Bioorganic & Medicinal Chemistry Letters
Identification of indole inhibitors of human hematopoietic
prostaglandin D synthase (hH-PGDS)
2
Fredrik Edfeldt a, , Johan Evenäs , Matti Lepistö , Alison Ward , Jens Petersen , Lisa Wissler ,
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a
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Mattias Rohman , Ulf Sivars , Karin Svensson , Matthew Perry , Isabella Feierberg , Xiao-Hong Zhou ,
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Thomas Hansson , Frank Narjes
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Discovery Sciences, Innovative Medicines, AstraZeneca R&D, 431 83 Molndal, Sweden
Respiratory, Inflammation and Autoimmunity, Innovative Medicines, AstraZeneca R&D, 431 83 Molndal, Sweden
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a r t i c l e i n f o
a b s t r a c t
Article history:
Human H-PGDS has shown promise as a potential target for anti-allergic and anti-inflammatory drugs.
Here we describe the discovery of a novel class of indole inhibitors, identified through focused screening
of 42,000 compounds and evaluated using a series of hit validation assays that included fluorescence
polarization binding, 1D NMR, ITC and chromogenic enzymatic assays. Compounds with low nanomolar
potency, favorable physico-chemical properties and inhibitory activity in human mast cells have been
identified. In addition, our studies suggest that the active site of hH-PGDS can accommodate larger struc-
tural diversity than previously thought, such as the introduction of polar groups in the inner part of the
binding pocket.
Received 10 March 2015
Revised 19 April 2015
Accepted 20 April 2015
Available online 28 April 2015
Keywords:
Prostaglandin D
PGDS inhibitors
Indole
2
synthase
Ó 2015 Elsevier Ltd. All rights reserved.
Focused screening
Hit validation
Prostaglandin D
2
(PGD
2
) is an allergic and inflammatory medi-
and more ligand efficient hits. A fluorescence polarization assay10
was used to screen two library subsets of 25,000 and 17,000 com-
ator produced by mast cells and Th2 cells after cross-linking of
an allergen with the specific IgE antibody on the cell membrane.
1
pounds at 125
determined for compounds showing a >30% displacement effect,
resulting in 1040 compounds with IC50 values <100 M. To further
validate the actives a 1D NMR binding assay was deployed. K val-
ues were determined using the competitive displacement of repor-
lM and 250 lM, respectively. IC50 values were
PGD
that have been associated with inflammatory conditions.
Production of PGD in the peripheral tissues and in immune and
inflammatory cells from the precursor prostaglandin H (PGH ) is
2 1
acts on two G-protein-coupled receptors, DP and CRTH2
2
l
2
d
2
2
1
1
mainly catalyzed by human hematopoietic prostaglandin D syn-
thase (hH-PGDS). This has led hH-PGDS to be envisioned as a target
for the treatment of asthma and inflammatory diseases. hH-PGDS
is a 26-kDa cytosolic homodimer of the sigma class glutathione-S-
transferase (GST) family. It depends on glutathione (GSH) for cat-
alytic activity, which is further increased by divalent metal ions.
ter ligands with known affinity.
The hits were clustered using structural similarity and 216 clus-
ter representatives were selected for testing in the NMR assay. 187
of these hits were confirmed in the NMR assay, corresponding to a
validation rate of 87%. Validated hits were then tested in a GST
enzymatic assay using CDNB (1-chloro-2,4-dinitrobenzene) as
3
4
1
2
Several inhibitors of hH-PGDS, most of them containing a signature
bis-aryl amide motif, have been disclosed in recent years by a
chromogenic substrate and IC50 values were generated. We chose
to monitor the GST enzymatic activity of hH-PGDS rather than its
5
number of organizations, such as (1 and 2) Pfizer, (3) Sanofi-
PGD
2
synthase activity due to technical feasibility. Both enzymatic
Aventis,6 (4) Evotec, (5) AstraZeneca8 and (6) Taiho9 (Fig. 1). For
7
activities reside in the same binding pocket.
4b,13
For a set of four
a recent review of hH-PGDS inhibitors, see Ref. 3.
reference compounds (1–3, and 5) activity in the NMR and GST
Our previous efforts led to the identification of compound (5) as
an hH-PGDS inhibitor. The current study reports our continued
work on identifying novel inhibitors of this enzyme applying a
fragment based screening approach, aiming to identify smaller
assays, as well as isothermal titration calorimetry (ITC),14 corre-
lated overall well with the inhibition of PGD
2
production in
1
5
megakaryoblast cells (Table 1). We therefore felt confident that
our screening set-up was fit for purpose in driving hit expansion
chemistry in a high throughput manner. We observed excellent
correlation between the NMR binding assay and inhibition of GST
activity for our screening hits. 168 compounds had an IC50
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960-894X/Ó 2015 Elsevier Ltd. All rights reserved.
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