Influence of acid surrogates toward potency of VLA-4 antagonist

Article abstract of DOI:10.1016/j.bmcl.2005.06.034

A series of VLA-4 antagonist were synthesized wherein carboxylic acid was replaced by various acid surrogates. The effect of these acid surrogates toward potency was evaluated in a binding assay. A number of acid surrogates were potent antagonist of VLA-4

Full text of DOI:10.1016/j.bmcl.2005.06.034

Bioorganic & Medicinal Chemistry Letters 15 (2005) 4053–4056
Influence of acid surrogates toward potency of VLA-4 antagonist
Shankar Venkatraman,a,* Jongwon Lim,a Merryl Cramer,a Michael F. Gardner,a
Joyce James,a Kenneth Alves,b Russell B. Lingham,b
Richard A. Mumfordb and Benito Munoza
aDepartment of Chemistry, Merck Research Laboratories, 3535 General Atomics Court, San Diego, CA 92121, USA
bDepartment of Immunology, Merck Research Laboratories, Rahway, NJ 07065, USA
Received 26 April 2005; revised 2 June 2005; accepted 6 June 2005
Available online 20 July 2005
Abstract—A series of VLA-4 antagonist were synthesized wherein carboxylic acid was replaced by various acid surrogates. The effect
of these acid surrogates toward potency was evaluated in a binding assay. A number of acid surrogates were potent antagonist of
VLA-4, albeit significantly less potent than the corresponding carboxylic acid. Heterocyclic acid surrogate, oxadiazolidinone 3, dem-
onstrated an improved pharmacokinetic property when dosed intravenously.
Ó 2005 Elsevier Ltd. All rights reserved.
The adhesion molecule VLA-4 (very late activating anti-
gen-4, a4b1, CD49d/CD29) is a member of the integrin
family and is expressed on all circulating leukocytes ex-
cept platelets.1 VLA-4 is a receptor for vascular cell
adhesion molecule-1 (VCAM-1), which is expressed on
endothelial surface. This interaction is important for
the activation, migration, and proliferation of leuko-
cytes during normal and pathophysiological processes.2
It has been shown that anti-VLA-4 antibodies or VLA-4
antagonists3 inhibit leukocyte infiltration to extravascu-
lar tissue and prevent tissue damage in animal models of
asthma,4 multiple sclerosis (MS),5 rheumatoid arthritis
(RA), and inflammatory bowel disease (IBD).6 In addi-
tion, a humanized monoclonal anti-a4 antibody (Natal-
izumab) has shown efficacy for MS and CrohnÕs disease
in clinical trials.7 Hence, there is a significant interest in
the development of small molecules VLA-4 antagonist
for the treatment of these diseases.
albumin. There are several examples in the literature
from other targets, wherein acid surrogates have provid-
ed inhibitors with similar potency with improved PK
properties.9 In our laboratory, we undertook efforts to
evaluate the effect of acid surrogates toward potency
of VLA-4 antagonists.
The acid surrogates were chosen based on pKa properties
that were similar to carboxylic acids.10 Oxadiazolidinone
3 was synthesized from the corresponding dipeptide car-
boxylic acid 1 by converting the acid to nitrile 2 under
dehydrating conditions. The nitrile 2 was then allowed
to react with hydroxylamine and subsequent cyclization
with 2-ethylhexyl chloroformate to afford oxadiazolidi-
none 3 (Scheme 1). Tetrazole 4 was obtained by reaction
of the nitrile intermediate with an azide.
The 3-hydroxyisooxazole acid surrogate 7 was synthe-
sized from the corresponding dipeptide b-ketoester 6.11
The b-ketoester 6 was cyclized with hydroxyl amine
which upon treatment with concentrated hydrochloric
acid yielded compound 7 (Scheme 2). The b-ketoester
was also condensed with hydrazine in ethanol to yield
3-pyrrazol 8. In addition, N-trifluoroacetamide 9 and
N-trifluoromethanesulfonamide 10 were synthesized
by reaction of the corresponding amide with trifluoroace-
tic anhydride and trifluoromethanesulfonyl chloride,
respectively.
To date, the small molecule VLA-4 antagonists reported
in literature are primarily characterized by carboxylic
acids (Fig. 1).8 Removal of carboxylic acid moiety or
replacement by alcohol leads to complete loss of activi-
ty. Furthermore, these carboxylic acids are often charac-
terized by poor PK, high protein binding, and significant
shifts in potency in the presence of human serum
Keywords: Acid surrogates; VLA-4 antagonist.
*
The seldom used acid surrogate 1,2,4-oxadiazolidone-
2,5-dione was easily incorporated into a a-amino acid
Corresponding author. Tel.: +8582025779; fax:+8582025753; e-mail:
shankar_venkatraman@merck.com
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.06.034

