Discovery of 5-(2-amino-[1,2,4]triazolo[1,5-a]pyridin-7-yl)-N-(tert-butyl) pyridine-3-sulfonamide (CZC24758), as a potent, orally bioavailable and selective inhibitor of PI3K for the treatment of inflammatory disease

Article abstract of DOI:10.1016/j.bmcl.2012.05.090

Herein, we disclose the discovery of a series of 7-substituted triazolopyridines which culminated in the identification of 14 (CZC24758), a potent, orally bioavailable small-molecule inhibitor of PI3Kγ, an attractive drug target for inflammatory and autoimmune disorders. Compound 14 has excellent selectivity across the kinome, demonstrates good potency in cell based assays and furthermore exhibits in vivo efficacy in a collagen induced arthritis model in mouse after oral dosing.

Full text of DOI:10.1016/j.bmcl.2012.05.090

Bioorganic & Medicinal Chemistry Letters 22 (2012) 4613–4618
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery of 5-(2-amino-[1,2,4]triazolo[1,5-a]pyridin-7-yl)-N-(tert-butyl)
pyridine-3-sulfonamide (CZC24758), as a potent, orally bioavailable and
selective inhibitor of PI3K for the treatment of inflammatory disease
Mihiro Sunose a, , Kathryn Bell a, Katie Ellard a, Giovanna Bergamini b, Gitte Neubauer b, Thilo Werner b,
⇑
Nigel Ramsden a
a Cellzome Ltd, Chesterford Research Park, Little Chesterford CB10 1XL, UK
b Cellzome AG, Meyerhofstr. 1, 69117 Heidelberg, Germany
a r t i c l e i n f o
a b s t r a c t
Article history:
Herein, we disclose the discovery of a series of 7-substituted triazolopyridines which culminated in the
Received 4 May 2012
Revised 24 May 2012
Accepted 28 May 2012
Available online 5 June 2012
identification of 14 (CZC24758), a potent, orally bioavailable small-molecule inhibitor of PI3Kc, an attrac-
tive drug target for inflammatory and autoimmune disorders. Compound 14 has excellent selectivity
across the kinome, demonstrates good potency in cell based assays and furthermore exhibits in vivo effi-
cacy in a collagen induced arthritis model in mouse after oral dosing.
Ó 2012 Elsevier Ltd. All rights reserved.
Keywords:
PI3K
c
Inflammation
Triazolopyridine
Collagen induced arthritis
Rheumatoid arthritis
Phosphoinositide 3-kinases (PI3Ks) represent a family of dual
specificity enzymes that, by acting as both lipid and protein
kinases, regulate numerous biological processes, including cell
growth, differentiation, survival, proliferation, migration, and
metabolism.1 These kinases are activated by a wide variety of dif-
ferent stimuli such as growth factors, inflammatory mediators, and
antigens. Deregulation of class I PI3Ks, is involved in the pathogen-
esis of various diseases, representing attractive targets for oncol-
ogy, inflammatory and cardiovascular diseases, with several
molecules being evaluated in early clinical trials in oncology and
inflammation.2
also play a role in immune function and therefore may have a syn-
ergistic anti-inflammatory effect,4 we were particularly interested
in achieving high selectivity over PI3Ka, a target with known anti-
proliferative effects,5 and hence more of interest in the cancer
area.6
Several PI3K inhibitors such as AS-605240,7 the first small-mol-
ecule known to inhibit PI3K
The suboptimal pharmacokinetics and the limited selectivity win-
dow of this inhibitor, particularly over the PI3K isoform as well as
other kinases such as DNAPK, severely limited its developability.
