Low molecular weight indole fragments as IMPDH inhibitors

Article abstract of DOI:10.1016/j.bmcl.2006.01.089

The study of non-oxazole containing indole fragments as inhibitors of inosine monophosphate dehydrogenase (IMPDH) is described. The synthesis and in vitro inhibitory values for IMPDH II are discussed.

Full text of DOI:10.1016/j.bmcl.2006.01.089

Bioorganic & Medicinal Chemistry Letters 16 (2006) 2535–2538
Low molecular weight indole fragments as IMPDH inhibitors
Rebekah E. Beevers, George M. Buckley, Natasha Davies, Joanne L. Fraser,
Francis C. Galvin, Duncan R. Hannah,* Alan F. Haughan, Kerry Jenkins,
Stephen R. Mack, William R. Pitt, Andrew J. Ratcliffe, Marianna D. Richard,
Verity Sabin, Andrew Sharpe and Sophie C. Williams
UCB Celltech, Granta Park, Great Abington, Cambridge CB1 6GS, UK
Received 21 November 2005; revised 19 January 2006; accepted 19 January 2006
Available online 17 February 2006
Abstract—The study of non-oxazole containing indole fragments as inhibitors of inosine monophosphate dehydrogenase (IMPDH)
is described. The synthesis and in vitro inhibitory values for IMPDH II are discussed.
Ó 2006 Elsevier Ltd. All rights reserved.
Proliferation of T and B lymphocytes is dependent on
access to a large cellular pool of guanine nucleotides.
Within the de novo purine biosynthetic pathway a key
rate-limiting step is oxidation of inosine-50-monophos-
phate to xanthosine-50-monophosphate by the NAD-de-
pendent enzyme inosine monophosphate dehydrogenase
(IMPDH).1 Two isoforms of the enzyme have been iden-
tified and designated type I and type II.2 Of these it is
IMPDH type II that is upregulated in actively prolifer-
ating cell types.3,4 As a consequence inhibition of
IMPDH II has become an attractive immunology target
for the treatment of transplant rejection, psoriasis, sys-
temic lupus erythematosus and rheumatoid arthritis.5
OH
Me
Me
MPA, R=H
MMF, R=
O
O
OR
N
O
O
OMe
From protein/inhibitor crystallographic studies
a
binding model for VX497 complexed to IMPDH II
has been reported.8 Key interactions include forma-
tion of H-bonds between the NH of Gly 326 and
the oxazole moiety, and carboxylate of Asp 274 with
the urea NH’s. In addition, favourable hydrophobic
interactions result from binding of the methoxy in a
pocket defined by the side chains of Asn 303 and
Arg 332, and p-stacking of the phenyl oxazole with
the bound nucleotide precursor. Based on this similar
binding interactions have been suggested for BMS-
337197 and used in the design of new IMPDH II
inhibitors.10
Mycophenolic acid (MPA) is a potent uncompetitive
reversible inhibitor of both IMPDH I and II, which
has been approved for clinical utility in transplant rejec-
tion in the form of CellceptÒ or mycophenolate mofetil
(MMF), an ester prodrug of MPA.6
Despite clinical efficacy of CellceptÒ its use in other dis-
ease end points is compromised by dose-limiting gastro-
intestinal (GI) side effects and efforts have focused on
discovery of new IMPDH inhibitors with an improved
therapeutic window.7 Towards this end Vertex and
BMS have reported N-[3-methoxy-4-(5-oxazolyl)phen-
yl-containing compounds, VX4978 and BMS-3371979
as potent IMPDH II inhibitors.
H
N
O
O
N
O
O
O
O
MeO
N
H
N
VX-497
H
O
N
N
O
Me
N
N
BMS-337197
Keywords: IMPDH; Inosine monophosphate dehydrogenase;
Fragment.
