6-alkoxy-5-aryl-3-pyridinecarboxamides, a new series of bioavailable cannabinoid receptor type 1 (CB1) antagonists including peripherally selective compounds

Article

pubs.acs.org/jmc

6‑Alkoxy-5-aryl-3-pyridinecarboxamides, a New Series of

Bioavailable Cannabinoid Receptor Type 1 (CB1) Antagonists

Including Peripherally Selective Compounds

Stephan Rover,* Mirjana Andjelkovic, Agnes Benardeau, Evelyne Chaput, Wolfgang Guba, Paul Hebeisen,

̀

́

̈

Susanne Mohr, Matthias Nettekoven, Ulrike Obst, Wolfgang F. Richter, Christoph Ullmer,

Pius Waldmeier, and Matthew B. Wright

Pharma Research and Early Development, F. Hoﬀmann-La Roche AG, Grenzacherstrasse 124, Basel CH-4070, Switzerland

S

* Supporting Information

ABSTRACT: We identiﬁed 6-alkoxy-5-aryl-3-pyridinecarbox-

amides as potent CB1 receptor antagonists with high

selectivity over CB2 receptors. The series was optimized to

reduce lipophilicity compared to rimonabant to achieve

peripherally active molecules with minimal central eﬀects.

Several compounds that showed high plasma exposures in rats

were evaluated in vivo to probe the contribution of central vs

peripheral CB1 agonism to metabolic improvement. Both

rimonabant and 14g, a potent brain penetrant CB1 receptor

antagonist, signiﬁcantly reduced the rate of body weight gain.

However, 14h, a molecule with markedly reduced brain

exposure, had no signiﬁcant eﬀect on body weight. PK studies

conﬁrmed similarly high exposure of both 14h and 14g in the

periphery but 10-fold lower exposure in the brain for 14h. On the basis of these data, which are consistent with reported eﬀects

in tissue-speciﬁc CB1 receptor KO mice, we conclude that the metabolic beneﬁts of CB1 receptor antagonists are primarily

centrally mediated as originally believed.

INTRODUCTION

■

The CB1 receptor is part of the endocannabinoid system that

derives its name from cannabis. While the use of cannabis can be

traced back thousands of years,¹the molecular targets of its

pharmacology have been explored in detail only in the last few

decades. Starting with the isolation of the main psychoactive

constituent of Cannabis sativa, Δ9-THC by Mechoulam in

1964,²the next decades saw the isolation and characterization of

both the CB1³⁻⁵and CB2 receptors⁶as well as several of their

physiological ligands such as anandamide⁷and 2-AG.⁸The ﬁeld

Figure 1. CB1 receptor antagonists.

has matured, with over 400 new publications on the cannabinoid

system appearing in PubMed each year. Recent reviews have

indicated that this breadth of information has uncovered as many

questions as it has solved.⁹⁻¹²The CB1 receptor is abundantly

expressed in both peripheral and the neuronal tissues.¹³Target-

based drug discovery has progressed and delivered many

diﬀerent series of CB1-receptor antagonists.¹⁴These include

rimonabant (SR141716A, 1), which received marketing author-

ization in the European Union as an antiobesity medication but

was later withdrawn due to increased incidence of anxiety,

depression, and suicidality, and taranabant (MK-0364, 2, Figure

1) and otenabant (CP-945,598), which entered late clinical

development but were never launched. Although taranabant and

otenabant were derived from diﬀerent structural classes, similar

increased incidence of psychiatric side eﬀects led to termination

after or during phase III clinical studies.¹⁵⁻¹⁸

The observed psychiatric side eﬀects appear to preclude the

use of CB1 receptor antagonists for non-life-threatening diseases

or chronic treatment. However, it has become apparent that the

CB1 receptor has peripheral functions, suggesting that

peripherally active and selective agents may have therapeutic

potential with reduced psychiatric side eﬀects. Indeed, several

groups have reported series of peripherally active CB1 receptor

Received: July 16, 2013

A

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

Figure 2. Peripherally selective CB1 receptor antagonists.

Figure 3. Superposition of low energy conformers of 3 (orange) and 4 (yellow) (upper panel); structures and data for 3 and 4 (lower panel).

antagonists in the past few years,^19,20with the most common

strategy to derive these molecules being the addition of polar

groups onto known inhibitor backbones (Figure 2). This either

increased polar surface area or conferred eﬀective Pgp-mediated

transport leading to reduced CNS exposure.

Our group has pursued a relatively small chemotype that has

evolved into more polar and less brain penetrating molecules. We

originally explored a series of rimonabant bioisosteres that

evolved into compounds such as 3 (Figure 3).²¹Such substituted

pyrroles had good CB1 receptor aﬃnity and selectivity but

exhibited low metabolic stability especially in rat microsomes.

Metabolite proﬁling revealed multiple oxidative transformations;

basically all substituents on the pyrrole were eﬃciently oxidized

in rat microsomes.

Rather than trying to block all metabolically labile sites we

employed a scaﬀold hop, replacing the electron rich pyrrole with

B

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

a

Scheme 1. Synthesis of Compounds

a

Reagents and conditions: (a) Br₂, AcOH; (b) POCl₃, reﬂux; (c) MeOH, reﬂux; (d) POCl₃, quinoline reﬂux, then water RT; (f) arylboronic acid,

Pd(dppf)Cl₂·CH₂Cl₂, Na₂CO₃, toluene, water, 90 °C; (g) LiOH·H₂O, THF, water, reﬂux; (h) R²-OH, KOH, DMSO, MW, 100 °C; (i) arylboronic

acid, (Ph₃P)₄Pd, Na₂CO₃, DME, water, 85 °C; (j) various amide coupling methods.

a six-membered electron deﬁcient heterocycle. We hypothesized

that this strategy would eliminate the need for a methyl

substituent on the core and additionally reduce the propensity

for oxidation of the phenyl residue by rendering it less electron

rich. In addition, the reduction in overall lipophilicity should

decrease the susceptibility to oxidative attack by cytochrome

P450s. As an initial prototype we chose to make nicotinamide 4

because modeling suggested that a low energy conformer of 4

would superpose suﬃciently well onto the energy-minimized

structure of 3.

Compound 4 was still very lipophilic (ClogP of 5.9) with low

CB1 receptor aﬃnity (∼40-fold less potent compared to 3),

however it was 4 times more stable than 3 in rat liver microsomes.

Following these encouraging results, SAR exploration based on

compound 4 was initiated by varying the alkoxy and aryl residues.

Because subsequent molecules showed both improved aﬃnity

and metabolic stability, we also explored variations of the core

and amide residue. In the course of this work, we identiﬁed

several orally active 5-aryl-6-alkoxy-3-pyridinecarboxamides,

including peripherally selective compounds.

Miyaura reaction of 7 with arylboronic acids furnished the 5-

aryl substituted 3-pyridine carboxylic acid esters 8, which were

saponiﬁed to the acids 9. This was followed by a S_NAr reaction to

replace the 6-chloro substituent with alkoxides to give 6-alkoxy-

5-aryl-3-pyridine carboxylic acids 10. This was done on small

scale with potassium hydroxide as base in DMSO in a microwave

reactor. During up-scaling tests, we measured the thermal

stability of potassium hydroxide/DMSO mixtures in an

accelerating rate calorimeter and found exothermic decom-

position above 145 °C. Consequently, for larger batches, we

switched to a low temperature process that employed stirring of

the above potassium hydroxide/DMSO mixtures for extended

times between room temperature and 40 °C.²³Similar high

yields can be obtained by using this low-temperature variant of

the reaction. Conventional amide coupling methods converted

acids 10a−l to the corresponding amides 14, 15, and 16. This

ﬁrst synthetic variant allowed installing the alkoxide and amide in

the last two steps of the synthesis. Alternatively, the alkoxide

addition via the S_NAr reaction was done as ﬁrst step with the acid

11 to give 6-alkoxy derivatives 12. These, in turn, could be

converted in a second step to the amides 13 and in the last step by

Suzuki−Miyaura arylation to compounds 4, 14, 15, and 16. A

ﬁnal synthetic variant utilized the Suzuki−Miyaura arylation of 6-

alkoxy derivatives 12 to aﬀord an alternative route to 6-alkoxy-5-

aryl-3-pyridine carboxylic acids 10.

CHEMISTRY

■

We explored several variations of the synthesis of 6-alkoxy-5-aryl-

3-pyridinecarboxamides and ﬁnally selected the route depicted in

Scheme 1, which allowed either amide or aryl variations to be

performed in the ﬁnal synthetic step.

Some of the 6-alkoxy-5-aryl-3-pyridine carboxylic acids

(10m−q) were produced by an alternative synthesis that started

from 3-bromo-2-chloro-5-methyl-pyridine 22 (Scheme 2) using

a series of steps beginning with nucleophilic aromatic

substitution of 22 with alkoxides to furnish 23a−e, followed by

Suzuki−Miyaura arylations to give picolines 24a−e. These were

then transformed into the corresponding carboxylic acids 10m−

q by bromination, hydrolysis, and permanganate oxidation.

All variants of this synthesis started with commercially

available 6-hydroxy-nicotinic acid 5, which was brominated in

acetic acid, leading to 6 that was then converted with POCl₃to 6-

chloro-5-bromo-nicotinic acid chloride, which was then either

hydrolyzed to the acid 11 with water or transformed to the

corresponding methyl ester 7 by methanolysis.²²Suzuki−

C

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

a

5-Alkoxy-4-aryl-2-pyridine carboxamides (32a,b) were pre-

pared from commercially available 2-chloro-3-pyridinol 25 by

formylation (to give 26) followed by iodination to 27 as

described by Wishka et al.²⁴The corresponding phenolate was

selectively alkylated with alkylhalogenides to produce 28a and

28b in 86% and 92% yield, respectively. Suzuki−Miyaura

coupling with the Pd(dppf) catalyst was regioselective and

produced 29a and 29b with yields of greater than 97%.

Chemoselective reduction with zinc in acetic acid provided the

intermediates 30a,b, which were oxidized to the acids 31a,b with

tetrabutylammoniumpermanganate in pyridine. The ﬁnal step

was an amide coupling that yielded the desired 5-alkoxy-4-aryl-2-

pyridine carboxamides 32a,b.

The synthesis of the isosteric 5-alkoxy-6-aryl-2-pyridinecar-

boxamides 37a,b started from commercially available 2-chloro-3-

ﬂuoro-pyridine 33. Suzuki−Miyaura coupling to produce 34 in

65% yield was followed by nucleophilic aromatic substitution

with appropriate alkoxides and oxidation with hydrogen peroxide

in acetic acid to yield the N-oxides 35a,b. The N-oxides could be

converted to the nitriles 36a,b by treatment with dimethylcarba-

mic chloride and trimethylsilyl cyanide in acetonitrile. Two-step

hydrolysis of the nitriles followed by amide coupling aﬀorded the

desired 5-alkoxy-6-aryl-2-pyridinecarboxamides 37a,b.

Scheme 2. Synthesis of Compounds 10m−q

a

Reagents and conditions: (a) R²-OH, NaH, DMF; (b) arylboronic

acid, Pd(dppf)Cl₂·CH₂Cl₂, Na₂CO₃, toluene, water, 90 °C; (c) NBS,

AIBN, CCl₄, hν; (d) NH₄OH, ethanol, reﬂux; H₂O; (e) Bu₄NMnO₄,

pyridine, reﬂux.

The synthesis of 6-aza and 6-carbon analogues of 6-alkoxy-5-

aryl-3-pyridine carboxamides can be accomplished in only 3−4

steps starting from some of the above-described intermediates

(Scheme 3). As an example, nucleophilic aromatic substitution of

a

Scheme 3. Synthesis of Compounds 18, 21

Synthesis of pyrazines (Scheme 5) was realized from

commercially available 3,5-dibromo-2-pyrazinamine 38, which

could be arylated with good regioselectivity (80% yield) to give

the 3-arylisomer 39. Classic diazotization-bromination chemistry

furnished the 2,5-dibromo-3-aryl-pyrazine 40. The nucleophilic

aromatic substitution proceeded to 41 as the major isomer and

was followed by palladium catalyzed carbonylation to provide the

pyrazine carboxylate 42. 5-Alkoxy-6-aryl-pyrazine-2-carboxa-

mide 44 was obtained from 42 by saponiﬁcation and coupling

with (1R,2R)-2-amino-cyclohexanol.

Compound 51, the pyridazine analogue of 14g, was derived by

multistep synthesis from 3,6-dichloro- pyridazine 45. Nucleo-

philic aromatic substitution to give the monoalkoxylated product

46 was followed by iodination (to 47) and Suzuki−Miyaura

coupling to provide the 3-chloro-6-alkoxy-5-aryl-pyridazine 48.

Palladium-catalyzed carbonylation, saponiﬁcation and coupling

with (1R,2R)-2-amino-cyclohexanol provided the 6-alkoxy-5-

aryl-pyridazine-3-carboxamide 51.

a

Reagents and conditions: (a) N-methyl-cyclopropanemethanamine,

DBU, 90 °C then 2N NaOH; (b) B-4-chlorophenyl-boronic acid,

(Ph₃P)₄Pd, Na₂CO₃, DME, water, 85 °C; (c) (1R,2R)-2-amino-

cyclohexanol hydrochloride, DMF, TBTU, DIEA, RT; (d) amine,

TBTU, DIEA, DMF, rt; (e) (Ph₃)₂PdCl₂, Cu(I)I, TPP-resin, DMF,

MW, 120 °C; (f) Pd/C, H₂, EtOAc.

