Cyclic thioether peptide mimetics as VCAM-VLA-4 antagonists

Article abstract of DOI:10.1016/S0960-894X(00)00175-X

Selective substitution of a sulfur atom by carbon in a highly potent 13-membered cyclic disulfide was accomplished by intramolecular displacement of a bromide. The potency of the resulting thioethers in the VCAM/VLA-4 assay was dependant on ring size and the position of the sulfur atom. (C) 2000 Elsevier Science Ltd. All rights reserved.

Full text of DOI:10.1016/S0960-894X(00)00175-X

Bioorganic & Medicinal Chemistry Letters 10 (2000) 1167±1169
Cyclic Thioether Peptide Mimetics as VCAM±VLA-4 Antagonists
Nader Fotouhi,* Pramod Joshi, Je€erson W. Tilley, Karen Rowan,
Virginia Schwinge and Barry Wolitzky
Roche Research Center, Ho€mann-La Roche Inc, Nultey, NJ 07110, USA
Received 17 November 1999; accepted 9 March 2000
AbstractÐSelective substitution of a sulfur atom by carbon in a highly potent 13-membered cyclic disul®de was accomplished by
intramolecular displacement of a bromide. The potency of the resulting thioethers in the VCAM/VLA-4 assay was dependant on
ring size and the position of the sulfur atom. # 2000 Elsevier Science Ltd. All rights reserved.
Vascular cell adhesion molecule-1 (VCAM-1), is
expressed on activated, but not resting, endothelium.
The principal receptor for VCAM-1, the integrin very
late antigen 4 (VLA-4, a4b1), is expressed on many
lymphocytes including circulating eosinophils, baso-
phils, and monocytes, but not neutrophils. Antibodies
to either protein are e€ective at inhibiting leukocyte
in®ltration and preventing tissue damage in animal
models of in¯ammation, including asthma and arthri-
tis.1,2 Peptides derived from the connecting segment 1
(CS1) sequence of ®bronectin have also been shown to
block the VCAM±VLA4 interaction and to block aller-
gen induced airway responses in a sheep model of
asthma.3,4 Thus, we were interested in discovering orally
active VCAM±VLA-4 antagonists that might be useful
for the treatment of asthma or rheumatoid arthritis.
ther with the N-terminal modi®cation of the cyclic core
would reduce the metabolic liabilities that may be asso-
ciated with a structure such as 1, while retaining the
desired biological activity. We also reasoned that since
the geometry of the disul®de may be di€erent from that
of a thioether, the overall ring size may also a€ect the
activity. Molecular modeling suggested that carbon
replacement would have minimal impact on the N- and
C-terminal vectors as well as the position of the cyclo-
pentyl moiety. We were thus encouraged to prepare the
thioether analogues 2b, to determine their e€ect on the
VCAM/VLA-4 interaction.
Results and Discussions
The target compounds were prepared by an intramole-
cular bromide displacement by a thiolate. Two of the
proposed analogues were prepared according to Scheme
1. The starting protected cysteine or homocysteine, was
coupled to the azido acid 3 under standard conditions
to give the corresponding azides 4a,b. Reduction with
trimethylphosphine a€orded the corresponding amines.
Protection of the amines and puri®cation were necessary
for the success of the next two steps. The Boc groups
were removed and the resulting amines were treated
with benzyloxy homoserine lactone 5 in pyridine at
80 ꢀC to a€ord the alcohols 6a,b. Conversion of the
alcohols to bromides, followed by deprotection of the
thiols and intramolecular displacement a€orded the
cyclic thioethers in relatively poor yield (12±15%) for
the three steps. The low yield is at least partially due to
the relatively poor conversion of the alcohols to the
bromides (around 50% yields), but is mainly due to an
inecient intramolecular cyclization. The yield of the
bromide displacement could not be improved even
In the accompanying paper,5 we described the design of
a class of spirocyclic disul®des, typi®ed by 1, which are
potent inhibitors of the VLA-4/VCAM interaction.
Although these molecules are relatively small (MW
<600), they are still somewhat peptidic in nature and
contain a disul®de bond which may result in potential
metabolic liabilities. The group at Genentech has
demonstrated in a related series that the N-terminal tyr-
osine may be replaced by a more drug like moiety with-
out any loss in potency.6 Indeed, replacement of tyrosine
in 1 with (Æ)-trans-N-acetyl-3-(4-hydroxyphenyl) proline
resulted in the potent antagonist 2a (manuscript in
prep.).
We hypothesized that replacement of one of the sulfur
atoms in the cyclic disul®de with a carbon atom, toge-
*Corresponding author. Tel.: +1-973-235-4660; fax: +1-973-235-
7122; e-mail: nader.fotouhi@roche.com
0960-894X/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(00)00175-X

