Discovery of 3-piperidinyl-1-cyclopentanecarboxamide as a novel scaffold for highly potent CC chemokine receptor 2 antagonists

Article abstract of DOI:10.1021/jm070166b

Introduction of ring restrictions to a linear aminobutyramide CC chemokine receptor 2 (CCR2) antagonist lead (2) led to the discovery of a 1,3-disubstituted cyclopentane scaffold with enhanced hCCR2 receptor binding and antagonist activity. (1S,3R)-N-[3,5

Full text of DOI:10.1021/jm070166b

J. Med. Chem. 2007, 50, 2609-2611
2609
Discovery of
3-Piperidinyl-1-cyclopentanecarboxamide as a
Novel Scaffold for Highly Potent CC Chemokine
Receptor 2 Antagonists
Lihu Yang,* Gabor Butora, Richard X. Jiao, Alex Pasternak,
Changyou Zhou, William H. Parsons, Sander G. Mills,
Pasquale P. Vicario, Julia M. Ayala,
Margaret A. Cascieri, and Malcolm MacCoss
Figure 1. 4-Aminobutyramide based hCCR2 antagonists.
Merck Research Laboratories, Rahway, New Jersey 07065
ReceiVed February 13, 2007
given.9 Herein, we report our systematic and rational approach
that resulted in the identification of 3-amino-1-cyclopentane-
carboxamide as the optimal scaffold for CCR2 antagonism. The
rigid cyclopentane scaffold provided structural novelty and
increased potency against human and murine CCR2 over the
linear series.
Our approach to introduce structural rigidity was also partly
inspired by the success of the earlier CCR5 antagonist program
carried out at Merck, where a linear CCR5 antagonist lead
similar to 1 (both were prepared as part of the NK1 antagonist
effort) was transformed to a pyrrolidine scaffold with improved
potency and selectivity.10 Screening of the Merck CCR5
collection of more than 5000 compounds did not yield any
CCR2 hits, suggesting that the structural requirements for the
two receptors are quite different.
There are many ways of introducing ring restriction to 1,
especially if one considers potential attachment points on the
two benzene rings and the piperidine. Furthermore, additional
ring sizes are possible with 3-7 being the most used in
medicinal chemistry. If all of these factors are considered, there
are many structural possibilities with a large number being
synthetically unattractive. We took a rational, stepwise approach
to this problem by addressing substitution at the 2-position.
Initial compounds showed 2-substitution to be potency enhanc-
ing but not required for CCR2 antagonism activity (Figure 1).
This allowed us to work on the simple butyramide as the starting
point. After scaffold modification, the 2-substitution can be
reintroduced to further enhance potency. This allowed us to limit
our attachment points to the five backbone atoms (from the
benzylic position 2′ to the γ carbon). Second, we limited our
ring size to 5 or 6, which would cover diverse conformations
and made the problem much more manageable.
To introduce a five-membered ring between any two of the
five backbone atoms, there are eight possibilities as shown in
Figure 2 (2′-4 connection needs a minimum six-membered ring
and 1′-4 connection gives rise to an acylated aminal that is
generally considered unstable under mildly acidic conditions).
Although it is feasible to make these 8 five-membered ring and
10 six-membered ring compounds, we felt that we could further
triage them and set priorities for earlier success in finding an
active scaffold.
Abstract: Introduction of ring restrictions to a linear aminobutyramide
CC chemokine receptor 2 (CCR2) antagonist lead (2) led to the
discovery of a 1,3-disubstituted cyclopentane scaffold with enhanced
hCCR2 receptor binding and antagonist activity. (1S,3R)-N-[3,5-Bis-
(trifluoromethyl)benzyl]-1-methyl-3-[(1R,3′R)-methyl-1′H-spiro[indene-
1,4′-piperidin]-1′-yl]cyclopentanecarboxamide (16) had IC50 of 1.3 nM
(binding) and 0.45 nM (functional chemotaxis) against hCCR2. It also
showed activity against the mouse CCR2 receptor with an IC50 of 130
nM. Compound 16 is selective against other chemokine receptors,
including CCR5 (∼500-fold).