RESULTS AND DISCUSSION

■

All compounds were tested in triplicate in CB1 and CB2 receptor

binding assays as previously described.²⁵Compounds with K_i

values below 100 nM in CB1 receptor binding and lower than

20% inhibition at 3 μM at the CB2 receptor were further tested in

a GTPγ[³⁵S] assay in HEK cells overexpressing the human CB1

receptor. All compounds behaved as functional antagonists in

these assays. We used ClogP to rank compounds for lipophilicity

because the log D for the higher lipophilicity compounds could

not be easily determined. Rat microsomal clearance was

determined as described in the Experimental Section.

Compound 4 with its cyclopentyloxy substituent was a

moderately potent CB1 receptor antagonist. Systematic variation

of the alkoxy substituent (Table 1) revealed that substituents

larger than cyclopentyloxy lead to a 5−10-fold loss of potency, as

shown for 14a and 14b in comparison with 14c, while smaller

substituents as exempliﬁed by 14d−e and 14g−i lead to up to

30−70-fold improved potency compared to 14c. More detailed

examination of the SAR revealed that the size was not the

determining factor; rather substitution adjacent to the oxygen

atom is detrimental. Consistent with this observation, the 2-

7 with N-methyl-cyclopropanemethanamine in the presence of

DBU, followed by saponiﬁcation of the intermediate ester,

yielded 5-bromo-6-[(cyclopropylmethyl)methylamino]-3-pyri-

dinecarboxylic acid in 73% yield, which was then converted to

17 by Suzuki−Miyaura arylation followed by amide coupling to

furnish 18. The 6-carba analogue of 14g, compound 21, could be

prepared from 6-chloro-5-(4-chlorophenyl)-3-pyridinecarbox-

ylic acid (9a) by conversion to the amide 19, followed by

Sonogashira coupling with ethynylcyclopropane to aﬀord 20, and

ﬁnally palladium-catalyzed hydrogenation to yield 21.

To explore the eﬀect of varying the nitrogen atom position in

the core on CB1 receptor aﬃnity, we pursued the synthesis of

isosteric 2-pyridincarboxamides and also prepared the corre-

sponding pyrazine and pyridazine carboxamides (Schemes 4 and

5).

D

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

a

Scheme 4. Synthesis Substituted 2-Pyridine Carboxamides 32a,b and 37a,b

a

Reagents and conditions: (a) CH₂O, NaHCO₃, reﬂux; (b) I₂, NaHCO₃; (c) R²-X, NaH, DMF, 90 °C; (d) arylboronic acid, Pd(dppf)Cl₂·CH₂Cl₂,

Na₂CO₃, toluene, water, 90 °C; (e) Bu₄NBr, Zn, AcOH, 50 °C; (f) Bu₄NMnO₄, pyridine, 80 °C; (g) (1R,2R)-2-amino-cyclohexanol hydrochloride,

DMF, TBTU, DIEA, rt; (h) arylboronic acid, Pd(dppf)Cl₂·CH₂Cl₂, toluene, DMF, Na₂CO₃, water, 90 °C; (i) R²-OH, base, DMSO, rt; (j) H₂O₂,

AcOH, 70 °C; (k) N,N-dimethyl-carbamic chloride, TMSCN, Et₃N, CH₃CN; (l) AcCl, EtOH, 90 °C; (m) NaOH, THF, water, 80 °C.

propyloxy compound 14f is 20-fold less potent than the n-

butyloxy compound 14e, while compounds with bigger

substituents such as benzyloxy- 14j and the heterocyclylmethy-

loxy- compounds 14k−n are all potent CB1 receptor antagonists.

The alkoxy oxygen atom itself does not contribute to potency as

both the N-methyl nitrogen analogue 18 and the carbon

analogue 21 have potency similar to 14g.

Both 18 and 21 are, however, inferior to 14g with respect to

ClogP and metabolic stability in rat microsomes due to increased

susceptibility to either oxidative N-demethylation or benzylic

hydroxylation of 18 and 21, respectively. Of the compounds in

Table 1, 14g−j, 14l, and 14o were the most stable in rat

microsomes. The increased metabolic stability of these

compounds may be, in part, due to the reduced overall

lipophilicity compared to 14a. Alternatively, improved metabolic

stability may be due to a reduced rate of hydroxylation of the

CH₂-group in α-position to the alkoxy oxygen by interference of

neighboring groups (i.e., cyclopropyl for 14g, CF₃for 14i, γ-

heteroatoms for 14h, 14l, and 14o). Compound 14g serves as

reference to highlight the SAR of aryl- (Table 2), amide- (Table

3), and core- variations (Table 4) because we have the most

complete set of data for analogues of 14g.

Moving the 4-chloro-atom to the 2-position on the phenyl

ring, as in 15b, led to a 17-fold loss in potency compared to 14g,

which could be partially compensated for by reintroduction of

the 4-chloro-atom as in 15a, which led to only 11-fold loss of

potency compared to 14g. Results with compound 15e suggest

that double substitution in the 2- and 5-position partially restores

potency; the 5-position substituent compensating for the

negative impact of the 2-position substituent. Polar 4-position

substituents were detrimental for CB1-receptor aﬃnity (15k−

n), with the 4-methoxy group leading to about 6-fold loss of

aﬃnity compared to 14g. However, all halogens and

pseudohalogens in the 4-position seem to be equally acceptable,

leading to compounds with potent CB1 receptor aﬃnity.

Metabolic stability in rat microsomes for 15g (4-CF₃) and 15h

(4-OCF₃) was excellent, which, for 15h, was also corroborated by

its high and sustained plasma exposure in rat SDPK studies

(Supporting Information Figure S2). These data may indicate

that modiﬁcations that reduce the propensity for arylhydrox-

ylations yield metabolically more stable molecules suitable for in

vivo experiments. Alternatively, the two relatively lipophilic

compounds, 15g and 15h, may be protected from metabolic

turnover by high protein binding. In some cases, microsomes

actually under predicted the metabolic stability of these

E

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

a

Scheme 5. Synthesis of Compounds 44 and 51 with Pyrazine and Pyridazine Scaﬀold, Respectively

a

Reagents and conditions: (a) arylboronic acid, (Ph₃)₄Pd, Na₂CO₃, THF/water, 90 °C; (b) isoamylnitrite, TMSBr, CH₂Br₂, rt; (c)

cyclopropanemethanol, NaH, DMF, rt; (d) Pd(dppf)Cl₂·CH₂Cl₂, CO, NEt₃, MeOH, 70 bar, 120 °C; (e) LiOH, water, THF, rt; (f) (1R,2R)-2-

amino-cyclohexanol hydrochloride, DMF, TBTU, DIEA, rt; (g) cyclopropanemethanol, NaH, DMSO, rt; (h) n-BuLi, TMP, I₂, THF, −75 °C; (i)

arylboronic acid, (Ph₃)₄Pd, Na₂CO₃, THF/water, 90 °C; (j) Pd(dppf)Cl₂·CH₂Cl₂, CO, NEt₃, MeOH, 70 bar, 120 °C; (k) LiOH, water, THF, rt.

Table 1. In Vitro Pharmacology, ClogP, and Microsomal Stability for Compounds 14a−o

R²

hCB1-R K_i[nM]

hCB2-R K_i[nM]

ClogP

Cl mic rat @ 2 μM [μL min⁻¹kg⁻¹

]

14a

14b

14c

14d

14e

14f

14g

14h

14i

cyclohexyl

phenyl

3035 394

1657 625

374 223

208 130

43.8 11.0

1074 127

11.4 4.5

5.1 1.6

>10000

>7000

5.8

5.1

5.3

4.7

5.4

4.6

4.8

3.7

4.6

5.2

3.4

2.7

3.7

5.3

662

353

cyclopentyl

cyclobutyl

n-butyl

210

101

2-propyl

cyclopropylmethyl

methoxyethyl

64

98

>10000

>7000

2,2,2-triﬂuoroethyl

benzyl

12.4 6.7

40.2 14.7

10.6 4.8

51.6 24.5

58.7 49.2

23.5 8.5

139 93

23

14j

>10000

20

14k

14l

(3-methyl-isoxazol-5-yl)methyl

pyrimidin-2-ylmethyl

pyridin-4-ylmethyl

pyridin-3-ylmethyl

pyridin-2-ylmethyl

151

12

14m

14n

14o

18

118

374

44

51.1 20.3

16.7 3.4

1730

1503

21

compounds, while rat hepatocyte clearance seemed to be more

predictive for in vivo behavior. For example, rat hepatocyte

clearance for 14g was 3.3 μL min⁻¹(10⁶cells)⁻¹@ 10 μM, and in

vivo clearance and V_ssin rat at a dose of 1 mg/kg iv was 7.4 mL

min⁻¹kg⁻¹and 1.8 L/kg, respectively.

The SAR exploration of the amide side chain revealed that the

rimonabant-like side chain (piperidin-1yl) is inferior to the

(1R,2R)-2-hydroxycyclohexyl side chain of 14g, as shown by the

50-fold loss in potency observed with 16a. The enantiomer and

the diastereoisomers of 14g (cf. 16c−e) lose considerable

potency for the CB1 receptor. Suitable alternatives leading to

F

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

Table 2. In Vitro Pharmacology, ClogP, and Microsomal Stability for Compounds 15a−n

R¹

2,4-dichloro

hCB1-R K_i[nM]

hCB2-R K_i[nM]

ClogP

Cl mic rat @ 2 μM [μL min⁻¹kg⁻¹

]

15a

15b

15c

15d

15e

15f

122 49

190 150

305 201

129 60

67.7 21.4

42.8 26.0

18.5 1.9

23.6 11.4

66.4 11.2

23.6 12.7

1900 820

>10000

>5000

>10000

1260 320

>5000

5.2

4.5

5.4

4.8

5.4

4.2

5.0

5.2

4.1

3.5

3.0

2.4

3.9

2.7

4.8

2-chloro

3,4-dichloro

3-chloro

2-chloro-5-CF₃

4-ﬂuoro

2080 480

>10000

>7000

59

37

15g

15h

15i

4-CF₃

9

4-CF₃O

0−3

4-methoxy

4-cyano

15j

15k

15l

4-methanesulfonylamino

4-sulfamoyl

4-carboxy

4-carbamoyl

4-Cl

15m

15n

14g

>10000

>8000

11.4 4.5

64

Table 3. In Vitro Pharmacology, ClogP, and Microsomal Stability for Compounds 16a−e

R³

hCB1-R K_i[nM]

hCB2-R K_i[nM]

ClogP

Cl mic rat @ 2 μM [μL min⁻¹kg⁻¹

]

16a

16b

16c

16d

16e

piperidin-1-yl

563 242

34.3 27.7

247 52

186 75

104 48

>10000

5.2

4.5

4.8

(2R)-2-cyclopropyl-2-hydroxypropyl

(1S,2S)-2-hydroxycyclohexyl

(1R,2S)-2-hydroxycyclohexyl

(1S,2R)-2-hydroxycyclohexyl

2.4

2637 342

>5000

163

111

>7000

provides the additional steric bulk in this position that is

necessary to obtain high aﬃnity CB1 receptor ligands.

Table 4. Pharmacological Data for Core Variants

Compound 14g and some of its core variants (32a,b, 37a,b,

44, and 51) allow us to rationalize the CB1 binding

pharmacophore in this compound class. The energy-minimized

structure of pyrazine 44 in Figure 4 served as a template to

explore the CB1 pharmacophore. In the minimum energy

conformer, the orientation of the amide bond is locked via a

combination of reduced steric repulsion and the favorable

electrostatic interaction between the amide NH H-bond donor

with the aromatic nitrogen H-bond acceptor.²⁶The amide bond

in 14g has no preference for either of the two possible planar

orientations and, therefore, 14g is readily able to adopt the same

conformation as 44.

The loss in potency observed with the isostere 37a highlights

the relevance of the pyridine nitrogen in both 44 and 14g to

mediate an attractive interaction with the CB1 receptor. The

dramatic decrease of binding activity of the isosteric 2-

pyridinecarboxamide 32a and pyridazine 51 compared to 14g

can be rationalized by the diﬀerent relative orientation of the

phenyl and amide substituents. The change in vector arrange-

ment with respect to the aryl group and the amide carbonyl is

depicted by the orange arrows overlaid on 44 and 32a in Figure 4.

compd

core (A¹, A², A³)

hCB1-R K_i[nM]

hCB2-R K_i[nM]

a

32a

C, N, C

N, C, C

N, C, N

C, N, N

C, C, N

>7000

1830 1369

25.8 13.5

>5000

1526 701

863 546

801 171

>3000

a

37a

44

51

14g

11.4 4.5

>7000

a

Values measured for 32b and 37b conﬁrmed a drastic loss of aﬃnity

for the CB1 receptor for these isosters of 14i.

CB1 aﬃnity similar to the (1R,2R)-2-hydroxycyclohexyl side

chain are many other simple hydroxy-alkyl side chains (data not

shown). However, care must be taken to avoid introducing

metabolically labile sites. One useful example is compound 16b,

with its tertiary hydroxy group where the cyclopropyl group

G

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

Table 5. Rat PK Data of Selected Compounds Compared with

a

Rimonabant

rimonabant

5.9 4.9

14g

14h

K_ihCB1-R [nM]

11.4 4.5

5.1 1.6

c

d

EC₅₀GTPγ[³⁵S] hCB1-R [nM]

1.5 0.2

3.9 3.3

e

11.6 8.2

b

f

rat Cl [mL min⁻¹kg⁻¹

]

54

7.4

18.6

b

e

f

rat Vss [L/kg]

19.5

1.8

0.7

g

h

i

rat plasma C_max[ng/mL]

rat plasma T_max[h]

F [%]

244

729

1615

g

h

i

1.5

1.9

0.5

b

h

i

nd

23

84

B/P ratio

3−5

∼0.7

∼0.1

rat PPB [%]

99.7

>98.4

96.1

g

h

i

rat plasma C_unbound,max[ng/mL]

0.83

<11

63

g

h

i

C_unbound,max[nM]

1.8

<28

156

a

Cl, clearance; Vss, volume of distribution at steady state; F,

bioavailability in % at the given dose; B/P ratio, Brain to plasma

b

ratio; PPB, plasma protein binding. Reported values: Cl, ∼17 mL

c

min⁻¹kg⁻¹; Vss, 11.5 L/kg; T_max, 1−3 h; F, 12% (male rats).²⁷n = 7.

d

e

f

g

h

n = 3. @ 1 mg/kg iv. @ 3.3 mg/kg iv. @ 10 mg/kg po. @ 7.9 mg/

i

kg po. @ 9 mg/kg po

lipophilicity, most likely due to improved solubility and

dissolution rate, while plasma protein binding decreased. As

one would expect, lipophilicity, or to be more precise, polar

surface area, was directly related to brain/plasma ratio. Both 6-

alkoxy-5-aryl-3-pyridinecarboxamides have reduced brain to

plasma ratios in comparison to rimonabant. In the case of 14h,

the brain/plasma ratio of 0.1 suggests that doses can be identiﬁed

that selectively antagonize CB1 receptors outside the CNS.

