Novel inhibitors of IMPDH: A highly potent and selective quinolone-based series

Article abstract of DOI:10.1016/S0960-894X(02)00944-7

A series of novel quinolone-based small molecule inhibitors of inosine monophosphate dehydrogenase (IMPDH) was explored. The synthesis and the structure-activity relationships (SARs) derived from in vitro studies are described.

Full text of DOI:10.1016/S0960-894X(02)00944-7

Bioorganic & Medicinal Chemistry Letters 13 (2003) 543–546
Novel Inhibitors of IMPDH:
A Highly Potent and Selective Quinolone-Based Series
Scott H. Watterson,* Marianne Carlsen, T. G. Murali Dhar, Zhongqi Shen,
William J. Pitts, Junqing Guo, Henry H. Gu, Derek Norris, John Chorba, Ping Chen,
Daniel Cheney, Mark Witmer, Catherine A. Fleener, Katherine Rouleau,
Robert Townsend, Diane L. Hollenbaugh and Edwin J. Iwanowicz
Bristol-Myers Squibb PRI, PO Box 400, Princeton, NJ 08543-4000, USA
Received 12 August 2002; accepted 1 October 2002
Abstract—A series of novel quinolone-based small molecule inhibitors of inosine monophosphate dehydrogenase (IMPDH) was
explored. The synthesis and the structure–activity relationships (SARs) derived from in vitro studies are described.
# 2002 Elsevier Science Ltd. All rights reserved.
Inosine 50-monophosphate dehydrogenase (IMPDH), a
key enzyme in the de novo synthesis of guanosine
nucleotides, catalyzes the irreversible NAD dependent
oxidation of inosine 50-monophosphate (IMP) to xan-
thosine 50-monophosphate (XMP).1ꢀ3 Two distinct
cDNA’s encoding IMPDH have been identified and
isolated. These transcripts, labeled type I and type II,
possess 84% sequence identity.4ꢀ6 IMPDH type II
activity is markedly up-regulated in actively proliferat-
ing cell types including cancers and activated peripheral
blood lymphocytes.7,8 IMPDH type I, on the other
hand, is the predominant, constitutively expressed iso-
form in normal, non-proliferating cells. Selective inhibi-
tors of the up-regulated IMPDH type II isoform may
mitigate toxicity caused by the inhibition of the con-
stitutively expressed IMPDH type I isoform.8 IMPDH
type II has emerged as a target for therapies in immu-
nology, virology, and oncology.7,9
exhibited from administration of either CellCept1 or
MPA limits its potential for the treatment of other
autoimmune disorders, such as psoriasis and rheuma-
toid arthritis.12
The formation of conjugates at either the carboxylic
acid or phenolic residues is implicated as a contributing
factor in the poor therapeutic index observed for Cell-
Cept1.13,14 Structurally related analogues, devoid of
acid and phenolic functionalities, are reputed to have an
improved therapeutic window with regard to dose lim-
iting GI toxicity.15ꢀ17 An alternative approach to
IMPDH inhibition has been described by Pankiewicz.18
Our focus is on the identification and development of
potent inhibitors of IMPDH with selectivity for
IMPDH type II and with improved pharmacological
properties. Recently, we have disclosed several new
classes of inhibitors of IMPDH,15,19ꢀ22 of which three
are exemplified by structures 2, 3, and 4 (Fig. 1). As we
examined urea isosteres in an effort to define the opti-
mal linkage between the two aryl groups of urea 1, we
became increasingly more concerned about the potential
liberation of an aniline moiety in vivo. In order to alle-
viate this concern, we directed our efforts toward struc-
tures in which the amide isostere of structure 3 may be
additionally appended to the 3-methoxy-5-oxazolylani-
line residue. In this paper, we have outlined the synthesis
and biological evaluation of a new class of potent inhib-
itors in which a vinylogous amide moiety is constrained
Mycophenolic acid (MPA, Fig. 1) and some of its deri-
vatives have been shown to be potent, uncompetitive,
reversible inhibitors of human IMPDH type I and type
II.10,11 CellCept1 (mycophenolate mofetil, MMF), a
prodrug of MPA, has been used clinically for the treat-
ment of transplant rejection, due to its inhibition of
IMPDH. Dose-limiting gastrointestinal (GI) toxicity
*Corresponding author. Tel.: +1-609-252-6778; fax: +1-609-252-
6601; e-mail: scott.watterson@bms.com
0960-894X/03/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(02)00944-7