Chemokines, or chemotactic cytokines, are a large family of
small structurally related proteins that play an important role
in leukocyte migration and activation. Chemokines mediate their
effect through binding to the specific cell-surface chemokine
receptors, which belong to the superfamily of seven-transmem-
brane spanning G-protein-coupled receptors.1-3 Monocyte
chemoattractant protein (MCP-1,a CCL2) belongs to the CC
chemokine family and binds to CC chemokine receptor 2
(CCR2), which is most abundantly expressed on monocytes.
Studies have suggested that small-molecule CCR2 antagonists
may provide potential therapies for a variety of diseases
including rheumatoid arthritis, multiple sclerosis, and athero-
sclerosis.4,5
We have previously reported the identification of a class of
submicromolar glycinamide-based CCR2 antagonists.6 Optimi-
zation of the backbone led to the identification of a novel class
of γ-aminobutyramide-based CCR2 receptor antagonists (1,
Figure 1, IC50 shown as ability to displace MCP-1 binding to
human CCR2b in Chinese hamster ovary cell line).7 Extensive
SAR studies on the phenylpiperidine identified that spiroindene-
piperidine and particularly methyl substitutions improve potency
(2).7 Further modifications revealed that the 2-aryl substitution
on the butyramide core can be eliminated with some loss of
affinity (3) or replaced by small alkyl groups with improved
potency (4).8 While the butyramide leads were quite potent and
orally bioavailable, the flexible backbone was an obvious target
for medicinal chemistry manipulation to further improve potency
and specificity. Furthermore, these acyclic leads lacked activity
against the murine CCR2 receptor, which is critical for proof
of concept studies in murine models. There has been one report
on the pharmacological characterization of a rodent active CCR2
antagonist (INCB3344), but no structural information was
We have previously reported that incorporation of methyl
groups in 1 at positions 2 and 4 of the butyramide backbone
led to decreases in potency while 3-substitution rendered the
compound inactive.7 Similarly, methylation of the amide
nitrogen afforded more than a 40-fold decrease in potency and
the introduction of methyl at the benzylic position of the
benzylamide produced an inactive compound (unpublished data).
On the basis of these results, we assumed that ring tethering at
* To whom correspondence should be addressed. Phone: 732-594-4660.
Fax: 732-594-3007. E-mail: lihu_yang@merck.com.
a Abbreviations: CCL2, chemokine (C-C motif) ligand 2; CCR2,
chemokine (C-C motif) receptor 2; hCCR2, human CCR2; mCCR2, mouse
CCR2; MCP-1, monocyte chemoattractant protein 1; NOE, nuclear Over-
hauser enhancement.
10.1021/jm070166b CCC: $37.00 © 2007 American Chemical Society
Published on Web 04/27/2007

2610 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 11
Letters
Table 1. Binding Affinity to Human Monocyte
compd
n
stereochemistry
IC50
8a
8b
10a
10b
10a-1
10a-2
10a-3
10a-4
1
2
1
2
1
1
1
1
racemic
racemic
inactive
inactive
65 nM
23% @ 1 µM
30% @ 1 µM
36 nM
mixture of four
mixture of four
cis-(1R,3S)
cis-(1S,3R)
trans-(1S,3S)
trans-(1R,3R)
44% @ 1 µM
25% @ 1 µM
Table 2. Effect of a Methyl on the Binding Affinity to Human and
Mouse Monocyte
compd
R1
R2
R3
hCCR2 IC50 (nM)
mCCR2 IC50 (nM)
11
12
16
17
18
H
H
H
Me
H
Me
H
H
H
Me
H
7.5
1.4
1.3
>1000
47
inactive
27% @ 1 µM
130
inactive
27% @ 1µM
Me
Me
Me
H
Figure 2. Chemically feasible five-membered ring constraint.
