Mycophenolic anilides as broad specificity inosine-5’-monophosphate dehydrogenase (IMPDH) inhibitors

Article abstract of DOI:10.1016/j.bmcl.2020.127543

Inosine-5′-monophosphate dehydrogenase (IMPDH) is a potential target for microorganisms. However, identifying inhibitor design determinants for IMPDH orthologs continues to evolve. Herein, a series of mycophenolic anilide inhibitors of Cryptosporidium par

Full text of DOI:10.1016/j.bmcl.2020.127543

Bioorganic & Medicinal Chemistry Letters 30 (2020) 127543
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Bioorganic & Medicinal Chemistry Letters
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Mycophenolic anilides as broad specificity inosine-5’-monophosphate
dehydrogenase (IMPDH) inhibitors
a
c
a,b
a
a
c
Seungheon Lee , Angela F. Ku , Mohana Rao Vippila , Yong Wang , Minjia Zhang ,
c,d
a,⁎
Xingyou Wang , Lizbeth Hedstrom , Gregory D. Cuny
a
b
c
Department of Pharmacological and Pharmaceutical Sciences, University of Houston, Health Building 2, Houston, TX 77204, USA
Department of Chemistry, University of Houston, Health Building 2, Houston, TX 77204, USA
Departments of Biology, 415 South St., Waltham, MA 02454, USA
d
Chemistry, Brandeis University, 415 South St., Waltham, MA 02454, USA
A R T I C L E I N F O
A B S T R A C T
Keywords:
Inosine-5′-monophosphate dehydrogenase (IMPDH) is a potential target for microorganisms. However, identi-
fying inhibitor design determinants for IMPDH orthologs continues to evolve. Herein, a series of mycophenolic
anilide inhibitors of Cryptosporidium parvum and human IMPDHs are reported. Furthermore, molecular docking
of 12 (e.g. SH-19; CpIMPDH Ki,app = 0.042 ± 0.015 µM, HsIMPDH2 Ki,app = 0.13 ± 0.05 µM) supports
different binding modes with the two enzymes. For CpIMPDH the inhibitor extends into a pocket in an adjacent
subunit. In contrast, docking suggests the inhibitor interacts with Ser276 in the NAD binding site in HsIMPDH2,
as well as an adjacent pocket within the same subunit. These results provide further guidance for generating
IMPDH inhibitors for enzymes found in an array of pathogenic microorganisms, including Mycobacterium tu-
berculosis.
Cryptosporidium parvum
IMPDH
Inosine-5’-monophosphate dehydrogenase
Inhibitor
Binding mode
Inosine-5′-monophosphate dehydrogenase (IMPDH) catalyzes the
nicotinamide adenine dinucleotide (NAD)-dependent oxidation of in-
osine 5′-monophosphate (IMP) to xanthosine 5′-monophosphate (XMP)
nicotinamide subsite, but is a poor inhibitor of prokaryotic and C.
22
parvum IMPDHs (e.g. CpIMPDH K = 9.3 μM). Mycophenolic anilides
23–25
i
have also been found to inhibit HsIMPDH2,
but their activities
1
as the rate-limiting step in the biosynthesis of guanine nucleotides.
against CpIMPDH and bacterial orthologs have not been described.
Therefore, IMPDH regulates intracellular guanine nucleotide pools and
Herein, we report a structure–activity relationship study for a series
2
is critical for cell proliferation in both eukaryotes and prokaryotes.
of mycophenolic anilides that incorporate a molecular fragment
IMPDH inhibition has recently gained momentum as a potential
treatment of microbial infections. For example, blocking prokaryotic
IMPDH could provide a strategy for growth inhibition of bacteria such
common to several classes of CpIMPDH inhibitors (Fig. 1C). Further-
more, an analysis of these anilides was conducted to elucidate addi-
tional structural determinants required for selective inhibition of
CpIMPDH and related prokaryotic orthologs versus HsIMPDHs.
An initial set of MPA-anilide derivatives 8 – 16 were prepared via
EDC-mediated coupling (Scheme 1). In addition, 10 was further mod-
ified by phenol alkylation or alkene reduction to provide derivatives 17
3
,4
as Mycobacterium tuberculosis (Mtb) and Staphylococcus aureus (Sa). In
addition, this enzyme has been targeted in the protozoan Cryptospor-
idium parvum (Cp), which has a similar IMPDH to some prokaryotes,
5
,6
likely resulting from lateral gene transfer from bacteria. These pro-
karyotic IMPDHs are structurally distinct from their human counter-
2
8
and 18, respectively.
