Fragment-Based Discovery of 6-Arylindazole JAK Inhibitors

Article abstract of DOI:10.1021/acsmedchemlett.6b00087

Janus kinase (JAK) inhibitors are emerging as novel and efficacious drugs for treating psoriasis and other inflammatory skin disorders, but their full potential is hampered by systemic side effects. To overcome this limitation, we set out to discover soft drug JAK inhibitors for topical use. A fragment screen yielded an indazole hit that was elaborated into a potent JAK inhibitor using structure-based design. Growing the fragment by installing a phenol moiety in the 6-position afforded a greatly improved potency. Fine-tuning the substituents on the phenol and sulfonamide moieties afforded a set of compounds with lead-like properties, but they were found to be phototoxic and unstable in the presence of light.

Full text of DOI:10.1021/acsmedchemlett.6b00087

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Letter
Fragment-based discovery of 6-arylindazole JAK inhibitors
Andreas Ritzén, Morten Dahl Sørensen, Kevin N. Dack, Daniel R Greve,
Anders Jerre, Martin A Carnerup, Klaus A Rytved, and Jesper Bagger-Bahnsen
ACS Med. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/acsmedchemlett.6b00087 • Publication Date (Web): 14 Apr 2016
Downloaded from http://pubs.acs.org on April 18, 2016
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Fragment-based discovery of 6-arylindazole JAK inhibitors
Andreas Ritzén,*,† Morten D. Sørensen,† Kevin N. Dack,† Daniel R. Greve,† Anders Jerre,‡ Martin A.
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Carnerup,† Klaus A. Rytved,§ and Jesper Bagger-Bahnsen‖
†Drug Design, ‡In vitro Biology, §Skin PK and Early Safety, ‖Preformulation & Early Analytical Development, Global R&D,
LEO Pharma A/S, Industriparken 55, DK-2750 Ballerup, Denmark.
KEYWORDS: Fragments, JAK inhibitors, psoriasis, indazoles, phototoxicity
ABSTRACT: Janus kinase (JAK) inhibitors are emerging as novel and efficacious drugs for treating psoriasis and other inflamma-
tory skin disorders, but their full potential is hampered by systemic side effects. To overcome this limitation, we set out to discover
soft drug JAK inhibitors for topical use. A fragment screen yielded an indazole hit that was elaborated into a potent JAK inhibitor
using structure-based design. Growing the fragment by installing a phenol moiety in the 6-position afforded a greatly improved
potency. Fine-tuning the substituents on the phenol and sulfonamide moieties afforded a set of compounds with lead-like properties,
but they were found to be phototoxic and unstable in the presence of light.
Psoriasis is a debilitating and chronic skin disorder affecting
as many as one in 25 adults in Europe and one in 50 in the
US.1 A recent analysis of survey data from the National Psori-
asis Foundation in the US showed that 52.3% of patients with
psoriasis were dissatisfied with their treatment.2 An urgent
need for safe and effective therapies clearly exists. Psoriasis is
a T cell-mediated chronic skin inflammation resulting from a
complex interplay between immune and skin cells.3 These
cells communicate using soluble mediators e.g. cytokines.4
Intracellular signaling relevant to psoriasis is relayed from
cytokine receptors by the Janus kinase (JAK) family of tyro-
sine kinases. It consists of four members (JAK1, JAK2, JAK3
and TYK2) that, upon activation by cytokine receptors, phos-
phorylate signal transducers and activators of transcription
(STATs).5 JAK inhibitors have been extensively investigated
because of their potential to treat chronic inflammatory condi-
tions e.g. rheumatoid arthritis (RA), psoriasis and atopic der-
matitis (AD).6,7 Two selective JAK inhibitors have been ap-
proved by the FDA: Tofacitinib (Xeljanz®) for treating RA,
and Ruxolitinib (Jakafi®) for treating myelofibrosis and poly-
cythemia vera. Both drugs have shown significant benefit in
psoriasis in clinical trials,8 and Tofacitinib was also found to
be effective in AD.9,10 However, because of their immunosup-
pressant and anemic effects, systemic administration of these
drugs may cause serious adverse events, e.g. infections, infes-
tations and decreased hemoglobin levels.11 Topical administra-
tion for the treatment of psoriasis has been investigated for
both drugs.8 Percutaneous absorption into systemic circulation
may still limit the surface area that can be treated, although
lower systemic levels than after oral administration were
found.12 To avoid systemic side effects while delivering an
effective treatment for dermatology indications, we set out to
discover a topical JAK family inhibitor that would be ab-
sorbed into the skin and rapidly metabolized once in the blood
stream; i.e. a soft drug.13
ments was screened against JAK2 at a single concentration
(100 μM) using SPR. Hits were validated by determination of
KD using SPR and determination of IC50 in a biochemical
enzyme inhibition assay for JAK1. In general, a good correla-
tion was observed between these two measurements suggest-
ing that most fragments had little or no selectivity for JAK2
over JAK1. Because we targeted pan-JAK inhibitors, this
result made us confident in the validity of the hits, and several
fragments were progressed to X-ray crystallography. In partic-
ular, compound 1 appeared to afford an excellent starting point
because of its compact size, available vectors and good ligand
efficiency. Although indazoles had been identified as viable
fragments for JAK2 previously,14 the sulfonamide moiety in
the 4-position of 1 offered an unexplored handle. We envi-
sioned that this feature in combination with fragment growth
from other positions would yield potent and selective JAK
inhibitors. Thus, we selected 1 for progression into hit-to-lead.