Scheme 1. Preparation of 2-Substituted Lactamsa
monocytes or mouse CCR2 expressed in Chinese hamster ovary
cell line. Functional antagonism and efficacy of key compounds
were studied in a calcium flux or in chemotaxis assays, both
using human primary monocytes.11
Neither five- nor six-membered-ring lactams (8a, 8b) showed
any activity (Table 1). On the other hand, the 1,3-disubstituted
cyclopentane 10a had an IC50 of 65 nM on the hCCR2 receptor
as a mixture of four isomers. Interestingly, its homologue 10b
was totally inactive, indicating very stringent stereo requirement
for the backbone. Compound 10a was a mixture of four
diastereomers in unequal ratios that proved to be inseparable
on chiral HPLC columns in a single run. Fortunately, the two
sets of enatiomers (3:1) can first be separated by a Chiralcel
OD column (hexane/ethanol 96:4). NOE measurements indicated
that the major isomer had a 1,3-cis arrangement. Single
enatiomers were then obtained using a Chiralcel AD column
(hexane/ethanol 96:4). The absolute stereochemistry of active
cis isomers (10a-2) was later established to be 1S,3R by
resynthesis using known (S)-3-oxocyclopentane carboxylic acid
as the starting material. Binding results for the four isomers
are listed in Table 1. 10a-2 was the only active isomer, showing
36 nM potency in the human monocyte binding assay.
In the linear butyramide series, we had previously demon-
strated that replacing the phenylpiperidine with spiroindene-
piperidine and methylspiroindene-piperidine enhanced the bind-
ing affinity.7,8 Incorporation of this feature in the cyclopentane
series, according to Scheme 2, resulted in 11 and 12, respec-
tively, after chiral separation. Compound 11 (single active
isomer) appeared to be 4-fold more active than 10a-2, while
single isomer 12 was more than 20-fold more active (Table 2).
Compound 12 binds to the hCCR2 receptor with high affinity
(IC50 ) 1.4 nM), and functionally, it inhibited MCP-1 induced
calcium flux (IC50 ) 2.3 nM) and chemotaxis (IC50 ) 57 nM)
in human monocytes.
a Reagents and conditions: Boc2O, DMAP, CH2Cl2; (b) LHMDS, allyl
bromide, THF, -78 °C to room temp; (c) HCl/dioxane-ether; (d) NaH,
3,5-bis-CF3BnBr, THF/DMF; (e) O3, CH2Cl2, -78 °C; NaB(OAc)3H,
4-phenylpiperidine, CH2Cl2.
Scheme 2. Preparation of 1,3-Disubstituted Cyclocarboxamidea
a Reagents: (a) 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydro-
chloride (EDC), 3,5-bis-CF3BnNH2, CH2Cl2; (b) NaB(OAc)3H, 4-phen-
ylpiperidine, CH2Cl2.
the 3 and 2′ positions would probably not be tolerated.
Therefore, these assumptions leave structures A and D (and their
six-membered ring homologues) with the highest probability
of yielding active compounds. We also prepared C and E, which
turned out to be inactive as initially hypothesized. Structure F
had been previously prepared and found to be inactive.
Analogues in the A series were prepared according to Scheme
1. The available lactam 5 was N-Boc protected, then alkylated
and deprotected to give the allyl analogue 6. Subsequent
N-benzylation gave N-benzyl lactam 7. The double bond was
cleaved with ozone, and the crude ozonide was decomposed in
the presence of 4-phenylpiperidine and sodium triacetoxyboro-
hydride to effect direct reductive amination.
Analogues in the D series were prepared in two steps from
the available keto acids as described in Scheme 2. Coupling of
the keto acid with 3,5-bis(trifluoromethyl)benzylamine followed
by reductive amination with a substituted piperidine provided
the desired compounds as a mixture of four isomers.
Having established 1,3-disubstitutedcyclopentane as the op-
timal template, we turned our attention to further increasing
potency through substitution. To take advantage of the observed
potency enhancing effects of the 2-substitutents in the linear
system, there are two possible positions for substitution in the
cyclopentane template, as shown in Figure 3. The 1-substituted
system is similar to the unsubsituted in terms of stereochemistry,
The synthesized compounds were screened for their ability
to displace 125I labeled MCP-1 in isolated human primary

Letters
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 11 2611
In counterscreens, 16 also showed good selectivity versus
neurokinin receptors NK1 and NK2, and it has ∼500-fold
selectivity against CCR5 receptor (IC50 ) 630 nM, MIP-1R
binding), a marked improvement over the linear analogues. In
addition, 16 has no affinity for other chemokine receptors tested
(no significant binding at 1 µM on CCR1, CCR3, CCR4,
CXCR3, CXCR4, and CCR8). It has a modest oral bioavail-
ability in rats (F ) 15%), moderately high clearance (Clp ) 50
mL min-1 kg-1), a large volume of distribution (Vdss ) 6 L
kg-1), and a long half-life (t1/2 ) 8.1 h).
