Pyridone derivatives as potent and selective VLA-4 integrin antagonists

Article abstract of DOI:10.1016/j.bmcl.2006.01.025

A novel series of pyridone inhibitors has been identified through pharmacophore analysis, as potent antagonists of VLA-4.

Full text of DOI:10.1016/j.bmcl.2006.01.025

Bioorganic & Medicinal Chemistry Letters 16 (2006) 2256–2259
Pyridone derivatives as potent and selective VLA-4
integrin antagonists
Jason Witherington,a,* Vincent Bordas,a Alessandra Gaiba,a Phil M. Green,b
Antoinette Naylor,a Nigel Parr,c David G. Smith,a
Andrew K. Taklea and Robert W. Warda
aDepartment of Medicinal Chemistry, Neurology & GI Centre of Excellence for Drug Discovery,
GlaxoSmithKline Research Limited, New Frontiers Science Park, Third Avenue, Harlow, Essex, CM19 5AW, UK
bDepartment of Assay Development and Compound Profiling, Medicines Research Centre, GlaxoSmithKline. Stevenage. SG1 2NY, UK
cDepartment of Medicinal Chemistry, Respiratory and Inflammation Centre of Excellence for Drug Discovery,
Medicines Research Centre, GlaxoSmithKline. Stevenage. SG1 2NY, UK
Received 9 December 2005; revised 6 January 2006; accepted 6 January 2006
Available online 7 February 2006
Abstract—A novel series of pyridone inhibitors has been identified through pharmacophore analysis, as potent antagonists of
VLA-4.
Ó 2006 Elsevier Ltd. All rights reserved.
VLA-4 (Very Late Antigen-4, a4b1, or CD49d/CD29) is
a member of the superfamily of transmembrane glyco-
protein integrins which is made up of a- and b-heterodi-
mers and is expressed on all leucocytes except
platelets.1–3 VLA-4 binds to an alternatively spliced seg-
ment of fibronectin (CS-1) on extracellular matrix and
to vascular cell adhesion molecule-1 (VCAM-1) on
endothelium. VCAM-1 is expressed on vascular endo-
thelial cells in response to proinflammatory cytokines
and the VCAM-1/VLA-4 binding interaction has been
shown to be critical for lymphocyte migration to extra-
vascular tissues.4 Antibodies against VLA-4 have been
shown to block leucocytes infiltration and prevent tissue
damage in inflammatory disease models of asthma,5
rheumatoid arthritis,6 multiple sclerosis (MS)7 and
inflammatory bowel disease.8 Orally active small mole-
cule inhibitors of VLA-4 might therefore serve as useful
agents in the treatment of these disease states. Indeed, a
small molecule antagonist of VLA-4 has recently dem-
onstrated efficacy in an acute model of MS.9
moiety, and the position and length of the carboxylate
linker were essential for potency (data not shown) the
SAR around the C-ring attracted most interest.
Although direct comparisons could not be made, initial
SAR around the C-ring heterocycle appeared to suggest
that a nitrogen at the 2-position was beneficial for inhib-
itory potency (e.g., 1 and 2). Replacement of the nitro-
gen atom for oxygen also appeared to be detrimental
for potency (cf. 5 and 6).
Given the structural similarity of 1 and 5 with the
known11 and potent VLA-4 antagonists 7 and 8, togeth-
er with the finding that both series demonstrate similar
B-ring SAR (cf. 1 and 5 with 7 and 8), we decided to
overlay the two series in an attempt to define a common
pharmacophore (Fig. 1).
The molecular overlay of 5 and 8 may explain the simi-
larity in SAR between the two series, with excellent
alignment of both the urea and carboxylate moieties.
Furthermore, the nitrogen at the 2-position of the
C-ring of 5 is orientated in the same direction, but not
identical position, to the lone pair of the carbonyl
moiety of 8. Although the published pharmacophore
of 8 postulates the amide moiety acts as a hydrogen
bond donor,12 given the excellent overlay with 5, we
decided to explore the possibility that the amide might
in fact behave as a H-bond acceptor (Fig. 2).
Recently, we identified a series of C-ring heterocyclic
ureas as antagonists of VLA-4 (Table 1). While the urea
Keywords: VLA-4; Molecular overlays; Pharmacophore; Pyridone;
Ureas; GASP.
*
Corresponding author. Tel.: +44 1279 627832; fax: +44 1279
627685; e-mail: Jason_Witherington@GSK.com
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.01.025

