Vanilloid and isovanilloid analogues as inhibitors of methionyl-tRNA and isoleucyl-tRNA synthetases

Article abstract of DOI:10.1016/S0960-894X(01)00096-8

As aminoacyl adenylate surrogates, a series of methionyl and isoleucyl phenolic analogues containing bioisosteric linkers mimicking ribose have been investigated. Inhibition of synthesized compounds to the aminoacylation reaction by the corresponding Escherichia coli methionyl-tRNA and isoleucyl-tRNA synthetases indicated that 18 was found to be a potent inhibitor of isoleucyl-tRNA synthetase. A molecular modeling study demonstrated that in 18, isovanillate and hydroxamate served as proper surrogates for adenine and ribose in isoleucyl adenylate, respectively.

Full text of DOI:10.1016/S0960-894X(01)00096-8

Bioorganic & Medicinal Chemistry Letters 11 (2001) 965–968
Vanilloid and Isovanilloid Analogues as Inhibitors
of Methionyl-tRNA and Isoleucyl-tRNA Synthetases
Jeewoo Lee,a,* Sang Uk Kang,a Su Yeon Kim,a Sung Eun Kim,a Yeong Joon Job
and Sunghoon Kimc
aLaboratory of Medicinal Chemistry, College of Pharmacy, Seoul National University, Shinlim-Dong, Kwanak-Ku, Seoul 151-742,
South Korea
bImaGene Co., Sung Kyun Kwan University, 300 Chunchundong, Jangangu, Suwon, Kyunggido 440-746, South Korea
cNational Creative Research Initiative Center for ARS Network, Sung Kyun Kwan University, 300 Chunchundong, Jangangu, Suwon,
Kyunggido 440-746, South Korea
Received 24 November 2000; accepted 31 January 2001
Abstract—As aminoacyl adenylate surrogates, a series of methionyl and isoleucyl phenolic analogues containing bioisosteric linkers
mimicking ribose have been investigated. Inhibition of synthesized compounds to the aminoacylation reaction by the corresponding
Escherichia coli methionyl-tRNA and isoleucyl-tRNA synthetases indicated that 18 was found to be a potent inhibitor of isoleucyl-
tRNA synthetase. A molecular modeling study demonstrated that in 18, isovanillate and hydroxamate served as proper surrogates
for adenine and ribose in isoleucyl adenylate, respectively. # 2001 Elsevier Science Ltd. All rights reserved.
Introduction
of sugar. These ribose surrogates have been successfully
utilized for pharmacologically active adenylates in drug
design.3
In the preceding communication,1 we described the
structure–activity relationshipof ester, and the hydrox-
amate analogues of methionyl and isoleucyl adenylate,
as methionyl-tRNA (MetRS) and isoleucyl-tRNA
synthetase (IleRS) inhibitors.
In this paper, we describe the synthesis of designed
methionyl and isoleucyl vanilloids (or isovanilloids)
containing ribose bioisosteres and their inhibitory
activities to Escherichia coli MetRS and IleRS.
In our continuing effort to find potent aminoacyl-tRNA
synthetases (aaRSs) inhibitors, based on the ester ana-
logue of aminoacyl adenylate (1) as a lead compound,2
we modified the adenine and ribose parts with their
appropriate bioisosteres, as depicted in Figure 1. Firstly,
the adenine moiety was replaced by vanilloid or iso-
vanilloid, in which phenolic hydroxyl and the methoxy
groupwas supposed to mimic 6-NH
of adenine as a
2
hydrogen bonding donor, and N-1 of adenine as a
hydrogen bonding acceptor, respectively. Secondly, the
ribose was substituted by its biosteres, such as acyclic
amide and hydroxamate (2), dihydroisooxazole (3) and
dihydrooxazole (4). In particular, the hydroxamate
grouphas been attractive in terms of its binding mode,
in which carbonyl oxygen is able to mimic the ring
oxygen of ribose, and N-hydroxyl serves as 20-hydroxyl
*Corresponding author. Tel.: +82-2-880-7846; fax: +82-2-888-0649;
e-mail: jeewoo@snu.ac.kr
Figure 1.
0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00096-8

