2′ Biaryl amides as novel and subtype selective M1 agonists. Part II: Further optimization and profiling

Article abstract of DOI:10.1016/j.bmcl.2010.04.127

Further optimization of the biaryl amide series via extensively exploring structure-activity relationships resulted in potent and subtype selective M ₁ agonists exemplified by compounds 9a and 9j with good rat PK properties including CNS penetration. Synthesis, structure-activity relationships, subtype selectivity for M₁ over M_2-5, and DMPK properties of these novel compounds are described.

Full text of DOI:10.1016/j.bmcl.2010.04.127

Bioorganic & Medicinal Chemistry Letters 20 (2010) 3545–3549
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
20 Biaryl amides as novel and subtype selective M1 agonists. Part II:
Further optimization and profiling
,
*
*
Brian Budzik , Vincenzo Garzya , Dongchuan Shi, Graham Walker, Yann Lauchart, Adam J. Lucas,
,à
*
Ralph A. Rivero, Christopher J. Langmead, Jeannette Watson, Zining Wu, Ian T. Forbes, Jian Jin
Discovery Research and Centers of Excellence for Drug Discovery, GlaxoSmithKline, 1250 South. Collegeville Road, Collegeville, PA 19426, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Further optimization of the biaryl amide series via extensively exploring structure–activity relationships
resulted in potent and subtype selective M1 agonists exemplified by compounds 9a and 9j with good rat
PK properties including CNS penetration. Synthesis, structure–activity relationships, subtype selectivity
for M1 over M2–5, and DMPK properties of these novel compounds are described.
Ó 2010 Elsevier Ltd. All rights reserved.
Received 20 March 2010
Revised 26 April 2010
Accepted 27 April 2010
Available online 17 May 2010
Keywords:
Subtype selective M1 muscarinic
acetylcholine receptor agonist
Subtype selective M1 agonist
CNS-penetrant and orally bioavailable M1
agonist
20 biaryl amides
Five muscarinic acetylcholine receptor (mAChR) subtypes, M1–
M5, are known to date.1–3 These seven-transmembrane (7TM)
receptors share a common orthosteric ligand-binding site with
an extremely high sequence homology, which explains why it
has been difficult historically to identify subtype selective ligands.3
Selective agonism of the M1 receptor has been suggested as a ther-
apeutic approach in dementia including Alzheimer’s disease and
age-associated memory impairment or cognitive impairment asso-
ciated with Schizophrenia.4 AC-42,5 and more recently TBPB and its
analogs,6 have been reported as M1 agonists which achieve sub-
type selectivity via binding to an allosteric site unique to M1.
We previously reported7 the identification, synthesis, and initial
structure–activity relationships (SAR) of biaryl amides derived
from 1 (Fig. 1) as novel and subtype selective M1 agonists.8,9 In
those studies, investigations of the lower aryl and diamine regions
of the template led to compounds of significantly improved M1
agonist potency (over 1) while maintaining up to 100-fold selectiv-
ity over M2–5. Herein we report further optimization and additional
SAR of this series, including exploration of the upper aryl and mid-
dle linker regions, and further exploration of the diamine region
that resulted in optimized compounds such as 9a and 9j possessing
excellent M1 agonist potency, intrinsic activity (IA) and subtype
selectivity for M1 over M2–5, as well as good rat PK properties
and CNS penetration.
Exploration of monosubstitution on the upper phenyl via the
solid phase route11 of Scheme 1, using the previously discovered
3-Cl phenyl moiety7 as the lower aryl, showed (Table 1) 4-substi-
tution (6e and 6f) and 5-substitution (6g and 6h) to be well toler-
ated.12,13 Although substitution at the 3 position led to large
activity erosion (6a–c), we were pleased to find that the introduc-
tion of a 6-fluoro group (6j14) resulted in a half-log potency (ꢀ5-
fold) increase over the reference compound7 with an unsubstituted
upper phenyl (last column). In general, agonist efficacy (intrinsic
activity) was improved along with agonist potency (pEC50).