Compound 1
6
JAK1 pIC50 = 4.5 (30 µM)
5
JAK2 pKD = 4.9 (13 µM)
3
4
JAK1 LE = 0.45
Figure 1. Fragment screen hit 1.
An X-ray crystal structure of 1 in complex with JAK2 was
obtained, see Figure 2. Compound 1 occupies the purine re-
gion of the active site and makes two hydrogen-bonded con-
tacts with the hinge residues Glu930 and Leu932. The binding
mode of 1 is similar to that reported for other indazoles, but
with an unfilled back pocket (BP) wider than previously ob-
served.14 The widened back pocket in the structure of 1 is
caused by a conformational change of Gly993 immediately
preceding the DFG residues. We speculated that growing into
this pocket might afford both potency and selectivity. To this
end, we surveyed published JAK crystal structures in the PDB
In order to identify novel JAK inhibitors we performed a
fragment screen. A kinase-targeted library of ca. 500 frag-
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and identified two potent JAK2 inhibitors from Vertex with
phenol moieties in the back pocket (PBD accession codes
3JY915 and 3KCK16). In contrast, a 6-phenylindazole had been
reported to be virtually inactive in JAK2.14 This discrepancy
can be attributed to a stabilizing hydrogen bond from the phe-
nol hydroxyl group to Glu898.
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3
Figure 3. Model of phenol 2 (green) docked into the JAK2 struc-
ture 3KCK16 (cyan; crystallographic ligand 3 shown as
wireframe) compared to the X-ray crystal structure of 1 (gray).
On the basis of this analysis, we proceeded to synthesize
phenol 2, see Scheme 1. The key intermediate 5 was readily
prepared from commercially available indazole 4 using stand-
ard conditions. Suzuki coupling with a TBS-protected phenol
boronic acid afforded 6, which upon nitro group reduction and
reaction with methanesulfonyl chloride yielded the protected
derivative 8. A global deprotection step furnished the desired
phenol 2.
Figure 2. X-ray structure of compound 1 bound to JAK2 (PDB:
5L3A).
Scheme 1. Synthesis of phenol 2a
As a first step to grow fragment 1 we set out to append a
phenol moiety to the 6-position. This proposal was supported
by docking a prototypical phenol-substituted indazole 2 into
the literature JAK2 structure 3KCK.16 We chose this structure
for two reasons: the kinase backbone and active site side
chains of 3KCK and the structure of 1 overlaid very well; and
the crystallographic ligand 3 in 3KCK has a phenol in the back
pocket. Comparison of the docked pose of 2 with the crystal
structures of 1 and 3KCK showed that both the indazole moie-
ty and the phenol moiety of 2 overlaid well with the corre-
sponding moieties in the crystallographic ligands (figure 3).