The set of compounds in Table 5 was selected to explore the

relative importance of central and peripheral antagonism of CB1

receptors on food intake and body weight. In the periphery, the

high plasma exposure of 14g and 14h after oral dosing in rat

compensates for their lower aﬃnity compared to rimonabant,

while at exposure levels expected to be relevant in the periphery,

brain/plasma ratios of these three compounds range from ∼4 for

rimonabant to ∼0.1 for 14h. This 1.5 log unit diﬀerence in brain/

plasma ratios should be suﬃcient to probe the relative

contributions of peripheral vs central CB1R antagonism to

therapeutic response. To exclude the possibility of unexpected

non-CB1R-mediated eﬀects, a CEREP binding proﬁle against 80

unrelated receptors and enzymes was performed. Both 14g and

14h were conﬁrmed as selective CB1 receptor binders, with 14g

showing no other relevant interactions and 14h identiﬁed only as

a weak NK2 receptor antagonist with an IC₅₀of 46 μM.²⁸

Compounds were tested in a rat model of diet induced obesity.

All compounds were given as food admix. Compounds 14g and

14h were administered to deliver 30 mg/kg/day for each, and

rimonabant was given at 10 mg/kg/day. Food admix was chosen

to ensure the new compounds maintained plasma exposure well

above their CB1 receptor K_is throughout the day, as well as to

ensure a smooth PK proﬁle by minimizing diﬀerences between

peak and trough levels. To conﬁrm exposure, the PK proﬁle for

14g and 14h was measured after 10 days of food admix in a

separate experiment in catheterized rats (Figure 5).

Figure 4. Deﬁnition of the pharmacophore for CB1 antagonists based

on the pyrazine template 44. The orientation of the amide group is

determined by a combination of reduced steric repulsion and attractive

electrostatic interaction between the amide NH and the pyridine

nitrogen (marked by the dashed line). CB1 receptor antagonists appear

to require optimal positioning of the pyridine nitrogen for eﬃcient

receptor interaction (highlighted in 44 and 14g as opposed to 37a). In

the low energy conformers of 32a and 51, the phenyl and amide group

adopt a diﬀerent relative orientation (highlighted by the orange arrows

on 44 and 32a).

The exit vector geometry between the aryl and amide groups

appears relevant for CB1 receptor binding. Exit vector geometry

in all isosteres except 14g is constrained by the strong preference

of the 2-pyridylamides, 2-pyrazineamides, and 2-pyridazylamides

for the depicted planar geometries.²⁶

Several compounds were selected based on their in vitro CB1

receptor aﬃnity proﬁles and stability in rat microsomes and

characterized further in functional assays and single dose PK

(SDPK) experiments in rats in comparison with rimonabant

(Table 5). Compounds 14g and 14h were approximately 2-fold

less potent with respect to binding and 2- (14g) to 6-fold (14h)

less potent as antagonists in the GTPγ[³⁵S] assay compared to

rimonabant. SDPK studies after oral administration showed both

14g and 14h to have better bioavailability than rimonabant,

leading to substantially higher C_maxand AUC values (latter are

not shown). Dose-normalized plasma exposure of 14g was 4-fold

and of 14h was 7-fold higher than rimonabant (C_max/dose:

rimonabant 24.4, 14g 92.3, 14h 179 ng/mL). The signiﬁcantly

higher dose-normalized exposure of 14g and 14h compensated

for their lower in vitro potency compared to rimonabant,

allowing us to use them in in vivo experiments at reasonable

doses. Plasma exposure of 14g and 14h in all in vivo studies was

signiﬁcantly above the K_ifor CB1 receptor binding for both total

as well as the protein unbound fraction. All three molecules are

lipophilic, but rimonabant is by far the most lipophilic of the

three, with a ClogP of 6.5 followed by 14g (ClogP 4.8) and 14h

(ClogP 3.7). This was reﬂected by signiﬁcant diﬀerences in

several parameters such as volume of distribution, which was

lowest for 14h. Bioavailability increased with decreasing

Interindividual variability in exposure was low for 14g, with

plasma C_maxof 1025 ng/mL and brain levels of 1400 ng/g after 10

days of administration. These were similar to the single dose PK

data but showing a slight accumulation in brain, which reached a

brain/plasma ratio of 1.3 at steady state. For 14h, interindividual

variability was also low. Plasma C_maxwas 1000 ng/mL and brain

levels were 130 ng/g at steady state, giving a calculated brain/

H

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

Figure 5. Plasma exposure data for 14g and 14h (three individual Sprague−Dawley rats each, 10 days of 30 mg/kg/day as food admix in chow, blood

sampling started after removal of food on day 10).

Figure 6. Body weight gain, body fat (determined by MRS) and plasma leptin levels in high fat diet fed Sprague−Dawley (N = 8−9) rats after 24 days of

dosing with rimonabant (10 mg/kg daily as food admix), 14g (30 mg/kg daily as food admix), and 14h (30 mg/kg daily as food admix). Data expressed

as mean SEM, *p < 0.05 or **p < 0.01 compared to vehicle (ANOVA followed by Dunnett’s posthoc test); nonﬁlled bars denote paired-feeding

groups for rimonabant, 14g, and 14h, respectively.

plasma ratio of 0.13. These PK studies conﬁrmed that peripheral

exposure levels were well above CB1 receptor K_ifor both

compounds and that the peripheral/brain selectivity for 14h was

sustained during chronic administration.

leptin. Notably, the pair-fed groups had signiﬁcantly higher leptin

levels vs their corresponding drug group. Moreover, in all cases,

leptin levels were even higher than in the vehicle group, albeit the

increases were not signiﬁcant if compared to vehicle.

Rimonabant performed as previously reported in high fat diet

fed rats,²⁹substantially and signiﬁcantly reducing body weight

gain at the dose of 10 mg/kg. Reduced body weight gain was

accompanied by a signiﬁcant reduction in body fat content and

decreased plasma leptin levels compared to vehicle-treated

animals. These results were similar in animals pair-fed to the

rimonabant group which had identical body weight loss, a

signiﬁcant reduction in body fat, but had no reduction in plasma

leptin levels. The 6-alkoxy-5-aryl-3-pyridinecarboxamides 14g

and 14h share similarly high plasma C_maxvalues, have similar

aﬃnities for the CB1 receptor, but diﬀer by a factor of 10 with

respect to brain/plasma ratios. This diﬀerence in brain/plasma

ratios is very succinctly mirrored in the results of the high fat

feeding experiment. Administration of 14g led to a strong and

signiﬁcant reduction in body weight gain and a signiﬁcant and

even stronger reduction in body fat content than was observed

with rimonabant as well as a signiﬁcant reduction of leptin levels

in plasma (Figure 6). Conversely, 14h had no signiﬁcant eﬀect on

body weight gain, produced a minor reduction in body fat

content and had no eﬀect on plasma leptin levels. Pair-fed groups

showed the same pattern of eﬀects that were observed in the

study with rimonabant. Treatment-related changes in body

weight were nicely reproduced in pair-fed animals, with slightly

less eﬀect to reduce body fat content but no reduction in plasma

Leptin is mainly expressed by adipose tissue.³⁰Lower plasma

leptin levels by reduced adipose mass is expected and has been

previously described.³¹The relationship between body fat and

leptin levels was fully reproduced in the three drug-treated

groups in comparison to vehicle-treated controls. However, this

relationship did not appear to hold for the pair-fed groups, which

was initially puzzling. It is tempting to ascribe the lower leptin

levels in the drug-treated groups compared to the pair-fed groups

to an additional metabolic beneﬁt and possibly peripheral eﬀect

of CB1 receptor antagonists; however, the plasma leptin levels

measured for 14h did not support this hypothesis. The

observation that all pair-fed groups had unexpectedly high

plasma leptin levels may have been due to the fact that drug-

treated groups were provided ad libitum access to food while

pair-fed groups were food restricted. This diﬀerence may have

inﬂuenced stress hormones and leptin levels. Consistent with this

notion is that behavioral stressors like restriction stress do

increase plasma leptin levels in cafeteria-diet induced obese

rats.³²

To test for eﬀects of CB1 receptor antagonist treatment on

glucose tolerance and insulin sensitivity, an oral glucose tolerance

test was performed. Both rimonabant and 14g improved insulin

sensitivity in an OGTT assay after 24 days of high fat diet fed

Sprague−Dawley rats, as reﬂected by decreased insulin AUC

I

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

Figure 7. Insulin excursion curves obtained after oral glucose challenge performed after 24 days of CB1 receptor antagonist treatment as detailed in

Figure 6 in overnight fasted obese SD (Sprague−Dawley) rats. A glucose load of 2 g/kg body weight was administered as bolus by gavage. Dotted lines

denote pair-fed groups. Rimonabant and vehicle curves are shown twice for clarity.

(Figure 7). Glucose excursion curves for all treatment groups

were unchanged compared to vehicle (Supporting Information

Figure S1). Groups pair-fed to both rimonabant and 14g showed,

despite reduction of body weight gain, no improvement in insulin

sensitivity compared to vehicle. Likewise, the peripherally

selective CB1 antagonist 14h, as well as its pair-fed group,

showed no improvement in insulin sensitivity compared to the

vehicle group. Our data (with 14g, rimonabant, and their

respective pair-fed groups) reinforce the notion that improve-

ment of insulin sensitivity is mainly driven by the central

mechanism of action of the compounds; an interpretation that is

corroborated by the data for 14h which did not have an eﬀect on

insulin sensitivity in this model.

suﬃcient to achieve metabolic beneﬁts, This conﬂicts with

several earlier reports, suggesting that peripherally selective CB1

receptor antagonists retain eﬀects on food intake and body

weight. However, two of the earlier compounds, LH-21 and

AM6545, were subsequently tested in CB1 receptor knockout

mice and were able to reduce body weight even in the absence of

CB1 receptors, pointing to oﬀ-target pharmacology.^33,34Results

with JD5037 varied depending on the experimental model. In

db/db and ob/ob mice, JD5037 improved insulin sensitivity

without aﬀecting body weight, while in DIO mice, JD5037

strongly reduced body weight. These data suggested an

interaction with leptin signaling that diﬀers between the genetic

and diet models.³⁵JD5037 has not been evaluated in CB1

receptor knockout mice.

Recently, new evidence from CNS-speciﬁc CB1 receptor

knockdown experiments in mice has added further support for

the relevance of central CB1 receptors for the antiobesity and

metabolic beneﬁts of rimonabant.^36,37These CNS-speciﬁc CB1

receptor knock-down animals show the typical body weight and

metabolic beneﬁts associated with global CB1 receptor knock

out. Moreover, the eﬀects of rimonabant on body weight,

glucose, insulin, leptin, and adiponectin were lost in these

animals, indicating an essential role for CB1 in the brain to

mediate the eﬀects of CB1 receptor antagonists.

On the basis of the preponderance of evidence derived from

experience with the molecules detailed above, as well as the

emerging literature in tissue-speciﬁc CB1 receptor KO animals,

we believe that the metabolic beneﬁts of CB1 receptor

antagonists are, indeed, and as originally believed, centrally

mediated. It is conceivable, however, that peripheral CB1

receptor antagonists may provide other beneﬁts perhaps via

anti-inﬂammatory mechanisms in the periphery and may have

relevance in treatment of speciﬁc conditions such as hepatic

steatosis¹¹or ischemia/reperfusion injury.³⁸

CONCLUSIONS

■

6-Alkoxy-5-aryl-3-pyridinecarboxamides are a new, relatively

small chemotype that has delivered potent CB1 receptor

antagonists with inherent high selectivity over CB2 receptors.

The series was optimized to obtain molecules with modest

lipophilicity compared to rimonabant, consequently lower brain

exposure and, as demonstrated for 14h, peripherally selective

molecules. Selected compounds were shown to be selective vs a

panel of unrelated receptors. Several molecules showed high and

persistent plasma exposures in rats and were selected as tools for

pharmacological investigations. Both rimonabant and 14g

signiﬁcantly reduced the rate of body weight gain and decreased

adiposity in high fat fed rats. However, the poorly brain penetrant

molecule 14h had no signiﬁcant eﬀect on body weight gain and

only a marginal eﬀect on body fat reduction. Both single dose PK

and PK after chronic administration as food admix conﬁrmed

high plasma exposure for 14h comparable to the levels reached

with 14g, an equipotent, structurally close analogue. Thus, the

lack of pharmacological activity for 14h could not be explained by

lack of peripheral exposure. Indeed, unbound plasma levels of

14h were achieved that would be expected to provide even

greater receptor occupancy than for pharmacologically eﬀective

levels of either 14g or rimonabant. The lack of metabolic eﬀects

for 14h is more likely consistent with its substantially lower

exposure in brain tissue, which was similarly low in both single

and chronic dosing studies. Although we did not measure

unbound fraction in brain, we feel that the low total exposure in

brain explains the lack of pharmacological activity for 14h. This

observation suggests that peripheral CB1 antagonism is not

EXPERIMENTAL SECTION

■

Compound Synthesis and Characterization. Chemistry.