NH
H
N
* Corresponding authors. Tel.: +1 610 754 9132 (B.B.); +1 44 1279 627488 (V.G.);
+1 919 843 8459; fax: +1 919 843 8465 (J.J.).
O
N
S
E-mail addresses: torqux@yahoo.com (B. Budzik), vincenzo.2.garzya@gsk.com
(V. Garzya), jianjin@unc.edu, jianjin19380@yahoo.com (J. Jin).
Cl
1, M1 FLIPR pEC50 = 6.6, IA = 84 %
2-5 FLIPR pIC50 < 5.0
Present address: 1031 Perkiomenville Road, Perkiomenville, PA 18074, USA.
à
Present address: Center for Integrative Chemical Biology
&
Drug Discovery,
M
Division of Medicinal Chemistry & Natural Products, Eshelman School of Pharmacy,
The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7363, USA.
Fig. 1. Structure and in vitro profile of 1.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.04.127

3546
B. Budzik et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3545–3549
Boc
Boc
NH
DMHB
DMHB
HN
a
b
N
DMHB
resin
N
N
R
O
N
N
NH
O
O
X
O
NH
Cl
Cl
H
R
N
c, d
6m, M1 pEC50 < 5.0
6n, M1 pEC50 < 5.0
Fig. 2. Bicyclic H-bond mimic compounds 6m and 6n.
Cl
Scheme 1. Solid phase exploration of upper aryl. Reagents and conditions: (a) 2,
6-dimethoxy-4-polystyrenebenzyloxybenzaldehyde (DMHB-resin10), 1-Boc-4-
(aminomethyl)piperidine, Na(OAc)3BH, DIEA, 10% AcOH in NMP, rt, 16 h; (b) 2-
haloaryl acids, DIC, NMP, rt, 16 h; (c) 3-Cl phenylboronic acid, Pd(PPh3)4, 2 M
Cs2CO3, DME, 80 °C, 16 h; (d) 50% TFA/DCE, rt, 1 h.
a
b, c
NBoc
N
N
H
N
7a
OH
Br
N
Br
O
O
N
OH
a, c
b, c
b
a
7b
7c
We wondered if the potency increase of 6-F 6j (Table 1) was due
to the stabilization of a favorable conformation via a hydrogen
bond between the 6-F and the amide hydrogen. To explore this
hypothesis, we synthesized 6m and 6n in which the ring structure
is conformationally-locked as in the H-bonded case above (Fig. 2).
The compounds proved inactive, therefore suggesting that the 6-F
increase in potency was likely the result of other interactions.
Heterocyclic replacements for phenyl in the upper aryl, includ-
ing all four possible pyridines, made via EDC amide formation and
Suzuki coupling, (7a–d, Scheme 2), a thiophene15 (7e), and a pyra-
zole15 (7f) were inactive (Table 2).
O
OH
OH
Cl
O
O
Cl
N
NBoc
NBoc
N
H
N
NH2
X
O
X = NH2
X = Br
d
b, c
a
H
N
7d
N
N
OH
We next examined the middle linker region (Table 3). A diverse
variety of groups were tried17 as alternatives to the original benz-
amide methylene linker (represented by 2d7 for comparison). Sev-
eral novel, structurally diverse linkers with varying M1 agonist
activities were discovered, including a hydroxyl and alkene (8g
and 8h), made via Aldol condensation of the appropriate ketone
with 1-Boc-4-formylpiperidine followed by hydrogenation of the
conjugated alkene, ketone reduction with NaBH4, and dehydration
of the benzylic alcohol (Scheme 3). Also discovered were various
amides18 (8l, 8m, 8o, and 8p), an 1,2,4-oxadiazole18 (8u), and a sul-
fone (8t), accessed via Mitsunobu reaction of the appropriate aryl
thiol and alcohol followed by m-CPBA oxidation and Suzuki cou-
pling (Scheme 4). However, all active compounds with novel link-
ers had invariably lost the desirable selectivity over M3 of the
original benzamide linker present in 2d. 1,2,4-Oxadiazole 8u had
the best selectivity of the group over M3 (eightfold), but had a low-
er potency compared to the corresponding amide analog (8v),
which contains the original benzamide methylene linker as in 2d.
