity as 73a
(Table 4). Like 73a , 73b has no growth inhibitory
effects on C. neoformans (EC50 > 100 µM). It is
pertinent to note that the tripeptide ester 73c which is
about 18-fold more potent than 73a did not display any
antifungal activity against either organism. This find-
ing suggests that installing a carboxyl group â to the
cyclohexyl moiety promotes cellular uptake of NMT
inhibitors. The (R,R,R) enantiomer (73d ) which had an
IC50 of >1000 µM against C. albicans NMT had no

2618 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16
Devadas et al.
Ta ble 4. Biological Activity of Imidazole-Substituted Tripeptide NMT Inhibitorsa
IC50, µM
EC50, µMc
compd
C. albicans NMT
human NMT
selectivityb
24 h, C. albicans
24 h, C. neoformans
73a
73b
73c
73d
1.45 ( 0.08
0.38 ( 0.03
0.081 ( 0.01
>1000
809.5 ( 33.2
840 ( 28.3
22.7 ( 2.7
>1000
560
2200
276
51 ( 17
33
>100
>100
>100
>100
>100
>100
a
Potency against the indicated NMT as assessed by IC50 using the peptide GNAASARR-NH2 at its apparent Km and myristoylCoA at
1 µM. Selectivity is the ratio of the IC50 for human NMT divided by the IC50 for C. albicans NMT. c See text.
b
purification. L-Amino acid derivatives were purchased from
growth inhibitory effect against C. albicans (or C.
neoformans) and did not produce any detectable reduc-
tion in C. albicans Arf N-myristoylation.
Sigma. Melting points were determined with a Melt-Temp
apparatus and are uncorrected. All reactions were performed
under anhydrous conditions in an atmosphere of argon.
Nuclear magnetic proton and carbon-13 spectra were recorded
on a Varian XL-300 spectrometer, and chemical shifts (δ) are
reported in ppm relative to tetramethylsilane. Splitting
patterns are designated as follows: s, singlet; d, doublet; t,
triplet; q, quartet; m, multiplet; br, broad peak. Low-resolution
mass spectra were recorded on a VG40-250T instrument, and
high-resolution mass spectra were recorded on a Finnigan
MAT 90 mass spectrometer operating in the FAB mode, unless
otherwise noted. Amino acid analyses were performed follow-
ing hydrolysis in 6 N HCl at 150 °C in vacuo for 1.5 h using a
Beckman 6300 high-performance analyzer. The purity of all
the reported compounds was found to be >96% by 1H-NMR
and HPLC techniques.
Analytical reverse-phase high-performance liquid chroma-
tography (HPLC) was carried out on a Waters Delta-Pak
cartridge (C-18, 8 × 100 mm), using a linear gradient of (A)
water containing 0.05% trifluoroacetic acid and (B) acetonitrile
containing 0.05% TFA, at a flow rate of 1 mL/min. The elution
was carried out with a linear gradient from 5 to 70% of B in
30 min, and the separation was monitored by UV absorbance
at 215 nm.
Together, these data confirm that the antifungal
activity exhibited by peptidomimetic inhibitors 73a and
73b is a consequence of attenuation of intracellular C.
albicans NMT activity. The fungistatic instead of
fungicidal activity of 73a may be attributed to its
observed susceptibility to degradation by cellular
carboxypeptidases: repeated addition of 73a to cultures
produces a more prolonged growth suppressive effect.
The antifungal effects of 73a and 73b are species-
specific. The lack of broad spectrum activity in these
compounds may reflect a number of mechanisms in-
cluding, for example, differences in their uptake by C.
albicans and C. neoformans, differences in their intra-
cellular stabilities or compartmentalization within these
organisms, and/or differences in their recognition by the
peptide binding sites of the two fungal NMTs.
Con clu sion s
We have discovered a new class of potent and selective
imidazole-substituted peptidomimetic inhibitors of C.
albicans NMT, starting from a weak octapeptide inhibi-
tor ALYASKLS-NH2 2. Remarkably, 2-methylimidazole
effectively mimics the pKa of the alanine residue in 2
and confers high affinity and selectivity in combination
with a phenacetyl moiety. We have also identified a
chiral recognition element which led to the discovery of
the most potent C. albicans NMT inhibitor 61a reported
to date. The high affinity and selectivity manifested by
31 and 61a toward fungal NMTs are notable since the
human enzyme recognizes a much wider variety of
protein substrates than its fungal counterpart. Fur-
thermore, we have demonstrated by synthesis and
chiral separation that incorporation of a methyl group
with (R) stereochemistry provides a favorable confor-
mational bias and results in a high-affinity ligand for
the fungal enzyme. The synthetic protocols described
above allow preparation of a variety of heteroatom or
heterocyclic substituted 4-phenylacetic acid analogs. The
solid-phase methodology adds synthetic diversity and
makes a combinatorial approach feasible as a future
alternative. The conceptual framework and the syn-
thetic efforts have led to the identification of selective
Candida NMT inhibitors 73a and 73b with a mecha-
nism-based antifungal activity. Incorporation of a car-
boxyl moiety â to the cyclohexyl group results in a dual
affect of enhancing selectivity and imparting antifungal
activity. The SAR studies reported in this paper should
serve as a basis for future efforts to develop nonpeptidal
and more potent inhibitors of NMT as novel antifungal
agents.
Final compounds were purified by reverse-phase HPLC
using a Waters Delta-Pak cartridge (C-18, 40 × 100 mm, 15
µm) and eluting with a linear gradient consisting of 5-70% of
B in 30 min. The flow rate was adjusted to 70 mL/min, and
the separation was monitored by UV absorbance at 215 nm.
The appropriate fractions were pooled and freeze-dried, and
the products were isolated as TFA salts.
Solid-phase peptide synthesis was carried out on polystyrene
resin using BOC-protected amino acid derivatives purchased
from Nova-Biochem.
Ser (OBn )-Lys(Cb z)-2-cycloh exylet h yla m id e H yd r o-
ch lor id e (4). A mixture of the lysine derivative 3b (6.0 g,
0.0158 mol), HOBt (2.07 g, 0.0158 mol), and EDC (3.03 g,
0.0158 mol) in dry DMF (40 mL) was stirred at room temper-
ature. After 1 h, a solution of 2-cyclohexylethylamine (2.01 g,
0.0158 mol) in dry DMF (10 mL) was added, and the mixture
was stirred at room temperature for 18 h. The reaction
mixture was diluted with dichloromethane (400 mL),washed
sequentially with saturated NaHCO3 (2 × 200 mL) and brine
(2 × 200 mL), and dried (MgSO4). Removal of the solvent
afforded 6.0 g (78%) of 3c as a white solid: 1H-NMR (CDCl3)
δ 7.31-7.34 (m, 5H), 6.15 (s, 1H), 5.28 (s, 2H), 4.90 (s, 1H),
3.17-3.26 (m, 4H), 0.86 -1.84 (m, 28H); HRMS (EI) m/ z calcd
for C27H43N3O5 489.3203(M+), found 489.3208.
This lysine derivative 3c (6.0 g, 0.012 mol) was stirred with
4 N HCl in dioxane (30 mL) for 1 h at room temperature. The
reaction mixture was concentrated under reduced pressure,
and the residue was dried in vacuo to afford the hydrochloride
salt (5.75 g) as a pale yellow solid: 1H-NMR (CD3OD) δ 7.33-
7.34 (m, 5H), 5.06 (s, 2H), 3.74 (m, 2H), 3.31 (m, 4H), 0.90-
1.89 (m, 19H); HRMS (EI) m/ z calcd for C22H35N3O3 389.2678
(M+), found 389.2658. This material was used without puri-
fication in the following step.
A mixture of Boc-Ser(OBn)-OH (3.81 g, 0.013 mol), HOBt
(1.70 g, 0.013 mol), and EDC (2.47 g, 0.013 mol) in dry DMF
(40 mL) was stirred at room temperature. After 1 h, a solution
of the hydrochloride salt (5.5 g, 0.013 mol) obtained as above
from 3c in dry DMF (20 mL), and triethylamine (1.30 g, 0.013
mol) were added, and the mixture was stirred at room
temperature for 18 h. The reaction mixture was diluted with
Exp er im en ta l Section
Unless otherwise stated, starting materials were obtained
from commercial suppliers and were used without further

Inhibitors of Candida albicans NMT
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16 2619
dichloromethane (400 mL) and washed successively with
saturated NaHCO3 (2 × 200 mL) and brine (2 × 200 mL) and
dried (MgSO4). After removal of the solvents under reduced
pressure, the resulting pale yellow solid (6.63 g) was stirred
with 4 N HCl in dioxane (30 mL) for 1 h at room temperature
and concentrated under reduced pressure. The resulting
hydrochloride salt was dried in vacuo to give 6.27 g of 4 as a
pale yellow solid which was used without purification: 1H-
NMR (CD3OD) δ 7.29-7.36 (m, 10H), 5.06 (s, 2H), 4.60 (s, 2H),
4.29-4.34 (m, 1H), 4.09-4.13 (m, 1H), 3.65-3.88 (m, 2H),
3.18-3.07 (m, 4H), 0.86-1.71 (m, 19H); HRMS (EI) m/ z calcd
for C32H46N4O5 566.3468 (M+), found 566.3502. Amino acid
analysis: Ser 1.00 (1.00), Lys 1.00 (1.00).
9-(1H-Im id a zol-1-yl)n on a n oyl-Ser -Lys-N-(2-cycloh ex-
yleth yl)a m id e (8). A mixture of the acid 5 (0.08 g, 0.34
mmol), HOBt (0.046 g, 0.34 mmol), and EDC (0.065 g, 0.34
mmol) in dry DMF (5 mL) was stirred at room temperature
for 40 min. It was then treated with a solution of 4 (0.2 g,
0.34 mmol) and triethylamine (0.034 g, 0.34 mmlo) in dry DMF
(2 mL), and the resulting mixture was stirred at room
temperature for 18 h. The reaction mixture was diluted with
dichloromethane (100 mL) and washed successively with
saturated NaHCO3 (2 × 100 mL) and brine (2 × 100 mL) and
dried (MgSO4). After removal of the solvent, the residue was
purified by silica gel flash chromatography using 1-4.5%
methanol in dichloromethane as the eluent to furnish 0.14 g
(52%) of the protected dipeptide as a white solid. This was
dissolved in methanol (10 mL), 1 N HCl (0.15 mL) and 5%
Pd-C (0.10 g) were added, and the resulting suspension was
stirred under H2 (45 psi) for 32 h at room temperature. The
catalyst was removed by filtration through Celite, the filtrate
was concentrated under reduced pressure, and the residue was
purified by reverse-phase HPLC. The appropriate fractions
were pooled and concentrated under reduced pressure to give
0.07 g (46.0%) of 8 as a glassy substance: 1H-NMR (CD3OD)
δ 8.95 (s, 1 H), 7.65 (d, 1 H, J ) 1.61 Hz), 7.56 (d, 1 H, J )
1.61 Hz), 4.37-4.30 (m, 2H), 4.24 (t, 2 H, J ) 7.35 Hz), 3.86-
3.81 (m, 1H), 3.74-3.64 (m, 1H), 3.18 (t, 2H, J ) 7.25 Hz),
2.92 (t, 2H, J ) 7.37 Hz), 2.26 (t, 2H, J ) 7.55 Hz), 1.97-0.85
(m, 31H); HRMS m/ z calcd for C29H52N6O4 549.4128 (M + H),
found 549.4106. Amino acid analysis: Ser 1.00 (0.97), Lys 1.00
(1.03).
