Imidazopyridines as VLA-4 integrin antagonists

Article abstract of DOI:10.1016/j.bmcl.2008.05.075

We describe a novel series of imidazopyridine substituted phenylalanines which are potent VLA-4 antagonists. A wide variety of substituents are tolerated as replacements for the pendant 3-pyridyl ring. A clear structure-activity relationship was identifie

Full text of DOI:10.1016/j.bmcl.2008.05.075

Bioorganic & Medicinal Chemistry Letters 18 (2008) 4146–4149
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Imidazopyridines as VLA-4 integrin antagonists
a,
David J. Phillips a, , Richard J. Davenport , Thierry A. Demaude b, Fiona P. Galleway a, Mark W. Jones a,
*
Laurent Knerr b,à, Benjamin G. Perry a, Andrew J. Ratcliffe a
a UCB, Granta Park, Great Abington, Cambridge CB21 6GS, UK
b UCB S.A., R&D, Chemin du Foriest, B-1420 Braine-l’Alleud, Belgium
a r t i c l e i n f o
a b s t r a c t
Article history:
We describe a novel series of imidazopyridine substituted phenylalanines which are potent VLA-4 antag-
onists. A wide variety of substituents are tolerated as replacements for the pendant 3-pyridyl ring. A clear
structure–activity relationship was identified around the substitution of the 3-amino-cyclobut-2-enone
portion of the molecule.
Received 8 May 2008
Revised 20 May 2008
Accepted 20 May 2008
Available online 23 May 2008
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords:
VLA-4
Integrin
Imidazopyridine
Phenylalanine
The integrin
a
4b1 (VLA-4) is predominantly expressed on the cell
propionic acid ethyl ester hydrochloride 3 was heated in the pres-
ence of an appropriately substituted cyclobuta-1,3-dione7 4 to af-
ford the vinylogous amide. The nitro group was reduced to the
aniline 5 using palladium on carbon and hydrogen. This was then re-
acted with 2-chloro-3-nitropyridine or 4-chloro-3-nitropyridine 6,
displacing the labile chloride in refluxing ethanol to afford the nitro-
pyridine. In the case of compound 9g, aniline 5 was reacted with 2,5-
difluoronitrobenzene. Hydrogenation of the nitro group gave the
corresponding aromatic amine 7 which was then reacted with nico-
tinic acid using EDC and HOBT as coupling reagents to yield the
amide 8. Ring closure to form the imidazopyridine was achieved in
acetic acid, with heating in a microwave reactor. Introduction of
the halogen, when required, was carried out by reaction with N-bro-
mosuccinimide (NBS) or N-chlorosuccinimide (NCS) in ethyl acetate.
Hydrolysis in sodium hydroxide gave carboxylic acids 9a–g.
surface of mononuclear leukocytes and eosinophils. VLA-4 is a hete-
rodimeric adhesion molecule which binds to two receptors, fibro-
nectin and vascular cell adhesion molecule 1 (VCAM-1) expressed
on endothelial cells.1 This binding interaction occurs in response to
inflammatory cytokines and enables rolling, adhesion and transmi-
gration of the cell through the endothelium to the site of inflamma-
tion.2 Natalizumab (TysabriÒ), is a therapeutic monoclonal IgG
antibody currently on the market which has validated VLA-4 antag-
onism as a therapeutic approach for the treatment of inflammatory
diseases such as multiple sclerosis3 and Crohn’s disease.4
Research into small molecule VLA-4 antagonists has been an ac-
tive area in the pharmaceutical industry for over 10 years.5 As a
part of our ongoing research programme we have previously re-
ported that compound 1 is a potent VLA-4 antagonist.6 We have
also shown that the cyclobutene 1,2-dione substituent can be re-
placed with functionalized 3-amino-cyclobut-2-en-1-ones as
shown by compound 2.7 Our search for a new chemical series led
to the discovery that replacement of the 4-pyridyl amide in com-
pound 2 with various imidazopyridines gives rise to VLA-4 antag-
onists of comparable potency.
Cl
Cl
N
N
O
O
Cl
HN
Cl
HN
Br
O
O
O
The compounds were readily prepared according to the methods
outlined in Scheme 1. In method A, (S)-2-amino-3-(4-nitro-phenyl)-
HO2C
N
HO2C
N
H
H
N
H
2
1
* Corresponding author. Tel.: +44 1223 896535; fax: +44 1223 896400.
E-mail
addresses:
david.phillips@ucb-group.com,
djp1979@yahoo.co.uk
(D.J. Phillips).
In method B, (S)-3-(4-amino-phenyl)-2-tert-butoxycarbonyla-
mino-propionic acid ethyl ester 10 was reacted with 2-chloro-3-
nitropyridine to give the nitro compound 11. The BOC protecting
Present address: Takeda Cambridge, 418 Cambridge Science Park, Cambridge CB4
0PA, UK.
à
Present address: AstraZeneca R&D Mölndal, SE-43183 Mölndal, Sweden.
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.05.075