All three ligands make similar hydrogen bonding interaction
with the hinge residues Glu930 and Leu932. The phenols of 2
and 3 also make very similar hydrogen bonding interactions
with the αC helix residue Glu898 and the backbone NH of
Phe995 in the DFG loop. The ligands do not directly interact
with the catalytic lysine Lys882, but this residue is in contact
with Glu898 in both structures. It is also worth noting that the
gatekeeper Met929 is in close contact with the aromatic ring
of the phenol: the Met S to aromatic ring centroid distance is
3.3 Å. The geometry of this complex suggests a stabilizing
S…π interaction.17
a
b
4
5
e
6: X = NO2
7: X = NH2
8: X = NHSO2Me
2
c
d
aReagents and conditions: (a) DHP, PPTS, CH2Cl2, reflux, 3 h,
77%; (b) 4-TBSO-C6H4-B(OH)2, K2CO3, Pd(dppf)Cl2, DMF,
water, rt, 1 h, 94%; (c) Pd/C, H2 (1 bar), EtOAc, rt, 6 h, 58%; (d)
MsCl, pyridine, CH2Cl2, rt, 3.5 h, 54%; (e) 4 M HCl in dioxane,
MeOH, rt, 2 h, 71%.
Gratifyingly, 2 exhibited greatly improved potency and also
improved LE compared with fragment 1, see Table 1. This
compound also showed low micromolar activity in a cellular
STAT6 reporter assay. Compound 2 appeared to be good
starting point for further optimization, and to this end, varia-
tions of the phenol and sulfonamide moieties were envisioned
as the next steps. Considerations of the binding mode and the
distance between these two moieties suggested that they could
be optimized independently. Based on previous experience
with other series of JAK inhibitors, we set criteria for a lead
compound to be cellular pIC50 ≥ 7 and LLE ≥ 3.
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Table 1. Optimization of the phenol moietya
1
3
2
3
4
4
5
2
6
7
8
1
9
JAK1
Cellular
pIC50
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
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34
35
36
37
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51
52
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56
57
58
59
60
Compound
R1
R2
R3
R4
JAK2
JAK3
TYK2
S+ log Pb
pIC50
LE
LLE
3.5
3.5
3.4
3.8
3.4
2.9
3.1
3.3
3.4
1.9
3.3
3.3
Me
Et
Et
Et
Et
Et
Et
Et
Et
Et
Me
Et
H
H
H
H
H
H
H
H
H
H
H
H
H
7.47 ± 0.09 (2)
7.43 ± 0.06 (2)
7.26 ± 0.05 (3)
7.78 ± 0.15 (2)
7.72 ± 0.11 (2)
6.53 ± 0.03 (3)
7.15 ± 0.11 (2)
7.54 ± 0.11 (3)
6.07 ± 0.17 (2)
0.50
0.47
0.44
0.45
0.47
0.40
0.44
0.46
0.35
0.32
0.49
0.47
7.7
8.0
7.5
7.7
7.7
6.9
7.7
8.1
6.0
5.7
8.1
8.3
7.9
8.1
7.6
8.1
7.8
7.2
7.9
8.0
6.6
5.6
8.6
8.6
n.d.
6.4
6.1
6.3
6.5
6.1
6.6
6.6
5.6
5.2
n.d.
6.8
5.4
5.7
1.9
2.3
2.4
2.7
2.8
2.6
2.9
2.5
2.3
2.7
3.1c
3.5c
2
9
Me
Et
Cl
H
H
5.78 ± 0.02 (2)
6.5
10
11
12
13
14
15
16
17
18
19
H
H
6.15 ± 0.05 (2)
5.5
Me
Cl
F
H
6.0
H
5.80 ± 0.01 (2)
5.67 ± 0.02 (2)
4.60 ± 0.03 (2)
6.40 ± 0.03 (3)
6.77 ± 0.03 (3)
H
OMe
H
Me
Et
Et
Me 5.54 ± 0.10 (3)
F
H
H
8.4
F
8.38 ± 0.01 (3)
aData given as pIC50 determined from single experiments with 10-point titration curves unless otherwise noted. Values from multiple
determinations are given as mean ± SEM (N), where SEM is the standard error of the mean and N is the number of replicates. Enzymatic
assays: HTRF (Homogenous Time-Resolved Fluorescence) based kinase assay with ATP concentration at Km. Cellular assay: STAT6
reporter gene assay in a human B lymphocyte cell line (Ramos). Calculated using Simulations Plus, Inc. MedChem DesignerTM cExperi-
b
mentally determined log D at pH 7.4.
