Structure-activity relationship studies and biological characterization of human NAD+-dependent 15-hydroxyprostaglandin dehydrogenase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2013.11.081

The structure-activity relationship (SAR) study of two chemotypes identified as inhibitors of the human NAD⁺-dependent 15-hydroxyprostaglandin dehydrogenase (HPGD, 15-PGDH) was conducted. Top compounds from both series displayed potent inhibition (IC₅₀ <50 nM), demonstrate excellent selectivity towards HPGD and potently induce PGE ₂ production in A549 lung cancer and LNCaP prostate cancer cells.

Full text of DOI:10.1016/j.bmcl.2013.11.081

Bioorganic & Medicinal Chemistry Letters 24 (2014) 630–635
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Structure–activity relationship studies and biological
characterization of human NAD+-dependent
15-hydroxyprostaglandin dehydrogenase inhibitors
Damien Y. Duveau, Adam Yasgar, Yuhong Wang, Xin Hu, Jennifer Kouznetsova, Kyle R. Brimacombe,
⇑
Ajit Jadhav, Anton Simeonov, Craig J. Thomas, David J. Maloney
National Center for Advancing Translational Sciences, National Institutes of Health, 9800 Medical Center Drive, Rockville, MD 20850, United States
a r t i c l e i n f o
a b s t r a c t
Article history:
The structure–activity relationship (SAR) study of two chemotypes identified as inhibitors of the human
NAD+-dependent 15-hydroxyprostaglandin dehydrogenase (HPGD, 15-PGDH) was conducted. Top com-
pounds from both series displayed potent inhibition (IC50 <50 nM), demonstrate excellent selectivity
towards HPGD and potently induce PGE2 production in A549 lung cancer and LNCaP prostate cancer cells.
Published by Elsevier Ltd.
Received 19 November 2013
Accepted 27 November 2013
Available online 4 December 2013
Keywords:
HPGD
15-PGDH
Prostaglandin
PGE2
Inhibitor
Prostaglandins are members of the eicosanoid family of signal-
ing molecules, along with lipoxins and leukotrienes. They are syn-
thesized from arachidonic acid (AA), which following its release
from membrane phospholipids, undergoes a series of transforma-
tions mediated by enzymes such as cyclooxygenases (COX) and
prostaglandin synthases. Prostaglandins are also inactivated
through chemical modification. One key metabolizing enzyme is
15-hydroxyprostaglandin dehydrogenase (HPGD, also referred to
as 15-PGDH), which catalyzes the oxidation of the prostaglandin
hydroxyl group at C-15 to the corresponding ketone.1
A number of prostaglandins of particular biological interest
have been identified as substrates of HPGD. Prostaglandin E2
(PGE2), a product of COX-2-mediated AA metabolism via prosta-
glandin H2 (PGH2) is converted by HPGD to its biologically inactive
metabolite, 15-keto-PGE2. Over-production of PGE2 has been re-
ported to promote tumor-associated neovascularization, cell pro-
liferation, and tumor growth.2 It has also been suggested that
decreased levels of PGE2 resulting from amplified HPGD activity
through pharmacological means or enhanced expression may mit-
igate these phenotypes and result in a chemoprevention strategy.3
Conversely, HPGD was found to be highly expressed in androgen
receptor-overexpressing advanced tumors, metastatic prostate
cancers,4 and metastatic breast cancer, where it has been found
to contribute to epithelial–mesenchymal transition (EMT).5
15-deoxy-D
12,14-prostaglandin J2 (15d-PGJ2) is a product of albu-
min-mediated transformation of prostaglandin D2 (PGD2), itself
derived from PGH2.6 15d-PGJ2 was shown to inhibit IKKb, a positive
regulator of NF-j
B,7,8 and to covalently bind Keap1, a negative reg-
ulator of Nrf2.9 As such, the metabolism of PGD2 by HPGD provides
a possible link between HPGD activity and levels of 15d-PGJ2.