10-(1H-Im id a zol-1-yl)d eca n oyl-Ser -Lys-N-(2-cycloh ex-
yleth yl)a m id e (9) was prepared in a manner similar to 8 by
coupling 6 with 4 in 42% yield as a white glassy substance:
1H-NMR (CD3OD) δ 8.95 (s, 1 H), 7.65 (s, 1 H), 7.55 (s, 1 H),
4.36-4.31 (m, 2H), 4.24 (t, 2 H, J ) 7.34 Hz), 3.86-3.82 (m,
1H), 3.74-3.69 (m, 1H), 3.18 (t, 2H, J ) 7.25 Hz), 2.92 (t, 2H,
J ) 7.39 Hz), 2.26 (t, 2H, J ) 7.52 Hz); 1.98-0.85 (m, 33H);
HRMS m/ z calcd for C30H54N6O4 563.4285 (M + H), found
563.4290. Amino acid analysis: Ser 1.00 (0.92), Lys 1.00
(1.08).
12-(1H -Im id a zol-1-yl)d od eca n oyl-Ser -Lys-N-(2-cyclo-
h exyleth yl)a m id e (10) was prepared in a manner similar to
8 by coupling 7 with 4: 1H-NMR (CD3OD) δ 8.95 (s, 1 H), 7.65
(d, 1 H, J ) 1.61 Hz), 7.56 (d, 1 H, J ) 1.61 Hz), 4.36-4.31
(m, 2H), 3.86-3.81 (m, 1H), 4.24 (t, 2 H, J ) 7.25 Hz), 3.74-
3.68 (m, 1H), 3.18 (t, 2H, J ) 7.25 Hz), 2.92 (t, 2H, J ) 7.35
Hz), 2.56 (t, 2H, J ) 7.55 Hz), 1.98-0.85 (m, 37H); HRMS m/ z
calcd for C32H58N6O4 591.4598 (M + H), found 591.4649.
Meth yl 4-(4-Hyd r oxy-1-bu tyn yl)p h en yla ceta te (15). To
1.78 (t, 1H, J ) 6.3 Hz, hydroxyl); 13C-NMR (CDCl3) δ 172.59,
133.57, 131.68, 129.06, 122.17, 86.59, 81.83, 60.97, 51.96,
40.81, 23.64; FAB-MS m/ z 219 (M + H); HRMS calcd for
C13H15O3 (M + H) 219.1021, found 219.1008.
Met h yl 4-(5-h yd r oxy-1-p en t yn yl)p h en yla cet a t e (16)
was prepared in a manner analogous to 15 by coupling
4-pentyn-1-ol with 14 in 83% yield: 1H-NMR (CDCl3) δ 7.34
(d, 2H, J ) 8.4 Hz), 7.19 (d, 2H, J ) 8.4 Hz), 3.83 (q, 2H, J )
6.0 Hz), 3.69 (s, 3H), 3.6 (s, 2H), 2.54 (t, 2H, J ) 6.9 Hz), 1.86
(m, 2H); FABMS m/ z 233 (M + H); HRMS calcd for C14H17O3
233.1178, found 233.1190.
Meth yl 4-(4-Hyd r oxybu tyl)p h en yla ceta te (17). The
ester 15 (10.6 g, 0.0485 mol) was dissolved in methanol (200
mL), 10% Pd-C (1.1g) was added, and the mixture was stirred
under a hydrogen (50 psi) atmosphere. After 6 h, additonal
10% palladium on carbon (1 g) was added, and the hydrogena-
tion continued overnight. The catalyst was filtered through
Celite, and the filtrate was concentrated and dried in vacuo
to give 10.20 g (94.5%) of 17 as a pale yellow liquid: 1H-NMR
(CDCl3) δ 7.15 (q, 4H, J ) 8.1 Hz), 3.69 (s, 3H), 3.65 (t, 2H, J
) 8.1 Hz), 3.6 (s, 2H), 2.62 (t, 2H, J ) 7.2 Hz), 1.65 (m, 4H),
1.2 (br, 1H); 13C-NMR (CDCl3) δ 172.16, 141.05, 131.08, 128.96,
128.44, 62.32, 51.84, 40.57, 35.06, 32.06, 27.33; FAB-MS m/ z
223 (M + H); HRMS calcd for C13H19O3 (M + H) 223.1334,
found 223.1328.
Meth yl 4-(5-h yd r oxyp en tyl)p h en yla ceta te (18) was pre-
pared in a manner similar to 17 starting from 16 in 90%
yield: 1H-NMR (CDCl3) δ 7.14 (d, 4H, J ) 8.1 Hz), 3.69 (s,
3H), 3.65 (m, 2H), 3.59 (s, 2H), 2.6 (t, 2H, J ) 7.5 Hz), 1.62
(m, 4H), 1.42 (m, 2H), 1.18 (t, 1H, J ) 5.1 Hz); FAB-MS m/ z
237 (M + H); HRMS calcd for C14H21O3 (M + H) 237.1491,
found 237.1486.
Meth yl 4-(4-Iod obu tyl)p h en yla ceta te (19). To a solution
of the hydroxymethyl ester 17 (3.88 g, 17 mmol) in dry
acetonitrile (10 mL) was added a solution of the methyl
triphenoxyphosphonium iodide (10.26 g, 23 mmol) in dry
acetonitrile (100 mL), and the mixture was stirred at 0 °C.
The reaction mixture was warmed to room temperature in 12
h and then quenched with excess methanol at 0 °C. Solvents
were removed under reduced pressure, and the residue was
dissolved in ethyl acetate (500 mL), washed successively with
cold 0.2 N NaOH (2 × 500 mL), water (2 × 500 mL), and
saturated NaCl (2 × 500 mL), dried over MgSO4, and concen-
trated. The crude material thus obtained was purified by flash
chromatography (10% EtOAc in hexane) to give 4.12 g (71%)
of 19 as a clear liquid: 1H-NMR (CDCl3) δ 7.17 (q, 4H, J )
7.92 Hz), 3.69 (s, 3H), 3.60 (s, 2H), 3.20 (t, 2H, J ) 6.86 Hz),
2.62 (t, 2H, J ) 7.52 Hz), 1.70-1.89 (m, 4H); 13C-NMR (CDCl3)
δ 172.02, 140.50, 131.40, 129.15, 128.50, 51.93, 40.67, 34.27,
32.81, 32.03, 6.71; FABMS m/ z 333 (M + H); HRMS calcd for
C13H18O2I (M + H) 333.0351, found 333.0347.
Meth yl 4-(5-iod op en tyl)p h en yla ceta te (20) was pre-
pared in a manner similar to 19 starting from 18 in 70%
yield: 1H-NMR (CDCl3) δ 7.16 (dd, 4H, J ) 8.4 Hz), 3.69 (s,
3H), 3.59 (s, 2H), 3.18 (t, 2H, J ) 6.9 Hz), 2.6 (t, 2H, J ) 6.9
Hz), 1.85 (m, 2H), 1.52 (m, 2H), 1.45 (m, 2H); FABMS m/ z
347 (M + H); HRMS calcd for C14H20O2I (M + H) 347.0508,
found 347.0511.
4-(4-Iod obu tyl)p h en yla cetic Acid (21). Ester 19 (21.28
g, 0.064 mol) was dissolved in methanol (160 mL). Lithium
hydroxide (6.72 g, 0.16 mol) and water (10 mL) were added,
and the reaction mixture was stirred for 18 h at room
temperature. The solvents were removed under reduced
pressure, the residue was treated with ethyl acetate (600 mL),
and the organic solution was washed with 1 N HCl (3 × 300
mL) and brine (3 × 300 mL), dried over MgSO4, and filtered.
The filtrate was concentrated, and the residue was dried in
vacuo to give 17.5 g (85.9%) of 21 as a pale yellow solid: 1H-
NMR (CD3OD) δ 7.16 (q, 4H, J ) 8.26 Hz), 3.55 (s, 2H), 3.24
(t, 2H, J ) 6.75 Hz), 2.61 (t, 2H, J ) 7.35 Hz), 1.68-1.84 (m,
4H); 13C-NMR (CDCl3) δ 178.24, 140.84, 130.70, 129.32, 128.59,
40.63, 34.31, 32.85, 32.02, 6.63; FABMS m/ z 319 (M + H);
HRMS calcd for C12H16IO2 (M + H) 319.0194, found 319.0208.
4-(5-Iod op en tyl)p h en yla cetic a cid (22) was prepared in
a manner similar to 21 starting from 20 in 89% yield: 1H-
NMR (CDCl3) δ 7.16 (dd, 4H, J ) 8.1 Hz), 3.6 (s, 2H), 3.18 (t,
2H, J ) 7.2 Hz), 2.6 (t, 2H, J ) 7.5 Hz), 1.85 (m, 2H), 1.63 (m,
a
solution of butyn-1-ol (0.77 g, 0.011 mol) and methyl
4-iodophenylacetate (1.5 g, 0.0054 mol) in acetonitrile (10 mL)
at 0 °C was added triethylamine (1.5 mL, 0.01 mol), followed
by the addition of bis(triphenylphosphine)palladium chloride
(0.25 g, 0.36 mmol) and CuI (0.025 g). The reaction mixture
was stirred at 0 °C under an argon atmosphere for 30 min
and at room temperature for 3 h. The dark-colored reaction
mixture was concentrated under reduced pressure, and the
residue was partitioned between 5% citric acid (50 mL) and
EtOAc (50 mL). The organic phase was washed with 5% citric
acid (3 × 15 mL) and water, dried (Na2SO4), and concentrated.
The resulting material was purified by silica gel flash chro-
matography using 35% EtOAc in hexane as the eluent to afford
1.0 g (85%) of 15 as a dark-colored liquid: 1H-NMR (CDCl3) δ
7.36 (d, 2H, J ) 8.1 Hz), 7.21 (d, 2H, J ) 8.1 Hz), 3.82 (q, 2H,
J ) 6.3 Hz), 3.69 (s, 3H), 3.61 (s, 2H), 2.69 (t, 2H, J ) 6.3 Hz),

2620 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16
Devadas et al.
2H), 1.46 (m, 2H), 13C-NMR δ 178.20, 141.28, 130.48, 129.19,
128.50, 40.57, 35.19, 33.29, 30.02, 6.76; FABMS m/ z 333 (M
+ H); HRMS calcd for C13H18IO2 (M + H) 333.0350, found
333.0368.
(R,S)-Meth yl 2-(4-Iod op h en yl)p r op ion a te (41). Sodium
hydride (0.095 g, 3.96 mmol, 80% suspension) was added to a
solution of the methyl 4-iodophenylacetate 14 (1.0 g, 3.62
mmol) in dry THF (10 mL), and the mixture was stirred at 5
°C. After 30 min, iodomethane (0.68 g, 4.8 mmol) was added,
and the mixture was stirred at room temperature. Additional
iodomethane (0.46 g, 3.2 mmol) was added after 2 h, and the
mixture was stirred at room temperature overnight for 16 h.
Acetic acid (0.2 mL) was added, and the reaction mixture was
concentrated under reduced pressure. The residue was par-
titioned between EtOAc (30 mL) and 5% citric acid (25 mL).
The organic phase was washed with water (2 × 20 mL), dried
(Na2SO4), and concentrated, and the resulting substance was
purified by silica gel flash chromatography using 20% EtOAc
in hexane to give 0.7 g of 41 (70%) as a colorless liquid: 1H
NMR (CDCl3) δ 7.64 (d, 2H, J ) 8.4 Hz), 7.03 (2H, J ) 8.4
Hz), 3.66 (m over s, 4H), 1.46 (d, 3H, J ) 7.2 Hz); 13C-NMR
(CDCl3) δ 174.31, 140.09, 137.61, 129.46, 92.52, 52.05, 44.89,
18.35; FABMS m/ z 291 (M + H); HRMS calcd for C10H12IO2
(M + H) 290.9882, found 290.9861.