Therefore we,8 and others9 set out to primarily target PI3K
inhib-
itory activity, with selectivity particularly over PI3K as well as the
c, are known in the literature (Fig. 1).
a
c
In our efforts to target the PI3K family for the treatment of
inflammatory disease, we were interested in developing selective
a
rest of the kinome, with novel compounds possessing drug-like
ligands for the PI3Kc isoform. This PI3Kc subtype has been shown
properties that would be suitable for oral dosing in vivo.
to play a role in several immune functions such as granulocyte
migration, activation of mast cells, and dendritic cells, as well as
the development and differentiation of lymphocytes, thereby sug-
gesting that its inhibition might be beneficial in both inflammatory
and autoimmune conditions.3 Given the high sequence homology
within the PI3K family we were aware that achieving substantial
As part of our efforts to identify potent and selective PI3Kc
inhibitors, we recently described the 6-substituted triazolopyri-
dine 1 (Fig. 1) which showed efficacy in a number of inflammatory
models.10 The X-ray crystal structure of the analogous triazolopyri-
dine 210,11 (Fig. 2) showed that the scaffold occupies the ATP
binding pocket with the aminotriazole portion making the usual
selectivity (e.g., >100-fold) for PI3K
c
alone would be challenging.
donor/acceptor interaction with the Val882 of the PI3Kc protein
However, as literature evidence suggests that both PI3Kb and PI3Kd
backbone (hinge). Furthermore, the triazolopyridine scaffold and
the pyridine ring efficiently fill the hydrophobic pocket formed
by a number of residues that include Ile963 and Tyr867. The
methylsulfone and the acetamide groups extend towards the sol-
vent front. We envisioned that the regioisomeric 7-substituted
⇑
Corresponding author. Tel.: +44 179 953 2832; fax: +44 179 953 2801.
E-mail address: sunosem@hotmail.co.jp (M. Sunose).
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.05.090

4614
M. Sunose et al. / Bioorg. Med. Chem. Lett. 22 (2012) 4613–4618
Figure 1. AS-605240, 6-substituted triazolopyridine 1 and 7-substituted triazolopyridine 14.
Figure 2. X-ray cocrystal structure (PDB ID: 4AOF) of 2 bound to the ATP pocket of the PI3Kc kinase domain.
triazolopyridine scaffold should keep all the key interactions
shown by the 6-substituted series and hence embarked on explor-
ing the SAR in this scaffold (Fig. 3). This work culminated in the
discovery of 14 (CZC24758) an orally bioavailable, potent and
selective PI3K inhibitor which showed efficacy in a mouse collagen
induced arthritis (CIA) model.
The convergent route for the preparation of 7-substituted triaz-
olopyridines which involved coupling of boronic acids with the key
intermediate 6 met with little success12 (Scheme 1). A more robust
synthetic route for the preparation of these analogues involved
Suzuki coupling prior to the cyclisation to form the triazolopyri-
dines 9–21 (Scheme 2). The synthesis began by coupling commer-
cially available sulfonyl chlorides with amines to form
sulfonamides 23. We observed inconsistent yields (29% to 60%)
and significant impurities in the coupling when the HCl salt of
the sulfonyl chloride 22 was used. Using the free base, the sulfon-
amide intermediates 23 were isolated in almost quantitative yield
after simple precipitation from the reaction mixture. Boronic esters
were prepared in situ followed by subsequent Suzuki coupling
reaction which yielded the intermediates 24. Thiourea formation
and cyclisation with hydroxylamine yielded 9–21 in 79% yield with
high purity. This synthetic approach was developed to enable the
preparation of multigram quantities required for later-stage exper-
iments in vivo. Compound 9 was synthesized using commercially
available 3-bromo-5-(methylsulfonyl)pyridine. The 5-fluorinated
triazolopyridine 21 was synthesized in an analogous manner using
commercially available 2-amino-4-bromo-6-fluoropyridine.
The compounds 9–21 were initially tested using the Kinobeads
assay developed for lipid kinases10 to determine potency and selec-
tivity followed by their cellular potency in a PI3Kc/PI3Kd depen-
dent neutrophil migration assay (Table 1).10 SAR analysis was
carried out with a focussed set of compounds using the prior
knowledge from the 6-substituted series.10 As in the previous ser-
ies the 7-substituted series showed good potency for PI3Kc. The
methyl sulfonamide showed a small increase in potency compared
to the methyl sulfone (pIC50 9: 7.1 vs pIC50 11: 7.6), but larger sub-
stituents on the sulfonamide, although tolerated, failed to give a
significant increase in potency (pIC50: 7.5–7.9). The diethylsulfona-
mide 19 was the most potent compound identified (pIC50: 8.1)
although disappointingly it showed a lower potency in the cellular
assay (pIC50: 5.9). The majority of analogues gave a similar selec-
tivity profile across the PI3K family. As was observed in the
Figure 3. Cartoon representation showing hydrogen bonding interactions of 2 in
PI3Kc (A) and proposed binding of 7-substituted triazolopyridine scaffold (B).