J. Witherington et al. / Bioorg. Med. Chem. Lett. 16 (2006) 2256–2259
2257
Table 1. Heterocyclic C-ring SAR
Table 2. SAR for a series of pyridone analogues
R1
R
CO2H
H
H
H
H
N
N
N
N
O
D
2
A
B
3
O
R2
O
Y
N
X
C
Z
1
W
4
5
Compound
R1
R2
Ph–4-(CH2)2CO2H
pKi a4b1
Compound
W
X
Y
Z
R
pKi a4b110
9
10
11
12
13
14
15
H
<6
8.7
9.3
8.4
6.4
6.6
6.5
1
2
3
4
5
6
C
N
N
C
C
N
N
N
N
N
N
C
N
N
C
C
N
O
C
C
C
N
C
N
H
6.6
6.3
<4
<6
7.6
<6
H
CH2–Ph–4-(CH2)2CO2H
CH2–Ph–4-(CH2)2CO2H
CH2–Ph–3-(CH2)2CO2H
CH2–Ph–2-(CH2)2CO2H
CH2–Ph–4-CH2CO2H
CH2–Ph–4-CO2H
H
OMe
H
H
H
H
MeO
MeO
H
H
A small subset of compounds, with variations in linker
length and position, were designed and analysed for
molecular overlap with compound 8. Several of these
compounds were selected for synthesis based on align-
ment of the proposed pharmacophoric features (Fig. 3).
Table 3. Cross screening data for pyridone 10
Integrin
pKi
a4b1
a4b7
a5b1
aVb3
aVb6
aIIb3
8.7
<5
<5
<5
<5
<5
Disappointingly, despite the excellent overlay the direct-
ly linked pyridone analogue 9 was considerably less ac-
tive, possibly due to the rigid nature of this molecule and
its inability to achieve the required bioactive conforma-
tion (Table 2). Gratifyingly, however, analogue 10
R
H
H
CO2H
N
N
O
O
N
H
7, R = H, α4β1 pKi 7.2
8, R = OMe, 4 1 pKi 9.1
α β
Figure 1. (a) Amide antagonists 7 and 8. (b) GASP molecular overlay of 8 (yellow) and 5 (green).
R
R
H
H
H
H
CO2H
N
N
N
N
O
O
cyclisation
( )n
O
O
N
N
( )n
CO2H
H
Figure 2. Cyclisation of amides to interrogate a hydrogen bond acceptor hypothesis.
Figure 3. (a) Molecular overlay of 8 (yellow) and 9 (green). (b) Molecular overlay of 8 (yellow) and 10 (green).