(R,S)-Meth yl 2-(4-iod op h en yl)bu tyr a te (42) was pre-
pared in a manner similar to 41 using ethyl iodide and 14 in
43% yield as a colorless liquid: 1H-NMR (CDCl3) δ 7.64 (d,
2H, J ) 8.1 Hz), 7.05 (d, 2H, J ) 8.1 Hz), 3.66 (s, 3H), 3.4 (t,
1H, J ) 6.7 Hz), 2.1 (m, 1H), 1.78 (m, 1H), 0.88 (t, 3H, J ) 7.5
Hz); FABMS m/ z 305 (M + H); HRMS calcd for C11H14IO2 (M
+ H) 305.0038, found 305.0030.
(R,S)-Meth yl 2-(4-Iod op h en yl)p en ta n oa te (43). To a
solution of 14 (2.5 g, 9.05 mmol) in THF (35 mL) at 0 °C was
added NaH (0.23 g, 9.6 mmol). After for 30 min of stirring, a
solution of iodopropane (1.1 mL) in 10 mL of THF was added
dropwise. The reaction mixture was stirred at room temper-
ature for 1 h, heated at 65 °C for 2 h, and concentrated under
reduced pressure. The residue was partitioned between cold
5% citric acid (30 mL) and ethyl acetate (50 mL). The organic
phase was washed with water (2 × 25 mL), dried (Na2SO4),
and concentrated. The residue was purified by silica gel flash
chromatography using 20% EtOAc in hexane to to afford 2.1
g (73%) of the title compound as a pale yellow liquid: 1H-NMR
(CDCl3) δ 7.65 (dd, 2H, J ) 6.6, 1.8 Hz), 7.04 (dd, 2H, J ) 6.6,
1.8 Hz), 3.65 (s, 3H), 3.50 (t, 1H, J ) 7.2 Hz), 2.0 (m, 1H), 1.65
(m, 1H), 1.24 (m, 2H), 0.9 (t, 3H, J ) 7.2 Hz); FABMS m/ z
318 (M+); HRMS calcd for C12H15IO2 (M+) 318.0116, found
318.0065.
(R,S)-Meth yl 2-[4-(4-h yd r oxy-1-bu tyn yl)p h en yl]p r op i-
on a te (44) was prepared in a manner similar to 15 starting
from 41 and 3-butyn-1-ol to afford 96% yield of the title
compound as an orange liquid: 1H-NMR (CDCl3) δ 7.37 (d, 2H,
J ) 8.1 Hz), 7.22 (d, 2H, J ) 8.1 Hz), 3.8 (q, 2H, J ) 6.9 Hz),
3.69 (m, 1H), 3.66 (s, 3H), 2.69 (t, 2H, J ) 6.0 Hz), 1.78 (t,
1H), 1.48 (d, 3H, J ) 6.9 Hz); 13C-NMR (CDCl3) δ 174.81,
140.39, 132.01, 127.55, 122.39, 86.68, 82.16, 61.25, 52.20,
45.35, 23.91, 18.49; FABMS m/ z 233 (M + H); HRMS calcd
for C14H17O3 (M + H) 233.1178, found 233.1149.
(R,S)-Meth yl 2-[4-(4-h ydr oxy-1-bu tyn yl)ph en yl]bu tyr ate
(45) was prepared in a manner similar to 15 starting from 42
and 3-butyn-1-ol in 81% yield as an orange liquid: 1H-NMR
(CDCl3) δ 7.36 (d, 2H, J ) 7.2 Hz), 7.36 (d, 2H, J ) 7.2 Hz),
3.81 (q, 2H, J ) 6.0 Hz), 3.65 (s, 3H), 3.44 (t, 1H, J ) 6.8 Hz),
2.69 (t, 2H, J ) 6.0 Hz), 2.15 (m, 1H), 1.78 (m, 2H), 0.87 (t,
3H, J ) 7.2 Hz); 13C-NMR (CDCl3) δ 174.36, 138.97, 131.97,
128.03, 122.41, 86.67, 81.92, 61.26, 53.28, 52.10, 26.71, 23.93,
12.19; FABMS m/ z 247 (M + H); HRMS calcd for C15H19O3
(M + H) 247.1334, found 247.1331.
4-[4-(2-Meth yl-1H-im idazol-1-yl)bu tyl]ph en ylacetic Acid
(24). To a suspension of sodium hydride (3.17 g, 0.132 mol)
in dry DMF (30 mL) at 5 °C was added a solution of
2-methylimidazole (10.11 g, 0.123 mol) in dry DMF (50 mL).
The reaction mixture was stirred for 30 min at 5 °C, a solution
of the iodo acid 21 (14 g, 0.044 mol) in dry DMF (15 mL) was
added, and the mixture was stirred for 1.5 h. The reaction
mixture was allowed to warm to room temperature and stirred
for 5 h, cooled to 0 °C, and quenched with 1 N HCl. The
solution was concentrated, and the residue was dissolved in
water and washed several times with ethyl acetate. Water
was removed under reduced pressure, and the residue was
dried in vacuo and treated with ethyl acetate/acetonitrile (1:1
v/v). The solid was filtered and washed with ethyl acetate
several times. The solid thus obtained was then washed with
absolute ethanol. The ethanol washings were concentrated
and dried to obtain 7.63 g (64%) of 24: 1H-NMR (CD3OD) δ
7.47 (d, 1H, J ) 2.01 Hz), 7.40 (d, 1H, J ) 2.01 Hz), 7.17 (q,
4H, J ) 8.16), 4.12 (t, 2H, J ) 7.25 Hz), 3.56 (s, 2H), 2.67 (t,
2H, J ) 7.35 Hz), 2.58 (s, 3H), 1.65-1.88 (m, 4H); FABMS
m/ z 273 (M + H); HRMS calcd for C16H21N2O2 (M + H)
273.1603, found 273.1635.
4-[4-(1H-Im id a zol-1-yl)bu tyl]p h en yla cetic a cid (23) was
prepared in a manner similar to 24 using imidazole and 21:
1
tR ) 14.3 min; H-NMR (CD3OD) δ 8.7 (s, 1H), 7.54 (d, 1H, J
) 1.5 Hz), 7.46 (d, 1H, J ) 1.5 Hz), 7.16 (dd, 4H, J ) 8.4 Hz),
4.22 (t, 2H, J ) 7.2 Hz), 3.56 (s, 2H), 2.66 (t, 2H, J ) 7.2 Hz),
1.91 (m, 2H), 1.66 (m, 2H); FAB-MS m/ z 259 (M + H); HRMS
calcd for C15H19N2O2 (M + H) 259.1447, found 259.1429.
4-[4-(1H-Im id a zol-1-yl)p en tyl]p h en yla cetic a cid (29)
was prepared in a manner similar to 24 using imidazole and
1
22: tR ) 15.46 min; H-NMR (CD3OD) δ 8.6 (s, 1H), 7.49 (d,
1H, J ) 1.5 Hz), 7.4 (d, 1H, J ) 1.5 Hz), 7.14 (dd, 4H, J ) 8.1
Hz), 4.18 (t, 2H, J ) 7.2 Hz), 3.55 (s, 2H), 2.6 (t, 2H, J ) 7.2
Hz), 1.88 (m, 2H), 1.68 (m, 2H), 1.3 (m, 2H); FABMS m/ z 273
(M + H); HRMS calcd for C16H21N2O2 (M + H) 273.1684, found
273.1727.
4-[4-(2-Iod o-1H-Im id a zol-1-yl)bu tyl]p h en yla cetic a cid
(25) was prepared in a manner similar to 24 starting from
2-iodoimidazole21 and 21: tR ) 15.4 min; 1H-NMR (CD3OD) δ
7.64 (d, 1H, J ) 2.1 Hz), 7.53 (d, 1H, J ) 2.1 Hz), 7.16 (dd,
4H, J ) 8.1 Hz), 4.16 (t, 2H, J ) 7.5 Hz), 3.56 (s, 2H), 2.67 (t,
2H, J ) 7.5 Hz), 1.87 (m, 2H); FABMS m/ z 259 (M + H);
HRMS calcd for C15H19N2O2 (M + H) 259.1447, found 259.1429.
4-[4-(2-Eth yl-1H-im id a zol-1-yl)bu tyl]p h en yla cetic a cid
(26) was prepared in a manner similar to 24 using 2-ethylimi-
dazole and 21 in 45% yield: 1H-NMR (DMSO-d6) δ 7.65 (d, 1H,
J ) 2.1 Hz), 7.57 (d, 1H, J ) 2.1 Hz), 7.16 (dd, 4H, J ) 8.1
Hz), 4.13 (t, 2H, J ) 7.5 Hz), 3.53 (s, 2H), 2.94 (q, 2H, J ) 7.5
Hz), 2.52 (t, 2H, J ) 7.5 Hz),1.75 (m, 2H), 1.58 (m, 2H), 1.26
(t, 3H, J ) 7.5 Hz); FAB-MS m/ z 293 (M + Li); HRMS calcd
for C17H22N2O2Li (M+Li) 293.1841, found 293.1859.
4-[4-(2-Isop r op yl-1H -im id a zol-1-yl)b u t yl]p h en yla ce-
tic a cid (27) was prepared in a manner similar to 24 using
2-isopropylimidazole and 21: 1H-NMR (CD3OD) δ 7.46 (d, 1H,
J ) 1.88 Hz), 7.43 (d, 1H, J ) 2.15 Hz), 7.17 (q, 4H, J ) 8.06
Hz), 4.16 (t, 2H, J ) 7.52 Hz), 3.55 (s, 2H), 3.38 (q, 1H, J )
6.98 Hz), 2.68 (t, 2H, J ) 7.25 Hz), 1.67-1.86 (m, 4H), 1.34
(d, 6 H); HRMS m/ z calcd for C18H24N2O2 301.1916 (M + H),
found 301.1859.
(R,S)-Met h yl 2-[4-(4-h yd r oxy-1-b u t yn yl)p h en yl]p en -
ta n oa te (46) was prepared in a manner similar to 15 starting
from 43 and 3-butyn-1-ol in 77% yield as an orange viscous
4-[4-(2-Meth yl-1H-ben zim id a zol-1-yl)bu tyl]p h en yla ce-
tic Acid (28). A mixture of 2-methylbenzimidazole (0.19 g,
1.44 mmol) and 21 (0.2 g, 0.63 mmol) in dry THF (10 mL)
containing NaH (0.038 g, 1.58 mmol) was heated to reflux for
1 h under a nitrogen atmosphere. The reaction mixture was
acidified with acetic acid (0.2 mL) and concentrated to dryness,
and the residue was purified by reverse-phase (C18) HPLC to
obtain 0.18 g (62%) of the title compound 28 as a hygroscopic
1
liquid: H-NMR (CDCl3) δ 7.35 (d, 2H, J ) 8.4 Hz), 7.23 (d,
2H, J ) 8.4 Hz), 3.79 (q, 2H, J ) 6.6 Hz), 3.65 (s, 3H), 3.54 (t,
1H, J ) 6.7 Hz), 2.69 (t, 2H, J ) 6.3 Hz), 2.1 (m, 1H), 1.78 (m,
2H), 1.22 (m, 1H), 0.89 (t, 3H, J ) 7.2 Hz); FABMS m/ z 261
(M + H); HRMS calcd for C16H21O3 (M + H) 261.1491, found
261.1523.
1
substance: tR ) 18.0 min; H-NMR (CD3OD) δ 7.75 (m, 2H),
7.57 (2d, 2H, J ) 3.0 Hz), 7.16 (q, 4H, J ) 8.4 Hz), 4.42 (t, 2H,
J ) 7.5 Hz), 3.55 (s, 2H), 2.81 (s, 3H), 2.68 (t, 2H, J ) 7.5 Hz),
1.92 (m, 2H), 1.78 (m, 2H); HRMS calcd for C20H21N2O2 (M -
H) 321.1603, found 321.1582.