Moreover, Blair et al. recently demonstrated that HPGD catalyzed
the transformation of 11-hydroxy-eicosatetranoic acid (11-HETE),
a product of COX-2/HPGD-mediated AA transformation, into 11-
oxo-eicosatetranoic acid (11-oxo-ETE),10 and that 11-oxo-ETE
inhibits NF-j
B signaling through inhibition of IKKb.11 Therefore,
down regulation of HPGD may lead to a decrease in the NF-
jB sig-
naling pathway and to an increase in Nrf2 signaling.
As a consequence of prostaglandins’ roles in a number of pro-
cesses such as inflammation, differentiation, and cellular signaling,
their cellular levels are tightly regulated. The central role of HPGD
to inactivate prostaglandins such as PGE2 or PGD2 makes it an
important target for pharmacological intervention. Prior to our ef-
forts, few inhibitors of HPGD had been identified, and all are based
off ciglitazone, a PPAR
c
inhibitor possessing a thiazolidinedione
scaffold. The parent compound was found to have an IC50 against
HPGD of 2.7
l
M12 whereas an optimized analogue, CT-8, displayed
an IC50 of 51 nM.13 More recently, the same group continued their
SAR studies around ciglitazone and related analogues, with dem-
onstrated activity in vivo for their top compounds,14 but the selec-
tivity of CT-8 toward other dehydrogenases was not reported. In
addition, the thiazolidinedione chemotype is likely to have
⇑
Corresponding author. Tel.: +1 301 217 4381; fax: +1 301 217 5736.
E-mail address: maloneyd@mail.nih.gov (D.J. Maloney).
0960-894X/$ - see front matter Published by Elsevier Ltd.
http://dx.doi.org/10.1016/j.bmcl.2013.11.081

D. Y. Duveau et al. / Bioorg. Med. Chem. Lett. 24 (2014) 630–635
631
O
halogen group led to analogues with slightly improved potencies
[4a (3,4-dichloro-phenyl), 4c (4-Cl-phenyl and 4d (4-Br-phenyl)],
with bis-substituted analogue 4a being the most potent (IC50 = 22 nM).
However, introduction of other electron-withdrawing groups at
the para position such as F (4e), CF3 (4g), CN (4j) and NO2 (4l), re-
sulted in a slight loss of potency. We did find that the phenyl ring
could not be replaced by a heterocycle, such as a pyridine ring
(compounds 4o–q) without a more significant drop in potency.
Of note, a number of phenyl analogues bearing ortho substituents
could not be successfully synthesized, likely because the steric
hindrance of the two pyrrole methyl groups prevent the Ullman-
type coupling reaction from taking place.
The synthetic sequence outlined in Scheme 1 is ideal for late-
stage diversification of the pendant aryl ring but is less optimal
for investigations of the triazoloazepine core. For facile preparation
of these analogues we utilized common intermediate 2, followed
by cyclization with various 1-aza-2-methoxy derivatives 5a–e to
ultimately provide five analogues (6a–d) with rings of various sizes
fused to the triazole moiety (Scheme 2).
N
N
N
N
Br
N
S
N
N
H
NC
N
ML147
ML148
ML149
Figure 1. Structures of chemotypes ML147, ML148, and ML149.
undesired off-target effects given the well documented PPAR
activity for this class of molecules.
c
In an effort to identify selective inhibitors of HPGD, we con-
ducted a quantitative high throughput screen (qHTS) of ꢀ160,000
compounds as part of the Molecular Libraries Small Molecule
Repository (MLSMR). These studies led to the identification of
three structurally distinct small molecule chemical probes as po-
tent and selective inhibitors of HPGD (denoted as ML147, ML148,
and ML149, Fig. 1).15 The first scaffold, ML147, is characterized
by an azabenzimidazole core appended with a thiobenzyl group
at C-1. The second compound, ML148 possesses a benzimidazole
core substituted with a cyclohexylamide moiety and 3-methyl-
phenyl group. Finally, ML149 contains a triazoloazepine core with
2,5-dimethylpyrrole group linked to the triazole ring. Given the po-
tential liabilities (e.g., glutathione reactivity) of the thiol ether link-
age on ML147 we decided to focus our initial SAR efforts on the
optimization of ML148 and ML149. In addition to our SAR efforts,
we also sought to define key in vitro ADME properties, cellular
efficacy (PGE2 levels) and selectivity against several related dehy-
drogenases (vide infra).