D. J. Phillips et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4146–4149
4147
Y
Y
O2N
H2N
O
X
6
H2N
4
R2
X
HN
O
R2
O2N
R1
i, ii
O
Cl
O
O
O
NH2.HCl
N
H
R1
R2
O
O
O
iii, iv
5
3
N
R1
H
Method A
O
7
Y
Y
X
O
X
N
N
N
H
vi, vii, viii
R3
v
HN
N
O
N
R2
O
R2
HO
N
R1
O
O
H
N
O
R1
9a-g
H
8
N
N
H2N
O2N
O2N
HN
HN
O
ii, iii
i
O
O
O
O
N
H
O
O
O
10
O
N
H
N
H
11
O
O
12
Method B
O
N
N
N
N
R4
N
H
N
N
iv, v
vi
vii or viii, ix
N
HN
R4
O
R4
O
O
R3
O
HO
O
N
N
H
N
H
H
14
O
O
O
13
15a-e
16a-w (R3 = Br)
Scheme 1. Reagents and conditions: Method A (i) 4, Et3N, EtOAc, 80 °C, 18 h, 87–100%; (ii) H2, Pd-C, EtOH, 100%; (iii) 6 or 2,5-difluoronitrobenzene, EtOH, DIPEA, reflux,
18–96 h, 16–100%; (iv) H2, 10% Pd-C, EtOH, 100%; (v) nicotinic acid, EDC, HOBT, DCM, 89%; (vi) AcOH, microwave, 125 °C, 10 min, 75–100%; (vii) NBS or NCS, EtOAc, 38–100%;
(viii) 2 M NaOH, THF, H2O, 6–54%. Method B (i) 2-chloro-3-nitropyridine, EtOH, DIPEA, reflux, 18 h, 47%; (ii) TFA, DCM, 94%; (iii) spiro[3.5]nonane-1,3-dione, EtOAc, 80 °C,
18 h, 100%; (iv) H2, Pd-C, EtOH, 100%; (v) R4COOH, EDC, HOBT, DCM, 21–100%; (vi) AcOH, microwave, 125 °C, 10 min, 32–98%; (vii) NBS or NCS, EtOAc, 39–100%; (viii) (iPrS)2
or (MeS)2, SO2Cl2, THF, 53–55%; (ix) 2 M NaOH, THF, H2O, 12–82%.
Table 1
group was removed to yield the amine which was reacted with
Human whole blood assay data (IC50
)
spiro[3.5]nonane-1,3-dione to furnish the vinylogous amide 12.
Reduction of the nitro group followed by reaction with a variety
of carboxylic acids gave the amide 13. Ring closure afforded the
imidazopyridine 14. Reaction with NBS or NCS when required fol-
lowed by hydrolysis gave access to compounds 15a–c and 16a–w.
Alternatively, treatment of compound 14 with a cooled premixed
solution of the appropriate alkyl disulfide and sulfuryl chloride fol-
lowed by hydrolysis furnished carboxylic acids 15d,e.
Compound
Method
R1 and R2
R3
X
Y
Human whole
blood8 IC50 (nM)
9a
9b
9c
9d
9e
9f
A
A
A
A
A
A
A
Me
Me
Me
H
N
N
N
CH
CH
CH
CH
CH
CH
CH
N
N
N
126
19
10
1616
504
236
2683
Cl
Br
H
Cl
Br
Br
Cyclohexyl
Cyclohexyl
Cyclohexyl
Cyclohexyl
9g
CF
Initially, we looked at the SAR around the 3-amino-cyclobut-2-
enone portion of the molecule. Substitution was investigated at the
spiro centre (R1 and R2) and at the double bond (R3). When gem di-
methyl groups were attached to the ring at the spiro centre we dis-
covered an interesting trend. Compound 9b (Table 1) with a
chlorine atom at the double bond was approximately 6-fold more
active in the whole blood assay8 compared to the unsubstituted
compound (9a). There was a further small increase with a bromine
atom as the substituent (9c). When a cyclohexyl ring was attached
at the spiro centre (15a) (Table 2) the whole blood activity was
approximately 5-fold higher than gem dimethyl (9a). We observed
Table 2
Human whole blood assay data (IC50
)
Compound
Method
R1 and R2
R3
R4
Human whole
blood8 IC50 (nM)
15a
15b
15c
15d
15e
B
B
B
B
B
Cyclohexyl
Cyclohexyl
Cyclohexyl
Cyclohexyl
Cyclohexyl
H
Cl
Br
S-i-Pr
SMe
3-Pyridyl
3-Pyridyl
3-Pyridyl
3-Pyridyl
3-Pyridyl
24
7
3
2
2