(R,S)-Meth yl 2-[4-(4-h yd r oxybu tyl)p h en yl]p r op ion a te
(47) was prepared in a manner similar to 17 starting from 44
1
in 98% yield as a pale yellow liquid: H-NMR (CDCl3) δ 7.21
(d, 2H, J ) 7.8 Hz), 7.13 (d, 2H, J ) 7.8 Hz), 3.68 (m, 3H),

Inhibitors of Candida albicans NMT
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16 2621
3.66 (s, 3H), 2.62 (t, 2H, J ) 7.5 Hz), 1.63 (m, 4H), 1.48 (d,
2H, J ) 7.2 Hz); 13C-NMR (CDCl3) δ 175.14, 141.14, 137.82,
128.57, 127.28, 62.76, 51.9, 44.94, 35.15, 32.23, 27.4, 18.52;
FABMS m/ z 237 (M + H); HRMS calcd for C14H21O3 (M + H)
237.1491, found 237.1491.
(R,S)-Meth yl 2[4-(4-h ydr oxybu tyl)ph en yl]bu tyr ate (48)
was prepared in a manner similar to 17 starting from 45 in
96% yield as a colorless liquid: 1H-NMR (CDCl3) δ 7.22 (d,
2H, J ) 7.8 Hz), 7.12 (d, 2H, J ) 7.8 Hz), 3.56 (s over m, 5H),
3.42 (t, 1H, J ) 7.5 Hz), 2.62 (t, 2H, J ) 7.5 Hz), 2.11 (m, 1H),
1.72 (m, 5H), 1.29 (t, 1H, J ) 5.3 Hz), 0.88 (t, 3H, J ) 7.5 Hz);
FABMS m/ z (M + H) 251; HRMS calcd for C14H26O3 (M + H)
251.1647, found 251.1646.
(R,S)-Meth yl r-p r op yl-4-(4-h yd r oxy-1-bu tyl)ben zen e-
a ceta te (49) was prepared in a manner similar to 17 starting
from 46 to afford 97% yield of the title compound as a colorless
liquid: 1H-NMR (CDCl3) δ 7.21 (d, 2H, J ) 8.1 Hz), 7.13 (d,
2H, J ) 8.1 Hz), 3.67 (t, 2H, J ) 8.5 Hz), 3.52 (t, 1H, J ) 7.5
Hz), 2.62 (t, 2H, J ) 7.5 Hz), 2.05 (m, 1H), 1.64 (m, 5H), 1.20
(m, 3H), 0.90 (t, 2H, J ) 7.5 Hz); FABMS m/ z 264 (M+); HRMS
calcd for C16H24O3 (M + H) 264.1725, found 264.1693.
(R,S)-Meth yl 2-[4-(4-iod obu tyl)p h en yl]p r op ion a te (50)
was prepared in a manner similar to 19 starting from 47 in
85% yield as a pale yellow viscous liquid: 1H-NMR (CDCl3) δ
7.2 (d, 2H, J ) 8.1 Hz), 7.14 (d, 2H, J ) 8.1 Hz), 3.69 (q, 1H,
J ) 6.9 Hz), 3.66 (s, 3H), 3.2 (t, 2H, J ) 6.9 Hz), 2.6 (t, 2H, J
) 7.5 Hz), 1.85 (m, 2H), 1.7 (m, 2H), 1.48 (d, 3H, J ) 7.2 Hz);
13C-NMR (CDCl3) δ 175.02, 140.57, 138.02, 128.55, 127.37,
51.94, 44.93, 34.28, 32.87, 32.05, 18.57, 6.71; FABMS m/ z 353
(M + Li); HRMS calcd for C14H20IO2 (M + H) 347.0508, found
347.0509.
(R,S)-Meth yl 2-[4-(4-iod obu tyl)p h en yl]bu tyr a te (51)
was prepared in a manner similar to 19 starting from 48 in
32% yield as a colorless liquid: 1H-NMR (CDCl3) δ 7.21 (d,
2H, J ) 8.1 Hz), 7.12, (d, 2H, J ) 8.1 Hz), 3.62 (s, 3H), 3.43 (t,
1H, J ) 7.8 Hz), 3.2 (t, 2H, J ) 6.6 Hz), 2.61 (t, 2H, J ) 7.5
Hz), 2.06 (m, 1H), 1.83 (m, 2H), 1,78 (m, 3H), 0.89 (t, 3H, J )
7.5 Hz); 13C-NMR (CDCl3) δ 174.70, 140.75, 136.73, 128.64,
127.98, 53.04, 51.96, 34.45, 33.03, 32.19, 26.86, 12.30, 6.87;
FABMS m/ z 361 (M + H); HRMS calcd for C15H22IO2 (M +
H) 361.0664, found 361.0647.
Meth yl (R,S)-2-[4-(4-iod obu tyl)p h en yl]p en ta n oa te (52)
was prepared in a manner similar to 19 starting from 49 in
82% yield as a colorless liquid: 1H-NMR (CDCl3) δ 7.22 (d,
2H, J ) 8.1 Hz), 7.12 (d, 2H, J ) 8.1 Hz), 3.65 (s, 3H), 3.53 (t,
1H, J ) 7.8 Hz), 3.12 (t, 2H, J ) 6.6 Hz), 2.61 (t, 2H, J ) 7.5
Hz), 2.02 (m, 1H), 1.82 (m, 2H), 1.74 (m, 3H), 1.25 (m, 2H),
0.9 (t, 2H, J ) 7.5 Hz); FABMS m/ z 374 (M+); HRMS calcd
for C16H23IO2 (M+) 374.0742, found 374.0762.
7.5 Hz), FABMS m/ z 346 (M+); HRMS calcd for C14H19IO2 (M+)
346.0430, found 346.0410.
(R,S)-2-[4-(4-Iodobu tyl)ph en yl]pen tan oic Acid (55). The
ester 52 (0.7 g, 1.9 mmol) was stirred with 1 M LiOH (3.00
mL) containing THF (5.00 mL) at room temperature for 4 h
under a nitrogen atmosphere. The reaction mixture was
diluted with water (20 mL) and washed with EtOAc (2 × 15
mL). The aqueous portion was acidified with 5% citric acid
and extracted with EtOAc (2 × 15 mL). The combined EtOAc
extracts were washed with water (3 × 15 mL), dried (Na2SO4),
and concentrated to give 0.23 g (50%) of 55 as a colorless
liquid: 1H-NMR (CDCl3) δ 7.23 (d, 2H, J ) 8.1 Hz), 7.12 (d,
2H, J ) 8.1 Hz), 3.62 and 3.54 (t, 1H, J ) 7.5 Hz), 3.38 and
3.19 (t, 2H, J ) 7.2 Hz), 2.61 (t, 2H, J ) 7.5 Hz), 2.15 (m, 1H),
1.83-1.6 (m, 5H), 1.25 (m, 2H), 0.91 (t, 3H, J ) 7.2 Hz);
FABMS m/ z 367 (M + Li); HRMS calcd for C15H21IO2Li (M +
Li) 367.0745, found 367.0816.
(R,S)-r-Meth yl-4-[4-(2-m eth yl-1H-im id a zol-1-yl)bu tyl]-
ben zen ea cetic Acid (56). Sodium hydride (0.05 g, 80%
suspension, 2.1 mmol) was added to a solution of 2-methylimi-
dazole (0.092 g, 1.12 mmol) at 0 °C. After 30 min, a solution
of 12 (0.25 g, 0.75 mmol) in DMF (1.5 mL) was added, and the
mixture was stirred at 0 °C for 30 min and at room temper-
ature for 1.5 h under an argon atmosphere. After the DMF
was distilled in vacuo, the residue was treated with 2 N HCl
(3 mL) and water (10 mL) and washed with EtOAc (3 × 5 mL).
The aqueous phase was freeze-dried, and the residue was
washed with 10% CH3CN in EtOAc (20 mL) and dried to afford
1
0.14 g (58%) of 56: tR ) 14.9 min; H-NMR (CD3OD) δ 7.46
(d, 1H, J ) 2.1 Hz), 7.39 (d, 1H, J ) 2.1 Hz), 7.22 (d, 2H, J )
7.8 Hz), 7.13 (d, 2H, J ) 7.8 Hz), 4.12 (t, 2H, J ) 7.2 Hz), 3.66
(q, 1H, J ) 6.8 Hz), 2.66 (t, 2H, J ) 7.5 Hz), 2.57 (s, 3H), 1.85
(m, 2H), 1.67 (m, 2H), 1.41 (d, 3H, J ) 7.5 Hz); FABMS m/ z
287 (M + H); HRMS calcd for C17H23N2O2 (M + H) 287.1760,
found 287.1754.
(R,S)-r-Meth yl-4-[4-(2-ch lor o-1H-im id a zol-1-yl)bu tyl]-
ben zen ea cetic Acid (59). A mixture of 2-chloroimidazole21
(0.056 g, 0.55 mmol), 53 (0.12 g, 0.36 mmol), and NaH (0.025
g, 1.04 mmol) in THF (5.00 mL) was heated at 70 °C for 1 h,
under a nitrogen atmosphere with vigorous stirring. The
reaction mixture was acidified with acetic acid and concen-
trated under reduced pressure, and the resulting material was
purified by reverse-phase HPLC. The appropriate fractions
were combined and freeze-dried to give 0.1 g (66%) of the title
compound as a glassy solid: tR ) 22.0 min; 1H-NMR (CD3OD)
δ 7.29 (s, 1H), 7.22 (d, 2H, J ) 7.8 Hz), 7.06 (s, 1H), 4.05 (t,
2H, J ) 7.2 Hz), 3.66 (q, 1H, J ) 7.2 Hz), 2.64 (t, 2H, J ) 7.5
Hz), 1.79 (m, 2H), 1.66 (m, 2H), 1.42 (d, 3H, J ) 7.2 Hz);
FABMS m/ z 307 (M + H); HRMS calcd for C16H19 ClN2O2 (M
+ H) 307.1042, found 307.1021.
(R,S)-2-[4-(4-Iodobu tyl)ph en yl]pr opion ic Acid (53). The
ester 50 (1.0 g) was stirred with 1 M LiOH (5.00 mL)
containing MeOH (3.75 mL) at 60 ˚C for 1 h, under a nitrogen
atmosphere. The reaction mixture was concentrated under
reduced pressure, water (25 mL) was added, and the mixture
was extracted with ether (3 × 15 mL). The aqueous extract
was acidified with 5% citric acid and extracted with EtOAc (3
× 15 mL). The combined organic extracts were washed with
water (3 × 15 mL), dried (Na2SO4) and concentrated to give
53 (0.7 g, 73%) as a colorless liquid: 1H-NMR (CDCl3) δ 7.22
(d, 2H, J ) 8.1 Hz), 7.15 (d, 2H, J ) 8.1 Hz), 3.72 (q, 1H, J )
7.2 Hz), 3.2 (t, 2H, J ) 7.2 Hz), 2.6 (t, 2H, J ) 7.5 Hz), 1.85
(m, 2H), 1.7 (m, 2H), 1.5 (d, 3H, J ) 7.2 Hz); 13C-NMR (CDCl3)
δ 181.19, 140.84, 137.14, 128.57, 127.50, 44.89, 34.25, 32.81,
32.00, 17.98, 6.73; FABMS m/ z 339 (M + Li); HRMS calcd for
C14H20IO2 (M + Li) 339.0432, found 339.0424.