1168
N. Fotouhi et al. / Bioorg. Med. Chem. Lett. 10 (2000) 1167±1169
The third analogue was prepared using a similar intra-
molecular cyclization, as indicated in Scheme 2. The
azido acid 3 was coupled with homoserine lactone under
standard conditions to give the lactone 10. The azide
was reduced with hydrogen over Pd/C and protected in
situ with Boc2O to facilitate isolation. Hydrogen bromide
gas was bubbled through an ethanolic solution of the
resulting lactone at 0 ꢀC, followed by warming to room
temperature, to give the aminobromide 11, in excellent
yield. The amine was coupled with Boc(t-butylthio)cys-
teine using standard DCC coupling to a€ord 12. The t-
butylthio protecting group was removed with triethyl-
phosphine, and the resulting bromothiol was cyclized
directly in the presence of DBU to give the core cyclic
derivative 13 in excellent yield (86%). Unlike the two
examples above, the intramolecular bromide displacement
is quite facile and occurs cleanly in 1 h, even at 10 mM
concentration. Removal of the Boc group, followed by
coupling with (Æ)-trans-N-acetyl-3-(4-hydroxyphenyl)
proline (8), and ®nal deprotection with LiOH a€orded the
cyclic thioether 14 (95% yield over three steps).
Figure 1.
under high dilution conditions. The carbobenzyloxy
group was then removed cleanly in re¯uxing TFA/
dimethylsul®de, and the cyclic thioether core was cou-
pled with (Æ)-trans-N-acetyl-3-(4-hydroxyphenyl)pro-
line 86 under standard conditions. Final deprotection
with LiOH a€orded 9a and 9b.
Compounds were assayed for VLA-4 antagonist activity
using a solid-phase, dual antibody ELISA in which
VLA-4 derived from Ramos cells was allowed to com-
pete for bound recombinant human VCAM in the pre-
sence of serial dilutions of test compound. VLA-4 bound
to VCAM-1 was detected by a complex of anti-b1 anti-
Scheme 1.
Scheme 2.

N. Fotouhi et al. / Bioorg. Med. Chem. Lett. 10 (2000) 1167±1169
Table 1. Activity of cyclic thioethers VCAM±VLA-4 assays
1169
in the ELISA assay. Although the di€erences are more
signi®cant in the cell-based assay, a similar trend is
observed. It is possible that the C-terminal sulfur atom
makes a key hydrophobic contact with the receptor, and
consequently, its replacement with a carbon atom
results in a suboptimal interaction. In conclusion, the
substitution of the disul®de with a thioether and replace-
ment of the N-terminal tyrosine was found to be well tol-
erated, resulting in a very potent relatively non-peptidic
small molecule antagonist of VLA-4.
Solid phase assay
IC50 (nM)
Ramos cell assay
No.
X
IC50 (nM)
References and Notes
2a
9a
9b
14
S-S
CH2S
CH,SCH2
SCH2
0.1
2.7
35
1.5
220
Ð
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0.5
20
body and HRP-conjugated anti-mouse IgG: chromo-
genic substrate (K-Blue).7 A secondary, cell based assay
was also run in which ¯uorescently labeled Ramos cells
were allowed to compete for immobilized VCAM. As
the data summarized in Table 1 indicate, increasing the
ring size by one atom had a detrimental e€ect on
potency. Similarly decreasing the ring size by one car-
bon resulted in a signi®cant loss in potency (unpub-
lished result). In contrast, replacement of a single sulfur
atom by carbon, while maintaining the ring size resulted
in only a relatively modest decrease in potency. More
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the e€ects of substitution of the N-terminal sulfur atom
and the C-terminal sulfur. In fact, the thioether 14 was
at least 5-fold more potent than its positional isomer 9a,
while only 4±5-fold less potent than the parent disul®de
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