Figure 3. Possible substitution patterns.
Scheme 3. Synthesis of 1-Substituted Analoguesa
In summary, we have described SAR studies that have
resulted in the identification of 1,3-disubstituted cyclopentane
as a novel scaffold for the preparation of potent and selective
CCR2 antagonist. This lead showed modest mCCR2 activity
and served as a starting point for all of our subsequent efforts
at finding potent and selective CCR2 antagonists.
a Reagents and conditions: (a) 2-[(trimethylsilyl)methyl]-2-propen-1-yl
acetate, Pd(OAc)2, (iPrO)3P, THF, reflux; (b) aqueous LiOH, dioxane, water,
80 °C; (c) EDC, 3,5-bis-CF3BnNH2; (d) O3, CH2Cl2/MeOH, -78 °C; (e)
NaB(OAc)3H, 4-phenylpiperidine, CH2Cl2; (f) chiral column HPLC separa-
tion.
Acknowledgment. The authors thank Dr. George Doss for
performing NOE studies on 10a and 16, and Drs. A. Pinkerton
and D. Huang for the synthesis of 8a and 8b.
while the 5-substituted system introduced an extra chiral center
with a possibility of eight isomers.
Supporting Information Available: Experimental details for
the synthesis of 16. This material is available free of charge via
the Internet at http://pubs.acs.org.
Both series were prepared using similar 2 + 3 cycloaddition
chemistry with the appropriate acrylate as the starting olefin.12
Only the synthesis of the 1-substituted analogue is shown in
Scheme 3. The introduction of 1-methyl began with a palladium-
catalyzed cycloaddition of 2-[(trimethylsilyl)methyl]-2-propen-
1-yl acetate to ethyl 1-methylacrylate 13 as shown. The volatile
ester 14 was used without any further purification and was
hydrolyzed and converted to the amide 15 as described in
Scheme 2. Ozonolysis and reductive amination in one pot, as
described in Scheme 1, afforded a mixture of four diastereo-
isomers, which were separated using chiral column HPLC as
described for 10a to yield the single isomer 16. Its relative
stereochemistry was established through NMR analysis and
comparisons with compound 10a-2, and the absolute stereo-
chemistry was confirmed by an alternative synthetic route
starting from (S)-3-oxocyclopentanecarboxylic acid. Similarly,
the 5-methyl analogue 17 was prepared starting from crotonic
acid derivatives.
As in the series described earlier, only one (16) of the four
isolated isomers was active. Although the introduction of
1-methyl did not significantly increase the hCCR2 binding
affinity (16, IC50 ) 1.3 nM) over 12, it showed substantial
improvement in the functional chemotaxis assay (0.45 nM).
Such discrepancies between the binding and functional assay
may be due to the binding assay having bottomed out in the
low nanomolar range, thus making the functional chemotaxis
assay a better predictor with this class of hCCR2 antagonists.
It had been a project goal to discover compounds with murine
CCR2 receptor antagonism to validate the target in murine
models; however, our previous reported compounds were devoid
of mCCR2 activity. Therefore, the discovery that 16 binds to
the mCCR2 with an IC50 of 130 nM was very encouraging.
Table 2 illustrates the important role that a methyl group played
in CCR2 activity. The introduction of a methyl to the 5-position
totally eliminated CCR2 activity (all eight isomers were
prepared). Removal of the methyl group on the piperidine (18)
resulted in loss of human and mouse CCR2 activity. Later
publications from our group will detail the SAR that led to more
potent mCCR2 compounds for proof of concept studies in
mouse.
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