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J. Lee et al. / Bioorg. Med. Chem. Lett. 11 (2001) 965–968
Scheme 1. Reagents and conditions: (a) MOMCl, i-Pr2NEt, DMF, 85%; (b) NH2CH2CH2OH, 100 ꢀC, 90%; (c) N-Boclle or N-BocMet, DCC,
DMAP, CH2Cl2, 94%; (d) TFA, anisole, 90%; (e) NaOH, THF–H2O, 85%; (f) t-BuOCOCl, 4-MNO, CH2Cl2; then HClNH2OPMB, 60%; (g) NaH,
BrCH2CH2OTBS, DMF, 35%; (h) Bu4NF, THF, 92%.
Scheme 2. Reagents and conditions: (a) MOMCl, i-Pr2NEt, DMF; (b) NaOH, THF–H2O; (c) pentafluorophenol, DCC, DMAP, CH2Cl2, 75% in
three steps; (d) NH2CH2CH2OH, DMF, 95%; (e) N-Bocelle or N-BocMet, DCC, DMAP, CH2Cl2, 93%; (f) TFA, anisole, 92%; (g) NH2OPMB,
DMF, 92%; (h) HOCH2CH2OTBS, PPh3, DEAD, THF, 84%; (i) Bu4NF, THF, 50%; (j) NaH, BnBr, DMF, 95%; (k) NaBH4, MeOH; (l) MsCl,
NEt3, CH2Cl2; (m) NaCN, DMF, 20% in three steps; (n) 30% NaOH, reflux, 70%.
amide (15, 16) and hydroxamate analogues (17, 18) were
prepared from isovanillic acid (14) by following the
same method described previously. The synthetic
Synthesis
The syntheses of vanillate and isovanillate analogues are
outlined in Scheme 1. Commercially available vanillic
acid (5) was protected by the methoxymethyl group on
both acid and hydroxyl to give 6, which reacted directly
with 2-aminoethyl alcohol to afford amide 7. The con-
densation of alcohol 7 with N-Boc methionine or N-Boc
isoleucine was followed by acidic hydrolysis to provide
vanillic amides 8 and 9, respectively. For the synthesis
of vanillic hydroxamate analogues, ester 6 was hydro-
lyzed and then condensed with O-4-methoxybenzylhy-
droxylamine4 to give hydroxamate 10. The N-alkylation
of 10 with O-TBS 2-bromoethanol followed by desilyla-
tion afforded alcohol 11, which was converted into 12
and 13 by following the amide procedure. Isovanillic
sequence employed for homovanillate and homo-
isovanillate analogues was depicted in Scheme 2.
Homovanillic acid (19) was converted into the corre-
sponding pentafluoroester 20 by three steps. The con-
densation of 20 with 2-aminoalcohol gave alcohol 21,
which was then converted into homovanillic amide ana-
logues 22 and 23 by following the sequence of reactions
described in Scheme 1. The condensation of 20 with O-4-
methoxybenzyl hydroxylamine, the Mitsunobu reaction
with monoprotected ethylene glycol followed by desilyla-
tion, produced alcohol 24, which was conventionally
converted into homovanillic hydroxamate analogues 25
and 26, respectively. For the synthesis of homo-
isovanillate analogues, isovanillin was transformed into