Upon biological testing of the compounds, no direct correlation
was found between the sizes of the ring fused to the triazole moiety
and HPGD activity (Table 1). However, the five-membered ring
analogue 6a resulted in a significant loss of potency (IC50 = 3.04 lM)
suggesting the importance of having at least a 6-membered fused
ring. Whereas, the 6-(6b), 8-(6c) and 9-membered (6d) ring ana-
logues were less active than ML149. Having completed our initial
SAR investigations of the core and pendant phenyl ring, we then
wanted to probe the influence of the substituents on the pyrrole ring
(Table 1). The synthesis of these compounds was achieved in a man-
ner similar to the analogues described previously, except the starting
carboxylic acid was varied (Scheme 3).
The unsubstituted pyrrole 7a displayed a six-fold drop in activ-
ity (IC50 = 607 nM) as compared to ML149, highlighting the impor-
tance of substitution on the pyrrole. The indole derivative (7c) was
less active (IC50 = 681 nM), and the addition of other heteroatoms
Optimization of ML149: In order to support compound scale-up
for more detailed biological testing and to facilitate SAR explora-
tions, we designed concise synthetic routes to ML148, ML149
and related analogues. Our initial efforts were focused on ML149
as shown in Scheme 1.
The carboxylic acid functionality of commercially available pyr-
role 1 (Scheme 1) was activated using carbonyldiimidazole (CDI) in
THF and the resulting acylimidazole was then treated with hydra-
zine monohydrate to afford hydrazide 2. This reaction was
performed on a two-gram scale, and the product was isolated from
the imidazole (reaction by-product) by recrystallization in ethyl
acetate. Treatment of hydrazide 2 with 1-aza-2-methoxy-1-cyclo-
heptene in chlorobenzene in a microwave (MW) reactor led to for-
mation of triazole 3 in 71% yield. Finally, Ullman-type coupling
between 3 and iodobenzene afforded ML149. The use of 4,7-dime-
thoxy-1,10-phenanthroline (L) as a copper ligand allowed for the
efficient activation of the air-stable metal catalyst;16 furthermore,
using polyethylene glycol (PEG) as a co-solvent improved the sol-
ubilization of the inorganic base and of the copper salt, accounting
for the improved performance of the reaction.17 Using this
synthetic sequence, 18 analogues (4a–r) with differentially substi-
tuted N-pyrrole aromatic rings were synthesized (Table 1).
Overall, the substitution at the meta or para position of the N-
pyrrole phenyl ring with small substituents provided analogues
with activities comparable to that of ML149, however, some subtle
SAR trends emerged. Modification of the para position with a
were not tolerated [e.g., pyrrazole derivative 7b (IC50 = 3.83 lM)].
Optimization of ML148: Following the optimization of ML149,
which led to the identification of compounds 4a (IC50 = 22 nM)
and 4b (IC50 = 34 nM), we turned our attention to benzimidazole-
based chemotype ML148. The synthetic route was optimized in or-
der to provide rapid access to a variety of analogues as shown in
Scheme 4.