*
Corresponding author. Fax: +44 1223 896400; e-mail: duncan.
hannah@ucb-group.com
O
MeO
N
H
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.01.089

2536
R. E. Beevers et al. / Bioorg. Med. Chem. Lett. 16 (2006) 2535–2538
We have disclosed two new classes of oxazole-contain-
ing IMPDH II inhibitors.11,12 Further detailed investi-
gation of these series revealed the oxazole moiety as a
potential source of reactive metabolites. Recently, a
number of non-oxazole nitrile containing inhibitors
have been reported in VX14813 and indole 1.14 This
report describes our preliminary efforts to discover
non-oxazole containing IMPDH II inhibitors using a
fragment optimisation approach.
H
N
N
a
N
H
N
H
5
Scheme 1. Reagents and condition: (a) 1,1-Diphenylethylene, Pd/C,
250 °C (30%).
H
OH
N
O
R1
CN
HO
B
NC
O
a, b
O
N
O S O
Ph
N
H
MeO
N
N
H
VX-148
H
6, 7
NC
Scheme 2. Reagents and condition: (a) R1-Br, Pd(PPh3)4, Na2CO3,
DME, H2O, microwave, 120–150 °C (74–81%); (b) NaOH, dioxane,
reflux; or KOH, MeOH, reflux (86%).
N
1
N
O
N
H
In a fragment study, commercially available 3-cyanoin-
dole (2) has been reported to inhibit IMPDH II at
approximately 30 lM.15 GOLD16 docking of 2 using
an in-house crystal structure of IMPDH II was under-
taken to define potential binding modes. Interestingly,
the best fit predicts the indole NH forming a H-bond
to Asp 274 with the nitrile potentially forming a hydro-
gen bond with Gly 326 (Fig. 1).
Br
R3
a, b
R
R
R3
N
O S O
Ph
N
H
I
R=H
R=Me
R3=NO2
R3=NH2
g
c
R1
e
R3
d
R
N
To further explore this concept with a view to improv-
ing the potency of 2 for IMPDH II, a series of indole
fragments containing hydrogen bond acceptor groups
at the 3-position were synthesised as described in
Schemes 1–4 or obtained from commercial sources.
A number of fragments substituted on the indole core
were also prepared, to establish precise structural
requirements and to assess potential sites for further
elaboration.
O S O
Ph
f
O
O
B
h, f
R1
R
N
O S O
Ph
R3
R
R3
N
H
II
8-14
Scheme 3. Reagents and conditions: (a) N-Bromosuccinimide, DCM,
rt (77–93%); (b) PhSO2Cl, TBAB, NaOH, toluene, H2O; or NaH,
THF; then PhSO2Cl (84–100%); (c) LDA, THF, ꢀ40 °C; then MeI,
ꢀ40 °C to ꢀ20 °C; (d) SnCl2, EtOH, H2O, reflux (37–83%); (e) R1-
B(OH)2, Pd(PPh3)4, Na2CO3, DME, H2O, microwave, 120–150 °C
(75–90%); (f) NaOH, dioxane, reflux; or, KOH, MeOH, reflux (12–
86%); (g) bis(pinacolato)diboron, Pd(dppf)Cl2, KOAc, DME, 80 °C
(or THF, microwave, 150 °C) (46%); (h) R1-Br or R1-Cl, Pd(PPh3)4,
Na2CO3, DME, H2O, microwave, 120–150 °C (51–90%).