Br
Br
O
O
Scheme 2. Syntheses of pyridyl compounds 7a–d16 Reagents and conditions: (a) 1-
Boc-4-(aminomethyl)piperidine, EDC, HOAt, CH2Cl2, rt, 16 h; (b) 3-Cl phenylboronic
acid, Pd(PPh3)4, 2 M Cs2CO3, DME, 80 °C, 4–16 h; (c) 4 M HCl, MeOH, rt, 16 h; (d) t-
Bu nitrite, CuBr2, CH3CN, 0 °C to rt, 16 h.
Table 2
Upper aryl heterocyclic SAR
NH
H
N
O
Cl
Compound
Structure
M1 pEC50 IA (%)
<5.0
18
N
7a
Table 1
Mono-substitution SAR of upper phenyl
N
<5.0
20
5
7b
7c
4
3
6
NH
H
N
<5.0
3
N
O
Cl
<5.0
0
Compound M1
pEC50 IA (%)
F
Cl
Me
6c
6f
H
N
7d
7e
3
4
5
6
6a
6d
6g
6j
5.3
45
6b
6e
6h
6k
<5.0
14
<5.0
34
7.2
86
S
<5.0
0
<5.0
24
7.0
79
6.8
77
7.3
78
7.0
74
6i
6.0
67
<5.0
0
N
N
7f
7.8
90
7.2
109
6l
<5.0
37

B. Budzik et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3545–3549
3547
Table 3
SAR of middle linker region
Table 3 (continued)
Compound
X=
Y=
Cl
Linker structure
M1 pEC50 IA (%)
M3 pIC50
X
NH
H
N
7.5
92
8v
F
5.9
Linker
O
N
Y
8w
8x
F
F
H
H
<5.0
<5.0
—
—
N
O
O
Cl
Y=
N
Compound
X=
H
Linker structure
M1 pEC50 IA (%)
M3 pIC50
5.4
N
H
N
7.2
86
2d7 (Ref.)
H
H
H
O
NH
H
N
5.5
52
8a
8b
H
H
—
—
a
b, c, d, e
O
O
O
O
5.1
43
Br
O
Cl
Cl
N
H
8f
O
f
g
8c
H
H
H
H
<5.0
<5.0
—
—
N
OH
N
8g
8h
8d
O
O
Scheme 3. Syntheses of 8f, 8g, and 8h. Reagents and conditions: (a) 3-Cl
phenylboronic acid, Pd(PPh3)4, 2 M Cs2CO3, DME, 80 °C, 16 h; (b) i—1-Boc-4-
formylpiperidine, LHMDS, THF, ꢂ78 °C to rt, 1 h; (c) AcOH dropwise to prior rxn
until slightly acidic; (d) 50 psi H2, PtO2, 15 min; (e) 4 M HCl, MeOH, rt, 16 h; (f)
NaBH4, EtOH, rt, 15 min; (g) p-TsOH, toluene, reflux 2 h, Dean–Stark trap (steps b, c,
d done consecutively on same reaction mixture).
8e
8f
H
H
H
H
<5.0
<5.0
—
—
O
O
9.0
100
8g
8h
H
H
H
H
8.7
7.9
OH
NH
6.0
55
a
b, c, d
S
NBoc
8i
8j
H
H
H
H
<5.0
<5.0
—
—
O
O
O
O
S
Br
H
N
Cl
8k
H
H
<5.0
—
8t
O
6.4
70
Scheme 4. Synthesis of 8t. Reagents and conditions: (a) 2-bromobenzenethiol,
N-Boc-4-piperidineethanol, DIAD, PPh3, THF, 0 °C to rt, 16 h; (b) m-CPBA, CH2Cl2, rt,
16 h; (c) 3-Cl phenylboronic acid, Pd(PPh3)4, 2 M Cs2CO3, DME, 80 °C, 16 h; (d) 4 M
HCl, MeOH, rt, 16 h.