Benzoic acid 8 was treated with m-toluidine in N-methylpyro-
lidinone (NMP) to afford intermediate 9. The nitro group was re-
duced using hydrazine and Raney nickel (Ra-Ni) in methanol to
afford the crude 3-amino-4-(m-tolylamino)benzoic acid which
was then converted to benzimidazole 10 via treatment with tri-
ethylorthoformate and catalytic p-toluenesulfonic acid (TsOH) in
THF. This synthetic sequence was carried out on a gram-scale,
and no flash chromatography or HPLC purification was required
to isolate any of the synthetic intermediates. Finally, treatment of
carboxylic acid 10 with a range of amines (R1R2NH), using HATU
and diisopropylethylamine (DIPEA) in DMF provided a library of
14 analogues of ML148 (compounds 11a–n) possessing differing
amide moieties. Unfortunately, none of these compounds resulted
H
H
N
N
N
HN
a
b
c
O
O
N
N
N
N
OH
N
HN
N
NH2
1
2
3
ML149
Scheme 1. Reagents and conditions: (a) CDI, THF, then hydrazine-hydrate, 23 °C, 3.5 h, 65%; (b) 1-aza-2-methoxy-1-cycloheptene, PhCl, MW, 180 °C, 45 min, 71%; (c) PhI,
Cs2CO3, Cu2O, 4,7-dimethoxy-1,10-phenanthroline, NMP-PEG, 100 °C, 16 h, 44%.

632
D. Y. Duveau et al. / Bioorg. Med. Chem. Lett. 24 (2014) 630–635
Table 1
SAR of ML149 and related analogues
H
N
H
n
N
a
Compound
R1
IC50 (nM)
N
N
a
n
b
O
N
N
+
n
OMe
R1
N
N
N
N
HN
NH2
N
n
n
2
6a
6b
1
2
3
4
5a
5b
5c
5d
5e
1
2
3
4
5
N
N
ML149
6c
N
ML149
4a
4b
4c
4d
4e
4f
4g
4h
4i
Phenyl
54
22
34
38
48
121
48
68
34
76
96
86
108
136
38
1078
271
383
108
6d
5
3,4-Cl-phenyl
3-Cl-phenyl
4-Cl-phenyl
4-Br-phenyl
4-F-phenyl
3-CF3-phenyl
4-CF3-phenyl
4-Me-phenyl
3-CN-phenyl
4-CN-phenyl
4-OMe-phenyl
4-NO2-phenyl
4-tBu-phenyl
4-CO2Et-phenyl
2-Pyridine
Scheme 2. Reagents and conditions: (a) PhCl, MW, 180 °C, 45 min; (b) PhI, Cs2CO3,
Cu2O, L, NMP-PEG, 80–100 °C, 16 h.
H
N
H
N
N
X
X
X
4j
a
b
N
O
R
O
R
R
N
N
4k
41
4m
4n
4o
4p
4q
4r
+
MeO
HO
HN
N
NH2
7a-c
5c
Scheme 3. Reagents and conditions: (a) CDI, THF, then hydrazine-hydrate, 23 °C;
(b) (1) 1-aza-2-methoxy-1-cycloheptene, PhCl, MW, 180 °C; (2) PhI, Cs2CO3, Cu2O, L,
NMP-PEG, 100 °C.
3-Pyridine
4-Pyridine
1-Naphthalene
a
Compound
R2
IC50 (nM)
Ph
N
much less active than cyclic ones, and secondary amides (11k
and 11l) were less active than tertiary amides (11h). In order to ex-
plore SAR around the N-linked phenyl ring of ML148, we wanted to
devise a route in which the varying phenyl groups could be intro-
duced in the last step. As such, benzoic acid 8 was treated with
oxalyl chloride and a small amount of DMF in dichloromethane,
and the resulting acyl chloride was treated with piperidine–hydro-
chloride in dichloromethane to afford amide 12 (Scheme 4). The
remainder of the synthesis proceeded as planned, starting with
substitution of the aromatic fluorine with various amines
(R3NH2) in the presence of DIPEA in DMF to provide anilines
13a–n. The aromatic nitro group was reduced, and the resulting
anilines were treated with triethylorthoformate as described above
to afford analogues 14a–n.
n
N
N
N
6a
6b
6c
6d
1
2
4
5
3040
192
242
383
R2
N
N
N
a
Compound
n
IC50 (nM)
Analogues 14a–n displayed activities similar to that of ML148
(Table 2) with relatively flat SAR being observed. Introduction of
a small substituents in para- and in meta-position afforded com-
pounds with similar activities, whereas electron-withdrawing
groups (14e and 14k) and bulkier substituents (14f, 14g and
14l) in para- and in meta-position resulted in a loss of potency.