M. Sunose et al. / Bioorg. Med. Chem. Lett. 22 (2012) 4613–4618
4615
Scheme 1. Reagents and conditions: (a) ethoxycarbonyl isothiocyanate, DCM, 35 °C; (b) NH2OHÁHCl, DIPEA, MeOH/EtOH, 90 °C; (c) bis(pinacolato)diboron, Pd(dppf)Cl2ÁDCM,
KOAc, dioxane, 120 °C, microwave; (d) Pd(dppf)Cl2ÁDCM, EtOH, 2 M Na2CO3, 140 °C, microwave.
Scheme 2. Reagents and conditions: (a) RNH2, pyridine, rt; (b) bis(pinacolato)diboron, Pd(dppf)Cl2ÁDCM, KOAc, dioxane, 120 °C, microwave; (c) 2-amino-4-bromopyridine or
2-amino-4-bromo-6-fluoropyridine, Pd(dppf)Cl2ÁDCM, EtOH, 2 M Na2CO3, 140 °C, microwave; (d) ethoxycarbonyl isothiocyanate, DCM, 35 °C; (e) NH2OHÁHCl, DIPEA, MeOH/
EtOH, 90 °C.
6-substituted series, we found that simple alkyl sulfonamides gave
the best balance of PI3K potency and cellular activity.
compounds would exhibit reasonable metabolic stability in vivo.
Indeed, they showed low clearance (12–22 mL/min/kg) when
dosed intravenously (i.v.) in rats and furthermore were orally bio-
available (56–81%) with good exposure. In a pharmacokinetic
study in a Cynomolgus monkey, 14 had an in vivo clearance of
1.6 mL/min/kg, a half life of 6.1 h and was 88% orally bioavailable.
Encouraged by the good cell potency and superior oral expo-
sure, we fully characterized 14 in other off-target activities and
broad selectivity across the kinome. In a panel of 158 kinases, 14
displayed greater than 100-fold selectivity against 155 kinases.
Within the PI3K family 14 showed some in vitro selectivity over
the other potential inflammatory targets PI3Kb and PI3Kd and
c
Substitution of the parent 6-substituted triazolopyridine core
with a fluorine at the C-8 position gave an increase in selectivity
for PI3Kc 10
As expected the fluoro analogue of 14, the 5-fluori-
.
nated triazolopyridine 21, showed improved selectivity, compara-
ble potency and selectivity profile to 6-substituted analogue 1.10
However, the 5-fluorinated compound 21 was not pursued further
due to instability in human/rat/mouse hepatocytes (2.5/7.5/
36.1 ml/min/g liver).
Having identified leads showing good in vitro PI3Kc inhibitory
activity and cellular potency, we carried out an evaluation of the
pharmacokinetic (PK) properties of selected compounds in rat.