Expansion of the methyl sulfonamide to the ethyl derivative
9 afforded a slight increase in cellular potency with no change
in LLE, see Table 1. Variations of the synthetic route shown in
Scheme 1 were used to prepare the compounds in tables 1 and
2; see Supporting Information for details. Compound 9 was
chosen as the reference point for optimization of the phenol
moiety. Inspection of the 3KCK structure showed that the
chloro substituent of compound 3 filled a lipophilic sub-pocket
lined by Val863 and the Cβ-Cδ methylenes of Lys882. Thus,
the small, lipophilic substituents methyl, ethyl and chloro were
installed at position R2 to afford compounds 10, 11 and 12. All
three compounds showed slight improvements in cellular
potency with the ethyl analog 11 being the best in terms of
both potency and LLE. Next, the R3 position was probed with
methyl, chloro, fluoro and methoxy substituents (compounds
13-16). For reasons of synthetic expediency, the R2 position
was left unsubstituted in the hope that optimum substituents
could be combined later. Compared with 9, moderate varia-
tions in enzymatic JAK1 and cellular STAT6 potency were
observed for these derivatives with the halides 14 and 15
emerging as the best of these substituents.
we became aware of a patent application claiming a series of
indazole JAK inhibitors with no 4-substituents, but with sub-
stituted bicyclic heterocycles in the 3-position and phenols in
the 6-position.18 Assuming that these indazoles bind with a
similar binding mode to that of 2, the phenol substituent would
be expected to overlap well between these two series. Interest-
ingly, this publication suggested the combination of an R2
ethyl and an R3 fluoro substituent. Because this is consistent
with the SAR we had already derived for these substituents,
we proceeded to prepare this combination, both with a methyl
(18) and ethyl sulfonamide (19). Compared with compound
11, the extra fluoro substituent of 19 afforded a slight im-
provement in both enzymatic JAK1 and cellular STAT6 po-
tency. Similar to that which was observed when comparing 2
with 9, the ethyl derivative 19 had a slight edge over the me-
thyl analog 18 in terms of cellular potency with no change in
enzymatic potency or LLE.
Armed with these results, we undertook an optimization of
the sulfonamide substituent using 19 as the new reference
compound, see Table 2. First, methylation of the free NH of
the sulfonamide was investigated, but the resulting compound
20 showed a marked drop in potency compared with 19. Next,
the small alkyl sulfonamides 21-24 and the trifluoromethane
sulfonamide 25 were prepared and tested. Of these, the iso-
propyl (21) and cyclopropyl (22) analogs showed similar
Having explored the R2 and R3 positions, we set out to try
various combinations of substituents. Because the phenol ring
plane was twisted out of the plane of the indazole moiety in
the docking pose of 2 (Figure 3), we prepared the dimethyl
analog 17, which has a conformational preference for this
geometry. Unfortunately, this compound showed poor activity,
probably due to steric clashes with the protein. At this point,
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Table 2. Optimization of the sulfonamide moietya
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JAK1
pIC50
Cellular
pIC50
Compound
X
JAK2
JAK3 TYK2
S+ log Pb
10
11
12
13
14
15
16
17
18
19
20
21
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LE
LLE
2.8
N(Me)SO2Me
NHSO2i-Pr
NHSO2c-Pr
7.5
0.42
0.44
0.43
7.1
8.1
8.0
7.5
8.6
8.5
n.d.
n.d.