Reactions were conducted under argon atmosphere. Unless otherwise

mentioned, all reagents and chemicals were obtained from commercial

suppliers and used without further puriﬁcation. Flash column

chromatography was carried out on a Jones Chromatography

Flashmaster II system using Telos Flash silica or Isolute FlashSi

prepacked columns. Compounds were characterized by a combination

of TLC, NMR, and MS analytical techniques as described below. Purity

J

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

of compounds used in bioassays was determined by reverse-phase high

performance liquid chromatography (HPLC) analysis to be ≥95% by

using an Agilent LC-system consisting of Agilent 1290 high pressure

system with DAD detection, a CTC PAL auto sampler, and a Agilent

6520 QTOF. The separation was achieved on a Zorbax Eclipse Plus C18

1.7 μm 2.1 mm × 50 mm column at 55 °C; A = 0.01% formic acid in

water; B = 0.01% formic acid in acetonitrile at ﬂow 1 mL/min. Gradient:

0 min 5%B, 0.3 min 5%B, 4.5 min 99%B 5 min 99%B; UV wavelength for

purity assessment 265 nm. ¹H and ¹³C NMR spectra were recorded on a

Bruker DRX 400, Bruker AVANCE II 400, or on a Bruker AVANCE III

600 spectrometer and were determined in CHCl₃-d or DMSO-d₆with

tetramethylsilane (TMS) (0.00 ppm) as the internal reference unless

otherwise noted. Chemical shifts are reported in ppm relative to TMS,

and coupling constants (J) are reported in hertz (Hz). Thin-layer

chromatography (TLC) was performed on EMD precoated silica gel 60

F254 plates, and spots were visualized with UV light or chemical

detection. Low-resolution mass spectra were obtained using an Applied

Biosystem API300, and high resolution LC-MS data were obtained

using an Agilent Q-TOF 6520 system.

5-Bromo-1,6-dihydro-6-oxo-3-pyridinecarboxylic Acid (6). To a

suspension of 5 (Acros, 50 g, 0.359 mol) in acetic acid (100 mL) was

added bromine (27.7 mL, 0.539 mol) dropwise during 45 min. The

temperature rose to 45 °C during the addition, and the resulting mixture

was stirred for 18 h at 50 °C. The mixture was concentrated and dried in

vacuo (45 °C/20 mbar) to aﬀord an orange solid that was digested with

toluene (350 mL). The solid was isolated by ﬁltration, washed with

toluene (4 × 50 mL), and dried (40 °C/20 mbar) to produce 6 (78.9 g,

quant) as yellow−orange solid.

5-Bromo-6-chloro-3-pyridinecarboxylic Acid Methyl Ester (7). To 6

(50 g, 0.229 mol) was added phosphorus oxychloride (Aldrich, 170 mL,

1.86 mol), and the resulting mixture was slowly warmed and stirred for 5

h at 70 °C. After cooling volatiles were removed in vacuo, toluene (2 ×

100 mL) was added to the residue and the mixture was concentrated

twice. The residue was combined with DCM (80 mL) and transferred to

a reaction vessel. Methanol (250 mL, 6.18 mol) wad added dropwise

during 15 min (caution: very exothermic). The mixture was stirred for 1

h at 70 °C, cooled to room temperature, and concentrated. The residue

was dissolved in diethyl ether and neutralized (ﬁnal pH of 8) by addition

of saturated sodium bicarbonate (100 mL) and sodium hydroxide

solution (2 N, 390 mL). Phases were separated, the water phase was

extracted with additional diethyl ether, and all organic phases were, after

a ﬁnal wash with sodium bicarbonate, combined. The organic phase was

dried over magnesium sulfate, ﬁltered, and concentrated. The crude

product was puriﬁed by ﬂash chromatography, eluting with 35% ethyl

acetate in heptane and crystallized from heptane to aﬀord 7 (42.0 g, 73%

yield) as white solid; mp 78−79 °C. ¹H NMR (600 MHz, CDCl₃) δ 3.97

(s, 3 H), 8.53 (d, J = 2.0, 1 H), 8.93 (d, J = 2.0, 1 H).

resulting mixture was stirred at reﬂux temperature for 1 h, cooled,

acidiﬁed with hydrochloric acid (2 N, 400 mL), and partitioned into

diethyl ether. The organic phases were combined, dried over magnesium

sulfate, ﬁltered, and concentrated to yield suﬃciently pure 9a (50 g,

quant) as oﬀ-white solid; mp 189 °C dec. ¹H NMR (400 MHz, DMSO-

d₆) δ 7.58 (s, 4 H), 8.20 (d, J = 2.1, 1 H), 8.90 (d, J = 2.1, 1 H), 13.70 (br

s, 1H). HRMS (m/z): [M − H⁻] calcd for C₁₂H₆Cl₂NO₂265.9776;

found 265.9787.

6-Chloro-5-(4-ﬂuorophenyl)-3-pyridinecarboxylic Acid (9b). Fol-

lowing a procedure similar to the preparation of 9a, compound 8b (49.0

g, 184 mmol) was used as starting material to produce 9b (46.5 g, quant)

as oﬀ-white solid; mp 182 °C dec. ¹H NMR (600 MHz, DMSO-d₆) δ

7.36 (t, J = 8.9, 2 H), 7.61 (dd, J = 8.9, 5.3, 2 H), 8.20 (d, J = 2.3, 1 H),

8.90 (d, J = 2.3, 1 H), 13.73 (br s, 1H). HRMS (m/z): [MH⁺] calcd for

C₁₂H₈ClFNO₂252.0228; found 252.0221.

5-(4-Chlorophenyl)-6-(cyclopropylmethoxy)-3-pyridinecarboxylic

Acid (10a). General Procedure A. To a solution of 9a (30.0 g, 112

mmol) and cyclopropanemethanol (13.5 mL, 168 mmol) in DMSO

(225 mL) was added potassium hydroxide powder (25.1 g, 448 mmol).

The resulting suspension was divided in 10 portions, heated with stirring

for 15 min in the microwave at 100 °C, cooled, poured onto ice/water

(500 mL), and citric acid solution (10%, 3.7 L) added. The resulting

precipitate was collected by ﬁltration, washed thoroughly with water,

and dissolved in ethyl acetate. The ethyl acetate solution was dried over

magnesium sulfate, ﬁltered, and concentrated to a ﬁnal volume of ∼200

mL. The product precipitated from this solution and 10a (30.0 g, 88%)

was obtained by ﬁltration, washing with heptane/ethyl acetate (2:1) and

drying in vacuo; mp 191 °C dec. ¹H NMR (400 MHz, DMSO-d₆) δ 0.33

(m, 2H), 0.53 (m, 2H), 1.25 (m, 1H), 4.26 (d, J = 7, 2H), 7.52 (d, J = 8.6,

2 H), 7.66 (d, J = 8.6, 2H), 8.12 (d, J = 2.1, 1 H), 8.70 (d, J = 2.1, 1 H),

13.10 (br s, 1H). HRMS (m/z): [MH⁺] calcd for C₁₆H₁₅ClNO₃

304.0741; found 304.0734.

5-(4-Chlorophenyl)-6-(2,2,2-triﬂuoroethoxy)-3-pyridinecarboxylic

Acid (10b). General procedure A, with 9a (32.0 g, 119 mmol) and

triﬂuoroethanol (10.3 mL, 143 mmol), was used to produce 10b (37.5 g,

95%) as white solid. ¹H NMR (600 MHz, DMSO-d₆) δ 5.13 (q, J = 9.0,

2H), 7.56 (d, J = 8.7, 2 H), 7.64 (d, J = 8.7, 2 H), 8.23 (d, J = 2.2, 1 H),

8.74 (d, J = 2.2, 1 H), 13.37 (br s, 1H). HRMS (m/z): [MH⁺] calcd for

C₁₄H₈ClF₃NO₃330.0145; found 330.0156.

6-(Cyclopropylmethoxy)-5-(4-ﬂuorophenyl)-3-pyridinecarboxylic

Acid (10c). General procedure A, with 9b (48.4 g, 192 mmol) and

cyclopropanemethanol (23.3 mL, 289 mmol), was used to produce 10c

1

(41.0 g, 74%) as white solid; mp 182−185 °C. H NMR (400 MHz,

DMSO-d₆) δ 0.34 (m, 2H), 0.53 (m, 2H), 1.25 (m, 1H), 4.26 (d, J = 7,

2H), 7.30 (t, J = 8.9, 2 H), 7.68 (dd, J = 8.6, 5.6, 2 H), 8.10 (d, J = 1.9, 1

H), 8.68 (d, J = 1.9, 1 H), 13.10 (br s, 1H). HRMS (m/z): [MH⁺] calcd

for C₁₆H₁₅FNO₃288.1036; found 288.1032.

6-Chloro-5-(4-chlorophenyl)-3-pyridinecarboxylic Acid Methyl

Ester (8a). To a solution of 7 (70 g, 0.28 mol) in toluene (1200 mL)

was added with stirring the [1,1-bis(diphenylphos-phino)ferrocene]

dichloropalladium DCM complex (11.4 g, 14 mmol), 4-chlorophe-

nylboronic acid (45.05 g, 0.28 mol), and sodium carbonate solution

(280 mL, 2M). The resulting mixture was stirred at 90 °C for 1 h, cooled,

and partitioned between water and ethyl acetate. The organic portion

was dried over magnesium sulfate, ﬁltered, and concentrated. The crude

product was puriﬁed by ﬂash chromatography, eluting with DCM/

heptane to produce 8a (52.2 g, 66%) as white solid; mp 105−107 °C. ¹H

NMR (400 MHz, CDCl₃) δ 3.97 (s, 3 H), 7.40 (d, J = 8.8, 2 H), 7.46 (d, J

= 8.8, 2 H), 8.24 (d, J = 2.2, 1 H), 8.89 (d, J = 2.2, 1 H). HRMS (m/z):

[MH⁺] calcd for C₁₃H₁₀Cl₂NO₂282.0089; found 282.0083.

5-(4-Chlorophenyl)-6-(pyrimidin-2-ylmethoxy)-3-pyridinecarbox-

ylic Acid (10d). General procedure A, with 9a (6.5 g, 24.2 mmol) and 2-

pyrimidinemethanol (2.94 g, 26.7 mmol), was used to produce 10d (8.3

g, quant) as oﬀ-white solid. ¹H NMR (600 MHz, DMSO-d₆) δ 5.70 (s,

2H), 7.42 (t, J = 4.9, 1H), 7.54 (d, J = 8.7, 2 H), 7.83 (d, J = 8.7, 2 H),

8.18 (d, J = 2.3, 1 H), 8.57 (d, J = 2.3, 1 H), 8.78 (d, J = 4.8, 2 H), 13.16

(br s, 1H). HRMS (m/z): [MH⁺] calcd for C₁₇H₁₃ClN₃O₃342.0646;

found 342.0644.

5-(4-Chloro-phenyl)-6-(pyridin-2-ylmethoxy)-3-pyridinecarbox-

ylic Acid (10e). General procedure A, with 9a (0.4 g, 1.5 mmol) and 2-

pyridinemethanol (0.22 mL, 2.2 mmol), was used to produce 10e (0.39

g, 76%) as white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 5.57 (s, 2H),

7.32 (t, J = 5.4, 1 H), 7.38 (d, J = 7.8, 1 H), 7.53 (d, J = 8.3, 2 H), 7.73 (d, J

= 8.3, 2 H), 7.80 (t, J = 7.8, 5.6, 1 H), 8.18 (d, J = 2.0, 1 H), 8.54 (d, J =

4.8, 1 H), 8.71 (d, J = 2.0, 1 H), 13.17 (br s, 1H). HRMS (m/z): [MH⁺]

calcd for C₁₈H₁₄ClN₂O₃341.0693; found 341.0693.

5-(4-Chloro-phenyl)-6-(pyridin-4-ylmethoxy)-3-pyridinecarbox-

ylic Acid (10f). General procedure A, with 9a (0.3 g, 1.1 mmol) and 4-

pyridinemethanol (0.147 g, 1.3 mmol), was used to produce 10f (0.27 g,

76%) as oﬀ-white solid. MS (ESI) m/z = 339.1 [M − H⁻].

5-(4-Cyanophenyl)-6-(cyclopropyl-methoxy)-3-pyridinecarbox-

ylic Acid (10g). To a solution of 12a (1.15 g, 4.2 mmol) in toluene (25

mL) and DMF (2.5 mL) was added with stirring the [1,1-

6-Chloro-5-(4-ﬂuorophenyl)-3-pyridinecarboxylic Acid Methyl

Ester (8b). Following a procedure similar to the preparation of 8a, 4-

ﬂuorophenylboronic acid (46.1 g, 319 mmol) was used to produce 8b

1

(49.5 g, 58%) as white solid; mp 95−97 °C. H NMR (600 MHz,

CDCl₃) δ 3.97 (s, 3 H), 7.18 (t, J = 8.7, 2 H), 7.45 (dd, J = 8.9, 5.3, 2 H),

8.25 (d, J = 2.3, 1 H), 8.98 (d, J = 2.3, 1 H). HRMS (m/z): [MH⁺] calcd

for C₁₃H₁₀ClFNO₂266.0384; found 266.0382.