Incorporation of nonaromatic groups (14m and 14n) had mini-
mal effect on the potency. These data suggest that this region
could be exploited for future modulation of ADME properties
since it appears very tolerant to change. However, it remains
unclear whether introduction of a hydrophilic group will be
tolerated since all analogues prepared thus far are generally
lipophilic in this region.
Ph
N
7a
607
Ph
N
7b
7c
3827
N
Ph
N
681
a
Determined using the HTS enzymatic assay and tested in triplicate.
in improved potency (Table 2); however, it did provide key infor-
mation regarding the steric requirements around the cyclohexyla-
mide functionality. A smaller ring (11i) or an exocyclic amine (11k)
led to compounds with decreased activity (20-fold change and 50-
fold change respectively). Substituents at the 3-position on the
piperidine ring were well tolerated (11a and 11c–e), including
the relatively bulky isopropyl group (11c). While adding a 4-Br
group (11f) only led to a modest decrease in potency, the 4-CF3
group led to a drastic loss of potency (80-fold) compared to the
corresponding 3-CF3 (11e). Moreover, the presence of heteroatoms
(N or O) at the 4-position was also not well tolerated, with activity
in the micromolar range (11m: N–Me–piperazine) or inactivity
(11n: morpholine). Finally, acyclic amides (11h and 11l) were
ADME profile: To assess the potential for these compounds to be
used in vivo, we obtained in vitro ADME properties of a few se-
lected analogues of the ML149 and ML148 chemotypes (Table 3).
In general, ML149 and related analogues showed good kinetic
solubility (>28 lM in phosphate buffered saline (PBS) at pH 7.4).
Results from the PAMPA assay, which measures passive (artificial)
membrane permeability, showed that halogen substituents seem
to improve permeability (e.g., compounds 4a and 4b). On the other
hand, substitution with a nitrile moiety led to a 10-fold drop in
permeability (4i). Unfortunately, ML149 and its analogues
degraded quickly in the presence of rat liver microsomes, as
evidenced by their short half-life (<10 min). ML148 and its
analogues also displayed good kinetic solubility (>47 lM), except

D. Y. Duveau et al. / Bioorg. Med. Chem. Lett. 24 (2014) 630–635
633
O
O
O
R1
N
N
N
N
N
R2
NO2
NH
d
b, c
HO
HO
a
O
9
10
ML148, 11a-n
NO2
F
HO
O
O
O
8
N
N
NO2
NO2 b, c
f
e
N
N
N
NH
R3
F
R3
ML148, 14a-n
13a-n
12
Scheme 4. Reagents and conditions: (a) o-toluidine, NMP, 60 °C; (b) hydrazine, Ra-Ni, MeOH, 23 °C; (EtO)3CH, TsOH, THF, MW, 120 °C; (d) R1R2NH or R3NH2, HATU, DIPEA,
DMF, 23 °C; (e) (COCl)2, DMF, DCM, 23 °C, then piperidine–HCl, DCM, 23 °C; (f) R3NH2, DIPEA, NMP, 120 °C.
Table 2
SAR of ML148 and related analogues
a
a
Compound
R1
IC50 (nM)
Compound
R2
IC50 (nM)
O
O
N
N
N
N
R1
N
R2
14a-n
ML148, 11a-n
N
ML148
11a
19
22
14a
14b
Phenyl
22
27
N
N
2-OMe-phenyl
3-Cl-phenyl
11b
11c
271
48
14c
14d
12
19
N
N
N
N
3-OMe-phenyl
11d
11e
68
68
14e
14f
3-CF3-phenyl
48
68
F3C
3-iPr-phenyl
11f
76
14g
14h
3-tBu-phenyl
171
19
Br
N
11g
1524
4-Me-phenyl
F3C
N
N
N
11h
11i
383
383
14i
14j
4-Cl-phenyl
22
19
4-OMe-phenyl
Ph
11j
429
961
14k
14l
4-CF3-phenyl
86
54
N
H
11k
4-tBu-phenyl
NH
N
11l
1358
2710
14m
14n
tBu
34
68
N
O
11m
nBu
N
11n
3040
a
Determined using the HTS enzymatic assay and tested in triplicate.