3-Formylindole (3) was commercially available. The 3-
carbamoyl derivative 4 was prepared directly from 2,
by reaction with hydrogen peroxide and sodium hydrox-
ide in methanol. A series of heteroaromatic derivatives
were also prepared. The 4-pyridyl indole 5 was synthe-
sised by dehydrogenation of a commercially available
precursor (Scheme 1). 1-Phenylsulfonylindol-3-yl boron-
ic acid was reacted with aromatic bromides to prepare,
after basic hydrolysis of the indole protecting group, in-
doles 6 and 7 (Scheme 2). Using similar chemistry, a set
of related derivatives, additionally substituted on the in-
dole core, were prepared starting from 6- and 7-nitroin-
dole and 2-methylindole, (Scheme 3). The key step
employed either Suzuki reaction of bromoindoles I with
heteroaromatic boronic acids in the synthesis of 8, 10, 12
and 14, or Suzuki reaction of the pinacolboronate ester
II with heteroaromatic bromides (or chloride towards
11) in the synthesis of 9, 11 and 13. The 2-methyl group
of 14 was successfully introduced by deprotonation of
the bromo nitroindole I followed by methyl iodide
Gly
326
Asp274
Figure 1. Indole 2 docked into IMPDH II, with potential H-bonds.

R. E. Beevers et al. / Bioorg. Med. Chem. Lett. 16 (2006) 2535–2538
2537
as pyridine 5 and the five-membered furan 7, thiophene
8 and thiazole 9. With an IC50 of 1.15 lM, pyridine 5
demonstrated a significant improvement over 2. The
affinity was very sensitive to the position of heteroatom
in the pendant heterocycles, for example, the pyrid-3-
ylindole 6 was inactive, illustrating the importance of
this interaction with the enzyme. In the case of the 3-
(pyrid-4-yl)indoles, simple substitution was tolerated at
the indole 6- and 7-position (see 10, 12 and 15), with a
slight increase in affinity. Methylation at the 2-position
in 13 also gave an improvement in activity
(IC50 = 343 nM), however, these benefits were not addi-
tive (compare 14 with 10 and 13). Substitution on the
pyridine, however, did not appear to be tolerated (com-
pare 11 and 10).
I
a, b
N
Br
N
Br
H
O S O
Ph
N
c, d
N
Br
H
15
Scheme 4. Reagents and conditions: (a) I2, KOH, DMF, rt (quant.);
(b) NaH, THF; then PhSO2Cl (85%); (c) pyridin-4-ylboronic acid,
Pd(PPh3)4, K3PO4, DME, H2O, 80 °C (87%); (d) KOH, MeOH, reflux
(74%).
Indole nitrogen methylation had a dramatic effect on the
observed affinities of the fragments. Potency of the cya-
no and formylindoles was increased 3-fold on N-methyl-
ation (compare 16 and 17 with 2 and 3, respectively).
With five-membered heterocycles, illustrated with the
furan 19, potency was largely unaffected, but in the case
of 4-pyridylindoles, N-methylation reduced activity at
least 100-fold (compare 20 and 15).
quench. In a variant of the above, 6-bromoindole was
iodinated at the 3-position, allowing for the chemoselec-
tive Suzuki reaction towards pyridylindole 15 (Scheme
4). Indoles 2, 7 and 15 were N-methylated by treatment
with potassium hydroxide and methyl iodide in acetone
at room temperature to provide indoles 16, 19 and 20,
respectively. 3-Formyl-1-methylindole (17) was com-
mercially available, and amide 18 was prepared from
the nitrile 16 as in the synthesis of 4 above.