6-Chloro-5-(4-chlorophenyl)-3-pyridinecarboxylic Acid (9a). To a

solution of 8a (52.2 g, 185 mmol) in THF (1300 mL) was added water

(400 mL) and lithium hydroxide monohydrate (23.3 g, 555 mmol). The

K

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

bis(diphenylphosphino)ferrocene] dichloropalladium DCM complex

(0.173 g, 0.21 mmol), B-4-cyanophenyl-boronic acid (0.62 g, 4.2 mmol),

and sodium carbonate solution (16.9 mL, 2M). The resulting mixture

was stirred at 85 °C for 1.5 h, cooled, and partitioned between citric acid

solution (10%) and ethyl acetate. The organic portion was dried over

Na₂SO₄, ﬁltered, and concentrated. The crude was crystallized from

ethyl acetate/heptane (3:1; ∼8 mL) to produce 10g (0.83 g, 67%) as

light-brown solid. ¹H NMR (400 MHz, DMSO-d₆) δ 0.34 (m, 2 H), 0.53

(m, 2 H), 1.25 (m, 1 H), 4.28 (d, J = 7.0, 2 H), 7.85 (d, J = 8.5, 2 H), 7.93

(d, J = 8.5, 2 H), 8.18 (d, J = 2.2, 1 H), 8.74 (d, J = 2.2, 1 H), 13.12 (br s, 1

H). HRMS (m/z): [M − H⁻] calcd for C₁₇H₁₃N₂O₃293.0932; found

293.0939.

6-(Cyclopropylmethoxy)-5-[4-(triﬂuoromethoxy)phenyl]-3-pyridi-

necarboxylic Acid (10h). To a solution of 12a (2.19 g, 8.05 mmol) in

DME (57 mL) was added with stirring tetrakis(triphenylphosphine)-

palladium(0) (0.94 g, 0.805 mmol). To this mixture was added B-4-

(triﬂuoromethoxy)phenyl-boronic acid (1.99 g, 9.66 mmol) dissolved in

ethanol (27 mL) and sodium carbonate solution (7.25 g in 37 mL

water). The resulting mixture was stirred at 85 °C for 3.5 h, cooled,

ﬁltered through Celite, and concentrated. The residue was partitioned

between brine and ethyl acetate. The organic portion was dried over

magnesium sulfate, ﬁltered, and concentrated. The crude product was

puriﬁed by ﬂash chromatography, eluting with a DCM/methanol

gradient to produce 10h (1.79 g, 63%) as oﬀ-white solid. ¹H NMR (600

MHz, DMSO-d₆) δ 0.33 (m, 2 H), 0.53 (m, 2 H), 1.25 (m, 1 H), 4.25 (d,

J = 7.2, 2 H), 7.45 (d, J = 8.7, 2 H), 7.77 (d, J = 8.7, 2 H), 8.16 (d, J = 2.1, 1

H), 8.68 (d, J = 2.1, 1 H), 13.45 (br s, 1H). HRMS (m/z): [M − H⁻]

calcd for C₁₇H₁₃F₃NO₄352.0802; found 352.0807.

6-(Cyclopropylmethoxy)-5-(4-triﬂuorophenyl)-3-pyridinecarbox-

ylic Acid (10i). To a solution of 12a (61 mg, 0.224 mmol) in dioxane (1

mL) was added the [1,1-bis(diphenylphosphino)ferrocene] dichlor-

opalladium DCM complex (8.2 mg, 11 μmol), B-4-triﬂuorophenyl-

boronic acid (63.9 mg, 0.336 mmol), and sodium carbonate solution

(0.34 mL, 2M). The resulting mixture was shaken at 80 °C for 2 h,

cooled, ﬁltered through Celite, and concentrated. The crude was

puriﬁed by preparative HPLC (Zorbax XDB, acetonitrile/water and

0.05% triﬂuoroacetic acid) to produce 10i (15 mg, 20%) as white solid.

MS (ESI) m/z = 336.1 [M − H⁻].

5-[4-Chlorophenyl]-6-(cyclobutoxy)-3-pyridinecarboxylic Acid

(10j). Following a procedure similar to the preparation of 10h, 12d

(0.22 g, 0.816 mmol) and B-4-chlorophenyl-boronic acid (0.145 g, 0.90

mmol) were used to produce 10j (0.215 g, 89%) as white solid. ¹H NMR

(300 MHz, DMSO-d₆) δ 1.65 (m, 1 H), 1.78 (m, 1 H), 2.06 (m, 2 H),

2.40 (m, 2 H), 5.29 (m, 1 H), 7.53 (d, J = 8.4, 2 H), 7.66 (d, J = 8.4, 2 H),

8.12 (d, J = 2.2, 1 H), 8.68 (d, J = 2.2, 1 H), 13.16 (br s, 1 H). MS (ESI)

m/z = 302.2 [M − H⁻].

6-(Cyclopropylmethoxy)-5-(2,4-dichlorophenyl)-3-pyridinecar-

boxylic Acid (10k). Following a procedure similar to the preparation of

10h, 12a (0.135 g, 0.496 mmol) and B-2,4-dichlorophenyl-boronic acid

(0.114 g, 0.597 mmol) were used to produce 10k (85 mg, 51%) as

colorless oil. ¹H NMR (600 MHz, DMSO-d₆) δ 0.27 (m, 2 H), 0.47 (m,

2 H), 1.17(m, 1 H), 4.21 (d, J = 7.0, 2 H), 7.48 (d, J = 8.3, 1 H), 7.53 (dd,

J = 8.3, 2.1, 1 H), 7.76 (d, J = 2.1, 1 H), 8.01 (d, J = 2.3, 1 H), 8.76 (d, J =

2.3, 1 H), 13.08 (br s, 1 H). HRMS (m/z): [M − H⁻] calcd for

C₁₆H₁₂Cl₂NO₃336.0194; found 336.0207.

5-[4-Chlorophenyl]-6-[(3-methyl-5-isoxazolyl)methoxy]-3-pyridi-

necarboxylic Acid (10l). Following a procedure similar to the

preparation of 10h, 12c (0.41 g, 1.32 mmol) and B-4-chlorophenyl-

boronic acid (0.213 g, 1.36 mmol) were used to produce 10l (0.248 g,

54%) as light-brown solid. ¹H NMR (400 MHz, DMSO-d₆) δ 2.22 (s, 3

H), 5.59 (s, 2 H), 6.41 (s, 1 H), 7.53 (d, J = 8.6, 2 H), 7.64 (d, J = 8.6, 2

H), 8.17 (d, J = 2.2, 1 H), 8.73 (d, J = 2.2, 1 H), 13.12 (br s, 1 H). HRMS

(m/z): [MH⁺] calcd for C₁₇H₁₄ClN₂O₄345.0637; found 345.0641.

5-[2-Chloro-5-(triﬂuoromethyl)phenyl]-6-(cyclopropylmethoxy)-

3-pyridinecarboxylic Acid (10m). To a solution of 24a (1.15 g, 3.4

mmol) in CCl₄(40 mL) was added with stirring 1-bromo-2,5-

pyrrolidinedione (1.38 g, 7.7 mmol) and 2,2′-azobis(2-methylpropioni-

trile) (6 mg). The mixture was irradiated for 1 h with a 500 W UV lamp.

After cooling, the reaction mixture was washed sodium hydrogen sulﬁte

solution (38−40%, 30 mL). The water phase was extracted with

dichloromethane; organic phases were washed three times with water,

combined, dried over MgSO₄, ﬁltered, and concentrated. The residue,

consisting mostly of 3-[2-chloro-5-(triﬂuoromethyl)phenyl]-2-cyclo-

propylmethoxy-5-dibromomethyl-pyridine, was dissolved in ethanol

(28 mL), a concentrated ammonia solution (7 mL) was added, and the

mixture was stirred for 1 h at reﬂux temperature and poured after cooling

into a cold hydrochloric acid solution (1 N, 150 mL). Workup

proceeded by extraction with diethyl ether and washing the organic

phases with ice water and brine. Organic phases were combined, dried

over Na₂SO₄, ﬁltered, and concentrated. The residue, consisting mostly

of 5-[2-chloro-5-(triﬂuoromethyl)phenyl]-6-cyclopropylmethoxy-pyri-

dine-3-carbaldehyde, was dissolved in pyridine (17 mL), a solution of

tetrabutylammonium permanganate (3.87 g, 10.7 mmol) in pyridine (17

mL) was added, and the resulting mixture was stirred at 80 °C for 5 h.

After cooling, the reaction mixture was added to sodium hydrogen

sulﬁte solution (38−40%, 50 mL), hydrochloric acid (2 N; 300 mL) was

added, and the mixture was extracted with diethyl ether. Organic phases

were combined, dried with Na₂SO₄, ﬁltered, and concentrated. The

residue was puriﬁed by ﬂash chromatography, eluting with an ethyl

acetate/heptane gradient to produce 10m (0.37 g, 30%) as light-brown

solid. ¹H NMR (600 MHz, DMSO-d₆) δ 0.27 (m, 2 H), 0.46 (m, 2 H),

1.16 (m, 1 H), 4.23 (d, J = 7.1, 1 H), 7.85 (m, 3 H), 8.11 (d, J = 2.3, 1 H),

8.79 (d, J = 2.3, 1 H), 13.19 (br s, 1 H). HRMS (m/z): [M − H⁻] calcd

for C₁₇H₁₂ClF₃NO₃370.0463; found 370.0466.

5-(4-Chloro-phenyl)-6-phenoxy-3-pyridinecarboxylic Acid (10n).

Following a procedure similar to the preparation of 10m, 24b (0.64 g,

2.2 mmol) was used to produce 10n (0.33 g, 47%) as yellow foam. ¹H

NMR (600 MHz, DMSO-d₆) δ 7.21 (m, 2 H), 7.25 (t, J = 7.5, 1 H), 7.43

(t, J = 8.0, 2 H), 7.57 (d, J = 8.6, 2 H), 7.78 (d, J = 8.6, 2 H), 8.27 (d, J =

2.2, 1 H), 8.62 (d, J = 2.2, 1 H), 13.29 (br s, 1 H). HRMS (m/z): [M −

H⁻] calcd for C₁₈H₁₁ClNO₃324.0433; found 324.0439.

5-(4-Chlorophenyl)-6-cyclopentyloxy-3-pyridinecarboxylic Acid

(10o). Following a procedure similar to the preparation of 10m, 24c

(0.75 g, 2.6 mmol) was used to produce 10o (0.43 g, 55%) as yellow

solid. ¹H NMR (600 MHz, DMSO-d₆) δ 1.60 (m, 2 H), 1.65 (m, 2 H),

1.72 (m, 2 H), 1.93 (m, 2 H), 5.56 (m 1 H), 7.52 (d, J = 8.7, 2 H), 7.61

(d, J = 8.7, 2 H), 8.11 (d, J = 2.3, 1 H), 8.71 (d, J = 2.3, 1 H), 13.11 (br s,

1H). HRMS (m/z): [M − H⁻] calcd for C₁₇H₁₅ClNO₃316.0746; found

316.0751.

6-Butoxy-5-(4-chlorophenyl)-3-pyridinecarboxylic Acid (10p). Fol-

lowing a procedure similar to the preparation of 10m, 24d (0.86 g, 3.1

mmol) was used to produce 10p (0.57 g, 65%) as yellow solid. ¹H NMR

(600 MHz, DMSO-d₆) δ 0.90 (t, J = 7.4, 3 H), 1.39 (m, 2 H), 1.69 (m, 2

H), 4.39 (t, J = 6.6, 2 H), 7.52 (d, J = 8.7, 2 H), 7.63 (d, J = 8.7, 2 H), 8.12

(d, J = 2.2, 1 H), 8.71 (d, J = 2.2, 1 H), 13.13 (br s, 1 H). HRMS (m/z):

[M − H⁻] calcd for C₁₆H₁₅ClNO₃304.0746; found 304.0753.

5-(4-Chlorophenyl)-6-(cyclohexyloxy)-3-pyridinecarboxylic Acid

(10q). Following a procedure similar to the preparation of 10m, 24e

(0.92 g, 3.0 mmol) was used to produce 10q (0.67 g, 64%) as yellow

solid. ¹H NMR (300 MHz, CDCl₃) δ 1.50 (m, 4 H), 1.69 (m, 2 H), 1.94

(m, 2 H), 5.30 (m, 1 H), 7.46 (d, J = 8.6, 2 H), 7.53 (d, J = 8.6, 2H), 8.21

(d, J = 2.2, 1 H), 8.86 (d, J = 2.2, 1 H), 10.67 (br s, 1 H). MS (ESI) m/z =

330.2 [M − H⁻].

5-[2-Chloro-5-(triﬂuoromethyl)phenyl]-6-cyclopentyloxypyridine-

3-carboxylic Acid (10r). Following a procedure similar to the

preparation of 10h, 12g (0.21 g, 0.734 mmol) and B-2-chloro-5-

(triﬂuoromethyl)-phenyl-boronic acid (0.181 g, 0.807 mmol) were used

to produce 10r (67 mg, 24%) as oﬀ-white solid. ¹H NMR (300 MHz,

DMSO-d₆) δ 1.50−1.77 (m, 6 H), 1.91 (m, 2 H), 5.58 (m, 1 H), 7.57 (m,

3 H), 8.15 (d, J = 2.2, 1 H), 8.96 (d, J = 2.2, 1 H), 13.0 (br s, 1 H). MS

(ESI) m/z = 384.3 [M − H⁻].

5-Bromo-6-chloro-3-pyridinecarboxylic Acid (11). Quinoline (15.3

mL) was added dropwise to phosphorus oxychloride (28.8 mL, 0.308

mol) and to this solution was added 6 (56 g, 0.257 mol). The mixture

was stirred for 2 h at 120 °C, cooled to below 100 °C, and water (231

mL, 12.8 mol) was added to give a suspension. The solid was collected

by ﬁltration and partitioned between diethyl ether and sodium

hydroxide solution (1 N, ﬁnal pH 12.2). The water phase was collected

and acidiﬁed with hydrochloric acid (1 N, ﬁnal pH 1). The product

precipitated, was collected by ﬁltration, dissolved in ethyl acetate, dried

L

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

over magnesium sulfate, ﬁltered, and concentrated to yield 11 (49.1 g,

81% yield) as beige solid; mp 156−158 °C. H NMR (600 MHz,

NH), 8.20 (d, J = 2.3, 1 H), 8.45 (d, J = 2.3, 1 H). HRMS (m/z): [MH⁺]

calcd for C₁₅H₂₂BrN₂O₄375.0743; found 375.0744.