8l
H
H
—
N
H
N
6.3
81
8m
8n
H
H
H
H
5.7
—
O
N
Introducing an alkene (9c) (Scheme 6) also gave outstanding
potency20 and selectivity while eliminating the chiral centers pres-
ent in 9a. The necessary diamine was accessed via a Henry reaction
to give nitroalkene, followed by successive reduction first to oxime
then to amine, followed by protection of amine as a phthalimide,
<5.0
O
O
6.8
74
8o
H
H
6.9
N
H
debenzylation with
a-chloroethyl chloroformate, reprotection as
O
7.3
70
Boc, and deprotection of phthalimide to finally give the needed
alkenyl diamine which was then coupled with biaryl acid to give
9c. An alternate alkene18 (9d) was less potent compared to 9c
and the corresponding saturated analog 9e. trans ethyl 9f18 was
substantially less potent than trans methyl 9a.
8p
8q
8r
H
H
H
H
H
H
7.8
—
N
H
N
<5.0
<5.0
S
O
O
O
3-Fluoro substitution18 in piperidine 9g or in fluoroalkene 9h
led to potency losses, but was reasonably tolerated at the 4 posi-
tion (9i21). Combining the modifications of 9a and 9i gave 9j with
outstanding potency and selectivity over M3. Synthesis of 9j
(Scheme 7) began with reprotection of 1-benzyl-3-methylpiperi-
din-4-one as benzoyl followed by Wittig reaction giving the termi-
nal alkene, which was converted to the epoxide via the
bromohydrin. Regioselective epoxide opening with HFꢁpyridine
gave a fluoro alcohol, in which benzoyl was reprotected as Boc.
The fluoro alcohol was transformed to the needed amine by Mits-
unobu reaction with phthalimide followed by cleavage with hydra-
zine. Coupling with the biaryl acid finally yielded 9j.
N
—
S
S
O
O
5.8
51
S
N
8s
F
H
—
O
7.2
71
8t
H
F
H
7.7
5.4
O
O
N
6.3
65
8u
Cl
O
N

3548
B. Budzik et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3545–3549
Table 4
Final generation SAR
Bn
Boc
e, f, g
a
h, i
N
N
9c
H2N
X
X
F
X = CH2NO2
(b) X = HC=N(OH)
(c) X = CH2NH2
H
N
R
O
(d) X = CH2NPhth
Scheme 6. Synthesis of 9c.22 Reagents and conditions: (a) 1-benzyl-3-methylpi-
peridin-4-one, CH3NO2, ethylenediamine (cat.), 100 °C, 3 h; (b) CS2, Et3N, CH3CN, rt,
1 h; (c) i—LiAlH4, THF, reflux 2 h; ii—Na2SO4ꢁ10H2O; (d) neat phthalic anhydride,
Cl
Compound
X=
H
R=
M1 pEC50 IA (%)
M3 pIC50
5.4
170 °C, 0.5 h; (e) i—a-chloroethyl chloroformate, K2CO3, DCE, reflux 1 h; ii—MeOH,
7.8
90
NH
reflux 1 h; (f) Boc2O, CH2Cl2, rt, 0.3 h; (g) NH2NH2ꢁH2O, EtOH, reflux, 2 h; (h) 30-
chloro-3-fluoro-1,10-biphenyl-2-carboxylic acid, EDC, HOAt, CH2Cl2, rt, 16 h; (i) 4 M
HCl, MeOH, rt.
6j (Ref.)