We were interested in establishing whether our small
fragments were actually binding at the IMPDH active
site, rather than exerting their effects through an alloste-
ric interaction. Classical steady-state kinetic analysis
with the most potent fragment, pyridylindole 13, dem-
onstrated uncompetitive inhibition with respect to
NAD and IMP, analogous to the behaviour of MPA
and VX497,8 supporting binding at the active site at
The in vitro potencies are shown in Table 1. The first
observation is the sensitivity to variations of the cyano
group for other small H-bond accepting groups, the
aldehyde 3 conferring similar potency, while the primary
amide 4 was inactive. A number of heteroaromatic
replacements of the nitrile also showed activity, such
Table 1. SAR of indoles
R1
R3
N
R2
17
IMPDH II IC50 (lM)
Compound
R1
R2
R3
2
3
CN
CHO
H
H
20.9
22.7
H
H
4
CONH2
Pyrid-4-yl
Pyrid-3-yl
Furan-3-yl
Thiophen-3-yl
Thiazol-5-yl
Pyrid-4-yl
2-Me-pyrid-4-yl
Pyrid-4-yl
Pyrid-4-yl
Pyrid-4-yl
Pyrid-4-yl
CN
H
H
IAa
5
H
H
1.15
IA
6
H
H
7
H
H
5.84
2.32
8
H
6-NH2
6-NH2
6-NH2
6-NH2
7-NH2
2-Me
2-Me–6-NH2
6-Br
H
9
H
27.9
0.637
IAa
10
11
12
13
14
15
16
17
18
19
20
21
H
H
H
0.524
0.343
0.419
0.408
7.66
H
H
H
Me
Me
Me
Me
Me
—
CHO
H
6.84
50% at 50 lM
6.21
CONH2
Furan-3-yl
Pyrid-4-yl
—
H
H
6-Br
—
62% at 50 lM
8.20
a Inactive at 50 lM.

2538
R. E. Beevers et al. / Bioorg. Med. Chem. Lett. 16 (2006) 2535–2538
Acknowledgment
Gly
326
Gly
326
The authors thank the Molecular and Cellular Systems
department at Granta Park for in vitro measurements.
References and notes
1. Jackson, R. C.; Weber, G. Nature 1975, 256, 331.
2. Collart, F. R.; Huberman, E. J. Biol. Chem. 1988, 263,
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Asp274
Asp274
3. Jayaram, H. N.; Grusch, M.; Cooney, D. A.; Krupitza, G.
Curr. Med. Chem. 1999, 6, 561.
Figure 2. Docking of 16 (left) and 5 (right) in IMPDH II.
4. Carr, S. F.; Papp, E.; Wu, J. C.; Natsumeda, Y. J. Biol.
Chem. 1993, 268, 27286.
the same point in the enzymatic path as these known
inhibitors.
5. Dhar, T. G. M.; Shen, Z.; Gu, H. H.; Chen, P.; Norris, D.;
Watterson, S. H.; Ballentine, S. K.; Fleener, C. A.;
Rouleau, K. A.; Barrish, J. C.; Townsend, R.; Hollenb-
augh, D. L.; Iwanowicz, E. J. Bioorg. Med. Chem. Lett.
2003, 13, 3557, and references cited therein.
6. Anderson, W. K.; Boehm, T. L.; Makara, G. M.; Swann,
R. T. J. Med. Chem. 1996, 39, 46.
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Nishimura, P.; Kawano, M.; Holt, C. D. Pharmacotherapy
1997, 17, 1178.
Further docking studies were performed to understand
the observed SAR, comparing the cyanoindoles 2 and
16 and looking at the pyridylindole 5. The results from
the docking of 2 and 16 suggest two different binding
modes: although a H-bond between cyano and the NH
of Gly 326 is a common feature, in the case of N-methy-
lindole 16 the loss of a H-bond to Asp 274, as suggested
for 2, is compensated by placing the methyl group in the
same hydrophobic region as the methoxy group of
VX497 (Figs. 1 and 2).
8. Sintchak, M. D.; Nimmesgern, E. Immunopharmacology
2000, 47, 163.
9. Dhar, T. G. M.; Shen, Z.; Guo, J.; liu, C.; Watterson, S.
H.; Gu, H. H.; Pitts, W. J.; Fleener, C. A.; Rouleau, K. A.;
Sherbina, N. Z.; McIntyre, K. W.; Witmer, M. R.;
Tredup, J. A.; Chen, B.-C.; Zhao, R.; Bednarz, M. S.;
Cheney, D. L.; MacMaster, J. F.; Miller, L. M.; Berry, K.
K.; Harper, T. W.; Barrish, J. C.; Hollenbaugh, D. L.;
Iwanowicz, E. J. J. Med. Chem. 2002, 45, 2127.