(R,S)-2-[4-(4-iod obu tyl)p h en yl]bu tyr ic Acid (54). The
ester 51 (0.35 g, 0.97 mmol) was stirred with 1 M LiOH (2.00
mL) containing THF (1.00 mL) at room temperature for 16 h
under a nitrogen atmosphere. The reaction mixture was
concentrated under reduced pressure, acidified with 5% citric
acid, and extracted with EtOAc (2 × 10 mL). The combined
EtOAc extracts were washed with water (3 × 15 mL), dried
(Na2SO4), and concentrated to give 0.26 g (73%) of 54 as a
colorless liquid: 1H-NMR (CDCl3) δ 7.21 (d, 2H, J ) 8.1 Hz),
3.38 (2t, 2H, J ) 6.8 Hz), 3.19 (t, 1H, J ) 6.8 Hz), 2.61 (t, 2H,
J ) 7.5 Hz), 2.1 (m, 1H), 1.85-1.5 (m, 5H), 0.91 (t, 3H, J )
(R,S)-r-E t h yl-4-[4-(2-m et h yl-1H -im id a zol-1-yl)b u t yl]-
ben zen ea cetic Acid (57). A mixture of 54 (0.12 g, 0.35
mmol) and 2-methylimidazole (0.15 g, 1.85 mmol) in dry THF
(5 mL) was heated at 60 °C for 4 h under
a nitrogen
atmosphere. The reaction mixture was acidified with acetic
acid and concentrated under reduced pressure. The resulting
product was purified by reverse-phase HPLC. The appropriate
fractions were pooled and freeze-dried to give 0.07 g (50%) of
1
57 as a glassy solid: tR ) 16.6 min; H-NMR (CD3OD) δ 7.47
(d, 1H, J ) 2.4 Hz), 7.39 (d, 1H, J ) 2.4 Hz), 7.22 (d, 2H, J )
8.1 Hz), 7.15 (d, 1H, J ) 8.1 Hz), 4.12 (t, 2H, J ) 7.2 Hz), 3.39
(t, 2H, J ) 7.5 Hz), 2.67 (t, 2H, J ) 7.5 Hz), 2.56 (s, 3H), 2.1
(m, 1H), 1.85 (m, 2H), 0.88 (t, 2H, J ) 7.5 Hz); FABMS m/ z
301 (M + H); HRMS calcd for C18H25N2O2 (M + H) 301.1916,
found 301.1927.
(R,S)-r-P r op yl-4-[4-(2-m eth yl-1H-im id a zol-1-yl)bu tyl]-
ben zen ea cetic a cid (58) was prepared in a manner similar
to 56 using 2-methylimidazole and 55 in 80% yield as a yellow
1
syrup: tR ) 18.25 min; H-NMR (CD3OD) δ 7.47 (d, 1H, J )
2.1 Hz), 7.41 (d, 1H, J ) 2.1 Hz), 7.23 (d, 2H, J ) 8.4 Hz),
7.15 (d, 1H, J ) 8.4 Hz), 4.13 (t, 2H, J ) 7.5 Hz), 3.51 (t, 2H,
J ) 8.1 Hz), 2.68 (t, 2H, J ) 7.5 Hz), 2.59 (s, 3H), 2.1 (m, 1H),
1.85 (m, 2H), 1.69 (m, 3H),1.25 (m, 2H), 0.93 (t, 2H, J ) 7.5
Hz); FABMS m/ z 315 (M + H); HRMS calcd for C19H27N2O2
(M + H) 315.2072, found 315.2072.
(R,S)-r-Met h yl-4-[4-(2-m et h yl-1H -b en zim id a zol-1-yl)-
bu tyl]ben zen ea cetic a cid (60) was prepared in a manner

2622 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16
similar to 56 using 2-methylbezimidazole and 53 in 87%
Devadas et al.
(R)-N-(1-Oxo-2-p h en ylp r op yl)-L-ser yl-N-(2-cycloh ex-
yleth yl)-L-lysin a m id e (68) was prepared in a manner similar
to 13 by coupling (R)-phenylacetic acid with 12. After the
workup, the crude product was treated with trifluoroacetic acid
and the desired product was isolated by reverse-phase HPLC
to afford 27% yield of the title compound as a white powder:
1
yield: tR ) 18.6 min; H-NMR (CD3OD) δ 7.79 (m, 2H), 7.73
(m, 2H), 7.21 (d, 2H, J ) 8.1 Hz), 7.14 (d, 2H, J ) 8.1 Hz),
4.44 (t, 2H, J ) 7.2 Hz), 3.67 (q, 1H, J ) 7.2 Hz), 2.71 (s, 3H),
2.68 (t, 2H, J ) 7.2 Hz), 1.93 (m, 2H), 1.76 (m, 2H), 1.42 (d,
3H, J ) 7.2 Hz); FABMS m/ z 337 (M + H); HRMS calcd for
1
tR ) 15.67 min; H-NMR (CD3OD, 500 MHz) δ 7.33 (m, 4H),
C
21H25N2O2 (M + H) 337.1917, found 337.1903.
7.23 (m, 1H), 4.36 (dd, 1H, Lys R-CH, J ) 4.0, 10.0 Hz), 4.28
(dd, 1H, Ser R-CH, J ) 5.5, 7.5 Hz), 3.78 (dd, 1H, J ) 5.5,
10.5 Hz), 3.75 (q, 1H, J ) 7.0 Hz), 3.66 (dd, 1H, J ) 7.5, 10.5
Hz), 3.19 (m, 2H), 2.92 (t, 2H, J ) 7.5 Hz), 1.95 (m, 1H), 1.71
(m, 8H), 1.5 (m, 1H), 1.44 (d, 3H, J ) 7.0 Hz), 1.38 (q, 2H, J
) 7.5 Hz), 1.31-1.12 (m, 5H), 0.92 (m, 2H); HRMS calcd for
C26H43N4O4 (MH+) 475.3284, found 475.3296. Amino acid
analysis: Ser 1.00 (1.00), Lys 1.00 (1.00).
(R,S)-N-[2-[4-[4-(2-Meth yl-1H-im id a zol-1-yl)bu tyl]p h e-
n yl]-1-oxop r op yl]-L-ser yl-N-(2-cycloh exyleth yl)-L-lysin a -
m id e (61). A mixture of 56 (0.092 g, 0.29 mmol) and
hydroxybenzotriazole (0.085 g, 0.57 mmol) in dimethylaceta-
mide (1.5 mL) and dichloromethane (3.0 mL) was treated with
DCC (0.62 g, 0.29 mmol) and stirred at 0 °C. After 2 h, a
solution of the amine acetate 12 (0.13 g, 0.24 mmol)25 in
dimethylacetamide (0.5 mL) and N-methylmorpholine (0.08
mL) were added, and the mixture was stirred at 0 °C for 1 h
and then at room temperature for 16 h. The reaction mixture
was concentrated in vacuo, and the residue was partitioned
between cold 0.25 N NaOH (15 mL) and EtOAc (25 mL). The
organic phase was washed successively with water (2 × 10
mL), 5% citric acid (2 × 10 mL), and water, dried (Na2SO4),
and concentrated. The resulting material was washed with a
solvent mixture of ether-hexane (1:1 v/v), filtered, and dried
to give a pale yellow powder (0.10 g) which was stirred with
trifluoroacetic acid (1.5 mL) for 4 h at room temperature. After
the removal of TFA under reduced pressure, the residue was
purified by reverse-phase HPLC. The appropriate fractions
were pooled and freeze-dried to give 0.025 g of 61 as a
diastereomeric mixture: tR ) 17.4 min (40%), 17.7 min (60%);
1H-NMR (CD3OD) δ 7.47 (d, 1H, J ) 2.1 Hz), 7.41 (d, 1H, J )
2.1 Hz), 7.25 (d, 2H, J ) 8.4 Hz), 7.14 (d, 2H, J ) 8.4 Hz),
4.4-4.2 (m, 2H), 4.13 (t, 2H, J ) 7.5 Hz), 3.9-3.6 (m, 3H),
3.18 (m, 2H), 2.98-2.84 (2t, 2H, J ) 7.2 Hz), 2.66 (t, 2H, J )
7.5 Hz), 2.59 and 2.58 (2s, 3H), 1.85 (m, 3H), 1.8-1.55 (m,
(S)-N-(1-Oxo-2-p h en ylp r op yl)-L-ser yl-N-(2-cycloh ex-
yleth yl)-L-lysin a m id e (69) was prepared in a manner similar
to 13 by coupling (S)-phenylacetic acid with 12. After the
workup, the crude product was treated with trifluoroacetic acid
and the desired product was isolated by reverse-phase HPLC
to afford a 43% yield of the title compound as a white powder:
1
tR ) 15.35 min; H-NMR (CD3OD, 500 MHz) δ 7.33 (m, 2H),
7.30 (m, 2H), 7.22 (m, 1H), 4.36 (t, 1H, Ser R-CH, J ) 7.0 Hz),
4.25 (dd, 1H, Lys R-CH, J ) 4.5, 7.5 Hz), 3.84 (dd, 1H, Ser
â-CH, J ) 5.5, 10.5 Hz), 3.75 (q, 1H, J ) 7.0 Hz), 3.73 (dd,
1H, Ser â-CH, J ) 7.0, 10.5 Hz), 3.15 (m, 2H), 2.82 (t, 2H, J )
7.5 Hz), 1.82 (m, 1H), 1.85 (m, 1H), 1.74-1.5 (m, 8H), 1.45 (d,
3H, J ) 7.0 Hz), 1.4-1.12 (m, 8H), 0.92 (m, 2H); HRMS calcd
for C26H43N4O4 (M + H+) 475.3284, found 475.3296. Amino
acid analysis: Ser 1.00 (0.99), Lys 1.00 (1.01).
(R,S)-N-[2-[4-[4-(2-Meth yl-1H-im id a zol-1-yl)bu tyl]p h e-
n yl]-1-oxob u t yl]-L-ser yl-N-(2-cycloh exylet h yl)-L-lysin a -
m id e (62) was prepared in a manner similar to 61 by reacting
1
57 with 12: tR ) 17.3 min; H-NMR (CD3OD) δ 8.18 (br, 1H,
exchangeable), 7.9 (br, 1H, exchangeable), 7.82 (br, 1H,
exchangeable), 7.49 (d, 1H, J ) 2.1 Hz), 7.42 (d, 1H, J ) 2.1
Hz), 7.27 (d, 2H, J ) 7.8 Hz), 7.15 (d, 2H, J ) 7.8 Hz), 4.41-
4.21 (m, 2H), 4.14 (t, 2H, J ) 7.2 Hz), 3.9-3.6 (m, 2H), 3.45
(m, 1H), 3.18 (m, 2H), 2.85 (m, 2H), 2.64 (m, 2H), 2.61 and
2.60 (s, 3H), 2.08-1.1 (m, 23H), 0.91 (m, 5H); HRMS calcd for
C35H57N6O4 (M + H+) 625.4441, found 625.4429. Amino acid
analysis: Ser 1.00 (1.01), Lys 1.00 (0.99).
21H), 0.92 (m, 2H); HRMS calcd for
C
34H54N6O4 (MH+)
611.4285, found 611.4265. Amino acid analysis: Ser 1.00
(1.02), Lys 1.00(0.98).
Ch ir a l Sep a r a tion of 61a a n d 61b. The racemic material
61 (0.5 mg) was dissolved in acetonitrile (25.0 µL), loaded on
an ASTEC vancomycin column (5-µM silica gel, 4.6 × 250
mm), and eluted with 23% acetonitrile containing 0.8% of
ammonium phosphate at pH ) 4.2, at a flow rate of 1.0 mL/
min for 15 min. The separation was monitored by UV
absorbance at 220 nm. The separation was repeated by
making 19 more injections of 25.0 µL each, containing 0.5 mg
of the recemic material. The appropriate fractions were
collected, pooled, and desalted by passing through a short C18
column (4.6 × 150 mm) and eluting with acetonitrile contain-
ing 0.1% trifluoroacetic acid. The fractions were collected and
freeze-dried to obtain 5.0 mg of 61a and 3.6 mg of 61b as white
amorphous substances.