6.5
5.7
2.9
3.5c
3.3
20
21
22
8.16 ± 0.08 (2)
8.04 ± 0.07 (3)
6.79 ± 0.05 (2)
6.70 ± 0.02 (3)
3.3
3.4
7.85 ± 0.06 (3)
7.60 ± 0.04 (3)
0.39
0.38
7.7
7.6
8.4
8.4
6.1
6.4
6.4
2.3
2.3
4.1
4.1
23
24
6.39 ± 0.04 (3)
NHSO2CF3
7.16 ± 0.12 (2)
7.94 ± 0.13 (3)
8.03 ± 0.15 (3)
0.37
0.43
0.42
0.40
6.8
7.7
7.8
8.0
7.6
8.2
8.4
8.5
6.0
6.2
6.3
6.4
5.53 ± 0.04 (2)
1.2
4.0
3.6
3.7
4.4
1.9
2.8
2.7
25
26
27
28
NHSO2CH2CH2OH
NHSO2CH2CH2OMe
6.0
6.4
6.4
NHSO2CH2CH2CO2Me 8.26 ± 0.15 (3)
6.98 ± 0.05 (3)
0.32
0.30
6.8
6.7
7.5
7.2
5.2
5.1
5.2
5.1
1.2
2.0
4.1
3.2
29
30
6.77 ± 0.13 (3)
7.78 ± 0.07 (2)
0.36
8.1
8.5
6.5
6.41 ± 0.02 (3)
1.8
4.6
31
7.53 ± 0.04 (2)
7.46 ± 0.13 (2)
0.36
0.36
7.7
7.6
8.7
8.3
6.2
5.6
5.67 ± 0.01 (2)
5.3
2.3
2.1
3.3
3.2
32
33
NHSO2NMe2
NHP(O)Et2
8.36 ± 0.06 (3)
6.08 ± 0.01 (3)
0.45
0.33
8.2
5.5
8.5
6.0
6.8
4.7
7.00 ± 0.08 (3)
4.8
3.9
1.5
3.1c
3.3
34
35
aData given as pIC50 determined from single experiments with 10-point titration curves unless otherwise noted. Values from multiple
determinations are given as mean ± SEM (N), where SEM is the standard error of the mean and N is the number of replicates. Enzymatic
assays: HTRF (Homogenous Time-Resolved Fluorescence) based kinase assay with ATP concentration at Km. Cellular assay: STAT6
reporter gene assay in a human B lymphocyte cell line (Ramos). Calculated using Simulations Plus, Inc. MedChem DesignerTM cExperi-
b
mentally determined log D at pH 7.4.
potency and lipophilicity compared to 19, but none of these
had any obvious advantage over 19. Because the sulfonamide
is positioned at the solvent front of the active site, hydrophilic
substituents were appended to the ethyl sulfonamide to im-
prove solubility and reduce lipophilicity (compounds 26-30).
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Unfortunately, all these analogs lost cellular potency to var-
while monitoring the compound concentrations using LCMS.
Although indazole itself was reasonably stable in this test, 1,
19, and 34 were rapidly degraded, see Figure 5. For topical
use, excellent photostability is required, implying that a re-
design of the scaffold to improve this property would be need-
ed.
1
2
3
4
5
6
7
8
ious degrees. Aromatic (31) and heteroaromatic (32, 33) sub-
stituents were also tried, but again, a loss of potency and LLE
was observed for these analogs. On the other hand, the dime-
thylsulfamide 34, formally an aza analog of 21, showed im-
proved cellular potency and LLE compared with both 19 and
21. Finally, a phosphinamide analog 35 was prepared and
tested, but this compound was almost inactive in the cellular
STAT6 assay.
TRKA
JAK3
PDGFRα
FGFR1
KDR
Compounds 18, 19, 21 and 34 were selected for further pro-
filing on the basis of cellular potency and LLE considerations.
Because we were targeting a soft topical drug, we sought
compounds with a rapid hepatic clearance. Specifically, we
targeted an intrinsic clearance in human liver microsomes
greater than three times the liver blood flow rate (assumed to
be 20 mL/min/kg). The intrinsic clearances of the selected
compounds are shown in Table 3; although none satisfy the
softness criteria, compounds 21 and 34 were close enough to
merit further investigation.
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
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ABL
LCK
EPHA2
TIE2
FLT3
SRC
SYK
ITK
Table 3. In vitro PK and toxicology
TYRO3
PYK2
MET
a
Compound
Clint,H
3T3 Phototoxicityb
hERG IC50
+UV
-UV
31.9
>125
PIF
EPHB4
EGFR
18
19
21
34
13
17
47
47
Compound 19
CSK
14.2
11
2.3
>100 µM
>100 µM
Compound 34
IGF1R
0
25
50
75 100
>11.5
% Inhibition
aIntrinsic clearance in human liver microsomes (mg/mL/min).