Compounds 11 and 14 were chosen as they were two of the most
potent in the cellular assay. The in vitro metabolic stability of these
leads in rat liver microsomes (Table 2) predicted that the
excellent selectivity over the unwanted kinase PI3K
Further profiling showed that 14 was negative in Ames14, was
clean in a patch clamp hERG assay (>250 M) and devoid of CYP
enzymatic inhibitory activities (>20 M inhibition against CYP1A,
a
(Fig. 4).13
l
l

4616
M. Sunose et al. / Bioorg. Med. Chem. Lett. 22 (2012) 4613–4618
Table 1
PI3K family Kinobeads assay and cellular data for 9 to 21
Compound
R
PI3Kca (pIC50
)
PI3Kaa,b
(D
pIC50
)
PI3Kba,b
(D
pIC50
)
PI3Kda,b
(D
pIC50
)
NeuMiga (pIC50
)
9
Me
NH2
NHMe
NHPr
7.1
7.7
7.6
7.5
7.6
7.9
7.7
7.7
7.4
7.0
8.1
1.7
1.8
1.9
1.9
1.9
1.7
2.1
2.0
1.4
2.0
2.1
1.4
1.4
1.4
1.5
1.2
1.3
1.9
1.5
1.5
1.6
1.6
1.4
1.5
1.6
>1.7
1.5
1.3
2.1
1.5
1.6
1.7
1.9
10
11
12
13
14
15
16
17
18
19
6.1
6.3
6.2
6.3
6.3
5.9
5.6
5.0
NHiPr
NHtBu
NHCH2CF3
NHCMe2CF3
NHPh
NHBn
NEt2
5.9
20
21
7.6
7.8
1.9
2.4
1.3
1.5
1.5
2.1
5.7
6.1
N
a
The values are averages of at least two independent experiments.
Selectivity versus PI3Kc.
b
Table 2
Rat microsomal stability and PK properties of lead compounds
a
CLint
F (%)
CLobs (mL/min/kg)
Vdss (L/kg)
T1/2 (h)
Cmax
(lM)
AUC (lM h)
11b
14c
0.32
0.31
56
81
21.7
11.7
2.2
1.3
1.2
1.2
5.0
5.9
6.7
15.3
a
b
c
Rat microsomal clearance (ml/min/g liver).
Wistar rats male; 5 mg/kg p.o. (5% DMSO/5% Cremophor/0.5% carboxymethylcellulose); 1 mg/kg i.v. (5% DMSO/5% Cremophor).
Wistar rats male; 5 mg/kg p.o. (5% DMSO); 1 mg/kg i.v. (5% DMSO).
Figure 4. Kinase selectivity profiling of compound 14 showing 10-fold and 100-fold selectivity.
2D6, 2C9, 2C19 and 3A4 (midazolam and testosterone as sub-
strates), thereby indicating that it has very low potential for
drug-drug interactions.
On the basis of its good on-target potency, off-target selectivity,
and PK profiles, 14 was chosen for further in vivo evaluation. The
efficacy of 14 was assessed in a mouse IL-8 air-pouch model to
evaluate its ability to reduce the recruitment of inflammatory cells
in vivo.3b The compound was administered orally 0.5 h before IL-8
was injected into the air pouch. After a further 4 h the pouch was
lavaged and the granulocytes counted. The IL-8-induced migration
of granulocytes into the air pouch was inhibited >70% after dosing
at 3 mg/kg and 10 mg/kg, while no significant inhibition was

M. Sunose et al. / Bioorg. Med. Chem. Lett. 22 (2012) 4613–4618
4617
inhibitors. Some members of this series have shown good pharma-
cokinetic profiles in rat and compound 14 (CZC24758) demon-
strated efficacy in a therapeutic CIA model after oral dosing,
thereby confirming the potential use of PI3K inhibitors in inflam-
matory disease.
Acknowledgments
The authors thank Tammy Ladduwahetty and Alan Watt for
invaluable advice on the manuscript and Warren Miller for chemo-
informatics support.
Supplementary data
Supplementary data associated with this article can be found,
in the online version, at http://dx.doi.org/10.1016/j.bmcl.2012.
05.090.
References and notes
Figure 5. Compound 14 in the IL-8 air-pouch mouse model: N = 4 for vehicle
control group, N = 10 for IL-8 control group, N = 5 for 1 mg/kg group, N = 8 for 3 mg/
kg group, N = 7 for 10 mg/kg group. 14 administered orally. Vehicle: 5% DMSO, 0.5%
carboxymethylcellulose. ⁄p 60.05, one-way ANOVA analysis.
1. (a) Cantley, L. C. Science 2002, 296, 1655; (b) Wymann, M. P.; Marone, R. Curr.
Opin. Cell Biol. 2005, 17, 141; (c) Hawkins, P. T.; Anderson, K. E.; Davidson, K.;
Stephens, L. R. Biochem. Soc. Trans. 2006, 34, 647.