(R,S)-N-[2-[4-[4-(2-Meth yl-1H-im id a zol-1-yl)bu tyl]p h e-
n yl]-1-oxop en tyl]-L-ser yl-N-2-cycloh exyleth yl)-L-lysin a -
m id e (63) was prepared in a manner similar to 61 by reacting
58 with 12: tR ) 19.5 min; 1H-NMR (CD3OD) δ 7.25 (m, 3H),
7.12 (m, 3H), 4.4-4.21 (m, 2H), 4.03 (t, 2H), 3.8-3.5 (m, 3H),
3.2 (m, 2H), 2.85 (2t, 2H), 2.64 (m, 2H), 2.45 and 2.44 (s, 3H),
2.1-1.6 (m, 16H), 1.6-1.15 (m, 9H), 0.91 (m, 5H); HRMS calcd
for C36H58N6O4 (M + H+) 639.4598, found 639.4592. Amino
acid analysis: Ser 1.00 (1.00), Lys 1.00 (1.00).
(R,S)-N-[2-[4-[4-(2-Ch lor o-1H-im id a zol-1-yl)bu tyl]p h e-
n yl]-1-oxop r op yl]-L-ser yl-N-(2-cycloh exyleth yl)-L-lysin a -
m id e (64) was prepared in a manner similar to 61 by reacting
59 with 12: tR ) 22.5 min; 1H-NMR (CD3OD) δ 7.25 (d, 1H, J
) 3.3 Hz), 7.23 (d, 1H, J ) 3.3 Hz), 7.2 (d, 1H, J ) 1.5 Hz),
7.12 (d, 2H, J ) 8.1 Hz), 6.95 (d, 1H, J ) 1.5 Hz), 4.34 (m,
1H), 4.26 (m, 1H), 4.01 (t, 2H, J ) 7.2 Hz), 3.71 (m, 2H), 3.18
(m, 2H), 2.92, 2.85 (2t, 2H, J ) 7.2 Hz), 2.62 (t, 2H, J ) 7.2
Hz), 2.0-1.1 (m, 24H), 0.94 (m, 2H); HRMS calcd for C36H58N6O4
(M + H+) 631.3739, found 631.3729. Amino acid analysis: Ser
1.00 (1.02), Lys 1.00 (0.98).
(R)-N-[2-[4-[4-(2-Meth yl-1H-im idazol-1-yl)bu tyl]ph en yl]-
1-o x o p r o p y l]-L -s e r y l-N -(2-c y c lo h e x y le t h y l)-L -ly s in -
a m id e (61a ): tR ) 17.56 min; H-NMR (CD3OD, 500 MHz) δ
1
7.44 (br, 1Η), 7.37 (br, 1H), 7.25 (d, 2H, J ) 8.0 Hz), 7.14 (d,
2H, J ) 8.0 Hz), 4.41 (dd, 1H, Lys R-CH, J ) 4.5 Hz, 9.5 Hz),
4.36 (t, 1H, Ser R-CH, J ) 7.0 Hz), 4.19 (t, 2H, J ) 7.5 Hz),
3.84 (dd, 1H, J ) 6.0, 10.5 Hz), 3.78 (q, 1H, J ) 7.0 Hz), 3.71
(dd, 1H, J ) 7.5, 10.5 Hz), 3.10 (m, 2H), 2.94 (t, 2H, J ) 7.0
Hz), 2.65 (t, 2H, J ) 8.0 Hz), 2.57 (s, 3H), 1.94 (m, 1H), 1.85
(m, 2H), 1.75-1.62 (m, 10H), 1.48 (m, 1H), 1.41 (d, 3H, J )
7.0 Hz), 1.37 (q, 2H, J ) 7.0 Hz), 1.32-1.12 (m, 4H), 0.92 (m,
2H); HRMS calcd for C34H57N6O4 (M + H+) 611.4285, found
611.4273. Amino acid analysis: Ser 1.00 (1.00), Lys 1.00
(1.00).
(R,S)-N-[2-[4-[4-(2-Meth yl-1H-ben zim id a zol-1-yl)bu tyl]-
p h en yl]-1-oxop r op yl]-L-ser yl-N-(2-cycloh exyleth yl)-L-ly-
sin a m id e (65) was prepared in a manner similar to 61 by
1
reacting 60 with 12: tR ) 22.8 min; H-NMR (CD3OD) δ 7.8
(S)-N-[2-[4-[4-(2-Meth yl-1H-im idazol-1-yl)bu tyl]ph en yl]-
1-oxop r op yl]-L-ser yl-N-2-cycloh exyleth yl)-L-lysin a m id e
(61b): tR ) 17.35 min; 1H-NMR (CD3OD, 500 MHz) δ 7.45 (br,
2Η), 7.25 (d, 2H, J ) 8.0 Hz), 7.13 (d, 2H, J ) 8.0 Hz), 4.35 (t,
1H, Ser R-CH, J ) 6.25 Hz), 4.25 (dd, 1H, Lys R-CH, J ) 4.0,
9.75 Hz), 4.14 (br, 2H), 3.84 (dd, 1H, J ) 5.5, 10.5 Hz), 3.72
(q, 2H), 3.16 (m, 2H), 2.87 (t, 2H, J ) 7.5 Hz), 2.64 (t, 2H, J )
6.5 Hz), 2.56 (br, 3H), 1.87 (m, 3H), 1.74-1.54 (m, 10H), 1.43
(d, 3H, J ) 7.0 Hz), 1.42-1.14 (m, 8H), 0.91 (m, 2H); HRMS
calcd for C34H57N6O4 (M + H+) 611.4285, found 611.4254.
Amino acid analysis: Ser 1.00 (0.99), Lys 1.00 (1.01).
(m, 1H), 7.75 (m, 1H), 7.58 (m, 2H), 7.25 (d, 2H, J ) 8.1 Hz),
7.14 (m, 2H), 4.44 (2H, t, J ) 6.6 Hz), 4.35 (m, 2H), 3.85-3.6
(m, 2H), 3.16 (m, 2H), 2.93 (t, 2H, J ) 6.9 Hz), 2.83 (2s, 3H),
2.67 (t, 2H, J ) 7.2 Hz), 1.93 (m, 3H), 1.72 (m, 10H), 1.55-1.1
(m, 11H), 0.92 (m, 2H); HRMS calcd for C38H57N6O4 (M + H+)
661.4441, found 661.4465. Amino acid analysis: Ser 1.00
(1.02), Lys 1.00 (0.98).
4-[4-(2-Met h yl-1H -im id a zol-1-yl)b u t yl]p h en yla cet yl-
Ser -Lys-N-(2-cycloh exyleth yl)a m id e (31). To a mixture of
24 (0.095 g, 0.3 mmol) and HOBt (0.066 g, 0.32 mmol) in
dimethylacetamide (0.5 mL) and dichloromethane (3.0 mL)

Inhibitors of Candida albicans NMT
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16 2623
was added DCC (0.66 g, 0.32 mmol), and the mixture was
stirred at 0 °C. After 1.5 h, a solution of the amine acetate 12
(0.13 g, 0.24 mmol)25 in dimethylacetamide (0.5 mL) and
N-methylmorpholine (0.04 mL) were added and stirred at 0
˚C for 1 h and then at room temperature for 16 h. The reaction
mixture was concentrated in vacuo, and the residue was
partitioned between cold 0.25 N NaOH (10 mL) and EtOAc
(25 mL). The organic phase was washed successively with
water (2 × 10 mL), 5% citric acid (2 × 10 mL), and water,
dried (Na2SO4), and concentrated. The resulting material was
washed with a solvent mixture of ether-hexane (1:1 v/v),
filtered, and dried to give a pale yellow powder (0.125 g) which
was stirred with trifluoroacetic acid (1.5 mL) for 3.5 h at room
temperature. After the removal of TFA under reduced pres-
sure, the residue was purified by reverse-phase HPLC to give
31 (0.08 g) as a white hygroscopic substance: tR ) 18.7 min;
1H-NMR (CD3OD) δ 7.3-7.15 (m, 6H), 4.34 (m, 2H), 4.08 (t,
2H, J ) 7.25 Hz), 3.85 (m, 1H), 3.75 (m, 1H), 3.56 (s, 2H), 3.2
(m, 2H), 2.89 (t, 2H, J ) 9.25 Hz), 2-0.8 (m, 23H); 13C-NMR
(CD3OD) 173.59, 173.51, 172.89, 141.39, 134.14, 130.19, 129.50,
122.89, 118.99, 62.70, 56.95, 54.32, 49.67, 42.77, 40.34, 39.28,
37.47, 36.24, 35.51, 34.06, 31.69, 29.91, 28.87, 27.62, 27.46,
27.17, 23.66; HRMS calcd for C33H53N6O4 (M + H+) 597.4128,
found 597.4122. Amino acid analysis: Ser 1.00 (1.00), Lys 1.00
(1.00).
4-[4-(2-E t h yl-1-im id a zol-1-yl)b u t yl]p h en yla cet yl-Ser -
Lys-N-(2-cycloh exyleth yl)a m id e (33) was prepared in a
manner similar to 31 by coupling 26 with 12 as white glassy
1
substance: tR ) 20.8 min; H-NMR (CD3OD) δ 7.49 (d, 1H, J
) 1.8 Hz), 7.44 (d, 1H, J ) 1.8 Hz), 7.22 (d, 1H, J ) 8.1 Hz),
7.15 (d, 1H, J ) 8.1 Hz), 4.35 (m, 2H), 4.15 (t, 2H, J ) 7.2
Hz), 3.85 (m, 1H), 3.75 (m, 1H), 3.57 (s, 2H), 3.18 (m, 2H),
2.97 (q, 2H, J ) 7.5 Hz), 2.9 (t, 2H, J ) 7.5 Hz) 2-1.8 (m, 3H),
1.78-1.6 (m, 10 H), 1.55-1.15 (t over m, 11H), 0.92 (m, 2H);
HRMS calcd for C34H55N6O4 (M + H+) 611.4285, found
611.4282. Amino acid analysis: Ser 1.00 (1.00), Lys 1.00
(1.00).
4-[4-(2-Isop r op yl-1-im id a zol-1-yl)b u t yl]p h en yla cet yl-
Ser -Lys-N-(2-cycloh exyleth yl)a m id e (34). A mixture of the
acid 27 (0.047 g, 0.156 mmol), HOBt (0.021 g, 0.156 mmol),
and EDC (0.030 g, 0.156 mmol) was stirred in dry DMF (2
mL) at room temperature for 40 min. It was then treated with
a solution of 4 (0.085 g, 0.156 mmol) in dry DMF (2 mL)
containing N-methylmorpholine (0.016 g, 0.312 mmol). After
the reaction mixture was stirred at room temperature for 18
h, it was diluted with dichloromethane (100 mL) and washed
successively with saturated NaHCO3 (2 × 100 mL) and brine
(2 × 100 mL) and dried (MgSO4). Removal of the solvent
under reduced pressure provided 90 mg of a yellow oil which
was then treated with 4 N HCl in dioxane (5 mL) at room
temperature for 2.5 h. The resulting mixture was concentrated
under reduced pressure, and the residue was purified by
reverse-phase HPLC to give 9 mg (12.5%) of 34 as a glassy
substance: 1H-NMR (CD3OD) δ 7.48 (d, 1H, J ) 2.01 Hz), 7.45
(d, 1H, J ) 2.01 Hz), 7.18 (ab q, 4H, J ) 8.06 Hz), 4.38-4.28
(m, 2H), 4.17 (t, 2H, J ) 7.45 Hz), 3.91-3.81 (m, 2 H), 3.75-
3.69 (m, 1H), 3.55 (s, 2 H), 3.42 (q, 1 H, J ) 7.00 Hz), 3.23-
3.14 (m, 2H), 2.94-2.86 (m, 3H), 1.96-0.84 (m, 29H); HRMS
m/ z calcd for C35H56N6O4 625.4441 (M + H), found 625.4410.
Amino acid analysis: Ser 1.00 (0.95), Lys 1.00 (1.05).