2
parts. The most dramatic difference is in the adenosine subsite (A-site)
Replacement of the alkene with a cyclopropane bioisostere was also
pursued. The synthesis of both enantiomers is illustrated in Schemes 2
7
,8
of the NAD binding site , and several inhibitor scaffolds (e.g. 1 – 6 in
9
–17
23
Fig. 1A) exploit this divergence.
Mycophenolic acid (MPA, 7, Fig. 1B) is a prototypic human (Hs)
and 3. MPA (7) was converted to aldehyde 19, which was protected
and then reduced to provide alcohol 21 (Scheme 2). A chiral auxiliary
1
8,19
IMPDH inhibitor
(e.g. HsIMPDH1 K
i
= 33 nM and HsIMPDH2
was attached generating key intermediate 23a using the methodology
2
0
29,30
K
i
= 7 nM) used clinically as an ester prodrug (e.g. mycophenolate
of Charette et al.
Finally, anomerization generated a second crucial
mofetil) for immunosuppression in preventing rejection following
intermediate 23b. Both of these intermediates were partially de-pro-
tected generating 24a and 24b, which were subjected to
organ transplantation.2
1,26
Interestingly, MPA (7) binds in the
⁎
Corresponding author.
E-mail address: gdcuny@central.uh.edu (G.D. Cuny).
https://doi.org/10.1016/j.bmcl.2020.127543
Received 2 July 2020; Received in revised form 3 September 2020; Accepted 4 September 2020
Available online 12 September 2020
0960-894X/ © 2020 Elsevier Ltd. All rights reserved.

S. Lee, et al.
Bioorganic & Medicinal Chemistry Letters 30 (2020) 127543
followed by alcohol oxidation, Horner–Wadsworth–Emmons reactions
and alkene reduction to produce 29a and 29b. Finally, ester hydrolysis
and EDC-mediated aniline coupling provided 31a and 31b.
Several other derivatives were prepared that incorporated addi-
tional changes to the linker region of the hybrid molecules. MPA (7)
was cleaved to aldehyde 32, which was reduced and protected to give
3
4 (Scheme 5). Alkylation of the primary alcohol, ester hydrolysis,
aniline coupling and deprotection produced the ether linked derivative
3
5
3
8. MPA (7) was also converted to aldehyde 40a, which via a Wittig
+
-
36,37
reaction with 4-ClPhNH(C]O)CH
2
P
Ph
3
Cl gave 41a (Scheme 6).
Similarly, 34 was oxidized to aldehyde 40b, which was converted into
4
1b. These two intermediates were de-protected or reduced/de-pro-
tected to provide 42a and 42b, respectively. Finally, intermediate 32
was oxidized to carboxylic acid 43, which was coupled to 4-chlor-
oaniline to give the truncated derivative 44 (Scheme 7).
Fig. 1. (A) Six structurally distinct CpIMPDH inhibitors (CpIMPDH IC50 = 12 –
3
8
2
7
6
4 nM). The fragments found in these inhibitors that interact with the ade-
Recombinant CpIMPDH and HsIMPDH2 were purified from E. coli.
nosine subsite (A-site) of the NAD binding site are highlighted in red. This is
based on co-crystal structures of 1, 2, 4 and 5 with CpIMPDH and 6 with
Clostridium perfringens IMPDH (ClpIMPDH). The interactions of inhibitor 3 are
assumed based on structural similarity since it has not been co-crystalized with
an IMPDH. (B) Structure of mycophenolic acid (MPA, 7). (C) Mycophenolic
anilides with three regions explored herein shown in boxes.
Enzyme activity was determined by monitoring the production of
39
NADH. For CpIMPDH, enzyme (10 nM) and inhibitor (1 nM to 5 μM)
were incubated in the presence of 50 mM Tris-HCl, pH 8.0, 100 mM
KCl, 3 mM EDTA, 1 mM dithiothreitol at 25 °C for 5 min prior to ad-
dition of substrates NAD (300 μM) and IMP (250 μM). For HsIMPDH2,
enzyme (20 nM) and inhibitor (1 nM to 5 μM) were incubated in the
presence of 50 mM Tris-HCl, pH 8.0, 100 mM KCl, 3 mM EDTA, 1 mM
dithiothreitol at 25 °C for 5 min prior to addition of substrates NAD
cyclopropanation conditions to give 25a and 25b, respectively (Scheme
3
1,32
41
3
).