2. (a) Marone, R.; Cmiljanovic, V.; Giese, B.; Wymann, M. P. Biochim. Biophys. Acta
2008, 1784, 159; (b) Hirsch, E.; Ciraolo, E.; Ghigo, A.; Costa, C. Pharmacol. Ther.
2008, 118, 192; (c) Folkes, A. J.; Ahmadi, K.; Alderton, W. K.; Alix, S.; Baker, S. J.;
Box, G.; Chuckowree, I. S.; Clarke, P. A.; Depledge, P.; Eccles, S. A.; Friedman, L.
S.; Hayes, A.; Hancox, T. C.; Kugendradas, A.; Lensun, L.; Moore, P.; Olivero, A.
G.; Pang, J.; Patel, S.; Pergl-Wilson, G. H.; Raynaud, F. I.; Robson, A.; Saghir, N.;
Salphati, L.; Sohal, S.; Ultsch, M. H.; Valenti, M.; Wallweber, H. J.; Wan, N. C.;
Wiesmann, C.; Workman, P.; Zhyvoloup, A.; Zvelebil, M. J.; Shuttleworth, S. J. J.
Med. Chem. 2008, 51, 5522; (d) Maira, S. M.; Stauffer, F.; Brueggen, J.; Furet, P.;
Schnell, C.; Fritsch, C.; Brachmann, S.; Chène, P.; De Pover, A.; Schoemaker, K.;
Fabbro, D.; Gabriel, D.; Simonen, M.; Murphy, L.; Finan, P.; Sellers, W.; Garcia-
Echeverria, C. Mol. Cancer Ther. 1851, 2008, 7.
3. (a) Sasaki, T.; Irie-Sasaki, J.; Jones, R. G.; Oliveira-dos-Santos, A. J.; Stanford, W.
L.; Bolon, B.; Wakeham, A.; Itie, A.; Bouchard, D.; Kozieradzki, I.; Joza, N.; Mak,
T. W.; Ohashi, P. S.; Suzuki, A.; Penninger, J. M. Science 2000, 287, 1040; (b)
Hirsch, E.; Katanaev, V. L.; Garlanda, C.; Azzolino, O.; Pirola, L.; Silengo, L.;
Sozzani, S.; Mantovani, A.; Altruda, F.; Wymann, M. Science 2000, 287, 1049; (c)
Li, Z.; Jiang, H.; Xie, W.; Zhang, Z.; Smrcka, A. V.; Wu, D. Science 2000, 287, 1046;
(d) Condliffe, A. M.; Davidson, K.; Anderson, K. E.; Ellson, C. D.; Crabbe, T.;
Okkenhaug, K.; Vanhaesebroeck, B.; Turner, M.; Webb, L.; Wymann, M. P.;
Hirsch, E.; Rückle, T.; Camps, M.; Rommel, C.; Jackson, S. P.; Chilvers, E. R.;
Stephens, L. R.; Hawkins, P. T. Blood 2005, 106, 1432; (e) Laffargue, M.; Calvez,
R.; Finan, P.; Trifilieff, A.; Barbier, M.; Altruda, F.; Hirsch, E.; Wymann, M. P.
Immunity 2002, 16, 441; (f) Wymann, M. P.; Bjoerkloef, K.; Calvez, R.; Finan, P.;
Thomast, M.; Trifilieff, A.; Barbier, M.; Altruda, F.; Hirsch, E.; Laffargue, M.
Biochem. Soc. Trans. 2003, 31, 275.