4-[4-(2-Met h ylb en zim id a zol-1-yl)b u t yl]p h en yla cet yl-
Ser -Lys-N-(2-cycloh exyleth yl)a m id e (35) was prepared in
a manner similar to 31 by coupling 28 with 12 as a white
4-[4-(1H-Im id a zol-1-yl)bu tyl]p h en yla cetyl-Ser -Lys-N-
(2-cycloh exyleth yl)a m id e (30) was prepared in a manner
similar to 31 by coupling 23 with 12 as white hygroscopic
1
substance: tR ) 17.2 min; H-NMR (CD3OD) δ 8.42 (s, 1H),
7.8 (br s, 1H, exchangeable), 7.43 (s, 1H), 7.33 (s, 1H), 7.2 (m,
4H), 4.33 (m, 2H), 4.17 (t, 2H, J ) 7.5 Hz), 3.85 (m, 1H), 3.75
(m, 1H), 3.56 (s, 2H), 3.19 (m, 2H), 2.88 (t, 2H, J ) 7.5 Hz),
2.65 (t, 2H, J ) 7.2 Hz), 2-0.8 (m, 23H); HRMS calcd for
C32H51N6O4 583.3972 (M + H+), found 583.3961. Amino acid
analysis: Ser 1.00 (0.99), Lys 1.00 (1.00).
4-[4-(1H-Im id a zol-1-yl)p en tyl]p h en yla cetyl-Ser -Lys-N-
(2-cycloh exyleth yl)a m id e (36) was prepared in a manner
similar to 31 by coupling 29 with 12 as white hygroscopic
substance: 1H-NMR (CD3OD) δ 8.82 (s, 1H), 7.61 (s, 1H), 7.53
(s, 1H), 7.19 (d, 2H, J ) 8.1 Hz), 7.12 (d, 2H, J ) 8.1 Hz), 4.34
(m, 2H), 4.22 (t, 2H, J ) 7.2 Hz), 3.83 (m, 1H), 3.75 (m, 1H),
3.57 (s, 2H), 3.19 (m, 2H), 2.89 (t, 2H, J ) 7.2 Hz), 2.62 (t, 2H,
J ) 7.2 Hz), 1.92 (m, 3H), 1.72 (m, 10H), 1.8-1.1 (m, 10H),
0.92 (m 2H); HRMS calcd for C33H53N6O4 597.4128 (M + H+),
found 597.4079. Amino acid analysis: Ser 1.00 (0.99), Lys 1.00
(1.01).
1
substance: tR ) 19.6 min; H-NMR (CD3OD) δ 7.75 (m, 2H),
7.60 (m, 2H), 7.19 (d, 1H, J ) 8.1 Hz), 7.14 (d, 1H, J ) 8.1
Hz), 4.44 (m, 2H), 4.35 (m, 2H), 3.84 (m, 1H), 3.72 (m, 1H),
3.56 (s, 2H), 3.18 (m, 2H), 2.88 (t, 2H, J ) 7.2 Hz), 2.83 (s,
3H), 2.65 (t, 2H, J ) 7.2 Hz), 1.95 (m, 3H), 1.82-1.55 (m, 10H),
1.55-1.11 (m, 8H), 0.95 (m, 2H); HRMS calcd for C37H55N6O4
(M + H+) 647.4285, found 647.4329. Amino acid analysis: Ser
1.00 (0.96), Lys 1.00 (1.04).
4-[4-(2-Iod oim id a zol-1-yl)bu tyl]p h en yla cetyl-Ser -Lys-
N-(2-cycloh exyleth yl)a m id e (32) was prepared in a manner
similar to 31 by coupling 25 with 12 as a white hygroscopic
4-[4-(2-Meth yl-1H-im id a zol-1-yl)bu tyl]p h en yla cetyl-D-
Ser -D-Lys-(2-cycloh exyleth yl)a m id e (40). BOC(2-Cl-Cbz)-
D-Lys-Merrifield resin (0.4 g of 0.63 mmol/g substitution) was
treated with 50% TFA in CH2Cl2 for 30 min in a glass shaker
vessel to remove the BOC protecting group. The amino acid-
resin was then washed with CH2Cl2 (3 × 20 mL), 10% DIEA
in CH2Cl2 (3 × 20 mL), 2-propanol, and CH2Cl2 (3 × 20 mL).
A solution of BOC(O-Bn)-D-Ser (0.3 g, 1.0 mmol) in CH2Cl2 (3
× 20 mL) was treated with DCC (0.1 g, 0.5 mmol) and stirred
for 20 min at ambient temperature, and the resulting mixture
was then added to the amino acid-resin. After the resin was
shaken for 60 min, the resin was washed thoroughly with CH2-
Cl2 (3 × 20 mL), treated with 50% TFA in CH2Cl2 for 30 min,
and washed again with CH2Cl2 (3 × 20 mL), 10% DIEA in CH2-
Cl2 (3 × 20 mL), 2-propanol, and CH2Cl2 (3 × 20 mL). The
1
powder: tR ) 19.5 min; H-NMR (CD3OD) δ 7.60 (d, 1H, J )
1.8 Hz), 7.48 (d, 1H, J ) 1.8 Hz), 7.21 (d, 2H, J ) 8.1 Hz),
7.15 (d, 2H, J ) 8.1 Hz), 4.35 (m, 2H,), 4.16 (t, 2H, J ) 7.2
Hz), 3.80 (m, 2H), 3.56 (s, 2H), 3.18 (m, 2H), 2.89 (t, 2H, J )
7.2 Hz), 2.66 (t, 2H, J ) 7.5 Hz), 1.85 (m, 2H), 1.66 (m, 10H),
1.55-1.1 (m, 9H), 0.85 (m, 2H); HRMS calcd for C34H50IN6O4
(M + H+)709.2937, found 709.2884. Amino acid analysis: Ser
1.00 (1.00), Lys 1.00 (1.00).
11-(1H-Im id a zol-1-yl)u n d eca n oyl-Ser -Lys-(2-cycloh ex-
yleth yl)a m id e (13) was prepared in a manner similar to 31
by coupling 11 with 12 as a white glassy substance: tR ) 19.8
min; 1H-NMR (CD3OD) δ 8.55 (s, 1H), 7.5 (s, 1H), 7.39 (s, 1H),
4.35 (m, 2H), 4.19 (t, 2H, J ) 7.2 Hz), 3.8 (m, 2H), 3.2(m, 2H),
2.94 (m, 2H), 2.27 (t, J ) 7.2 Hz, 2H), 2-0.8 (m, 35H); HRMS
calcd for C31H57N6O4 (M + H+) 577.4442, found 577.4452.
Amino acid analysis: Ser 1.00 (0.98), Lys 1.00 (1.02).
Cbz-Lys(BOC)-Ch a -OMe (71). A solution of BOC-Cha-
OMe (1.0 g, 3.5 mmol) in dichloromethane (4.00 mL) was
treated with trifluoroacetic acid (1.00 mL), and the mixture
was stirred at room temperature for 1 h. After removal of the
solvents under reduced pressure, the residue was triturated
with ether and filtered. The solid material thus obtained was
washed thoroughly with ether and dried in a desiccator under
vacuum over NaOH pallets to give the trifluoroacetate salt
(0.87 g), which was used without further purification in the
following step.
peptide-resin thus obtained was suspended in
a solvent
mixture of DMF-DMA (5 mL), 24 (0.086 g, 0.25 mmol), DCC
(0.057 g, 0.28 mmol), HOBt (0.034 g, 0.25 mmol), and diiso-
propylethylamine (0.04 mL, 0.5 mmol) were added, and the
mixture was shaken for 18 h at room temperature. The
product was divided in half, and one portion reacted with a
mixture of cyclohexylethylamine (1 mL) in trifluroethanol (0.5
mL) at 50 ˚C for 5 h under N2. The free peptide was
concentrated under vacuum, and the amino acid side chain
protecting groups were removed with 90% HF/anisole for 60
min at 0 °C. The crude peptide was concentrated under
vacuum and purified to 99% purity on RP C-18, using a linear
gradient of 5-55% acetonitrile (0.05% TFA) in water (0.05%
TFA) over 25 min. The product was lyophilized to give 19 mg
of 40 as a white powder: tR ) 14.6 min; ES/MS m/ z 597 (M +
H). Amino acid analysis: Ser 1.00 (1.04), Lys 1.00 (0.96).
To a solution of Cbz-Lys(BOC)-OH (1.3 g, 3.4 mmol) (70)
and HOBt (0.6 g, 4.0 mmol) in dichloromethane (8.00 mL) and

2624 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16
Devadas et al.
dimethylacetamide (2.0 mL) was added dropwise a solution
of DCC (0.74 g, 3.6 mmol) in dichloromethane (10 mL), and
the mixture was stirred at 0 °C for 1 h at 0 °C. The reaction
mixture was filtered, and the filtrate was added to a solution
of the trifluoroacetate salt as prepared above in dichlo-
romethane (3.0 mL) and added N-methylmorpholine (0.37 g,
3.68 mmol). After being stirred at room temperature for 16
h, the reaction mixture was concentrated under vacuum. The
residue was partitioned between EtOAc (40 mL) and 5% citric
acid (20 mL). The organic phase was washed successively with
5% citric acid (2 × 20 mL), water (2 × 20 mL), 0.25 N NaOH
(2 × 20 mL) and brine and dried (Na2SO4). After removal of
the solvent, the product was crystallized from EtOAc/hexane
to give 1.4 g (73%) of the dipeptide ester 71 as a white
powder: mp 98-100 °C; 1H-NMR (CDCl3) δ 7.36 (m, 5H), 6.31
(br, 1H), 4.41 (br, 1H), 5.11 (s, 2H), 4.63 (m, 2H), 4.16 (m, 1H),
3.73 (s, 3H), 3.08 (m, 2H), 2-1.45 (m, 12H), 1.42 (s, 9H), 1.4-
0.8 (m, 7H); FAB-MS m/ z 554 (M + Li); HRMS calcd for
C29H45N3O7Li (M + Li) 554.3418, found 554.3447.
appropriate fractions were combined and freeze-dried to afford
73a (0.053 g, 34%) as a white powder: tR ) 19.8 min; 1H-NMR
(CD3OD) δ 7.45 (d, 1H, J ) 2.1 Hz), 7.38 (d, 1H, J ) 2.1 Hz),
7.22 (m, 4H), 4.41 (m, 3H), 4.13 (m, 2H), 3.81 (m, 2H), 3.58 (d,
2H, J ) 2.4 Hz), 2.92 (m, 2H), 2.67 (m, 2H), 2.57 (s, 3H), 1.95-
0.85 (m, 23H); FAB-MS m/ z 641 (M + H); HRMS calcd for
C34H53N6O6 (M + H), 641.4027, found 641.4041. Amino acid
analysis: Ser 1.00 (1.02), Lys 1.00 (1.00).
(R,S)-N-[2-[4-[4-(2-Meth yl-1H-im id a zol-1-yl)bu tyl]p h e-
n yl]-1-oxop r op yl]-L-Ser -L-Lys-L-Ch a -OH (73b) was pre-
pared in a manner similar to 73a , starting from 72 and 56:
yield 23% (a white powder); tR ) 21.6 min; 1H-NMR (CD3OD)
δ 7.37 (d, 1H, J ) 2.1 Hz), 7.30 (d, 1H, J ) 2.1 Hz), 7.17 (d,
1H, J ) 7.8 Hz), 7.05 (d, 1H, J ) 7.8 Hz), 4.35 (m, 2H), 4.23
(m, 1H), 4.03 (t, 2H, J ) 7.5 Hz), 3.64 (m, 3H), 2.84 (t, 2H, J
) 7.8 Hz), 2.57 (t, 2H, J ) 7.5 Hz), 2.49 (s, 3H), 2.0-1.05 (m,
24H), 0.85 (m, 2H); FAB-MS m/ z 655 (M + H); HRMS calcd
for C35H55N6O8 (M + H) 655.4183, found 655.4210. Amino acid
analysis: Ser 1.00 (1.00), Lys 1.00 (1.00).