H
H
9.1
116
NH
NH
9a
9b
7.4
5.6
7.7
92
Bz
Bz
Y
d, e
a, b, c
f
N
N
N
N
X
H
8.5
91
NH
NH
NH
O
F
9c
9d
9e
9f
5.5
—
X = OH,
Y = Bz
Y = H
Y = Boc
(g) X = OH,
(h) X = OH,
Cl
Cl
H
6.8
72
(i) X = NPhtH, Y = Boc
Boc
j
7.6
80
k, l
—
9j
H2N
F
7.4
92
NH
5.0
Scheme 7. Synthesis of 9j.22 Reagents and conditions: (a) i—1-benzyl-3-methylpi-
peridin-4-one, -chloroethyl chloroformate, K2CO3, DCE, reflux 1 h; ii—MeOH,
F
F
a
H
6.8
68
NH
reflux 1 h; (b) PhCOCl, Et3N, CH2Cl2, 1 h, rt; (c) Ph3PMe+Brꢂ, n-BuLi, THF, ꢂ78 °C to
rt; (d) NBS, THF/H2O, 16 h, rt; (e) NaOH, IPA/H2O, rt, 15 min; (f) 70% HFꢁpyridine,
CH2Cl2, 0 °C, 15 min; (g) LiBH4, THF, MeOH, 70 °C, 2 h; (h) Boc2O, CH2Cl2, rt, 0.3 h; (i)
phthalimide, DIAD, PPh3, THF, 0 °C to rt; (j) NH2NH2ꢁH2O, EtOH, reflux, 2 h; (k) 30,4-
dichloro-3-fluoro-1,10-biphenyl-2-carboxylic acid, EDC, HOAt, DCM, rt, 16 h; (l) 4 M
HCl, MeOH, rt.
9g
9h
9i
5.0
5.0
5.9
Cl
H
6.8
74
NH
NH
7.2
87
F
F
9a and cis 9b, trans 9a proving to be the most potent compound
discovered in this series.
Cl
8.0
85
NH
9j
5.3
We then evaluated our most potent compounds in M2–5 selec-
tivity assays and were pleased to find that compounds 9a, 9c,
and 9j not only possessed excellent M1 agonist activity, but also
had maintained excellent subtype selectivity against M2–5 (Table 5).
In rat PK studies, subcutaneous administration of compound 9a
(10 mg/kg) resulted in good exposure in the brain
(AUC = 1655 ng h/g), half life (T1/2 = 2.3 h), and CNS penetration
(brain–blood ratio = 0.9). Oral administration of compound 9j
(3 mg/kg) gave good exposure in the brain (AUC = 2221 ng h/g),
half life (T1/2 = 3.0 h) and estimated oral bioavailability
(F = 57%),24 and moderate CNS penetration (brain–blood ra-
tio = 0.3) and estimated clearance (Cl = 35 mL/min/kg).24 In addi-
tion, compound 9a showed excellent general selectivity—inactive
against all targets in the CEREP selectivity panel except M1–4 recep-
tors. The combination of high potency and intrinsic activity, excel-
lent subtype and general selectivity, and good rat PK parameters
makes these novel compounds valuable tool compounds for
in vivo pharmacodynamic studies.
c
d, e
a, b
NBoc
NBoc
H2N
H2N
O
2
3
NH
NH
F
9a
9b
NH
H
N
f
O
Cl
4
Scheme 5. Synthesis of trans 9a and cis 9b.19,22 Reagents and conditions: (a) 3-Me-
4-carboxyl piperidine HCl, Boc2O, Et3N, CH2Cl2, rt; (b) 2 M NH3 in MeOH, EDC, HOAt,
CH2Cl2, rt, 16 h; (c) i—LiAlH4, ꢂ78 °C to rt, THF; ii—Na2SO4ꢁ10H2O; (d) 30-chloro-3-
fluoro-1,10-biphenyl-2-carboxylic acid, EDC, HOAt, CH2Cl2, rt, 16 h; (e) 4 M HCl,
MeOH (f) separation, CN column.