bCellular viability IC50 (µg/mL) in the presence (+UV) and ab-
sence (-UV) of non-toxic doses of ultraviolet UVA irradiation,
and the ratio between the two (PIF). A PIF value in the range of 2-
5 predicts probable phototoxicity and PIF > 5 predicts phototoxi-
city.22
Figure 4. Tyrosine kinase panel selectivity results for compounds
19 and 34.19
0
-0.5
-1
Next, we undertook an in vitro kinome selectivity and toxi-
cology profiling of 19 and 34. The compounds were tested in a
panel of 20 tyrosine kinases19 at a single concentration chosen
to be 1000-fold their respective JAK1 IC50, see Figure 4. Both
compounds showed a similar selectivity profile with a number
of off-target kinases inhibited. To quantify and compare the
selectivity of these compounds, we calculated a Gini coeffi-
cient from this data. In this context, the Gini coefficient is a
measure of selectivity: a completely selective inhibitor (one
kinase 100% inhibited and all others 0% inhibited) has a Gini
coefficient of 1, and a completely non-selective inhibitor (all
kinases equally inhibited) has a value of 0.20 The Gini coeffi-
cient has been shown to predict the kinome-wide selectivity of
a compound from single-point panel data using as few as 20
kinases.20 We found a Gini coefficient of 0.27 for compounds
19 and 34 and a Gini coefficient of 0.71 for the highly selec-
tive JAK inhibitor tofacitinib21 in the same panel. The com-
pounds were devoid of binding to the hERG potassium chan-
nel at 100 µM. Unfortunately, 19 showed probable phototoxi-
city and 34 showed clear phototoxicity in the 3T3 NRU photo-
toxicity test.22 Because indazole is rapidly photodegraded in
aqueous solution,23 we hypothesized that the observed photo-
toxicity might be due to the formation of reactive or toxic
photodegradation products. To determine if these compounds
were degraded by light, we exposed solutions of indazole, 1,
19, and 34 to simulated outdoor light for a period of six hours
-1.5
Compound t½ (h)
Indazole
1
66
3.0
0.9
0.8
-2
19
34
-2.5
0
2
4
6
Time (h)
Figure 5. Photostability of indazole, 1, 19, and 34. Compounds
were dissolved at 0.1 mg/mL in 1:1 MeCN : aqueous phosphate
buffer (pH = 7.4) and irradiated with simulated outdoor light.
In conclusion, a hit from a fragment screen was elaborated
into a lead series of 6-arylindazoles. In two iterations, potency
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(8) Hsu, L.; Armstrong, A. W. JAK inhibitors: treatment efficacy
and safety profile in patients with psoriasis. J. Immunol. Res. 2014,
2014:283617.
(9) Levy, L. L.; Urban, J.; King, B. A. Treatment of recalcitrant
atopic dermatitis with the oral Janus kinase inhibitor tofacitinib cit-
rate. J. Am. Acad. Dermatol. 2015, 73, 395-399.
was increased by four orders of magnitude while preserving
the excellent LE of the fragment hit. The lead compounds
were moderately selective for the JAK family of tyrosine
kinases and possessed in vitro PK properties consistent with
our targeted soft drug profile. Poor light stability and photo-
toxicity deriving from the indazole core point to a need for
further optimization of the scaffold.
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Raman, M.; Papacharalambous, J.; Wang, C.; Purohit, V.; Mamolo,
C.; Mallbris, L; Ports, W. Efficacy and safety of topical Janus kinase
inhibitor tofacitinib in the treatment of atopic dermatitis: results of a
Phase 2a randomized clinical trial. Poster presented at 23rd World
Congress of Dermatology, Vancouver, Canada, June 8-13, 2015.
(11) Papp, K. A.; Menter, A.; Strober, B.; Langley, R. G.; Buonan-
no, M.; Wolk, R.; Gupta, P.; Krishnaswami, S.; Tan, H.; Harness, J.
A. Efficacy and safety of tofacitinib, an oral Janus kinase inhibitor, in
the treatment of psoriasis: a Phase 2b randomized placebo-controlled
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(12) Ports, W. C.; Khan, S.; Lan, S.; Lamba, M.; Bolduc, C.; Bis-
sonnette, R.; Papp K. A randomized phase 2a efficacy and safety trial
of the topical Janus kinase inhibitor tofacitinib in the treatment of
chronic plaque psoriasis. Br. J. Dermatol. 2013, 169, 137-145.
(13) Bodor, N.; Buchwald, P. Soft drug design: general principles
and recent applications. Med. Res. Rev. 2000, 20, 58-101.