These two materials were then subjected to a similar series of
(60 μM) and IMP (250 μM). Production of NADH was monitored by
3
3,34
transformations (Scheme 4).
The chiral auxiliary was removed
fluorescence. K
i,app
values were determined for each inhibitor against
R3 R4
R3 R4
N
OH
OH
O
O
R2
R1
NH
R2
R1
HO
a or b
+
O
O
O
O
MeO
MeO
8
- 16
7
OMe
OH
O
H
N
F3C
Cl
c
O
O
O
OH
MeO
MeO
O
H
N
F3C
Cl
17
O
O
O
H
N
MeO
F3C
Cl
10
d
O
18
Scheme 1. Synthesis of 8 – 18. Reagents and conditions: (a) EDC•HCl, HOBt or HOAt or DMAP, TEA or DIPEA, DMF or DCM, rt or 0 °C to rt, 16 h, 17–80%. (b) i)
oxalyl chloride, DMF (cat.), DCM, rt. ii) aniline, TEA, rt, 6 h, 40%. (c) MeI, K
2
CO
3
, acetone, rt, 16 h, 64%. (d) 10% Pd/C, H (1 atm), EtOAc, rt, 3 h, 34%.
2
OH
OH
OTBS
Scheme 2. Synthesis of intermediates 23a and 23b.
O
O
O
HO
O
O
Reagents and conditions: (a) i) OsO , NMO, NaIO
THF/H O, rt, 1.5 h. ii) PPh =C(CH )CHO, benzene,
reflux, 24 h, 77%. (b) TBSCl, imidazole, DCM, rt,
6 h, 71%. (c) NaBH , MeOH, 0 °C to rt, 1 h, 92%. (d)
22, BF •OEt , DCM, −78 °C for 0.5 h then 0 °C for
4
4
,
a
b
O
O
O
2
3
3
O
MeO
MeO
19
MeO
20
1
4
7
3
2
OTBS
0.5 h, 88%. (e) TiCl
4
, DCM, −78 °C for 10 min then
O
BnO
BnO
BnO
OTBS
O
O
0 °C for 0.5 h, 99%.
HO
O
c
d
O
OAc
O
BnO
MeO
MeO
BnO
O
BnO
2
1
23a
AcO
O
CCl3
22
NH
BnO
BnO
BnO
O
OTBS
O
AcO
e
O
O
MeO
23b
2

S. Lee, et al.
Bioorganic & Medicinal Chemistry Letters 30 (2020) 127543
BnO
BnO
BnO
OH
BnO
BnO
BnO
OH
O
O
O
O
O
O
O
OH
O
OH
2
3a
or
3b
MeO
MeO
a
b
24a
25a
2
BnO
BnO
BnO
BnO
BnO
BnO
O
O
OH
OH
O
O
HO
HO
O
O
O
O
MeO
MeO
24b
25b
Scheme 3. Synthesis of intermediates 25a and 25b. Reagents and conditions: (a) NaOMe, MeOH, 0 °C to rt, 16 h, 55 and 77%. (b) For 25a: Et
2
Zn, CH
2
I , toluene,
2
−
20 °C, 3 h, 76%. For 25b: Et
2
Zn, CH
2
I
2
, toluene, −78 °C to rt, 16 h (dr 6:1), 98%.
OH
OH
OTBS
OTBS
O
O
O
O
HO
*
O
*
*
TBSO
O
a
b
b
25a or b
*
a
O
*
O
7
O
O
O
MeO
MeO
MeO
MeO
39
2
6a or b
27a or b
OH
40a
d
OH
O
O
O
OBn
OR
EtO
*
RO
O
O
c
d
H
N
O
*
*
O
O
O
c
d
3
4
O
O
MeO
MeO
O
MeO
Cl
MeO
28a or b
29a or b R = Et
e
4
1a R = TBS
41b R = Bn
30a or b R = H
40b
OH
O
H
N
OH
O
F3C
H
N
O
O
e
O
Cl
MeO
O
O
3
1a
1b
f
Cl
MeO
2a
30a or b
4
OH
O
H
N
OH
41a or b
O
F3C
Cl
H
N
O
O
O
MeO
f
3
Cl
MeO
2b
4
Scheme 4. Synthesis of 31a and 31b. Reagents and conditions: (a) i) Tf
pyridine, DCM, −20 °C to rt; ii) DMF, pyridine, H O, 120 °C, 10 min, 81% and
3%. (b) DMP, DCM, rt, 0.5 h, 99%, and 83%. (c) triethyl phosphonoacetate,
NaH, benzene, rt, 1 h, 57% and 51% (d) CoCl , NaBH , MeOH/DMF, rt, 0.5 h,
2
O,
Scheme 6. Synthesis of 42a and 42b. Reagents and conditions: (a) TBSCl,
2
imidazole, DMF, rt, 6 h, 66%. (b) OsO
4
, NMO, NaIO
4
, THF/H
2
O, rt, 1.5 h, 70%.