for compound 11d (5.6
l
M). PAMPA permeability of ML148 and
of five other dehydrogenases (Supplemental Table 1).18 Encourag-
ingly, none of the compounds tested showed any activity against
GAPDH, LDHA, LDHB and HSD17b4. Some inhibition was observed
against ALDH1A1, though all compounds displayed considerable
selectivity toward HPGD (greater than 28-fold selectivity). Ana-
logues 4b (142-fold selectivity), 14c (160-fold selectivity), and
14e (142-fold selectivity) were the most selective HPGD inhibitors.
related analogues was similar to the ML149 series. As with
ML149, the rat liver microsomal stability remained undesirable,
except for compound 14j (T1/2 = 23 min).
Selectivity toward HPGD: In order to evaluate the selectivity of
the prepared compounds toward HPGD, analogues of ML148 and
of ML149 were tested for their inhibitory activities versus a panel

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D. Y. Duveau et al. / Bioorg. Med. Chem. Lett. 24 (2014) 630–635
Table 3
In vitro ADME properties of selected analogues of ML149 and ML148
a
d
Compound
IC50 (nM)
Permeabilityb (10À6 cm/s)
Kinetic solubilityc
(l
M)
RLM T1/2 (min)
ML149
4a
4b
4f
4h
112
22
34
48
34
76
19
68
12
48
19
599
1646
1100
1512
1330
229
1196
2265
895
>45
32
37
33
28
>49
>47
5.6
>50
>55
>50
Nd
1.9
2.0
1.9
1.8
8.1
7.3
1.8
6.4
4.4
23
4i
ML148
11d
14c
14e
14j
1346
624
a
b
c
Represents the IC50 against HPGD in the HTS enzymatic assay.
Represents the PAMPA permeability at pH 7.4.
Represents the kinetic aqueous solubility in PBS buffer (pH 7.4) as measured by UV quantification.
Represents the T1/2 in rat liver microsomes (RLM) in the presence of NADPH.
d
Table 4
PGE2 produced in A549 and LNCaP cells
Compound
A549 cells
LNCaP cells
% Inductionc
AC50
(
lM)
% Inductionc
a
b
b
IC50 (nM)
AC50
(l
M)
ML149
4a
4b
4f
4h
54
0.743
0.296
0.590
2.09
0.166
2.64
0.469
0.743
0.372
2.64
106
110
114
145
107
350*
129
127
170
173
174
2.80
0.352
0.558
2.22
0.703
1.77
0.884
4.43
0.222
2.80
73
105
68
44
71
63
62
53
85
70
69
22
34
48
34
76
19
68
12
48
19
4i
ML148
11d
14c
14e
14j
0.662
0.884
a
b
c
Represents the IC50 against HPGD in the HTS enzymatic assay.
Represents the half-maximal concentration in the PGE2 production assay.
Represents the percentage induction of PGE2 production.
*
The titration curve was incomplete.
PGE2 production: A selection of top actives around the ML148
data obtained in the PGE2 production assay showed that treatment
with our inhibitors leads to increased PGE2 production in A549
lung cancer and in LNCaP prostate cancer cells, consistent with
inhibition of HPGD in the cell.
and ML149 chemotypes were tested for their ability to increase
PGE2 levels. Lung cancer A549 cells and androgen-sensitive human
prostate adenocarcinoma LNCaP cells were titrated with the se-
lected HPGD inhibitors; the half-maximal activation concentration
(AC50) and percentage induction (% induction) of PGE2 production
are reported in Table 4.