1

5-Bromo-6-(cyclopropylmethoxy)-N-[(1R,2R)-2-hydroxycyclohex-

yl]-3-pyridinecarboxamide (13b). General procedure C, with 12a

(3.89 g, 14.3 mmol) was used to produce 13b (5.02 g, 95%) as oﬀ-white

solid. ¹H NMR (600 MHz, DMSO-d₆) δ 0.41 (m, 2 H), 0.51 (m, 2 H),

1.21 (m, 4 H), 1.27 (m, 1 H), 1.64 (m, 2 H), 1.80 (m, 1 H), 1.91 (m 1

H), 3.36 (m, 1 H), 3.58 (m, 1 H), 3.89 (m, 2H), 4.66 (d, J = 4.8, OH),

7.98 (bd, J = 8.2, NH), 8.41 (d, J = 2.4, 1 H), 8.44 (d, J = 2.4, 1 H).

HRMS (m/z): [MH⁺] calcd for C₁₆H₂₂BrN₂O₃369.0814; found

369.0802.

5-Bromo-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(3-pyridinylme-

thoxy)-3-pyridinecarboxamide (13c). General procedure C, with 12b

(0.25 g, 0.81 mmol) in DMF, was used to produce 13c (0.21 g, 64%) as

white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 1.22 (m, 4 H), 1.64 (m, 2

H), 1.81 (m, 1 H), 1.89 (m 1 H), 3.39 (m, 1 H), 3.61 (m, 1 H), 4.64 (d, J

= 4.8, OH), 5.54 (s, 2 H), 7.43 (dd, J = 7.8, 4.8, 1 H), 7.89 (dt, J = 8.0, 1.9,

1 H), 8.22 (bd, J = 8.3, NH), 8.50 (d, J = 2.0, 1 H), 8.55 (dd, J = 4.8, 1.6, 1

H), 8.64 (d, J = 2.0, 1 H), 8.71 (bd, J = 1.9, 1 H). MS (ESI) m/z = 406.2

[MH⁺].

DMSO-d₆) δ 8.55 (d, J = 2.1, 1 H), 8.86 (d, J = 2.1, 1 H), 13.9 (br s, 1 H).

HRMS (m/z): [M − H⁻] calcd for C₆H₂BrClNO₂235.8937; found

235.8946.

5-Bromo-6-(cyclopropylmethoxy)-3-pyridinecarboxylic Acid

(12a). General Procedure B. To a solution of 11 (2.4 g, 10.1 mmol)

and cyclopropanemethanol (1.23 mL, 15.2 mmol) in DMSO (8 mL)

was added potassium hydroxide powder (2.28 g, 40.6 mmol). The

resulting suspension was heated with stirring for 8 min in the microwave

at 100 °C, cooled, and poured onto ice/water (30 mL), and citric acid

solution (10%, 40 mL) was added. The resulting precipitate was

collected by ﬁltration, washed thoroughly with water, and dissolved in

ethyl acetate. The ethyl acetate solution was dried over magnesium

sulfate, ﬁltered, and concentrated to aﬀord 12a (2.4 g, 87%) as white

solid. ¹H NMR (400 MHz, DMSO-d₆) δ 0.37 (m, 2H), 0.58 (m, 2H),

1.29 (m, 1H), 4.27 (d, J = 7, 2H),8.35 (d, J = 2.1, 1 H), 8.65 (d, J = 2.1, 1

H), 13.28 (br s, 1H). HRMS (m/z): [MH⁺] calcd for C₁₀H₁₁BrNO₃

273.9902; found 273.9905.

5-Bromo-6-(pyridin-3-yl-methoxy)-3-pyridinecarboxylic Acid

(12b). General procedure B, with 11 (1.0 g, 4.2 mmol) and 3-

pyridinemethanol (0.62 mL, 6.3 mmol), was used to produce 12b (1.05

g, 80%) as white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 5.55 (s, 2 H),

7.44 (m, 1 H), 7.90 (m, 1 H), 8.40 (d, J = 2.1, 1 H), 8.56 (m, 1 H), 8.71

(d, J = 2.1, 1 H), 8.72 (m, 1 H), 13.33 (br s, 1 H). HRMS (m/z): [MH⁺]

calcd for C₁₂H₁₀BrN₂O₃310.9855; found 310.9853.

5-Bromo-6-[(3-methyl-5-isoxazolyl)methoxy]-3-pyridinecarbox-

ylic Acid (12c). General procedure B, with 11 (0.50 g, 2.1 mmol) and 3-

methyl-5-isoxazolemethanol (0.36 g, 3.2 mmol), was used to produce

12c (0.43 g, 65%) as white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 2.24

(s, 3 H), 5.61 (s, 2 H), 6.48 (s, 1 H), 8.40 (d, J = 1.9, 1 H), 8.70 (d, J = 1.9,

1 H), 13.18 (br s, 1 H). HRMS (m/z): [M − H⁻] calcd for

C₁₁H₈BrN₂O₄310.9668; found 310.9679.

5-Bromo-6-cyclobutoxy-3-pyridinecarboxylic Acid (12d). General

procedure B, with 11 (0.25 g, 1.06 mmol) and cyclobutanol (0.115 g,

1.59 mmol), was used to produce 12d (0.23 g, 81%) as white solid. ¹H

NMR (600 MHz, DMSO-d₆) δ 1.67 (m, 1 H), 1.82 (m, 1 H), 2.11 (m, 2

H), 2.44 (m, 2 H), 5.25 (m, 1 H), 8.34 (d, J = 2.0, 1 H), 8.64 (d, J = 2.0, 1

H), 13.31 (br s, 1 H). HRMS (m/z): [M − H⁻] calcd for C₁₀H₉BrNO₃

269.9766; found 269.9778.

6-Benzyloxy-5-bromo-3-pyridinecarboxylic Acid (12e). General

procedure B, with 11 (1.00 g, 4.23 mmol) and benzenemethanol

(0.65 mL, 6.34 mmol), was used to produce 12e (0.31 g, 24%) as white

solid. ¹H NMR (400 MHz, DMSO-d₆) δ 2.54 (s, 3 H), 5.51 (s, 2 H),

7.3−7.5 (m, 5 H), 8.40 (d, J = 2.1, 1 H), 8.70 (d, J = 2.1, 1 H), 13.32 (br s,

1 H). MS (ESI) m/z = 308.3, 310.3 [M − H⁻].

5-Bromo-6-(2-methoxyethoxy)-3-pyridinecarboxylic Acid (12f).

General procedure B, with 11 (4.00 g, 17 mmol) and 2-methoxyethanol

(2.0 mL, 25 mmol), was used to produce 12f (2.90 g, 62%) as white

solid. ¹H NMR (600 MHz, DMSO-d₆) δ 3.32 (s, 3 H), 3.71 (t, J = 4.6, 2

H), 4.54 (t, J = 4.6, 2 H), 8.36 (d, J = 2.0, 1 H), 8.67 (d, J = 2.0, 1 H),

13.34 (br s, 1 H). HRMS (m/z): [MH⁺] calcd for C₉H₁₁BrNO₄

275.9872, 277.9851; found 275.9871, 277.9849.

5-Bromo-6-cyclopentyloxypyridine-3-carboxylic Acid (12g). Gen-

eral procedure B, with 11 (0.5 g, 2.11 mmol) and cyclopentanol (250 μL,

2.75 mmol), was used to produce 12g (321 mg, 53%) as white solid. ¹H

NMR (300 MHz, DMSO-d₆) δ 1.55−1.80 (m, 6 H), 1.96 (m, 2 H), 5.50

(m, 1 H), 8.33 (d, J = 2.2, 1 H), 8.67 (d, J = 2.2, 1 H), 13.2 (br s, 1 H). MS

(ESI) m/z = 284.2, 286.2 [M − H⁻].

5-Bromo-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(1-methylethoxy)-3-

pyridinecarboxamide (13d). General procedure C, with 5-bromo-6-(1-

methylethoxy)-3-pyridinecarboxylic acid³⁹(21 mg, 0.081 mmol) in

DMF, was used to produce 13d (21 mg, 73%) as white solid. MS (ESI)

m/z = 357.0, 359.0 [MH⁺].

5-Bromo-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(phenylmethoxy)-3-

pyridinecarboxamide (13e). General procedure C, with 12e (145 mg,

0.47 mmol) in DMA was used to produce 13e (93 mg, 49%) as orange

solid. ¹H NMR (400 MHz, DMSO-d₆) δ 1.21 (m, 4 H), 1.63 (m, 2 H),

1.85 (m, 1 H), 3.40 (m, 1 H), 3.61 (m, 1 H), 4.66 (d, J = 4.7, OH), 5.50

(s, 2 H), 7.3−7.5 (m, 5 H), 8.23 (bd, J = 8.3, NH), 8.49 (d, J = 2.2, 1 H),

8.63 (d, J = 2.2, 1 H). MS (ESI) m/z = 405.3, 407.3 [MH⁺].

5-(4-Chlorophenyl)-6-(cyclopropylmethoxy)-N-[(1R,2R)-2-hydrox-

ycyclohexyl]-3-pyridinecarboxamide (14g), General Procedure D. To

a solution of 10a (21.5 g, 70.8 mmol) in DMF (650 mL) was added

TBTU (25.0 g, 77.9 mmol), DIEA (60.6 mL, 354 mmol), and ﬁnally

(1R,2R)-2-amino-cyclohexanol hydrochloride (11.8 g, 77.9 mmol). The

reaction was stirred for 16 h at RT and afterward concentrated in vacuo

at 45 °C. The resulting residue was puriﬁed by ﬂash chromatography,

eluting with ethyl acetate/heptane (2:1), and product fractions were

concentrated to a ﬁnal volume of ∼200 mL. The product precipitated

from this solution, and 14g (21.7 g, 76%) was obtained by ﬁltration,

washing with heptane/ethyl acetate (1:1) and drying in vacuo; mp 193−

194 °C. ¹H NMR (400 MHz, CDCl₃) δ 0.33 (m, 2H), 0.58 (m, 2H),

1.32 (m, 5H), 1.77 (m, 2H), 2.11 (m, 2H), 3.28 (d, J = 5.1, OH), 3.44

(m, 1H), 3.85 (m, 1H), 4.26 (d, J = 7, 2H), 6.06 (bd, J = 7, NH), 7.40 (d,

J = 8.6, 2 H), 7.55 (d, J = 8.6, 2H), 8.01 (d, J = 2.4, 1 H), 8.51 (d, J = 2.4, 1

H). HRMS (m/z): [MH⁺] calcd for C₂₂H₂₆ClN₂O₃401.1632; found

401.1629. [α]²_D⁰= −27.3 (c = 1.03, MeOH).

5-[2-Chloro-5-(triﬂuoromethyl)phenyl]-6-(cyclopentyloxy)-N-

[(1R,2R)-2-hydroxycyclohexyl]-3-pyridinecarboxamide (4). General

procedure D, with 10r (0.135 g, 0.3 mmol), was used to produce 4 (108

mg, 64%) as white solid. ¹H NMR (400 MHz, CDCl₃) δ 1.36 (m, 4 H),

1.67 (m, 8 H), 1.89 (m, 2 H), 2.11 (m, 2 H), 3.21 (d, J = 5.1, OH), 3.44

(m, 1 H), 3.85 (m, 1H), 5.52 (m, 1 H), 6.02 (d, J = 7, NH), 7.57 (m, 3H),

7.92 (d, J = 2.7, 1 H), 8.64 (d, J = 2.7, 1 H). HRMS (m/z): [MH⁺] calcd

for C₂₄H₂₇ClF₃N₂O₃483.1657; found 483.1674.

5-(4-Chlorophenyl)-6-(cyclohexyloxy)-N-[(1R,2R)-2-hydroxycyclo-

hexyl]-3-pyridinecarboxamide (14a). General procedure D, with 10q

(0.1 g, 0.3 mmol), was used to produce 14a (79 mg, 61%) as white solid.

¹H NMR (400 MHz, CDCl₃) δ 1.36 (m, 8 H), 1.56 (m, 2 H), 1.73 (m, 4

5-Bromo-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(2-methoxyethoxy)-

3-pyridinecarboxamide (13a). General Procedure C. To a solution of

12f (5.0 g, 18.1 mmol) in THF (65 mL) was added TBTU (6.4 g, 19.9

mmol), DIEA (15.5 mL, 90.6 mmol), and ﬁnally (1R,2R)-2-amino-

cyclohexanol hydrochloride (3.0 g, 19.9 mmol). The reaction was stirred

for 4 h at RT and afterward concentrated in vacuo. Puriﬁcation of the

resulting residue was by ﬂash chromatography, eluting with ethyl

acetate, aﬀorded 13a (6.5 g, 96%). ¹H NMR (400 MHz, CDCl₃) δ 1.32

(m, 5H), 1.76 (m, 2H), 2.10 (m, 2H), 3.46 (m, 1H), 3.46 (s, 3 H), 3.79

(t, J = 4.8, 2 H), 4.56 (t, J = 4.8, 2H), 5.14 (br s, OH), 6.20 (bd, J = 7,

H), 1.94 (m, 2 H), 2.11 (m, 2 H), 3.31 (br s, OH), 3.44 (m, 1 H), 3.84

(m, 1H), 5.23 (1, 1 H), 6.03 (m, NH), 7.39 (d, J = 8.0, 2 H), 7.52 (d, J =

8.0, 2H), 7.99 (s, 1 H), 8.52 (s, 1 H). HRMS (m/z): [MH⁺] calcd for

C₂₄H₃₀ClN₂O₃429.1945; found 429.1953.