J. Lee et al. / Bioorg. Med. Chem. Lett. 11 (2001) 965–968
967
Scheme 3. Reagents and conditions: (a) MOMCl, i-Pr2NEt, CH2Cl2, 95%; (b) HCl–NH2OH, NaHCO3, EtOH, 95%; (c) NCS, DMF, 80%;
(d) CH2¼CHCO2CH3, NaOCl, CH2Cl2, 70%; (e) LiAlH4, ether, 80%; (f) N-Boclle, DCC, DMAP, CH2Cl2, 92%; (g) TFA, anisole, 97%.
Scheme 4. Reagents and conditions: (a) NaOH, THF–H2O, 85%; (b) t-BuOCOCl, 4-NMO, CH2Cl2; then l(or d)-serine methyl ester, 70%; (c) PPh3,
DEAD, THF, 91%; (d) NaBH4, MeOH, 60%; (e) N-Boclle (or N-BocMet), DCC, DMAP, CH2Cl2, 90–92%; (f) TFA, anisole, 90–94%.
the corresponding pentafluoroester 29 by six steps, which
was converted into 30–33 by following the same sequence
used for the synthesis of homovanillate analogues.
elongated hydroxamate, also proved less potent than 18
by ca. 3- and 4.5-fold, respectively. Obviously, the spe-
cific interaction of the two hydroxyls of phenolic and
hydroxmate in 18 would be required for its inhibitory
activity. Dihydroisooxazole (38, 40) and dihydrooxazole
analogues (44, 46) showed moderate inhibitory activities
(IC50=38.6–79 mM) as IleRS inhibitors.
The dihydroisooxazole analogues were synthesized as
represented in Scheme 3. Aldehyde of vanillin (or iso-
vanillin) was transformed into a dihydroisooxazole
derivative 36 by four steps via nitrile oxide 1,3-dipolar
addition.5 The sequence described in Scheme 1 provided
the dihydroisooxazoles 37–40. For the synthesis of
dihydrooxazole analogues, as shown in Scheme 4, ester
6 was hydrolyzed and then condensed with l(or d)-ser-
ine methyl ester to give 41, which was cyclized into 42
by intramolecular O-alkylation. The same method that
was described in Scheme 3 with 42 produced the
dihydrooxazoles 43–46.
Concerning the MRS inhibitors, although the vanillic
hydroxamate analogue 12 appeared to be optimal in
this series, most of the compounds displayed moderate
inhibition to E. coli MetRS (IC50=32.5–163.5 mM)
(Table 1).
A molecular modeling study of 18 as a potent E. coli
IleRS inhibitor was conducted by overlapping 18 onto
isoleucyl adenylate as a standard. The energy-minimized
structure of isoleucyl adenylate was obtained based on
reported X-ray structures of several aminoacyl
adenylates bound to aminoacyl-tRNA synthetases.7
Biological Results and Discussion
Synthesized vanillate and isovanillate analogues were
evaluated as inhibitors of E. coli MetRS or IleRS. Their
inhibitory activities were determined by measuring the
decrease of the aminoacylation product, the [35S]methio-
nyl tRNAMet or [3H]isoleucyl tRNAIle of E. coli, in the
presence of different chemical concentrations.6
Table 1. Enzyme inhibitory activities of synthesized compounds
toward MetRS and IleRS
MRS
Inhibitors
E. coli MRS
(IC50, mM)
IRS
inhibitors
E. coli IRS
(IC50, mM)
8
60
32.5
78
72
94
128.5
163.5
87
58
47
9
13
16
18
23
26
31
33
38
40
44
46
64.4
12.8
106
4.5
Among IleRS inhibitors tested, the isovanillic hydrox-
amate analogue 18 was found to be the most potent one
with IC50=4.5 mM. The fact that its amide surrogate 16
(IC50=106 mM) displayed a 24-fold decrease as inhi-
bitor, indicated that the hydroxyl of hydroxamate
played an important role in interacting with enzymes, and
probably acted as a mimic of 20-OH in isoleucyl adenylate
as expected. Vanillic hydroxamate 13 (IC50=12.8 mM)
with a phenolic hydroxyl at the para-position and homo-
isovanillic surrogates 33 (IC50=21 mM) with a one-carbon
12
15
17
22
25
30
32
37
39
43
45
45
228
56.5
21
79
57.6
38.6
50
33.5
39