4. (a) Rommel, C.; Camps, M.; Ji, H. Nat. Rev. Immunol. 2007, 7, 191; (b) Kulkarni,
S.; Sitaru, C.; Jakus, Z.; Anderson, K. E.; Damoulakis, G.; Davidson, K.; Hirose, M.;
Juss, J.; Oxley, D.; Chessa, T. A.; Ramadani, F.; Guillou, H.; Segonds-Pichon, A.;
Fritsch, A.; Jarvis, G. E.; Okkenhaug, K.; Ludwig, R.; Zillikens, D.; Mocsai, A.;
Vanhaesebroeck, B.; Stephens, L. R.; Hawkins, P. T. Sci. Signal 2011, 4, 23; (c)
Boyle, K. B.; Gyori, D.; Sindrilaru, A.; Scharffetter-Kochanek, K.; Taylor, P. R.;
Mócsai, A.; Stephens, L. R.; Hawkins, P. T. J. Immunol. 2011, 186, 2978; (d)
Guillermet-Guibert, J.; Bjorklof, K.; Salpekar, A.; Gonella, C.; Ramadani, F.;
Bilancio, A.; Meek, S.; Smith, A. J. H.; Okkenhaug, K.; Vanhaesebroeck, B. Proc.
Natl. Acad. Sci. U.S.A. 2008, 105, 8292.
Figure 6. CIA mouse model with 14: N = 10 mice for treatment and disease control
groups, N = 5 mice for Dexamethasone group, N = 4 mice for vehicle control group.
14 administered orally 10 mg/kg and 3 mg/kg BID. Vehicle: 5% DMSO, 0.5%
carboxymethylcellulose. ⁄p 60.05, Student’s t-test to vehicle.
5. (a) Liu, P.; Cheng, H.; Roberts, T. M.; Zhao, J. J. Nat. Rev. Drug Disc. 2009, 8, 627;
(b) Zhao, L.; Vogt, P. K. Oncogene. 2008, 27, 5486; (c) Ihle, N. T.; Powis, G. Curr.
Opin. Drug Discov. Devel. 2010, 13, 41; (d) Wee, S.; Lengauer, C.; Wiederschain,
D. Curr. Opin. Oncol. 2008, 20, 77; (e) Foukas, L. C.; Claret, M.; Pearce, W.;
Okkenhaug, K.; Meek, S.; Peskett, E.; Sancho, S.; Smith, A. J. H.; Withers, D. J.;
Vanhaesebroeck, B. Nature 2006, 441, 366.
observed at 1 mg/kg dose (Fig. 5). In the same model the 6-substi-
tuted-8-fluoro triazolopyridine 1 showed 29% inhibition at 3 mg/kg.
The efficacy of 14 was also evaluated in a mouse collagen-in-
duced arthritis (CIA) model, an autoimmune disease model which
shares pathogenic similarities to human rheumatoid arthritis.
Compound 14 was examined in a therapeutic model where treat-
ment started when each group of mice had reached a mean clinical
score of 1.5.15 Mice were scored in a blinded manner and dosed or-
ally twice daily for 14 days. Treatment with compound 14 signifi-
cantly inhibited the progression of disease compared to the vehicle
control at 3 and 10 mg/kg (Fig. 6).16 The Cmax following the final
6. Bowles, D. W.; Jimeno, A. Expert Opin. Investig. Drugs 2011, 20, 507.
7. (a) Camps, M.; Rückle, T.; Ji, H.; Ardissone, V.; Rintelen, F.; Shaw, J.; Ferrandi, C.;
Chabert, C.; Gillieron, C.; Francon, B.; Martin, T.; Gretener, D.; Perrin, D.; Leroy,
D.; Vitte, P. A.; Hirsch, E.; Wymann, M. P.; Cirillo, R.; Schwarz, M. K.; Rommel, C.
Nat. Med. 2005, 11, 936. Follow up program with improved selectivity is
published in:; (b) Pomel, V.; Klicic, J.; Covini, D. D.; Church, D.; Shaw, J. P.;
Roulin, K.; Burgat-Charvillon, F.; Valognes, D.; Camps, M.; Chabert, C.; Gillieron,
C.; Françon, B.; Perrin, D.; Leroy, D.; Gretener, D.; Nichols, A.; Vitte, P. A.;
Carboni, S.; Rommel, C.; Schwarz, M. K.; Rückle, T. J. Med. Chem. 2006, 49, 3857.