Table 5
Subtype selectivity profiles of select compounds23
Compound M1 pEC50 IA (%) M2 pIC50 M3 pIC50 M4 pIC50 M5 pIC50
For our final series of compounds (Table 4) we focused on care-
ful modifications to the diamine region, while fixing the biaryl re-
gion with optimal substitution of 6-F on upper phenyl and 3-Cl on
lower phenyl. In our previous report,7 introduction of a 3-methyl
substituent on the piperidine ring led to a half-log potency (five-
fold) increase. Combining this modification with our optimized
biaryl (Scheme 5, full experimentals given in Note 19) gave trans
9a
9c
9j
9.1
116
6.7
5.9
5.6
7.4
5.5
5.3
6.8
5.1
5.5
6.3
5.8
5.2
8.5
91
8.0
85

B. Budzik et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3545–3549
3549
1-carboxylate followed by deprotection with 4 M HCl. 8e was made via EDC,
DMAP coupling of 30-chlorobiphenyl-2-carboxylic acid and N-Boc-4-
piperidinemethanol followed by deprotection with 4 M HCl.
18. Please see Supporting Information for synthetic schemes for the following
compounds whose syntheses are not shown herein: 8b–d, 8i–s, 8u, 8w, 8x, 9d,
9f, 9g, and 9h. All other compounds were made via routes given in general
schemes with commercially available materials.
In conclusion, further optimization of the biaryl amide series via
extensively exploring SAR resulted in very potent and selective M1
agonists with good rat PK properties and CNS penetration. These
novel compounds exemplified by 9a and 9j are excellent tools for
elucidating and validating potential therapeutic benefits resulting
from selective M1 agonism.
19. Experimental details of the synthesis of 9a and 9b in Scheme 5:
N-Boc-3-methylpiperidine-4-carboxamide 2 (steps a, b):
3-Me-4-carboxyl piperidine HCl (2.0 g, 11 mmol, 1.2 equiv), Boc anhydride
(2.0 g, 9.3 mol, 1 equiv), and Et3N (1.4 g, 14 mmol, 1.5 equiv), were combined
in DCM (100 mL) and stirred overnight. The reaction was washed with 1 N HCl
(50 mL), H2O (100 mL), dried Na2SO4, and evaporated to yield N-Boc-3-Me-4-
carboxylpiperidine, to which was added 2 M NH3 in MeOH (23 mL, 46 mmol,
5 equiv), EDC (1.8 g, 9.3 mmol, 1 equiv), HOAt (1.26 g, 9.3 mmol, 1 equiv), and
DCM (100 mL), then stirred overnight. The reaction was then washed with H2O
(2ꢃ 75 mL), dried Na2SO4, and evaporated to yield 2 (1.55 g, 69%) which was
used without further purification.
Acknowledgments
The authors thank Bing Wang for NMR, Qian Jin for LC/MS, and
Karl Erhard for separations support.
Supplementary data
N-Boc-3-methyl-4-methylaminopiperidine 3 (step c):
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.04.127.
2 (1.55 g, 6.4 mmol, 1 eq) was dissolved in THF (75 mL), cooled ꢂ78 °C, and 1 M
LiAlH4 in THF (16 mL, 16 mmol, 2.5 equiv) added. The reaction was allowed to
warm to room temperature overnight, then surrounded by ice bath and
Na2SO4ꢁ10H2O (ꢀ3 g) added portion wise. The slurry was filtered and solids
washed 3ꢃ THF. Combined filtrate was evaporated to give 3 (0.79 g, 54%)
which was used without further purification.
References and notes
30-Chloro-3-fluoro-N-[(3-methylpiperidin-4-yl)methyl]-1,10-biphenyl-2-
carboxamide 4 (steps d and e):
1. Caulfield, M. P.; Birdsall, N. J. M. Pharmacol. Rev. 1998, 50, 279.
2. Eglen, R. M. Prog. Med. Chem. 2005, 43, 105.
30-Chloro-3-fluoro-1,10-biphenyl-2-carboxylic acid (see Note 22) (64 mg,
0.26 mmol, 1 equiv), 3 (58 mg, 0.26 mmol, 1 equiv), EDC (50 mg, 0.26 mmol,
1 equiv), and HOAt (35 mg, 0.26 mmol, 1 equiv) were combined in DCM
(10 mL) and stirred overnight. The reaction mixture was then washed with H2O
(2ꢃ 7 mL), dried Na2SO4, and evaporated. The residue was deprotected with
excess 4 M HCl in MeOH, evaporated, then purified by HPLC to give 4 (56 mg,
ꢀ50:50 cis:trans via methyl doublet integration) which was freebased, then
separated as follows:
3. Hulme, E. C.; Birdsall, N. J. M.; Buckley, N. J. Ann. Rev. Pharmacol. Toxicol. 1990,
30, 633.
