Conformational studies of 3-amino-1-alkyl-cyclopentane carboxamide CCR2 antagonists leading to new spirocyclic antagonists

Article abstract of DOI:10.1016/j.bmcl.2008.01.016

In an effort to shed light on the active binding conformation of our 3-amino-1-alkyl-cyclopentane carboxamide CCR2 antagonists, we prepared several conformationally constrained analogs resulting from backbone cyclization. Evaluation of CCR2 binding affini

Full text of DOI:10.1016/j.bmcl.2008.01.016

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Bioorganic & Medicinal Chemistry Letters 18 (2008) 1374–1377
Conformational studies of 3-amino-1-alkyl-cyclopentane
carboxamide CCR2 antagonists leading
to new spirocyclic antagonists
Alexander Pasternak,* Stephen D. Goble, George A. Doss, Nancy N. Tsou, Gabor Butora,
Pasquale P. Vicario, Julia Marie Ayala, Mary Struthers, Julie A. DeMartino,
Sander G. Mills and Lihu Yang
Merck Research Laboratories, PO Box 2000, Rahway, NJ 07065-0900, USA
Received 17 November 2007; revised 30 December 2007; accepted 3 January 2008
Available online 9 January 2008
Abstract—In an effort to shed light on the active binding conformation of our 3-amino-1-alkyl-cyclopentane carboxamide CCR2
antagonists, we prepared several conformationally constrained analogs resulting from backbone cyclization. Evaluation of CCR2
binding affinities for these analogs gave insight into the optimal relative positions of the piperidine and benzylamide moieties while
simultaneously leading to the discovery of a new, potent lead type based upon a spirocyclic acetal scaffold.
ꢀ 2008 Elsevier Ltd. All rights reserved.
Chemokines are secreted by proinflammatory cells, leu-
kocytes, and endothelial cells in response to a stimulus
to the immune system. Monocyte chemoattractant pro-
tein-1 (MCP-1, CCL2), included within the CC class of
chemokines,1 mediates chemotaxis of monocytes to
inflammatory sites primarily through interactions with
its receptor, CCR2,2 a member of the G-protein-coupled
seven-transmembrane receptor superfamily. Numerous
recent studies have linked CCL2 and CCR2 to various
inflammatory diseases3 including rheumatoid arthritis
(RA)4 and atherosclerosis.5 Consequently, the therapeu-
tic potential of CCR2 antagonists in treating inflamma-
tory diseases has stimulated considerable interest.6 Our
group has recently disclosed the discovery and proper-
ties of a new class of CCR2 antagonists based upon a
3-amino-1-alkyl-cyclopentane carboxamide core typified
by 1.7 A study of molecular models revealed that the
bond connecting the cyclopentane to the amide in com-
pound 1 can assume two types of lowest energy confor-
mations which have similar energies: one in which the
carbonyl projects over the plane of the cyclopentane
ring and where the benzyl moiety is extended away,
and one in which the carbonyl is roughly parallel to
the cyclopentane, leaving the benzyl moiety above the
plane of the cyclopentane. This letter describes the de-
sign, synthesis, and biological activity of analogs of 1
where our 3-fluoro-5-trifluoromethylbenzyl pharmaco-
phore is locked into each of these two possible
orientations.
O
N
CF3
N
H
F
1
After studying models, we reasoned that backbone cycli-
zation from the a-position to the carbonyl to the amide
nitrogen (generating a spirocyclic core, 2) would lock the
3-fluoro-5-trifluoromethylbenzyl group into an extended
trajectory away from the cyclopentane subunit (Fig. 1).
Compound 2 as a mixture of two syn-diastereomers was
prepared according to Scheme 1. Commercially avail-
able methyl (3-methylenecyclopentane)carboxylate 3
was deprotonated with LDA (ꢀ78 ꢁC) and alkylated
with allyl bromide (ꢀ78 ꢁC to rt, 76%) to give 4.
Ozonolysis of both alkenes, followed by treatment with
PPh3, afforded ketoaldehyde 5 in 84% yield. Reductive
Keywords: CCR2; Chemokine; Antagonist.
*
Corresponding author. Tel.: +1 732 594 3847; fax: +1 732 594
3007; e-mail: Alexander_pasternak@merck.com
0960-894X/$ - see front matter ꢀ 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.01.016