9
+
-
(
c) DMP, DCM, rt, 3 h, 80–88%. (d) LDA, 4-ClPhNH(C]O)CH
2
P
Ph
3
Cl (see
2
4
Supporting Information compound S2), THF/toluene, 0 °C to rt, 3 h, 73% and
6
2% and 87% (e) LiOH, THF/H O, rt, 3 h, 86% and 85%. (f) 4-chloro-3-(tri-
2
5
1
1% (e) TBAF, THF, 0 °C to rt, 0.5 h, 22% (f) 10% Pd/C, H
2
(1 atm), MeOH, rt,
fluoromethyl)aniline, EDC•HCl, HOAt, DMF, rt, 16 h, 81% and 71%.
h, 46%.
CpIMPDH and HsIMPDH2 using Dynafit.40
HsIMPDH2 (Ki,app = 0.35 µM), in contrast to all prior reported
CpIMPDH inhibitors. Since electron withdrawing groups were preferred
in previously developed CpIMPDH inhibitors, the methoxy group in 8
was replaced with a chloro (9). This compound showed similar in-
hibitory potency as 8 for both enzymes. However, replacing with a
trifluoromethyl (10) increased potency against both enzymes. Trans-
posing the para-chloro to the ortho-position (11) resulted in loss of
CpIMPDH inhibitory activity. This was consistent with the SAR of other
CpIMPDH inhibitors, possibly resulting from a clash with Y358′ and loss
Introducing an anilide as the common fragment to MPA improved
CpIMPDH inhibition. For example, 8, which incorporated the anilide
from 4 with MPA, potently inhibited CpIMPDH with Ki,app value of
0
.046 µM (Table 1). Interestingly, the compound also inhibited
OH
OH
OBn
O
O
O
HO
HO
a
R
c
O
O
O
O
MeO
MeO
MeO
9
of a halogen bond with G357′. A survey of various electron with-
7
32 R = CHO
3 R = CH2OH
34
b
drawing groups at the 4-position demonstrated that chloro (12) pro-
vided potent and balanced CpIMPDH and HsIMPDH2 inhibition.
3
Cl
O
OBn
O
OR
O
O
O
O
d
RO
f
N
H
O
O
OH
OH
O
O
H
MeO
MeO
N
a
HO2C
MeO
b
32
O
O
O
3
3
5 R = Et
6 R = H
37 R = Bn
38 R = H
e
g
Cl
MeO
Scheme 5. Synthesis of 38. Reagents and conditions: (a) OsO
THF/H O, rt, 1.5 h. (b) NaBH , EtOH, rt, 3 h, 70–81%. (c) benzyl bromide,
TBAF, rt, 6.5 h, 81–89%. (d) InCl , ethyl diazoacetate, DCM, rt, 16 h, 57%. (e)
LiOH, MeOH/H O, rt, 3 h, 76–95%. (f) EDC•HCl, HOAt, 4-chloroaniline, DMF,
rt, 16 h, 65–80%. (g) 10% Pd/C, H (1 atm), MeOH, rt, 1 h, 27–56%.
4
, NMO, NaIO
4
,
43
44
2
4
Scheme 7. Synthesis of 44. Reagents and conditions: (a) NaClO
methyl-2-butene, t-BuOH/H O, rt, 4 h (b) EDC, DIPEA, 4-chloroaniline, THF/
DMF, rt, 16 h, 38% over two steps.
2
, NaH
2
PO , 2-
4
3
2
2
2
3

S. Lee, et al.
Bioorganic & Medicinal Chemistry Letters 30 (2020) 127543
Table 1
in loss of CpIMPDH inhibition.