All the compounds tested increased PGE2 production levels
when compared to a DMSO control, in both A549 cells and LNCaP
cells. The percent induction was consistently higher in A549 cells
(from 106% for ML149 to 174% for 14j) than in LNCaP cells (from
44% for 4f to 105% for 4a). The majority of the compounds tested
displayed sub-micromolar activity in both cell lines, and a good
correlation in AC50 was observed between the two cell lines: low
AC50 values measured in A549 cells correlated with low AC50 val-
Conclusions: The structure–activity relationship (SAR) study of
two small molecule inhibitors of the human HPGD, ML148 and
ML149, was carried out. These studies revealed that while the
fused seven-membered ring and the presence of a 2,5-dimethyl-
pyrrole were required for activity of ML149, a certain tolerance
in the substitution pattern of the phenyl ring was observed. As a
result compound 4h was found to have improved potency toward
HPGD, improved cell permeability and induced PGE2 production
both in A549 and in LNCaP cells, when compared to ML149. For
the benzimidazole chemotype (ML148), the cyclohexylamide
group was found to be optimal for enzymatic activity, although
small substituents at the 3-position of the cyclohexylamine group
were tolerated. Moreover, substitution at the meta-position and
para-position of the phenyl ring was also well-tolerated, leading
to analogs 14c and 14j. Both compounds displayed similar ADME
properties, although 14j was more stable in the RLM assay, and
both analogues had comparable effects on PGE2 levels in A549
and in LNCaP cells. Unfortunately, none of the HPGD inhibitors
ues in LNCaP cells, with the exception of ML149 [AC50 = 0.743
l
l
M
M
(A549) and AC50 = 2.80
(A549) and AC50 = 4.43
lM (LNCaP)] and of 11d [AC50 = 0.743
lM (LNCaP)]. Unfortunately, no tractable
SAR emerged from the PGE2 production assay, a likely consequence
of the often diverging enzymatic potency and cell permeability
properties of the inhibitors. Indeed, compound 4i displayed partic-
ularly low cell permeability in the PAMPA assay (229 Â 10À6 cm/s),
probably accounting for the low activity measured in the A549
tested led to any observable down regulation of NF-jB, as mea-
(AC50 = 2.64
lM) and LNCaP (AC50 = 1.77
lM) cell lines. It is also
sured in our assay (Supplemental Fig. 1), perhaps due to compen-
possible that enzyme occupancy of different agents and the
kinetics of the sequential enzymatic events adds a level of
complexity that may account for these differences. Overall, the
satory mechanisms counteracting the effect of modulation of
HPGD activity on NF-jB regulation. In conclusion, we have
identified potent and selective inhibitors of HPGD (e.g., 4h and

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14j) and hope these compounds help better define the role of
HPGD in a variety of diseases. As such, we make these compounds
freely available to the research community upon request.
Acknowledgements
We thank Jim Bougie, Thomas Daniel and William Leister for
compound purification, as well as Danielle van Leer, Misha Itkin,
Crystal Mcknight, Christopher LeClair and Paul Shinn for assistance
with compound management. We also thank Udo Oppermann at
the Structural Genomics Consortium (SGC), Oxford, UK for provid-
ing the ALDH1A1 and HSD17b4 enzymes. This research was sup-
ported by the Molecular Libraries Initiative of the National
Institutes of Health Roadmap for Medical Research Grant
U54MH084681 and the Intramural Research Program of the Na-
tional Center for Advancing Translational Sciences at the National
Institutes of Health.
11. Snyder, N. W.; Revello, S. D.; Liu, X.; Zhang, S.; Arora, J. S.; Blair, I. A. Presented
at the AACR 103rd Annual Meeting, Chicago, IL, March 31–Apr 4, 2012.
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Supplementary data
15. Niesen, F. H.; Shultz, L.; Jadhav, A.; Bhatia, C.; Guo, K.; Maloney, D. J.; Pilka, E. S.;
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18. The activity data of ML148 and of ML149 versus ALDH1A1, HADH2 and
HSD17b4 were reported in Ref. 15.
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2013.
11.081.
References and notes
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