A. Pasternak et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1374–1377
1375
F
CF3
CF3
O
HN
N
NH
N
O
F
CF3
F
F
CF3
O
N
N
O
N
O
2
Figure 1. Cyclization locks benzyl in the extended conformation.
8
Figure 2. Spiroacetal locks benzyl above the cyclopentane.
CO2Me LDA, THF;
CO2Me
allyl bromide
76%
3
4
fragment. In addition, the acetal ring oxygen can serve
as an isoelectronic substitute for the original carbonyl
oxygen (Fig. 2).
O3, DCM, -78 oC;
NaB(OAc)3H,
O
CO2Me
CHO
PPh3, -78 oC-rt
84%
DCM, 1h @ rt
CF3
H2N
The synthesis of the two epimeric acetal isomers of 8 was
accomplished as described in Schemes 2 and 3. Accord-
ing to Scheme 2, the commercially available ketoacid 9 is
fully protected by treatment with trimethylorthoformate
and catalytic TsOH in MeOH/DCM to give 10 (86%).
Alkylation as described previously afforded 11 in 88%
yield. Ozonolysis followed by treatment with sodium
borohydride gave the corresponding alcohol, which
upon warming in toluene cyclized to provide the lactone
12 (85%, two steps). Treatment with TFA to give ketone
13 was followed by reductive amination with piperidine
7, as described previously, to give aminolactone 14 as a
4:4:1:1 mixture of 4 diastereomers in 64% yield (ratio
determined by chiral analytical HPLC). All four isomers
were separated by chiral preparative HPLC using a Chi-
ralcel OD column (2 cm · 25 cm, purchased from Chiral
Technologies, Inc), using 7% ethanol/hexanes as the elu-
ent. One of the two major isomers (2nd peak to elute)
was converted to its HCl salt and crystals grown from
DCM/heptane were subjected to X-ray crystallographic
analysis (see Scheme 2 for 3D structure, R = 3.3%). The
resulting coordinates (deposited with the Cambridge
crystallographic Data Centre for small molecules:
670426) indicated that the cyclopentane stereocenters
had the opposite configuration to what we required.
From this we deduced that the other major isomer
(3rd peak to elute) should have the correct syn-
(1R,3S)-stereochemistry corresponding to our original
lead 1 since the reductive amination is selective for the
syn-isomers.
5
F
50%
O
NaB(OAc)3H, Et3N
CF3
O
N
4 Å sieves, DCM, 3 days
6
F
NH HCl
7
70%
O
N
a
CF3
N
f
Ha-HfNOE
F
2a
Scheme 1. Synthesis of syn-spirolactams 2a (mix two syn-
diastereomers).
amination with 1.0 equivalent of 3-fluoro-5-trifluorom-
ethylbenzylamine took place selectively at the aldehyde
and occurred with concomitant cyclization to the dihy-
dropyrolidinone 6 in 50% yield. A second reductive ami-
nation reaction with (1R,30R)-30-methylspiro[indene-
1,40-piperidine]8 over 3 days at rt afforded the target
analog 2a as a mixture of two syn-isomers. The stereo-
chemistry was determined by a ROESY 2D NMR
experiment which revealed an NOE between the methine
hydrogen Ha and the methylene hydrogens Hf (see
Scheme 1).
The homochiral lactone 14a was treated with DIBAH
(1 M in hexanes) at ꢀ78 ꢁC to afford a 3:2 mixture of
diastereomeric lactols 15 in 71% yield (Scheme 3). Alkyl-
ation with 3-fluoro-5-trifluoromethylbenzyl bromide
(NaH, DMF, 0.5 h, 0 ꢁC) gave acetal 8 as a 3:2 mixture
of diastereomers (81%). Preparative chiral HPLC sepa-
Toward preparing the alternative locked analog we envi-
sioned a spirocyclic acetal scaffold 8, which according to
models, positions the benzyl subunit above the
cyclopentyl and in closer proximity to the piperidine