Cbz-Ser (OtBu )-Lys(Boc)-Ch a -OMe (72). A solution of 71
(0.6 g, 1.1 mmol) in MeOH (15 mL) and acetic acid (0.07mL)
was hydrogenated at atmospheric pressure, in the presence
of 5% Pd/C (0.2 g) for 1 h. The catalyst was removed by
filtration, the filtrate was concentrated under reduced pres-
sure, and the resulting amine acetate (FAB-MS 420 (M + Li))
was used without further purification in the next step.
To a solution of Cbz-Ser(OtBu)-OH (0.4 g, 1.35 mmol) in
dimethylacetamide (1 mL) and dichloromethane (5 mL) at 0
°C were added HOBt (0.23 g, 1.53 mmol) and DCC (0.3 g, 1.45
mmol), and the mixture was stirred for 1 h. The reaction
mixture was filtered, and the filtrate was added to a solution
of the amine acetate in dimethylacetamide (1 mL) containing
N-methylmorpholine (0.14 g, 1.36 mmol). After the resulting
mixture was stirred at room temperature for 16 h, the solvents
were removed in vacuo, and the residue was partitioned
between cold 0.25 N NaOH (25 mL) and ethyl acetate (25 mL).
The organic phase was washed successively with cold 0.25 N
NaOH (2 × 25 mL), water, 5% citric acid (2 × 15 mL), and
water, dried (Na2SO4), and concentrated under reduced pres-
sure. The resulting material was crystallized from EtOAc/
hexane to give 72 (0.48 g, 63%) as a white powder: m p 108-
4-[4-(2-Meth yl-1H-im id a zol-1-yl)bu tyl]p h en yla cetyl-D-
Ser -D-Lys-D-Ch a -OH (73d ). The peptide was assembled by
the solid-phase method of synthesis.22 To a suspension of
hydroxymethylpolystyrene resin (3.0 g, 0.67 mmol of OH/g of
resin) in dichloromethane (40 mL) at 0 °C were added Boc-â-
cyclohexyl-D-Ala (1.08 g, 4.0 mmol), DCC (0.82 g, 4.0 mmol),
and DMAP (0.05 g, 0.4 mmol). The mixture was allowed to
warm to room temperature and shaken in a glass vessel for
18 h. The amino acid-resin was washed successively with
dichloromethane (25 mL), methanol (25 mL), and dichlo-
romethane (25 mL). The resulting resin 74 was treated with
50% TFA (trifluoroacetic acid) in CH2Cl2 (25 mL) for 30 min
and washed successively with CH2Cl2 (25 mL), 2-propanol (25
mL), 10% DIEA, and dichloromethane (25 mL). This resin was
then treated with a stirred solution of BOC-(2-Cl-Cbz)-D-Lys
(3.32 g, 8.0 mmol) and DCC (0.82 g, 4.0 mmol) in dichlo-
romethane (40 mL) and shaken for 60 min. The peptide resin
was then washed with dichloromethane, methanol, and dichlo-
romethane to obtain dipeptide-resin product 75. This was
treated with 50% trifluoroacetic acid in dichloromethane for
30 min, followed by washing with dichloromethane (25 mL),
2-propanol (25 mL), 10% DIEA, and dichloromethane (25 mL).
The resulting resin was treated with a stirred solution of BOC-
(O-Bn)-D-Ser (2.36 g, 8.0 mmol) and DCC (0.82 g, 4.0 mmol)
in dichloromethane (40 mL) and shaken for 60 min. After the
resin was washed successively with dichloromethane (25 mL),
methanol (25 mL), and dichloromethane (25 mL), the resulting
tripeptide-resin product 76 was treated with 50% TFA in
dichloromethane (25 mL) and washed with dichloromethane
(25 mL), 2-propanol (25 mL), 10% DIEA, and dichloromethane
(25 mL). This resin was suspended in DMF/DME (9/1, v/v,
40 mL); 24 (0.2 g, 0.65 mmol), HOBt (0.09 g, 0.65 mmol), DCC
(0.15 g, 0.72 mmol), and DIEA (0.11 mL, 0.65 mmol) were
added; and the mixture was allowed to shake for 18 h. The
crude product was liberated from the resin, and the protecting
groups were removed, by treatment with 90% HF/anisole for
60 min at 0 °C. The resulting crude peptide was purified to
98% purity on RP C-18 HPLC, using a linear gradient of
5-55% acetonitrile (0.05% TFA) in water (0.05% TFA) over
25 min. The appropriate fractions were pooled and lyophilized
to give 280 mg of 73d as a white powder: tR ) 19.98 min; 1H-
NMR (CD3OD) δ 7.46 (d, 1H, J ) 2.1 Hz), 7.39 (d, 1H, J ) 2.1
Hz), 7.22 (d, 2H, J ) 8.1 Hz), 7.14 (d, 2H, J ) 8.1 Hz), 4.42
(m, 3H), 4.12 (t, 2H, J ) 7.5 Hz), 3.79 (m, 2H), 3.57 (s, 2H),
2.89 (t, 2H, J ) 7.5 Hz), 2.66 (t, 2H, J ) 7.5 Hz), 2.58 (s, 3H),
2.0-1.1 (m, 21 H), 0.95 (m, 2H); HRMS calcd for C34H53N6O6
(M + H+) 641.4027, found 641.4043. Amino acid analysis: Ser
1.00 (0.98), Lys 1.00 (1.00).
1
109 °C; H-NMR (CDCl3) δ 7.37 (m, 4H), 7.18 (d, 1H, J ) 9.6
Hz), 6.41 (br, 1H), 5.68 (br, 1H), 5,12 (s, 2H), 4.69 (m, 1H),
4.59 (m, 1H), 4.43 (q, 1H, J ) 7.2 Hz), 4.24 (m, 1H), 3.83 (m,
1H), 3.73 (s, 3H), 3.4 (t, 1H, J ) 7.5 Hz), 3.07 (m, 2H), 2-1.45
(m, 11H), 1.43-1.2 (m, 3H), 1.19 (s, 9H), 1.12-0.8 (m, 5H);
FAB-MS m/ z 697 (M + Li), 641, and 597; HRMS calcd for
C36H59N4O9 (M + H) 691.4282, found 691.4273.
4-[4-(2-Met h yl-1-im id a zol-1-yl)b u t yl]p h en yla cet yl-L-
Ser -L-Lys-L-Ch a -OH (73a ). A solution of 72 (0.2 g, 0.29
mmol) in MeOH (10.0 mL) was hydrogenated at room tem-
perature for 1 h in the presence of 5% Pd/C (0.05 g). The
catalyst was removed by filtration, the filtrate was concen-
trated under reduced pressure, and the residue was dried in
a desiccator for 2 h in vacuo to afford the amine (FAB-MS m/ z
563 M + Li) which was used as such in the following step.
To a solution of 24 (0.12 g, 0.39 mmol) and HOBt (0.065 g,
0.43 mmol) in dimethylacetamide (2.5 mL) was added DCC
(0.085 g, 0.41 mmol), and the mixture was stirred at 0 °C for
2 h. It was then treated with a solution of the amine as
prepared above in dimethylacetamide (0.5 mL) containing
N-methylmorpholine (0.041g, 0.41 mmol), and DMAP (0.005
g) was added. The reaction mixture was stirred at room
temperature for 24 h and concentrated in vacuo. The residue
was partitioned between EtOAc (25 mL) and cold 0.25 N NaOH
(10 mL). The organic phase was washed with water (3 × 15
mL), dried (Na2SO4), and concentrated under reduced pressure
to give an amorphous substance which was crystallized from
ether/hexane and stirred with 1 M LiOH, (0.3 mL), containing
MeOH (0.2 mL), for 2 h at room temperature. The mixture
was acidified with 5% citric acid and extracted with EtOAc (3
× 15 mL). The combined organic extracts were washed with
water (3 × 10 mL), dried (Na2SO4), and concentrated. The
residue was dried in vacuo for 16 h and treated with trifluo-
roacetic acid (1.5 mL). After being stirred for 4 h at room
temperature, the solution was concentrated under reduced
pressure and residue was purified by reverse-phase HPLC. The
4-[4-(2-Meth yl-1H-im id a zol-1-yl)bu tyl]p h en yla cetyl-L-
Ser -L-Lys-L-Ch a -OMe (73c) was prepared in
a manner
similar to 31 starting from 72 and 24: tR ) 21.3 min; 1H-NMR
(CD3OD) δ 7.36 (d, 1H, J ) 2.1 Hz), 7.28 (d, 1H, J ) 2.1 Hz),
7.21 (d, 2H, J ) 8.1 Hz), 7.14 (d, 2H, J ) 8.1 Hz), 4.41 (m,3
H), 4.09 (t, 2H, J ) 7.5 Hz), 3.81 (m, 2H), 3.69 (s, 3H), 3.57 (s,
2H), 2.89 (t, 2H, J ) 7.5 Hz), 2.68 (t, 2H, J ) 7.5 Hz), 2.53 (s,
3H), 2.0-1.1 (m, 21H), 0.85 (m, 2H); FAB-MS m/ z 655 (M +

Inhibitors of Candida albicans NMT
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 16 2625
(15) Rocque, W. J .; McWherter, C. A.; Wood, D. C.; Gordon, J . I. A
comparative analysis of the kinetic mechanism and peptide
substrate specificity of human and Saccharomyces cerevisiae
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1993, 268, 9964-9971.
(16) Rudnick, D. A.; Rocque, W. J .; McWherter, C. A.; Toth, M. V.;
J ackson-Machelski, E.; Gordon, J . I. Use of photoactivatable
peptide substrates of Saccharomyces cerevisiae myristoyl-CoA:
protein N-myristoyl-transferase (Nmt1p) to characterize a myris-
toyl-CoA-Nmt1p-peptide ternary complex and to provide evi-
dence for an ordered reaction mechanism. Proc. Natl. Acad. Sci.
U.S.A. 1993, 90, 1097-1091.
(17) McWherter, C. A.; Rocque, W. J .; Zupec, M. E.; Freeman, S. K.;
Brown, D. L.; Devadas, B.; Getman, D. P.; Sikorski, J . A.;
Gordon, J . I. Scanning alanine mutagenesis and de-peptidization
of a Candida albicans myristoyl-CoA:protein N-myristoyltrans-
ferase octapeptide substrate reveals three elements critical for
molecular recognition. J . Biol. Chem. 1997, 272, 11874-11880.
(18) Stearns, T.; Kahn, R. A.; Botstein, D.; Hayt, M. A. ADP
ribosylation factor is an essential protein in Saccharomyces
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(19) (a) Yoshimoto, T.; Yamamoto, S.; Hayaishi, O. Selective inhibi-
tion of prostaglandin endoperoxide thromboxane isomerase by
1-carboxyalkylimidazoles. Prostaglandins 1978, 16 (4), 529-540.
(b) Devadas, B.; Tinabao, Lu; Katoh, A.; Kishore, N. S.; Wade,
A. C.; Mehta, P. P.; Rudnick, D. A.; Bryant, M. L.; Adams, S. P.;
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H); HRMS calcd for C35H55N6O8 (M + H) 655.4183, found
655.4225. Amino acid analysis: Ser 1.00 (0.95), Lys 1.00
(1.05).
Ack n ow led gm en t. This work was supported in part
by a grant from the National Institute of Health (AI
38200). The authors thank Mr. J ames F. Zobel for
providing amino acid analyses and Mr. J ames P. Doom
and Charles A. Gloeckner of Monsanto Company for
obtaining FAB mass spectral data. We also thank Dr.
J ohn J . Likos for technical help in obtaining proton-
COSY NMR spectra.
Refer en ces
(1) J ohnson, D. R.; Bhatnagar, R. S.; Knoll, L. J .; Gordon, J . I.
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