4. Fisher, A.; Pittel, Z.; Haring, R.; Bar-Ner, N.; Kligerspatz, M.; Natan, N.; Egozi, I.;
Sonego, H.; Marcovitch, I.; Brandeis, R. J. Mol. Neurosci. 2003, 20, 349.
5. (a) Spalding, T. A.; Trotter, C.; Skjaerbaek, N.; Messier, T. L.; Currier, E. A.;
Burstein, E. S.; Li, D.; Hacksell, U.; Brann, M. R. Mol. Pharm. 2002, 61, 1297; (b)
Langmead, C. J.; Fry, V. A. H.; Forbes, I. T.; Branch, C. L.; Christopoulos, A.; Wood,
M. D.; Hedron, H. Mol. Pharm. 2006, 69, 236.
6. (a) Jones, C. K.; Brady, A. E.; Davis, A. A.; Xiang, Z.; Bubser, M.; Tantawy, M. N.;
Kane, A.; Bridges, T. M.; Kennedy, J. P.; Bradley, S. R.; Peterson, T.; Baldwin, R.
M.; Kessler, R.; Deutch, A.; Levey, A. I.; Lindsley, C. W.; Conn, P. J. J. Neurosci.
2008, 28, 10422; (b) Bridges, T. M.; Brady, A. E.; Kennedy, J. P.; Daniels, R. N.;
Miller, N. R.; Kim, K.; Breininger, M. L.; Gnetry, P. R.; Brogan, J. T.; Jones, C. K.;
Conn, P. J.; Lindsley, C. W. Bioorg. Med. Chem. Lett. 2008, 18, 5439; (c) Miller, N.
R.; Daniels, R. N.; Bridges, T. M.; Brady, A. E.; Conn, P. J.; Lindsley, C. W. Bioorg.
Med. Chem. Lett. 2008, 18, 5443.
7. Budzik, B.; Garzya, V.; Shi, D.; Foley, J. J.; Rivero, R. A.; Langmead, C. L.; Watson,
J.; Wu, Z.; Forbes, I. T.; Jin, J. Bioorg. Med. Chem. Lett. 2010, 20, 3540.
8. Fluorometric imaging plate reader (FLIPR) assays were used to measure M1
agonist potency and intrinsic activity, and M2–5 subtype selectivity. For FLIPR
assay details, see: Budzik, B. W.; Cooper, D. G.; Forbes, I. F.; Jin, J.; Shi, D.; Smith,
P. W.; Walker, G. R. W.O. Patent 2007036711-A1, 2007; Chem. Abstr. 2007, 146,
401966.
9a and 9b:
14 mg of 4 per 0.5 mL mobile phase was used per injection on a Berger Cyano
column (6 m, 20 ꢃ 150 mm), mobile phase 50:50:0.1 hexane/EtOH/i-PrNH2,
15 mL/min, and 280 nm UV detection, collecting racemic trans 9a with baseline
resolution at a retention time of 6.2 min. Racemic cis 9b was collected at
6.6 min:
trans-30-Chloro-3-fluoro-N-[(3-methyl-4-piperidinyl) methyl]-2-biphenyl carbox-
amide (freebase) 9a:
1H NMR (600 MHz, CD3OD) d 7.43–7.38 (m, 2H), 7.32–7.27 (m, 3H), 7.12 (m,
2H), 3.43 (m, 1H), 2.77 (m, 3H), 2.28 (m, 1H), 2.06 (m, 1H), 1.09–0.96 (m, 3H),
0.79 (m, 1H), 0.77 (d, J = 6.6 Hz, 3H). MS (ES+) 361 [M+H]+.