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J. Lee et al. / Bioorg. Med. Chem. Lett. 11 (2001) 965–968
is in progress to find more potent and selective inhibitors
of aminoacyl-tRNA synthetases based on 18.
Acknowledgements
This work was supported in part by a grant (HMP-00-
CH-15-0014) from the Ministry of Health & Welfare
and by the Brain Korea 21 Project to J. Lee, as well as a
grant from the National Creative Research Initiatives
from Ministry of Science and Technology to S. Kim.
Figure 2. Superposition of 18 (green) on isoleucyl adenylate (wheat):
only matched pharmacophores are colored: oxygen (red), nitrogen
(blue).
References and Notes
The low-energy structure of 18 was calculated using the
SYBYL 6.6 program.8 We performed a molecular
alignment and pharmacophore detection for two
energy-minimized molecules using GASP (Genetic
Algorithm Similarity Program, Tripos). During the cal-
culation, the conformation of isoleucyl adenylate was
fixed as a reference structure. In the result, 18 was
aligned to isoleucyl adenylate to match with five phar-
macophores (amino and carbonyl in isoleucine, oxygen
and 20-hydroxyl on ribose, and amine of adenine). In the
overlapped structure, shown in Figure 2, 18 aligns itself
with key pharmacophores of isoleucyl adenylate, pro-
viding that isovanillic and hydroxamate moieties are
adequate surrogates for adenine and ribose, respec-
tively. The overlay will be useful information for the
design of improved related compounds.
1. Lee, J.; Kang, S. U.; Kim, S. Y.; Kim, S. E.; Jo, Y. J.; Kim,
S. Bioorg. Med. Chem. Lett. 2001, 11, 961.
2. Lee, J.; Kang, S. U.; Kang, M. K.; Chun, M. W.; Jo, Y. J.;
Kwak, J. H.; Kim, S. Bioorg. Med. Chem. Lett. 1999, 9, 1365.
3. Kleinman, E. F.; Campbell, E.; Giordano, L. A.; Cohan,
V. L.; Jenkinson, T. H.; Cheng, J. B.; Shirley, J. T.; Pettipher,
E. R.; Salter, E. D.; Hibbs, T.; DiCapua, F. M.; Bordner, J. J.
Med. Chem. 1998, 41, 266.
4. Kim, M. G.; Jung, J. C.; Sung, M. J.; Choi, Y. K.; An,
S. G.; Lee, S.-J.; Yoon, G.-J.; Park, M. H. Bioorg. Med. Chem.
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5. Carruthers, W. In Cycloaddition Reactions in Organic
Synthesis; Pergamon: Oxford, 1990; p285.
6. Jo, Y. J.; Lee, S. W.; Jo, M. K.; Lee, J.; Kang, M.-K.;
Yoon, J. H.; Kim, S. J. Biochem. Mol. Biol. 1999, 32, 547.
7. We extracted and superimposed X-ray structures of
aminoacyl adenylates embedded in aminoacyl-tRNA synthe-
tase in order to investigate their conformational similarity.
Since the conformations of aminoacyl adenylates examined
were quite similar, the structure of isoleucyl adenylate was
built and optimized to adopt their common conformation.
8. The structure of 18 was built and converted to a three-
dimensional structure using the CONCORD program, Tripos.
All molecular mechanics calculations were carried out with
SYBYL 6.6, Tripos on a Silicon Graphics INDIGO 2
workstation.
In summary, we investigated acyclic surrogates of
methionyl adenylate and isoleucyl adenylate as MetRS
and IleRS inhibitors having vanilloid (or isovanilloid)
and ribose bioisosteres. Isovanillic hydroxamate 18 dis-
played potent inhibitory activity to E. coli IleRS, and its
pharmacophores matched well with those of isoleucyl
adenylate by molecular modeling. Further investigation