544
S. H. Watterson et al. / Bioorg. Med. Chem. Lett. 13 (2003) 543–546
7. The subsequent cyclization to form the quinolone
ring system (8) occurred during heating in xylene at
250 ꢁC. Alternatively, urethane 7 could be prepared
through a N-N exchange reaction25 with the N-methyl
enamino ester 9. Quinolone inhibitors were also pre-
pared utilizing a palladium-catalyzed carbonylation
sequence,26 as outlined in Scheme 1.
The in vitro inhibitory activity against IMPDH type II
for this series of quinolone inhibitors is summarized in
Tables 1 and 2. The phenyl substituted quinolone 5a
very effectively inhibited IMPDH type II with an IC50
of 0.008 mM.27 This compound exemplified an increase
in potency relative to the Vertex urea 1. With this high
binding affinity compound in hand, a study of various
simple residues for R1 and R2 was undertaken in order
to optimize the core structure. Replacement of the
phenyl ring with various heterocycles was explored.
Overall, substitution of the quinolone core with both
five- and six-membered heterocycles resulted in modest
losses in potency, as demonstrated by compounds 5b
and 5d–5h. When the quinolone was substituted with
the phenyl-like 3-thiophene substituent, activity was
maintained. When the phenyl group of 5a was removed
(5i), a loss in enzymatic activity of ꢂ37-fold was
observed (IC50=0.30 mM). Alkyl groups such as methyl
(5j) and t-butyl (5k) also resulted in significant reduction
in potency. Extension of the phenyl ring by one methy-
lene group was not as well tolerated (5l, IC50=0.18)
relative to 5a. Amino-quinolones 5m and 5n showed
moderate inhibitory activity against IMPDH type II,
although significantly less potent than 5a, with IC50
values of 0.22 and 0.20 mM, respectively. Introduction
of a methyl group at the 3-position of the quinolone
(5o) possibly perturbed the orientation of the phenyl
residue enough to result in an 8-fold decrease in
potency. Additionally, analogue 5a was determined to
be an uncompetitive, reversible inhibitor of IMPDH
type II with respect to IMP and NAD+ with a Ki=7ꢃ2
nM.19
Figure 1. Chemical structures of MPA, MMF, urea 1, cyano-guani-
dine 2, amide 3, and amino-oxazole 4 (BMS-337197).
to form a quinolone scaffold, as depicted in structure 5
(Fig. 2).
Through protein crystallographic studies, Vertex has
shown that analogues of 1 bind to IMPDH forming a
hydrogen bond between the urea NH and the carboxy-
late of Asp 274 (Fig. 2) in the active site of the enzyme.2
In developing the quinolone chemotype (5), we antici-
pated that the quinolone carbonyl would serve as a
hydrogen bond acceptor and engage in an interaction
with Gln 441 (Fig. 2). Additionally, the critical hydro-
gen bond interaction between the quinolone NH and
the carboxylate of Asp 27423 would be maintained.
The synthetic pathways utilized in the preparation of
the quinolones (5) are shown in Schemes 1 and 2. The
synthesis of 3-methoxy-4-(5-oxazolyl)-aniline 6 has been
previously described.16 The condensation of aniline 6
with various substituted acetoacetates24 was carried out
in toluene at reflux in the presence of catalytic p-tolue-
nesulfonic acid to give vinylogous urethane derivatives
As indicated in Table 1, our efforts to optimize the qui-
nolone core indicated that a phenyl substituent (5a) at
the 2-position was optimal. We subsequently focused
our attention on developing the basic SAR around the
phenyl ring, as outlined in Table 2. Exploration of the
ortho-, meta-, and para-tolyl analogues 12a, 12b, and
12g demonstrated that para- and meta-substitution is
favored over ortho-substitution with both 12b and 12g
showing an enhancement in potency relative to 5a.
Interestingly, the para-bromo analogue 12h (IC50=5
nM) was more potent than the corresponding meta-
derivative 12c. In general, highly polar residues such as
carboxylic acids 12e and 12k and methylsulfone 12f
were not as well tolerated, resulting in significant loss in
potency. Phenolic residues 12d and 12i provided a
modest loss in activity. A notable compound in this
series was the 4-methoxyphenyl quinolone 12j which
had a IC50 of <0.005 mM against IMPDH type II.
Additionally, the synthetic studies were expanded to
include di-substitution (12l–-12n). The 3,4-dimethyl-
phenyl quinolone (12l) was as potent as the mono-
methyl substituted analogues (12b and 12g).
Figure 2. Proposed binding model for the Vertex urea 1, and a
potential binding model for quinolone 5.