10. Dhar, T. G. M.; Shen, Z.; Fleener, C. A.; Rouleau, K. A.;
Barrish, J. C.; Hollenbaugh, D. L.; Iwanowicz, E. J.
Bioorg. Med. Chem. Lett. 2002, 12, 3305.
11. Buckley, G. M.; Davies, N.; Dyke, H. J.; Gilbert, P. J.;
Hannah, D. R.; Haughan, A. F.; Hunt, C. A.; Pitt, W. R.;
Profit, R. H.; Ray, N. C.; Richard, M. D.; Sharpe, A.;
Taylor, A. J.; Whitworth, J. M.; Williams, S. C. Bioorg.
Med. Chem. Lett. 2005, 15, 751.
12. Birch, H. L.; Buckley, G. M.; Davies, N.; Dyke, H. J.;
Frost, E. J.; Gilbert, P. J.; Hannah, D. R.; Haughan, A.
F.; Madigan, M. J.; Morgan, T.; Pitt, W. R.; Ratcliffe, A.
J.; Ray, N. C.; Richard, M. D.; Sharpe, A.; Taylor, A. J.;
Whitworth, J. M.; Williams, S. C. Bioorg. Med. Chem.
Lett. 2005, 15, 5335.
13. Jain, J.; Almquist, S. J.; Heiser, A. D.; Shlyakhter, D.;
Leon, E.; Memmott, C.; Moody, C. S.; Nimmesgern, E.;
Decker, C. J. Pharmacol. Exp. Ther. 2002, 302, 1272.
14. Dhar, T. G. M.; Shen, Z.; Gu, H. H.; Chen, P.; Norris, D.;
Watterson, S. H.; Ballentine, S. K.; Fleener, C. A.;
Rouleau, K. A.; Barrish, J. C.; Townsend, R.; Hollenb-
augh, D. L.; Iwanoxicz, E. J. Bioorg. Med. Chem. Lett.
2003, 13, 3557.
In the case of pyridylindole 5, H-bonds between both
the indole NH to Asp 274 and pyridine N to Gly 326
can be realized and a good contact with the active site
achieved, and it might be that this fragment can reach
a more favourable geometry than 2, resulting in the in-
creased potency observed (Fig. 2). Docking also predicts
the inactivity of an N-methyl-3-pyrid-4-ylindole, which
would result in a steric clash in the active site. Docking
studies also suggested the methyl group of 2-methylin-
dole 13 may be able to access the hydrophobic region
without disrupting the two H-bonds, which could ac-
count for the further increase in potency seen. Addition-
ally, this methyl substitution would affect the torsion
angle between pyridine and indole.
The most active fragments described here compare
favourably with 3-methoxy-4-(oxazol-5-yl)aniline (21)
from VX497 and BMS-337197 (IC50 = 8.2 lM). The
implications of the proposed binding modes are that
with the NH indoles, such as 2 and 5, a greater variety
of groups could be added to the carbocyclic ring without
the need for a H-bond donor to interact with Asp 274,
whereas with the N-methylindoles, such as 16, this
requirement still remains.
15. Pickett, S. D.; Sherborne, B. S.; Wilkinson, T.; Bennett, J.;
Borkakoti, N.; Broadhurst, M.; Hurst, D.; Kilford, I.;
McKinnell, M.; Jones, P. S. Bioorg. Med. Chem. Lett.
2003, 13, 1691.
16. Jones, G.; Willett, P.; Glen, R. C. J. Mol. Biol. 1995, 245,
43–53.
17. For IMPDH II inhibition protocol, see Ref. 12 (Ref. 14).
In conclusion, our fragment-based approach has deliv-
ered sub-micromolar, low molecular weight indole
inhibitors of IMPDH II, with a platform to develop into
new classes of IMPDH inhibitors. Our efforts to elabo-
rate these hits further are described in the following
paper.