5-(4-Chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(phe-

noxy)-3-pyridinecarboxamide (14b). General procedure D, with 10n

(80 mg, 0.25 mmol), was used to produce 14b (77 mg, 74%) as white

foam. ¹H NMR (400 MHz, CDCl₃) δ 1.36 (m, 4 H), 1.76 (m, 2 H), 2.10

(m, 2 H), 3.07 (br s, OH), 3.43 (m, 1 H), 3.84 (m, 1 H), 6.03 (m, NH),

7.11 (d, J = 7.8, 2 H), 7.23 (t, J = 7.4, 1 H), 7.41 (t, J = 8.3, 2 H), 7.44 (d, J

M

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

= 8.3, 2 H), 7.62 (d, J = 8.3, 2H), 8.17 (d, J = 2.4, 1 H), 8.47(d, J = 2.4, 1

H). HRMS (m/z): [MH⁺] calcd for C₂₄H₂₄ClN₂O₃423.1476; found

423.1475.

7.30 (t, J = 7.4, 1 H), 7.36 (7, J = 7.3, 2 H), 7.41 (d, J = 7.6, 2 H), 7.53 (d, J

= 8.7, 2 H), 7.69 (d, J = 8.7, 2 H), 8.21 (bd, J = 8.2, NH), 8.23 (d, J = 2.3,

1 H), 8.66 (d, J = 2.3, 1 H). HRMS (m/z): [M − H⁻] calcd for

C₂₅H₂₄ClN₂O₃435.1475; found 435.1483.

5-(4-Chlorophenyl)-6-(cyclopentyloxy)-N-[(1R,2R)-2-hydroxycy-

clohexyl]-3-pyridinecarboxamide (14c). General procedure D, with

10o (100 mg, 0.31 mmol), was used to produce 14c (82 mg, 63%) as

white solid. ¹H NMR (400 MHz, CDCl₃) δ 1.34 (m, 4 H), 1.58−1.84

(m, 8 H), 1.95 (m, 2 H), 2.11 (m, 2 H), 3.28 (d, J = 4.8, OH), 3.44 (m, 1

H), 3.85 (m, 1 H), 5.56 (m, 1 H), 6.02 (m, NH), 7.38 (d, J = 8.6, 2 H),

7.49 (d, J = 8.6, 2 H), 7.99 (d, J = 2.4, 1 H), 8.53(d, J = 2.4, 1 H). HRMS

(m/z): [MH⁺] calcd for C₂₃H₂₈ClN₂O₃415.1789; found 415.1790.

5-(4-Chlorophenyl)-6-cyclobutoxy-N-[(1R,2R)-2-hydroxycyclohex-

yl]-3-pyridinecarboxamide (14d). General procedure D, with 10j (210

mg, 0.69 mmol), was used to produce 14d (160 mg, 58%) as oﬀ-white

solid. ¹H NMR (600 MHz, DMSO-d₆) δ 1.22 (m, 4 H), 1.65 (m, 3 H),

1.77 (m, 1 H), 1.87 (m, 2 H), 2.05 (m, 2 H), 2.41 (m, 2 H), 3.41 (m, 1

H), 3.62 (m, 1 H), 4.69 (d, J = 4.9, OH), 5.27 (m, 1H), 7.55 (d, J = 8.7, 2

H), 7.69 (d, J = 8.7, 2 H), 8.21 (d, J = 2.3, 1 H), 8.23 (bd, J = 8, NH),

8.62(d, J = 2.3, 1 H). HRMS (m/z): [MH⁺] calcd for C₂₂H₂₆ClN₂O₃

401.1632; found 401.1629.

5-(4-Chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-[(3-meth-

yl-5-isoxazolyl)methoxy]-3-pyridinecarboxamide (14k). General pro-

cedure D, with 10l (58 mg, 0.168 mmol), was used to produce 14k (46

mg, 62%) as white solid. ¹H NMR (400 MHz, CDCl₃) δ 1.33 (m, 4 H),

1.78 (m, 2 H), 2.12 (m, 2 H), 2.28 (s, 3H), 3.44 (m, 1 H), 3.86 (m, 1 H),

5.52 (s, 2 H), 6.08 (s, 1 H), 6.12 (bd, J = 7, 1 H), 7.40 (d, J = 8.7, 2 H),

7.49 (d, J = 8.7, 2 H), 8.04 (d, J = 2.4, 1 H), 8.54 (d, J = 2.4, 1 H). HRMS

(m/z): [MH⁺] calcd for C₂₃H₂₅ClN₃O₄442.1534; found 442.1531.

5-(4-Chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(2-pyrimi-

dinylmethoxy)-3-pyridinecarboxamide (14l). General procedure D,

with 10d (9.25 g, 27.1 mmol), was used to produce 14l (8.5 g, 72%) as

white solid. ¹H NMR (400 MHz, CDCl₃) δ 1.32 (m, 4 H), 1.76 (m, 2

H), 2.10 (m, 2 H), 3.19 (d, J = 4.8, OH), 3.42 (m, 1 H), 3.83 (m, 1 H),

5.72 (s, 2 H), 6.04 (bd, J = 7.3, NH), 7.19 (t, J = 4.9, 1 H), 7.40 (d, J = 8.6,

2 H), 7.71 (d, J = 8.6, 2 H), 8.06 (d, J = 2.4, 1 H), 8.41 (d, J = 2.4, 1 H),

8.70 (d, J = 4.9, 2 H). [α]_D²⁰= −23.7 (MeOH). HRMS (m/z): [MH⁺]

calcd for C₂₃H₂₄ClN₄O₃439.1537; found 439.1533.

5-(4-Chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(4-pyridi-

nylmethoxy)-3-pyridinecarboxamide (14m). General procedure D,

with 10f (60 mg, 0.176 mmol), was used to produce 14m (59 mg, 77%)

as white solid. ¹H NMR (400 MHz, CDCl₃) δ 1.33 (m, 4 H), 1.77 (m, 2

H), 2.12 (m, 2 H), 3.99 (br s, OH), 3.44 (m, 1 H), 3.85 (m, 1 H), 5.51 (s,

2 H), 6.05 (bd, J = 6.7, NH), 7.24 (d, J = 6.0, 2 H), 7.44 (d, J = 8.6, 2 H),

7.54 (d, J = 8.6, 2 H), 8.06 (d, J = 2.4, 1 H), 8.52 (d, J = 2.4, 1 H), 8.57 (d,

J = 6.0, 2 H). HRMS (m/z): [MH⁺] calcd for C₂₄H₂₅ClN₃O₃438.1585;

found 438.1579.

5-(4-Chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(3-pyridi-

nylmethoxy)-3-pyridinecarboxamide (14n). General procedure E,

with 13c (99 mg, 0.244 mmol) and 4-chlorophenylboronic acid (43 mg,

0.276 mmol), was used to produce 14n (89 mg, 83%) as white foam. ¹H

NMR (400 MHz, CDCl₃) δ 1.34 (m, 4 H), 1.77 (m, 2 H), 2.11 (m, 2 H),

3.15 (d, J = 4.6, OH), 3.44 (m, 1 H), 3.85 (m, 1 H), 5.52 (s, 2 H), 6.07

(bd, J = 7, NH), 7.27 (dd, J = 8.0, 4.8, 1 H), 7.40 (d, J = 8.8, 2 H), 7.49 (d,

J = 8.8, 2 H), 7.70 (dt, J = 8.1, 1.9, 1 H), 8.04 (d, J = 2.4, 1 H), 8.55 (dd, J

= 4.8, 2.0, 1 H), 8.55 (d, J = 2.4, 1 H), 8.67 (bd, J = 1.9, 1 H). HRMS (m/

z): [M − H⁻] calcd for C₂₄H₂₃ClN₃O₃436.1428; found 436.1432.

5-(4-Chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(2-pyridi-

nylmethoxy)-3-pyridinecarboxamide (14o). General procedure D,

with 10e (100 mg, 0.293 mmol), was used to produce 14o (102 mg,

79%) as white solid. ¹H NMR (400 MHz, CDCl₃) δ 1.34 (m, 4 H), 1.77

(m, 2 H), 2.11 (m, 2 H), 3.18 (d, J = 5.1, OH), 3.44 (m, 1 H), 3.85 (m, 1

H), 5.61 (s, 2 H), 6.06 (bd, J = 7.0, NH), 7.20 (dd, J = 5.4, 7.3, 1 H), 7.29

(d, J = 7.8, 1 H), 7.41 (d, J = 8.6, 2 H), 7.58 (d, J = 8.6, 2 H), 7.65 (dt, J =

1.8, 7.8, 1 H), 8.08 (d, J = 2.4, 1 H), 8.52 (d, J = 2.4, 1 H), 8.58 (bd, J =

4.8, 1 H). HRMS (m/z): [MH⁺] calcd for C₂₄H₂₅ClN₃O₃438.1585;

found 438.1570.

6-(Cyclopropylmethoxy)-5-(2,4-dichlorophenyl)-N-[(1R,2R)-2-hy-

droxycyclohexyl]-3-pyridinecarboxamide (15a). General procedure

E, with 13b (100 mg, 0.270 mmol) and B-(2,4-dichlorophenyl)-boronic

acid (56.3 mg, 0.295 mmol), was used to produce 15a (49 mg, 42%) as

light-white solid. ¹H NMR (600 MHz, CDCl₃) δ 0.29 (m, 2 H), 0.52 (m,

2 H), 1.21 (m, 1H), 1.33 (m, 4 H), 1.77 (m, 2 H), 2.11 (m, 2 H), 3.27

(brd, OH), 3.44 (m, 1 H), 3.85 (m, 1 H), 4.23 (d, J = 7.0, 2 H), 6.03 (brd,

J = 7.0, NH), 7.24 (d, J = 8.2, 1 H), 7.26 (d, J = 2.1,1 H), 7.32 (dd, J = 8.2,

2.1, 1 H) 7.89 (d, J = 2.5, 1 H), 8.59 (d, J = 2.4, 1 H). HRMS (m/z):

[MH⁺] calcd for C₂₂H₂₅Cl₂N₂O₃435.1237; found 435.1244.

6-Butoxy-5-(4-chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-3-

pyridinecarboxamide (14e). General procedure D, with 10p (100 mg,

0.33 mmol), was used to produce 14e (95 mg, 72%) as light-yellow solid.

¹H NMR (400 MHz, CDCl₃) δ 0.95 (t, J = 7.4, 3 H), 1.25−1.45 (m, 6

H), 1.75 (m, 4 H), 2.11 (m, 2 H), 3.26 (d, J = 5.1, OH), 3.44 (m, 1 H),

3.85 (m, 1 H), 4.41 (t, J = 6.5, 1 H), 6.03 (m, NH), 7.40 (d, J = 8.6, 2 H),

7.51 (d, J = 8.6, 2 H), 8.00 (d, J = 2.4, 1 H), 8.52 (d, J = 2.4, 1 H). HRMS

(m/z): [MH⁺] calcd for C₂₂H₂₈ClN₂O₃403.1789; found 403.1789.

5-(4-Chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(1-methyl-

ethoxy)-3-pyridinecarboxamide (14f). General procedure A, with 19

(95.0 mg, 0.260 mmol) and 2-propanol (30 μL, 0.389 mmol), was used

1

to produce 14f (14 mg, 14%) as white solid. H NMR (600 MHz,

CDCl₃) δ 1.34(d, J = 6.0, 6 H), 1.35 (m, 5 H), 1.77 (m, 2 H), 2.11 (m, 2

H), 3.33 (d, J = 4.5, OH), 3.45 (m, 1 H), 3.85 (m, 1 H), 5.45 (spt, J = 6.0,

1 H), 6.03 (br. d, J = 6.2, NH), 7.39 (d, J = 8.6, 2 H), 7.51 (d, J = 8.6, 2

H), 7.99 (d, J = 2.4, 1 H), 8.53 (d, J = 2.5, 1 H). HRMS (m/z): [MH⁺]

calcd for C₂₁H₂₆ClN₂O₃389.1626; found 389.1635.

5-(4-Chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(2-me-

thoxyethoxy)-3-pyridinecarboxamide (14h). General Procedure E.

To a suspension of 13a (3.0 g, 8.06 mmol) in toluene (60 mL) was

added with stirring the [1,1-bis(diphenylphosphino)ferrocene] dichlor-

opalladium DCM complex (0.33 g, 0.403 mmol), 4-chlorophenylbor-

onic acid (1.33 g, 8.06 mmol), and sodium carbonate solution (8.1 mL,

2M). The resulting mixture was stirred at 90 °C for 16 h, cooled, and

partitioned between water and ethyl acetate. The organic portion was

dried over magnesium sulfate, ﬁltered, and concentrated. The crude

product was puriﬁed by ﬂash chromatography, eluting with ethyl

acetate/heptane (2:1) to produce 14h (1.79 g, 55%) as oﬀ-white solid;

mp 176−177 °C. ¹H NMR (400 MHz, CDCl₃) δ 1.34 (m, 4 H), 1.77

(m, 2 H), 2.11 (m, 2 H), 3.22 (d, J = 5.1, OH), 3.38 (s, 3 H), 3.44 (m, 1

H), 3.73 (t, J = 4.8, 2 H), 3.85 (m, 1 H), 4.57 (t, J = 4.8, 2 H), 6.05 (bd, J

= 7, NH), 7.39 (d, J = 8.6, 2 H), 7.55 (d, J = 8.6, 2 H), 8.02 (d, J = 2.4, 1

H), 8.52 (d, J = 2.4, 1 H). HRMS (m/z): [M − H⁻] calcd for

C₂₁H₂₄ClN₂O₄403.1425; found 403.1430. [α]²_D⁰= −26.2 (c = 1.00,

MeOH).