4148
D. J. Phillips et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4146–4149
Table 3
an increase in activity when the double bond was substituted as
Table 3 (continued)
Human whole blood assay data (IC50
)
Compound
R4
Human whole
blood8 IC50 (nM)
Compound
R4
Human whole
blood8 IC50 (nM)
~~~
N
~~~
N
16s
3
15c
3
Cl
~~~
16t
3
2
Me
~~~
16a
16b
570
95
~~~
16u
F3C
~~~
~~~
NH
16v
3
6
~~~
16c
16d
83
46
~~~
N
N
16w
N
H
~~~
N
N
~~~
Table 4
In vitro DMPK
16e
16f
16g
71
22
22
Compound
Microsomes CLint (
l
L/min/mg) @
CYP inhibition IC50 (lM)
CF
0.5
l
M
~~~3
Human
Rat
Mouse
2D6
1A2
3A4
2C9
~~~
15c
16o
16u
<1
4
6
48
38
52
11
14
15
>100
>100
>100
>100
>100
>100
62
79
10
10
18
8
N
N
16h
16i
16j
18
11
14
previously described with the gem dimethyl compounds. A bro-
mine atom at R3 (15c) was superior to chlorine (15b). The intro-
duction of S-i-Pr (15d) or SMe (15e) at R3 gave no significant
improvement over a bromine atom (15c).
We also investigated changing the position of the nitrogen atom
in the imidazopyridine ring (9d, 9e, 9f). This resulted in a marked
loss of activity, however, the trend in double bond substitution
continues across this series as described above. Removal of the
nitrogen altogether (9g) also resulted in a significant loss of activ-
ity. As the presence and position of the nitrogen is key for activity
we kept this fixed at position X in all further work.
In an attempt to find an alternative to the pyridine substituent
attached to the imidazopyridine ring, more analogues were syn-
thesized via Scheme 1, method B. This robust route allowed rapid
access to a wide range of compounds with various R4 substituents.
The data for these compounds are presented in Table 3 (R3 was
a bromine atom in all cases). The data show that a wide variety of
groups are tolerated at R4. We investigated substitution of the pyr-
idine substituent of 15c at the 6 position. These compounds (16c,
16g and 16p) were accessed by heating 16s and the appropriate
amine in a microwave using methanol or ethanol as the solvent.
Introduction of a 6-amino substituent was found to be detrimental
to activity. Substitution with NHMe (16p) reduced activity by 3-
fold compared to the unsubstituted pyridine (15c). Larger amino
substituents were not as well tolerated. For example, NMe2 (16g)
decreased activity by 2-fold compared to NHMe (16p). The larger
piperidine (16c) was 9-fold less potent than NHMe (16p).
Alkyl substituents exhibited a range of activities. None of these
brought any significant advantage over the pyridine of 15c. The
tert-butyl (16t) and trifluoromethyl cyclopropyl (16u) substituents
had comparable activities to 15c at 3 nM and 2 nM, respectively.
A simple phenyl substituent (16e) was not as effective as the
pyridine while 2 and 3-piperidines exhibited comparable activities
(16v and 16w, respectively).
~~~
~~~
~~~
H
N
16k
13
~~~
S
16l
11
10
~~~
~~~
16m
N
H
~~~
N
16n
16o
10
9
O
~~~
~~~
N
16p
9
N
H
~~~
N
16q
16r
8
8
F3C
~~~