CpIMPDH and HsIMPDH2 inhibitory activities of 8–17
Next, the central linker was explored using various bioisosteres and
by truncation (Table 2). 3-Trifluoromethyl-4-chloro or 4-chloro sub-
stituted anilides were chosen based on 10 and 12. Reduction of the
alkene (18) resulted in some loss of potency for CpIMPDH inhibition,
which was more dramatic for HsIMPDH2. Bioisosteric replacement of
the alkene with a cyclopropane revealed that one enantiomer (31b)
retained potent CpIMPDH inhibitory activity (Ki,app = 0.066 µM), al-
though HsIMPDH2 inhibition was reduced (Ki,app = 0.46 µM). The
eudysmic ratios of 31b and 31a for CpIMPDH and HsIMPDH inhibitions
were 8.6 and 3.1, respectively. Several additional changes to the linker
connecting the two aryl groups resulted in reduced CpIMPDH and
HsIMPDH2 inhibitory activities.
R2
N
OR3
O
R1
O
O
MeO
Compound
R1
R2
R3
CpIMPDH
HsIMPDH2
K
i,app (µM)
Ki,app (µM)
0.046 ( ± 0.013)σ
0.35 ( ± 0.09)
0.34 ( ± 0.11)
0.23 ( ± 0.08)
ND*
σ
σ
σ
8
9
3-OMe
,
4-Cl
H
H
H
H
H
H
H
H
Me
H
H
H
H
H
H
H
H
H
H
Me
σ
σ
3,4-di-Cl
3-CF3, 4-Cl
2,3-di-Cl
4-Cl
0.041 ( ± 0.010)
10
11
12
13
14
15
16
17
0.016 ( ± 0.007)
r
0.681 (0.143)
σ
r
r
0.13 ( ± 0.05)σ
Two critical differences between HsIMPDH2 and CpIMPDH are in a
loop structure of the NAD binding region and a proximal site in the
adjacent subunit. The A-site of HsIMPDH2 is largely within the same
monomer as the IMP site, but for CpIMPDH and related prokaryotic
0.042 ( ± 0.02)
0.180 ( ± 0.06)
0.151 ( ± 0.04)
0.11
4-F
0.27
0.40
0.35
ND*
ND*
4-CF
3
4-CN
4-Cl
0.87 ( ± 0.13)r
1.7
7
,8
IMPDHs, a portion of the A-site is located in the adjacent subunit.
3-CF3, 4-Cl
4
2
This hydrophobic pocket is absent in HsIMPDH. As illustrated in the
sequence alignments of HsIMPDH2 and CpIMPDH (Fig. 2), the NAD
loop of HsIMPDH contains a hydrophilic serine at residue 276, while
CpIMPDH has a hydrophobic alanine at the equivalent position (residue
*
ND: Not Determined.
σ: Standard deviation.
r: Range.
1
65′; note that CpIMPDH lacks an approximately 100 residue reg-
Reminiscent of other CpIMPDH inhibitors, methylation of the anilide
ulatory domain present in most IMPDHs). For the A-site, there are also
distinct residue differences with aspartic acid (e.g. 470′) in HsIMPDH
corresponding to tyrosine (e.g. Y358′) in CpIMPDH.42 Interaction with
(
(
16) resulted in loss of activity. Similarly, methylation of the phenol
17), known to eliminate MPA inhibition of HsIMPDH,4 also resulted
1
Table 2
CpIMPDH and HsIMPDH2 inhibitory activities of 18, 31a, 31b, 38, 42a, 42b and 44.
OH
O
R1
H
N
X
O
O
O
Me
Compound
R1
X
CpIMPDH
HsIMPDH2
K
i,app (µM)
K
i,app (µM)
1
3
8
3-CF3, 4-Cl
3-CF3, 4-Cl
0.060 ( ± 0.01)r
ND*
1a
0.57
1.40
3
1b
8
3-CF3, 4-Cl
4-Cl
0.066 ( ± 0.023)σ
0.405 ( ± 0.176)r
0.46 ( ± 0.28)σ
0.87
3
O
42a
42b
44
4-Cl
4-Cl
4-Cl
0.48
0.55
0.29
0.8
0.45 ( ± 0.13)r
2.8
*
ND: Not Determined.
σ: Standard deviation.
r: Range.
a
Fig. 2. Sequence alignment of IMPDH enzymes from human, C. parvum and several bacteria highlighting the NAD loop and A-site. Y358′ is based on numbering for
CpIMPDH and the ’ denotes the adjacent subunit.
4

S. Lee, et al.
Bioorganic & Medicinal Chemistry Letters 30 (2020) 127543
Fig. 3. (A) Docked model of 12 (pink) with
CpIMPDH (gray, PDB: 3KHJ) and IMP (yellow).