S. H. Watterson et al. / Bioorg. Med. Chem. Lett. 13 (2003) 543–546
545
Scheme 1. Methods for the preparation of the quinolones.
Scheme 2. Preparation of quinolone 5i.
Table 1. SAR of quinolone-based inhibitors of structure 5
Table 2. SAR of phenyl quinolone-based inhibitors of structure 12
R3
R4
IMPDH II IC50 (mM)
Compd
R1
R2
IMPDH II IC50 (mM)
Compd
MPA
1
NA
NA
H
H
H
H
H
H
H
H
H
H
H
H
H
H
Me
NA
NA
Phenyl
0.014
0.019
0.008a
0.032
0.009
0.063
0.034
0.043
0.070
0.046
0.30
5a
H
2-Me
3-Me
H
H
H
H
H
H
H
H
H
H
H
H
4-Me
5-Me
4-Me
0.008
0.065
0.005
0.022
0.018
0.16
12a
12b
12c
12d
12e
12f
12g
12h
12i
12j
12k
12l
12m
12n
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
5o
3-Faranyl
3-Thiophenyl
2-Thiophenyl
4-Thiazolyl
2-Pyridyl
3-Pyridyl
4-Pyridyl
H
3-Br
3-OH
3-CO2H
3-SO2Me
4-Me
0.16
<0.005
0.005
0.043
<0.005
>1
<0.005
0.029
0.019
4-Br
4-OH
4-OMe
4-CO2H
3-Me
2-Me
3-Br
Me
t-Bu
Benzyl
NHMe
N
Phenyl
0.11
>10
0.18
0.22
0.20
0.065
Me
2
aKi=7ꢃ2 nM (IMPDH-II).
Table 3. IMPDH type I versus type II and T-cell proliferation results
Compd
Type I
IC50 (mM)
Type II
IC50 (mM)
Ratio
TypeI/II
IC50
(mM)(CEM)
Several of the compounds which were potent inhibitors
of IMPDH type II were examined against IMPDH type
I in order to determine the relative selectivity.27 The
results are presented in Table 3. The parent phenyl qui-
nolone provided 12-fold selectivity for type II, com-
pared with urea 1 which was only ꢂ3-fold selective.
Compounds 12b, 12j, and 12l demonstrated higher
selectivity with ratios of type I to type II of 30, 18, and
17, respectively.
MPA
1
5a
0.055
0.055
0.099
0.055
0.15
0.046
0.090
0.083
0.014
0.019
0.008
0.009
0.005
<0.005
<0.005
<0.005
3.9
2.9
12
6.0
30
>9.0
>18
>17
0.39
0.32
0.69
0.34
1.1
0.56
0.37
0.41
5c
12b
12g
12j
12l

546
S. H. Watterson et al. / Bioorg. Med. Chem. Lett. 13 (2003) 543–546
These compounds were also examined in a T-cell pro-
liferation assay (CEM) (Table 3).29 In this assay, MPA
inhibited T-cell proliferation with an IC50 of 0.39 mM,
whereas the phenyl quinolone (5a) had an IC50 of 0.69
mM. Three compounds, 5c, 12j, and 12l, inhibited T-cell
proliferation comparable to MPA with IC50 values of
0.34, 0.37, and 0.41 mM, respectively.
17. Jain, J.; Almquist, S. J.; Shlyakhter, D.; Harding, M. W.
J. Pharm. Sci. 2001, 90, 625.
18. Pankiewicz, K. W.; Lesiak-Watanabe, K. B.; Watanbe,
K. A.; Patterson, S. E.; Jayaram, H. N.; Yalowitz, J. A.;
Miller, M. D.; Seidman, M.; Majumdar, A.; Prehna, G.;
Goldstein, B. M. J. Med. Chem. 2002, 45, 703.
19. Gu, H. H.; Iwanowicz, E. J.; Guo, J.; Watterson, S. H.;
Zhen, Z.; Pitts, W. J.; Dhar, T. G. M.; Fleener, C. A.; Rou-
leau, K.; Sherbina, N. Z.; Witmer, M.; Tredup, J.; Hollen-
baugh, D. Bioorg. Med. Chem. Lett. 2002, 12, 1323.
20. Pitts, W. J.; Guo, J.; Dhar, T. G. M.; Shen, Z.; Gu, H. H.;
Watterson, S. H.; Bednarz, M. S.; Chen, B.-C.; Barrish, J. C.;
Bassolino, D.; Cheney, D.; Fleener, C. A.; Rouleau, K. A.;
Hollenbaugh, D. L.; Iwanowicz, E. J. Bioorg. Med. Chem.
Lett. 2002, 12, 2137.
21. Watterson, S. H.; Liu; C.; Dhar; T. G. M.; Gu; H. H.;
Pitts; W. J.; Barrish; J. C.; Fleener; C. A.; Rouleau; K.; Sher-
bina; N. Z.; Hollenbaugh; D.; Iwanowicz; E. J. Bioorg. Med.
Chem. Lett. 2002, 12, 2879.
In summary, we have identified a novel quinolone-based
series of highly potent, selective inhibitors of IMPDH
type II. These compounds demonstrate that the urea
and amide isosteres can be effectively constrained to the
3-methoxy-5-oxazolylaniline through
a
vinylogous
amide to form a bicyclic quinolone scaffold. This series
has been the subject of further studies to enhance the
physiochemical properties, and the results are outlined
in the accompanying article.30
22. Iwanowicz; E. J.; Watterson; S. H.; Liu; C.; Gu; H. H.;
Barrish; J. C.; Fleener; C. A.; Rouleau; K.; Sherbina; N.
Z.; Hollenbaugh; D. Bioorg. Med. Chem. Lett. 2002, 12,
2931.
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