D. J. Phillips et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4146–4149
4149
On the basis of excellent whole blood potency and chemical
References and notes
diversity we selected 15c, 16o and 16u for drug metabolism and
pharmacokinetics (DMPK) profiling (Table 4). Of these three R4
substituents the isopropyl group (16o) had a slight advantage in
terms of the cleanest cytochrome P450 (CYP) profile. Compound
16u moderately inhibited both CYP3A4 and CYP2C9. Overall, 16o
had the best in vitro DMPK profile and a human whole blood po-
tency of 9 nM.
Initial studies suggest that these compounds have an encourag-
ing in vitro DMPK profile. However, there is evidence that biliary
excretion is a characteristic of this compound class.9 We are cur-
rently investigating this to determine the relationship between
the in vitro DMPK and the in vivo clearance.
In conclusion, we have identified a novel and extremely po-
tent series of VLA-4 antagonist. We have identified a clear SAR
around the substitution of the 3-amino-cyclobut-2-enone portion
of the molecule. Cyclohexyl substitution at the spiro centre cou-
pled with bromine at the double bond was found to be optimal.
It is clear that the position of the nitrogen in the imidazopyri-
dine ring is important for activity and that removing it alto-
gether is not tolerated. We have investigated over 20
substituents at position R4 and found that many of these retain
significant activity.
1. (a) Elices, M. J. In Cell Adhesion Molecules and Matrix Proteins: Role in Health and
Diseases; Mousa, S. A., Ed.; Springer and R.G. Landes Co: Berlin, 1998; p 133; (b)
Elices, M. J. Curr. Opin. Anti-inflamm. Immunomod. Invest. Drugs 1999, 1, 14.
2. Kitayama, J.; Fuhlbridge, R. C.; Puri, K. D.; Springer, T. A. J. Immunol. 1997, 159,
3929.
3. Miller, D. H. et al N. Eng. J. Med. 2003, 348, 15.
4. Sandborn, W. J. et al N. Eng. J. Med. 2005, 353, 1912.
5. Davenport, R. J.; Munday, J. R. Drug Discov. Today 2007, 12, 569.
6. Porter, J. R.; Archibald, S. C.; Childs, K.; Critchley, D.; Head, J. C.; Linsley, J. M.;
Parton, T. A.; Robinson, M. K.; Shock, A.; Taylor, R. J.; Warrellow, G. J.; Alexander,
R. P.; Langham, B. Bioorg. Med. Chem. Lett. 2002, 12, 1051.
7. Brand, S.; de Candole, B. C.; Brown, J. A. Org. Lett. 2003, 5, 2343.
8. Compounds are diluted in TBS to 50 times desired start concentration.
Compounds are titrated across a NUNC immunosorp plate (50
to give tripling dilutions). Ninety-five microlitres of blood are incubated with
3 mM MnCl2 (Sigma—M-9522) and 1 L streptavidin-FITC (Pierce—21224), with
or without 10 g/mL biotinylated dimeric hVCAM-mFc in FACS tubes (Falcon—
lL into 100 lL
l
l
352052). Two microlitres diluted compound are spiked into relevant tube and
left for 30 min at room temperature on a shaking platform. Red blood cells are
lysed with 2 mL FACS lysing solution (BD Biosciences—349202), and the
remaining leukocytes are pelleted by centrifugation at 1200 rpm for 5 min.
Leukocytes are washed twice with 3 mL TBS before resuspension in 100 lL TBS
for analysis by flow cytometry. Values are the mean of at least two experiments.
9. (a) Porter, J. R.; Archibald, S. C.; Brown, J. A.; Childs, K.; Critchley, D.; Head, J. C.;
Parton, T. A.; Robinson, M. K.; Shock, A.; Taylor, R. J.; Warrellow, G. J. Bioorg. Med.
Chem. Lett. 2003, 13, 805; (b) Tang, W.; Stearns, R. A.; Chen, Q.; Bleasby, K.;
Teffera, Y.; Colletti, A.; Hafey, M.; Evers, R.; Dean, D. C.; Magriotis, P. A.; Lanza, T.
J.; Lin, L. S.; Hagmann, W. K.; Baillie, T. A. Xenobiotica 2008, 38, 223.