8. (a) Wilson, F.; Ramsden, N.; Bell, K.; Cansfield, A.; Burckhardt, S.; Taylor, J.;
Sunose, M.; Middlemiss, D. PCT Int. Appl. WO2008025821, 2008.; (b) Ramsden,
N.; Bell, K.; Cansfield,; Taylor, J.; Sunose, M.; Middlesmiss, D.; Neubauer, G. PCT
Int. Appl. WO2009068482, 2009.; (c) Ramsden, N.; Bell, K.; Cansfield, A.; Taylor,
J.; Sunose, M.; Middlesmiss, D. PCT Int. Appl. WO2010007099, 2010.; (d)
Ramsden, N.; Bell, K.; Taylor, J.; Sunose, M.; Middlemiss, D.; Ellard, K. PCT Int.
dose was 2.5
respectively.
lM and 10.2 lM for the 3 mg/kg and 10 mg/kg doses,
In summary, we have described the synthesis and SAR of a ser-
ies of 7-substituted triazolopyridines, which are potent PI3K
c

4618
M. Sunose et al. / Bioorg. Med. Chem. Lett. 22 (2012) 4613–4618
Appl. WO2010092015, 2010.; (e) Ramsden, N.; Bell, K.; Taylor, J.; Sunose, M.;
Middlemiss, D.; Ellard, K. PCT Int. Appl. WO2010057877, 2010.
14. Compound 14 was tested against TA1535, TA100, TA98 and TA102 strains S9
metabolic activation.
9. Ameriks, M. K.; Venable, J. D. Curr. Top. Med. Chem. 2009, 9, 738.
15. Arthritis was induced by immunizing male DBA/1OlaHsd mice with Bovine
Type II collagen in Freund’s complete adjuvant. Disease severity was evaluated
by scoring all four paws for each animal, with a maximum possible score being
5.0: No arthritis, 1:1 hind or fore paw joint affected or minimal diffuse
erythema and swelling, 2:2 hind or fore paw joints affected or mild diffuse
erythma and swelling, 3:3 hind or fore paw joints affected or moderate diffuse
erythma and swelling, 4: Marked diffuse erythma and swelling, or 4 digit joints
affected, 5: severe diffuse erythma and severe swelling entire paw, unable to
flex digits. 21–35 days later, onset of arthritis occurred and mice were
randomized by clinical scores into groups once the mean clinical arthritis
score had reached 1.5. Mice were orally dosed with compound 14 at 3 or
10 mg/kg BID for 14 days. Disease progression was significantly inhibited by 14
compared to vehicle control (P <0.05, by Student’s t-test) at 3 and 10 mg/kg.
16. The histopathological analysis of the joints resulted in a significant inhibition
of cartilage damage, bone resorption, pannus formation and joint inflammation
at both 3 and 10 mg/kg, 41% and 46% respectively.
10. Bergamini, G.; Bell, K.; Shimamura, S.; Werner, T.; Cansfield, A.; Müller, K.;
Perrin, J.; Rau, C.; Ellard, K.; Hopf, C.; Doce, C.; Leggate, D.; Mangano, R.;
Mathieson, T.; O’Mahony, A.; Plavec, I.; Rharbaoui, F.; Reinhard, F.; Savitski, M.
M.; Ramsden, N.; Hirsch, E.; Drewes, G.; Rausch, O.; Bantscheff, M.; Neubauer,
G. Nat. Chem. Biol. 2012, 8, 576. Kinobeads assay data (pIC50) for 1; PI3K
c
: 7.6;
PI3Ka: <5.0; PI3Kb: 6.0; PI3Kd: 5.1; Neutrophil migration assay data (pIC50):
6.0..
11. PDB ID code: 4AOF.
12. Synthesis of 6-substituted triazolopyridine series was highly successful using
the convergent route as described previously.5 However, in the 7-substituted
triazolopyridine series, the last Suzuki step was capricious with variable yields
despite screening numerous conditions. There was no improvement when
boronic ester of the triazolopyridine 6 was made in situ to carry out the Suzuki
reaction.
13. Compound 14 was tested against Cellzome off-target panel, which includes
>150 kinases, as well as other proteins. List of kinases/proteins screened are
shown in Supplementary data.