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S. Venkatraman et al. / Bioorg. Med. Chem. Lett. 15 (2005) 4053–4056
Figure 1. VLA-4 antagonist.
Scheme 1.
Scheme 2.

S. Venkatraman et al. / Bioorg. Med. Chem. Lett. 15 (2005) 4053–4056
4055
Scheme 3.
Table 1. Potency and relative off-rate for acid surrogates as inhibitors
of VLA-4
11 to yield 16 as described in literature12 and is shown in
Scheme 3.
10
Compound
R
Calc. pKa
BURJ
(IC50nM
RORa
%
bound at 1 h
)
The acid surrogates were evaluated as inhibitors using
binding assay of the non-activated state of VLA-4 on
Jurkat cells (BURJ assay).13 A secondary assay measur-
ing the relative off-rate under the non-activating state at
1 h was carried out to further distinguish them (Table 1).
In general, acid surrogates 3, 6, 7, 9, and10 were found
to be potent, albeit significantly shifted compared to the
parent carboxylic acid 1. In the heterocyclic series, 1,2,4-
oxadiazolidinone 3 and 3-hydroxyisoxoazole 7 were
more potent than the most often used acid surrogate
tetrazole 4. In the acylic series, b-ketoester 6, trifluoro-
acetamide 9, and trifluoromethanesulfonamide 10 had
potency similar to 3 and 7 but significantly less potent
than 1. The inhibitory potency of 1,2,4-oxadiazoli-
done-3,5-dione 16 (BURJ, IC50 = 6 lM) was found to
be 3000-fold less than that of the corresponding carbox-
ylic acid (BURJ, IC50 = 2 nM).
1
4.0
0.03
12
66
3
4
6.5
5.3
8.6
4.6
6.2
8.1
0.35
0
0
60
6
13
10
8
7
20
8
300
10
6
When compared to the parent carboxylic acid 1, the acid
surrogates demonstrated poor affinity for the receptor as
evidenced by its relatively poor off-rate. The differences
between the volume, dipole moment and polarizability
of the carboxylic acid, and those of the acid surrogates
may account for the differences in the binding affinity.
9
0
10
11
0
a ROR refers to the amount of compound bound to the receptor and
measured as competition between binding ligand and the acid
surrogates.
The heterocyclic acid surrogates were further evaluated
for their pharmacokinetic properties in rats to determine
Table 2. PK of acid surrogates in rats
Compound
IV-AUC (lM h)
Vd (L/kg)
t1/2 (h)a
Cl (ml/min/kg)
% Fb
Cmax (nM)b
1
3
7
0.68
2.01
3.51
0.57
2.61
1.25
0.72
2.78
2.85
38.2
60
n.d.
0.19
n.d.
3.14
6.1
37.57
n.d.
a Data from intravenous (i.v.) studies.
b Data from postoperative (p.o.) studies. nd: compound not detected, below the level of limit of quantification (<300 pM). The compounds were
dissolved in 400 PEG/water (3:1) for i.v. and p.o. dosing.

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S. Venkatraman et al. / Bioorg. Med. Chem. Lett. 15 (2005) 4053–4056
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whether they offer any advantage over the parent acid 1
(Table 2). When dosed intravenously at 5 mpk, they
demonstrated an improved half-life and higher exposure
compared to acid 1. However, when dosed orally at
5 mpk, there was no significant improvement in oral bio-
availability or in exposure.
In summary, a series of carboxylic acid surrogates in
dipeptide series with moderate potency were identified,
as VLA-4 antagonists. The oxadiazolidinone acid surro-
gate 3 demonstrated an improved pharmacokinetic
property. These acid surrogates themselves represent
novel structures and will be further evaluated in animal
models of asthma and MS.
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