Adjacent monomer protein is shown as purple and
residue numbers are differentiate by prime (’).
Hydrogen bonds are shown as red dashes (< 3.0 Å).
Docking score of 12 with CpIMPDH•IMP was −9.86.
(
B) Docked structure of 12 (pink) with hamster
IMPDH2 (green, PDB: 1JR1) and IMP (yellow),
which has only eight non-binding site amino acid
4
3
differences compared to HsIMPDH2
. Hydrogen
bonds are shown as red dashes (< 3.5 Å). Docking
score of 12 with hamster IMPDH2•IMP was −8.83.
+
Water, K and other protein subunits were deleted
for docking and presentation for clarity.
this tyrosine residue has previously been shown to be critical for
In conclusion, mycophenolic anilides were found that inhibit both
CpIMPDH and HsIMPDH2 by incorporation of a molecular fragment
from previously reported CpIMPDH inhibitors. Prior studies combined
with molecular docking assessments revealed that selectivity for mi-
croorganism IMPDHs (e.g. those with alanine in the NAD loop and
Y358′ in the adjacent subunit) requires two distinct design elements: 1)
interaction with the adjacent subunit via π-π interactions with Y358′
and 2) lack of interactions with Ser276 in the NAD binding site of
HsIMPDH2. These two features provide further guidance for generating
selective IMPDH inhibitors for a subset of susceptible microorganisms.
9
–17, 27
achieving selective CpIMPDH inhibitors.
Interestingly, a number
of other microorganisms, including several pathogenic Gram-(+) and
Gram-(-) bacteria listed in Fig. 2, also have alanine and tyrosine re-
sidues in these two positions and are inhibited by CpIMPDH in-
2
7
hibitors.
In order to understand potential binding modes of MPA-anilides to
CpIMPDH and HsIMPDH, molecular docking studies were conducted
using Autodock Tools. Docking of 12 into CpIMPDH (PDB: 3KHJ)
provided a low energy pose that satisfied the three key interactions of
previously co-crystalized selective CpIMPDH inhibitors. Specifically,
the phenol portion of MPA had a π-π interaction with the hypoxanthine
of IMP, the anilide NH formed a hydrogen bond with Glu329 and the
anilide extended into the adjacent subunit forming a π-π interaction
with Tyr358′ (Fig. 3A). Additionally, the phenol OH formed an ionic-
dipole interaction with Glu329. This binding mode is also consistent
with several of the observed structure–activity relationship features,
such as alkylation of the anilide or phenol disrupting interactions with
Glu329. In addition, truncation of the linker would result in an inability
of the inhibitor to extend into the other subunit to provide the critical π-
π interaction with Tyr358′. Overlay of this docking model with the co-
crystal structure of CpIMPDH•IMP•5 (PDB: 4RVB) further illustrates
that 12 can readily be accommodated in a binding mode similar to
Declaration of Competing Interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influ-
ence the work reported in this paper.
Acknowledgements
This work was supported in part by a grant from the National
Institutes of Health (R01AI125362).
Appendix A. Supplementary data
1
0
selective CpIMPDH inhibitors (see Supporting Information, Fig. S1A).
Docking of 12 into hamster IMPDH2 (PDB: 1JR1), which has only
Supplementary data to this article can be found online at https://
doi.org/10.1016/j.bmcl.2020.127543.
4
3
eight non-binding site amino acid differences
compared to
HsIMPDH2, provided a low energy pose with similar interactions as
MPA (7), including H-bonding of the anilide NH and phenol to Ser276
in the NAD binding site and Gln441, respectively (Fig. 3B; for an
overlay of the docked structure of 12 with hamster IMPDH2•IMP•7 see
Supporting Information, Fig. S1B). The para-chlorophenyl occupies a
modestly large pocket created by Thr252, His253, Phe282 and Ser275,
as well as being within 3.2 Å of the methylene portion of this later
residue’s side-chain. Interestingly, attempts to dock a CpIMPDH selec-
tive inhibitor (e.g. 5) into hamster IMPDH2 did not produce reasonable
binding modes with low binding energies (data not shown). This could
result from the compound being less flexible and not being able to form
a productive interaction with Ser276 in the NAD site. Collectively, these
data elucidated an additional structural criterion for achieving
CpIMPDH inhibitor selectivity: the inability to form interactions with
Ser276.
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