1376
A. Pasternak et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1374–1377
MeO
MeO
O
CO2H TMOF, TsOH
CO2Me
O
DIBAH, THF, -78 o
C
MeOH/DCM
rt, 72 h, 86%
N
O
9
10
1 h, 71%
1) O3, MeOH, -78 oC;
LDA, THF;
MeO
14a
CO2Me
allyl bromide
88%
MeO
NaBH4
NaH, DMF, 0 oC, 0.5 h
2) toluene 60 oC, 0.5 h
85% (two steps)
OH
N
11
O
F
Br
O
O
O
TFA/DCM
81%
CF3
MeO
MeO
15
O
O
0.5 h @ rt
97%
F3C
F
12
13
NaB(OAc)3H, Et3N
O
oA sieves, DCM, 4 days
O
4
Preparative chiral HPLC
Chiralpak AD column
2% EtOH/hexanes
N
N
O
O
14
NH HCl
64%
4:4:1:1 ratio of isomers
8
7
F3C
F
F3C
F
Preparative chiral HPLC
Chiralcel OD column
7% EtOH/hexanes
NOE
N
Peaks 1 and 4:
H
O
O
H
O
N
O
O
O
N
H
N
O
O
H
NOE
8a, peak 2
8b, peak 1
5% from 13
5% from 13
Scheme 3. Completion of 8a and 8b.
O
O
O
N
N
O
Table 1 below. The extended analog 2a was inactive in
our binding assay. In contrast, both analogs 8a and 8b
were potent, with 8b in particular having a very high
binding affinity comparable to the original lead 1. More-
over, when 8b was evaluated in a functional assay mea-
suring inhibition of MCP1 mediated chemotaxis of
monocytes,9 it proved to be a potent functional antago-
nist with an IC50 of 2.5 nM. These data strongly support
the hypothesis that the active binding conformation of
CCR2 antagonists typified by 1 is one in which the ben-
zyl group is positioned above (and roughly perpendicu-
lar) to the cyclopentyl ring, forming a cup shaped
structure with the piperidine subunit in close proximity
to the benzyl subunit.
14a Peak 3, 20% from 13
14b Peak 2, 20% from 13
X-ray analysis of 14b HCl salt
Scheme 2. Synthesis of aminolactone 14a.
ration using a Chiralpak AD column (purchased from
Chiral Technologies, Inc., 2 cm · 25 cm), and eluting
with 2% ethanol/hexanes afforded the two homochiral
acetals 8a and 8b. The absolute configuration of the ace-
tal stereocenter in both 8a and 8b (shown in Scheme 3)
was determined by NOESY 2D NMR experiments after
all spectral assignments were obtained by a combination
of COSY, TOCSY, and HSQC 2D experiments to
assign all protons. The key NOE signals indicated in
Scheme 3 correlate with models to support the assigned
stereochemistry in each case.
In summary we have prepared two conformationally
constricted analogs of CCR2 antagonist 1 in an effort
to shed light on its active conformation. The first analog
prepared, 2b, in which the 3-fluoro-5-trifluoromethylb-
enzyl pharmacophore was projected away from the
Table 1. Binding affinities for original lead and new analogs
Compound
Human monocytes
a
IC50
1
1.4 nM
2a
8a
8b
18% at 1 lMb
115 nM
4.6 nM
The conformationally constrained analogs 2a, 8a, and
8b, along with our initial lead compound 1, were
evaluated in our CCR2 binding assay (inhibition of
125I-MCP-1 binding to endogenous receptor on human
monocyte whole cells).9 The results are listed in
a For details on this binding assay see Ref. 9.
b % Inhibition at 1 lM.

A. Pasternak et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1374–1377
1377
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cyclopentyl core, was inactive in our binding assay. The
second analog, based on a spiroacetal framework 8b,
and in which the 3-fluoro-5-trifluoromethylbenzyl phar-
macophore is positioned over and roughly perpendicu-
lar to the cyclopentyl core, proved to be a potent
CCR2 antagonist. This led us to infer that the active
conformation of 1 is analogous to 8b where the benzyl
group is over the cyclopentane core. In addition, use
of the spirocyclic acetal as a tool for studying the bind-
ing conformation of our CCR2 antagonists has led to
the discovery of a new structural class of CCR2 antago-
nist which shows potent binding and functional activity.
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Supplementary data
Supplementary data associated with this article can be
found, in the online version, at doi:10.1016/j.bmcl.
2008.01.016.
8. Pasternak, A.; Marino, D.; Vicario, P. P.; Ayala, J. M.;
Cascierri, M. A.; Parsons, W.; Mills, S. G.; MacCoss, M.;
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