(14) Antonysamy, S.; Hirst. G.; Park, F.; Sprengeler, P.; Stap-
penbeck, F.; Steensma, R.; Wilson, M.; Wong. M. Fragment-based
discovery of JAK-2 inhibitors. Bioorg. Med. Chem. Lett. 2009, 19,
279-282.
ASSOCIATED CONTENT
Supporting Information
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The Supporting Information is available free of charge on the
ACS Publications website.
Full experimental details for the compounds synthesized and
descriptions of biological assays (PDF)
AUTHOR INFORMATION
Corresponding Author
* E-mail: andreas.ritzen@leo-pharma.com.
Author Contributions
A.R., M.D.S., K.N.D., and D.R.G. contributed to the design of
compounds, computer modelling, and interpretation of results.
A.J. performed the enzymatic and cell-based JAK assays and
provided the kinase panel selectivity data. M.A.C. provided the
intrinsic clearance data, K.A.R. provided the in vitro toxicology
(hERG and phototoxicity) data, and J.B.-B. performed the photo-
stability study.
(15) Wang, T.; Duffy, J. P.; Wang, J.; Halas, S.; Salituro, F. G.;
Pierce, A. C.; Zuccola, H. J.; Black, J. R.; Hogan, J. K.; Jepson, S.;
Shlyakter, D.; Mahajan, S.; Gu, Y.; Hoock, T.; Wood, M.; Furey, B.
F.; Frantz, J. D.; Dauffenbach, L. M.; Germann, U. A.; Fan, B.;
Namchuk, M.; Bennani, Y. L.; Ledeboer, M. W. Janus Kinase 2
Inhibitors. Synthesis and Characterization of a Novel Polycyclic
Azaindole. J. Med. Chem. 2009, 52, 7938-7941.
Notes
(16) Wang, T.; Ledeboer, M. W.; Duffy, J. P.; Pierce, A. C.; Zuc-
cola, H. J.; Block, E.; Shlyakter, D.; Hogan, J. K.; Bennani, Y. L. A
novel chemotype of kinase inhibitors: Discovery of 3,4-ring fused 7-
azaindoles and deazapurines as potent JAK2 inhibitors. Bioorg. Med.
Chem. Lett. 2010, 20, 153-156.
(17) Beno, B. R.; Yeung, K.-S.; Bartberger, M. D.; Pennington, L.
D.; Meanwell, N. A. A Survey of the Role of Noncovalent Sulfur
Interactions in Drug Design. J. Med. Chem. 2015, 58, 4383-4438
(18) Coe, J. W.; Dehnhardt, C. M.; Jones, P.; Kortum, S. W.; Sab-
nis, Y. A.; Wakenhut, F. M.; Whitlock, G. A. Indazoles. PCT Pat.
Appl. WO2013014567, Jan. 31, 2013.
(19) Carna Biosciences, Inc. QuickScout™ TK Panel; assay ATP
concentration at KM of the respective enzyme.
(20) Graczyk, P. P. Gini Coefficient: A New Way To Express Se-
lectivity of Kinase Inhibitors against a Family of Kinases. J. Med.
Chem. 2007, 50, 5773-5779.
(21) Davis, M. I.; Hunt, J. P.; Herrgard, S.; Ciceri, P.; Wodicka, L.
M.; Pallares, G.; Hocker, M.; Treiber, D. K.; Zarrinkar, P. P. Com-
prehensive analysis of kinase inhibitor selectivity. Nat Biotechnol.
2011, 29, 1046-1051.
(22) A thorough description of the assay can be found in:
OECD (2004), Test No. 432: In Vitro 3T3 NRU Phototoxicity Test,
OECD Guidelines for the Testing of Chemicals, Section 4, OECD
Publishing, Paris.
The authors are employees of LEO Pharma A/S.
ACKNOWLEDGMENT
We gratefully acknowledge Thomas Vifian and Zhixing Wu for
synthesizing the compounds presented in this paper, and Paola
Lovato for helpful discussions during the preparation of this
manuscript.
ABBREVIATIONS
AD, atopic dermatitis; JAK, Janus kinase; LE, ligand efficiency;
LLE, ligand lipophilic efficiency; n.d., not determined; RA,
rheumatoid arthritis.
REFERENCES
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Fragment hit
JAK1 pIC50 = 4.5
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Cell pIC50 = 7.0
LE = 0.45
LE = 0.45
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