cis-30-Chloro-3-fluoro-N-[(3-methyl-4-piperidinyl) methyl]-2-biphenyl carboxam-
ide (freebase) 9b:
1H NMR (600 MHz, CD3OD) d 7.32 (m, 1H), 7.29 (m, 1H), 7.23–7.17 (m, 3H),
7.03 (m, 2H), 2.90 (m, 2H), 2.72 (m, 1H), 2.49 (m, 2H), 2.28 (m, 1H), 1.46 (m,
1H), 1.27 (m, 1H), 1.02 (m, 1H), 0.92 (m, 1H), 0.66 (d, J = 7.2 Hz, 3H).). MS (ES+)
361 [M+H]+.
9. The biological assay results in the paper are a mean of at least 2 determinations
with standard deviation of < 0.3. We report agonist potency in pEC50 defined as
the negative log of the EC50 value in molarity.
trans and cis were assigned via NOE studies.
10. (a) Jin, J.; Graybill, T. L.; Wang, M. A.; Davis, L. D.; Moore, M. L. J. Comb. Chem.
2001, 3, 97; (b) Available from Polymer Laboratories, part number: 1466-6689,
150–300 lm, 1.5 mmol/g loading.
11. Please see Ref. 7 for full experimental procedures exemplifying the use of the
solid phase route of Scheme 1.
20. Notably, an alkene (not shown) analogous to 9c, without 3-methyl on
piperidine and 6-F on upper phenyl, proved inactive.
21. 9i was made via EDC, HOAt coupling of 30-chloro-3-fluoro-1,10-biphenyl-2-
carboxylic acid (Note 22) and tert-butyl 4-(aminomethyl)-4-fluoro piperidine-
1-carboxylate followed by deprotection with 4 M HCl. For synthesis of the
above Boc 4-fluoropiperidine, see Barrow, J.; Lindsley, C.; Shipe, W.; Yang, Z.;
Wisnoski, D. Preparation of 4-fluoropiperidine derivatives as T-type calcium
antagonists. PCT Int. Appl. 2007, 89 pp. WO 2007002884 (page 25)
22. 30-Chloro-3-fluoro-1,10-biphenyl-2-carboxylic acid was easily made by Suzuki
coupling of methyl 2-bromo-6-fluorobenzoate (3-Cl phenylboronic acid,
Pd(PPh3)4, 2 M Cs2CO3, DME, 80 °C, 4 h) followed by hydrolysis (NaOH,
MeOH, reflux 1 h).
12. All new compounds in this paper were characterized via LC/MS and 1H NMR.
13. Many compounds made with disubstitution on the upper phenyl ring (not
shown in this paper) showed linearly additive SAR with potencies well
predictable from the trends shown for mono-substituted compounds in Table 1.
14. 30-Chloro-3-fluoro-N-(piperidin-4-ylmethyl)-1,10-bi phenyl-2-carboxamide tri-
fluoroacetate 6j: 1H NMR (600 MHz, CD3OD) d 8.60 (m, 1H), 7.54 (m, 1H),
7.51 (m, 1H), 7.46–7.39 (m, 3H), 7.25 (m, 2H), 3.32 (m, 2H), 3.13 (m, 2H), 2.85
(m, 2H), 1.66 (m, 1H), 1.59 (m, 2H), 1.24 (m, 2H). MS (ES+) 347 [M+H]+.
15. 7e and 7f both were made via the solid phase route of Scheme 1.
16. For method of step (d), see: Doyle, M.; Siegfried, B.; Dellaria, J. J. Org. Chem.
1977, 42, 2426.
23. Compounds were also tested in agonist mode for M2–5 and showed no agonist
activity.
24. Drug concentrations in hepatic porter vein and tailor vein at various time
points were measured and used to estimate in vivo clearance and oral
bioavailability of test compounds.
17. 8a was made via EDC, HOAt coupling of commercially available 30-
chlorobiphenyl-2-carboxylic acid and tert-butyl 4-(2-aminoethyl) piperidine-