2258
J. Witherington et al. / Bioorg. Med. Chem. Lett. 16 (2006) 2256–2259
CO2Me
Cl
O2N
O2N
O
18
c
a, b
N
NH
16
19
17
H
H
O2N
N
N
d-f
O
O
O
N
N
CO2Me
CO2R
20, R = Me
10, R = H
Scheme 1. Reagents: (a) mCPBA, THF, 25 °C (58%); (b) i—Ac2O, reflux; ii—HCl, reflux, (80%); (c) Cs2CO3, DMF, 25 °C (90%); (d) 10% Pd/C,
EtOH, H2 (1 atm), 25 °C (50%); (e) o-tolyl isocyanate, DCM, 25 °C (80%); (f) 0.5 N aq LiOH, THF, 25 °C (100%).
3. Elices, M. J. Curr. Opin. Anti-inflam. Immunomod. Inves-
tig. Drugs 1999, 1, 14.
4. (a) Gearing, A. J. H.; Newman, W. Immunol. Today 1993,
14, 506; (b) Helmer, M. E. Annu. Rev. Immunol. 1990, 8,
365.
5. (a) Lin, K.-C.; Ateeq, H. S.; Hsiung, S. H.; Chong, L. T.;
Zimmerman, C. N.; Castro, A.; Lee, W.-C.; Hammond,
C. E.; Kalkunte, S.; Chen, L.-L.; Pepinsky, R. B.; Leone,
D. R.; Sprague, A. G.; Abraham, W. M.; Gill, A.; Lobb,
R. A.; Adams, S. P. J. Med. Chem. 1999, 42, 920; (b) Lin,
K.-C.; Castro, A. C. Curr. Opin. Chem. Biol. 1998, 2, 453.
6. Seiffge, D. J. Rheumatol. 1996, 23, 2086.
displayed excellent inhibitory potency and introduction
of a methoxy group into the B-ring gave further
enhancement in potency (cf. 10 and 11). The increased
potency observed for the pyridone 10 relative to the
amide 7 is also noteworthy, suggesting that the carbonyl
of the pyridone may be held in its bioactive conforma-
tion. Although the meta isomer 12 displays comparable
potency to the para isomer 10, in general the position
and length of the linker is critical for optimal potency
(cf. 10 and 13, 14 and 15).
7. (a) Tubridy, N.; Behan, P. O.; Caplildeo, R.; Chaudhuri,
A.; Forbes, R.; Hawkins, C. P.; Hughes, R. A. C.; Palace,
J.; Sharrack, B.; Swingler, R.; Young, C.; Mosley, I. F.;
MacManus, D. G.; Donoghue, S.; Miller, D. H. Neurology
1999, 53, 466; (b) Keszthelyi, E.; Karlik, S.; Hyduk, S.;
Rice, A.; Gordon, G.; Yednock, T.; Horner, H. Neurology
1996, 47, 1053.
8. Podolsky, D. K.; Lobb, R.; King, N.; Benjamin, C. D.;
Pepinsky, B.; Sehgal, P.; deBaumont, M. J. Clin. Invest.
1993, 92, 972.
Cross screening of 10 against a range of integrin recep-
tors indicated excellent selectivity (Table 3). Additional-
ly, cross screening of 10 against a diverse panel of 50
receptors and ion channels13 indicated 615% inhibition
at 1 lM (data not shown).
The pyridone analogues were prepared via a short con-
cise synthesis14 starting with the oxidation of the known
pyridine 1615 with m-chloroperoxybenzoic acid which
afforded the corresponding pyridine N-oxide. Treatment
of the N-oxide with acetic anhydride at reflux afforded,
after hydrolysis, the corresponding pyridone 17. Selec-
tive N-alkylation of 17 with the chloride 1816 in the pres-
ence of caesium carbonate afforded the ester 19.
Hydrogenation of the nitro group afforded the corre-
sponding aniline, which was treated with o-tolyl isocya-
nate to afford the urea 20. Hydrolysis with lithium
hydroxide provided the desired analogue 10 in good
overall yield (Scheme 1).
9. Cannella, B.; Gaupp, S.; Tilton, R. G.; Raine, C. S.
J. Neurosci. Res. 2003, 71, 407.
10. A Jurkat J6 Scintillation Proximity Assay was used to
investigate the interaction of the integrin VLA-4 (Very
Late Antigen-4; a4b1; CD49d, CD29) expressed on the
Jurkat J6 cell membrane with test compounds. J6 cells (1
million cells/well) were allowed to coat wheat germ
agglutinin-coated SPA beads (Amersham, 1 mg/well) in
assay buffer containing 50 mM HEPES, 100 mM NaCl
and 1 mM MnCl2 (pH with 4 M NaOH to 7.5). Tritiated
3H Standard Compound A (1–3 nM final assay concen-
tration) and test compounds were dissolved in an appro-
priate solvent and diluted in assay buffer. Data are
presented as means of pKi. Standard compound A is
(2S)-3-[4-({[4-(aminocarbonyl)-1-piperidinyl]carbonyl}oxy)-
phenyl]-2-[((2S)-4-methyl-2-{[2-(2-methylphenoxy)ace-
tyl]amino}pentanoyl)amino] propanoic acid potassium
salt which is described in patent application WO 00/
37444. Compounds were assayed in duplicate, a four-
parameter curve fit being applied. The equilibrium disso-
ciation constant for each compound was calculated
according to the method of Cheng and Prusoff (Biochem.
Pharmacol. 1973, 22, 3099).
In summary, molecular overlay studies of our initial
leads with known VLA-4 antagonists have led to
hypothesis of a novel pharmacophore. Subsequent
design and synthesis of analogues exploiting this
hypothesis has afforded a novel series of potent and
selective VLA-4 antagonists.
References and notes
11. Astles, P. C.; Clark, D. E.; Collis, A. J.; Cox, P. J.;
Eastwood, P. J.; Harris, N. V.; Lai, J. Y. Q.; Morley, A.
D.; Porter, B. WO 992306.
1. Helmer, M. E.; Elices, M. J.; Parker, C.; Takada, Y.
Immunol. Rev. 1990, 114, 45.
2. Springer, T. A. Cell 1994, 76, 301.

J. Witherington et al. / Bioorg. Med. Chem. Lett. 16 (2006) 2256–2259
2259
12. Singh, J.; Vlihmen, H. V.; Liao, Y.; Lee, W.-C.; Corbenise,
M.; Harris, M.; Shu, I.-H.; Gill, A.; Cuervo, J. H.;
Abraham, W. M.; Adams, S. P. J. Med. Chem. 2002, 45,
2988.
14. (a) Witherington, J; Elliot, R. L. JP 2005255675.; (b)
Witherington, J; Elliot, R. L. WO 2004014859.
15. Ishikura, M.; Kamada, M.; Terashima, M. Heterocycles
1984, 22, 265.
ˆ
13. CEREP Poitiers, Le Bois l’Eveque, B.P.186 600 Celle
l’Evescault, FRANCE.
16. Fukuda, Y.; Chujo, I.; Fuchikami, T.; Hirosawa, C.;
Wakasa, N. WO 97/47581.