5-(4-Chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(2,2,2-tri-

ﬂuoroethoxy)-3-pyridinecarboxamide (14i). General procedure D,

with 10b (36.3 g, 109 mmol), was used to produce 14i (38.0 g, 81%) as

white solid; mp 186−187 °C. ¹H NMR (400 MHz, DMSO-d₆) δ 1.24

(m, 4 H), 1.65 (m, 2 H), 1.87 (m, 2 H), 3.40 (m, 1 H), 3.64 (m, 1 H),

4.66 (d, J = 4.8, OH), 5.11 (q, J = 9.0, 2 H), 7.58 (d, J = 8.6, 2 H), 7.66 (d,

J = 8.6, 2 H), 8.28 (bd, J = 8.1, NH), 8.31 (d, J = 2.4, 1 H), 8.66 (d, J = 2.4,

1 H). [α]²_D⁰= −23.6 (c = 1.00, MeOH). HRMS (m/z): [MH⁺] calcd for

C₂₀H₂₁ClF₃N₂O₃429.1193; found 429.1193.

5-(2-Chlorophenyl)-6-(cyclopropylmethoxy)-N-[(1R,2R)-2-hydrox-

ycyclohexyl]-3-pyridinecarboxamide (15b). General procedure E,

with 13b (100 mg, 0.271 mmol) and B-(2-chlorophenyl)-boronic acid

(46.6 mg, 0.298 mmol), was used to produce 15b (82 mg, 76%) as white

solid. ¹H NMR (600 MHz, CDCl₃) δ 0.29 (m, 2 H), 0.45 (m, 2 H), 1.21

(m, 1 H), 1.34 (m, 4 H), 1.76 (m, 2 H), 2.1 (m, 2 H), 3.36 (br s, OH),

3.43 (m, 1 H), 3.85 (m, 1 H), 4.24 (d, J = 7.0, 2 H), 6.04 (brd, J = 6.9,

NH), 7.32 (m, 3 H), 7.47 (m, 1H), 7.89 (d, J = 2.5, 1 H), 8.61 (d, J = 2.5,

1 H). HRMS (m/z): [MH⁺] calcd for C₂₂H₂₆ClN₂O₃401.1627; found

401.1639.

5-(4-Chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-6-(phenyl-

methoxy)-3-pyridinecarboxamide (14j). General procedure E, with

13e (85 mg, 0.210 mmol) and 4-chlorophenylboronic acid (37.2 mg,

0.231 mmol), was used to produce 14j (51 mg, 55%) as white solid. ¹H

NMR (600 MHz, DMSO-d₆) δ 1.23 (m, 4 H), 1.65 (m, 2 H), 1.87 (m, 2

H), 3.40 (m, 1 H), 3.64 (m, 1 H), 4.65 (d, J = 5.0, OH), 5.49 (s, 2 H),

N

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

6-(Cyclopropylmethoxy)-5-(3,4-dichlorophenyl)-N-[(1R,2R)-2-hy-

droxycyclohexyl]-3-pyridinecarboxamide (15c). General procedure E,

with 13b (49 mg, 0.134 mmol) and B-(3,4-dichlorophenyl)-boronic

acid (26 mg, 0.138 mmol), was used to produce 15c (35 mg, 60%) as oﬀ-

white solid. ¹H NMR (400 MHz, CDCl₃) δ 0.36 (m, 2 H), 0.61 (m, 2

H), 1.32 (m, 5 H), 1.78 (m, 2 H), 2.11 (m, 2 H), 3.30 (d, J = 5.0, OH),

3.46 (m, 1 H), 3.86 (m, 1 H), 4.27 (d, J = 7.1, 2 H), 6.08 (bd, J = 7, NH),

7.48 (m, 2 H), 7.75 (d, J = 1.9, 1 H), 8.03 (d, J = 2.6, 1 H), 8.52 (d, J = 2.4,

1 H). HRMS (m/z): [MH⁺] calcd for C₂₂H₂₅Cl₂N₂O₃435.1237; found

435.1243.

5-(3-Chlorophenyl)-6-(cyclopropylmethoxy)-N-[(1R,2R)-2-hydrox-

ycyclohexyl]-3-pyridinecarboxamide (15d). General procedure E,

with 13b (97 mg, 0.263 mmol) and B-(3-chlorophenyl)-boronic acid

(41 mg, 0.262 mmol), was used to produce 15d (53 mg, 50%) as white

solid. ¹H NMR (400 MHz, CDCl₃) δ 0.36 (m, 2 H), 0.60 (m, 2 H), 1.32

(m, 5 H), 1.78 (m, 2 H), 2.12 (m, 2 H), 3.32 (d, J = 4.8, OH), 3.43 (m, 1

H), 3.86 (m, 1 H), 4.28 (d, J = 7.1, 2 H), 6.04 (bd, J = 6.9, NH), 7.36 (m,

2 H), 7.49 (m, 1 H), 7.64 (m, 1 H), 8.02 (d, J = 2.4, 1 H), 8.54 (d, J = 2.4,

1 H). HRMS (m/z): [MH⁺] calcd for C₂₂H₂₆ClN₂O₃401.1626; found

401.1632.

5-[2-Chloro-5-(triﬂuoromethyl)phenyl]-6-(cyclopropylmethoxy)-

N-[(1R,2R)-2-hydroxycyclohexyl]-3-pyridinecarboxamide (15e).

General procedure D, with 10m (90 mg, 0.242 mmol), was used to

produce 15e (83.0 g, 73%) as white solid. ¹H NMR (400 MHz, CDCl₃)

δ 0.29 (m, 2 H), 0.52 (m, 2 H), 1.32 (m, 5 H), 1.77 (m, 2 H), 2.12 (m, 2

H), 3.18 (d, J = 5.1, OH), 3.45 (m, 1 H), 3.85 (m, 1 H), 4.24 (d, J = 7.0, 2

H), 6.03 (bd, J = 6.7, NH), 7.60 (m, 3 H), 7.94 (d, J = 2.4, 1 H), 8.62 (d, J

= 2.4, 1 H). HRMS (m/z): [MH⁺] calcd for C₂₃H₂₅ClF₃N₂O₃469.1506;

found 469.1504.

6-(Cyclopropylmethoxy)-5-(4-ﬂuorophenyl)-N-[(1R,2R)-2-hydrox-

ycyclohexyl]-3-pyridinecarboxamide (15f). General procedure D, with

10c (25.0 g, 87 mmol), was used to produce 15f (29.3 g, 88%) as white

solid. ¹H NMR (400 MHz, CDCl₃) δ 0.34 (m, 2 H), 0.58 (m, 2 H), 1.33

(m, 5 H), 1.76 (m, 2 H), 2.10 (m, 2 H), 3.44 (br s, OH), 3.44 (m, 1 H),

3.85 (m, 1 H), 4.26 (d, J = 7.0, 2 H), 6.11 (bd, J = 7.0, NH), 7.12 (t, J =

8.7, 2 H), 7.58 (dd, J = 5.5, 8.9, 2 H), 8.00 (d, J = 2.4, 1 H), 8.50 (d, J =

2.4, 1 H). [α]²_D⁰= −16.4 (MeOH). HRMS (m/z): [MH⁺] calcd for

C₂₂H₂₆FN₂O₃385.1928; found 385.1925.

H), 1.33 (m, 5 H), 1.77 (m, 2 H), 2.11 (m, 2 H), 3.15 (d, J = 5.1, OH),

3.45 (m, 1 H), 3.85 (m, 1 H), 4.28 (d, J = 7.3, 2 H), 6.09 (bd, J = 7.3,

NH), 7.72 (s, 4 H), 8.06 (d, J = 2.4, 1 H), 8.55 (d, J = 2.4, 1 H). HRMS

(m/z): [MH⁺] calcd for C₂₃H₂₆N₃O₃392.1974; found 392.1974.

6-(Cyclopropylmethoxy)-N-[(1R,2R)-2-hydroxycyclohexyl]-5-(4-

methanesulfonylamino-phenyl)-3-pyridinecarboxamide (15k). Gen-

eral procedure E, with 13b (99.1 mg, 0.268 mmol) and B-[4-

[(methylsulfonyl)amino]phenyl]-boronic acid (87.8 mg, 0.408 mmol)

in DME with Cs₂CO₃as base and tetrakis(triphenylphosphine)-

palladium(0), was used to produce 15k (86.2 mg, 70%) as white

solid. ¹H NMR (600 MHz, DMSO-d₆) δ 0.34 (m, 2 H), 0.53 (m, 2 H),

1.25 (m, 5 H), 1.64 (m, 2 H), 1.87 (m, 2 H), 3.05 (s, 3 H), 3.39 (m, 1 H),

3.63 (m, 1 H), 4.24 (d, J = 7.1, 2 H), 4.63 (d, J = 5.1, OH), 7.30 (d, J =

8.0, 2 H), 7.64 (d, J = 8.0, 2 H), 8.16 (d, J = 8.1, NH), 8.18 (d, J = 2.3, 1

H), 8.58 (d, J = 2.3, 1 H), 9.90 (s, 1 H). HRMS (m/z): [MH⁺] calcd for

C₂₃H₃₀N₃O₅S 460.1898; found 460.1909.

6-(Cyclopropylmethoxy)-N-[(1R,2R)-2-hydroxycyclohexyl]-5-(4-

sulfamoyl-phenyl)-3-pyridinecarboxamide (15l). General procedure

E, with 13b (99.5 mg, 0.269 mmol) and B-(4-sulfamoylphenyl)-boronic

acid (81.9 mg, 0.407 mmol) in DME with Cs₂CO₃as base and

tetrakis(triphenylphosphine)palladium(0), was used to produce 15k

(99.6 mg, 83%) as white solid. ¹H NMR (600 MHz, DMSO-d₆) δ 0.34

(m, 2 H), 0.53 (m, 2 H), 1.25 (m, 5 H), 1.65 (m, 2 H), 1.87 (m, 2 H),

3.40 (m, 1 H), 3.64 (m, 1 H), 4.25 (d, J = 7.1, 2 H), 4.64 (d, J = 5.0, OH),

7.43 (s, 2 H), 7.85 (d, J = 9.0, 2 H), 7.91 (d, J = 9.0, 2 H), 8.19 (d, J = 8.2,

NH), 8.25 (d, J = 2.3, 1 H), 8.65 (d, J = 2.3, 1 H). HRMS (m/z): [MH⁺]

calcd for C₂₂H₂₈N₃O₅S 446.1744; found 446.1755.

4-[2-Cyclopropylmethoxy-5-((1R,2R)-2-hydroxy-cyclohexylcarba-

moyl)-pyridin-3-yl]-benzoic Acid (15m). General procedure E, with

13b (99.4 mg, 0.269 mmol) and 4-boronobenzoic acid (67.6 mg, 0.407

mmol) in DME with Cs₂CO₃as base, was used to produce 15k (95.1

mg, 86%) as white solid. ¹H NMR (600 MHz, DMSO-d₆) δ 0.34 (m, 2

H), 0.53 (m, 2 H), 1.25 (m, 5 H), 1.64 (m, 2 H), 1.87 (m, 2 H), 3.39 (m,

1 H), 3.64 (m, 1 H), 4.25 (d, J = 7.1, 2 H), 4.65 (d, J = 4.9, OH), 7.80 (d, J

= 8.5, 2 H), 8.03 (d, J = 8.5, 2 H), 8.20 (d, J = 8.2, NH), 8.27 (d, J = 2.4, 1

H), 8.65 (d, J = 2.3, 1 H), 13.0 (br s, 1 H). HRMS (m/z): [MH⁺] calcd

for C₂₃H₂₇N₂O₅411.1914; found 411.1921.

6-(Cyclopropylmethoxy)-5-(4-triﬂuoromethylphenyl)-N-[(1R,2R)-

2-hydroxycyclohexyl]-3-pyridinecarboxamide (15g). General proce-

dure E, with 13b (100 mg, 0.271 mmol) and B-(4-triﬂuorophenyl)-

boronic acid (56.6 mg, 0.298 mmol), was used to produce 15g (85 mg,

72%) as white solid. ¹H NMR (600 MHz, CDCl₃) δ 0.34 (m, 2 H), 0.59

(m, 2 H), 1.32 (m, 5 H), 1.78 (m, 2 H), 2.12 (m, 2 H), 3.25 (br d, J = 3.8,

OH), 3.45 (m, 1 H), 3.86 (m, 1 H), 4.28 (d, J = 7.2, 2 H), 6.06 (br d, J =

7.2, NH), 7.69 (d, J = 7.9, 2 H), 7.73 (d, J = 7.9, 2 H), 8.06 (d, J = 2.4, 1

H), 8.55 (d, J = 2.4, 1 H). HRMS (m/z): [MH⁺] calcd for

C₂₃H₂₆F₃N₂O₃435.1906; found 435.1906.

6-(Cyclopropylmethoxy)-N-[(1R,2R)-2-hydroxycyclohexyl]-5-[4-

(triﬂuoromethoxy)phenyl]-3-pyridinecarboxamide (15h). General

procedure D, with 10h (428 mg, 1.211 mmol), was used to produce

15h (498 mg, 91%) as colorless foam. ¹H NMR (600 MHz, CDCl₃) δ

0.34 (m, 2 H), 0.59 (m, 2 H), 1.32 (m, 5 H), 1.77 (m, 2 H), 2.12 (m, 2

H), 3.30 (bd, J = 5.0, OH), 3.44 (m, 1 H), 3.86 (m, 1 H), 4.27 (d, J = 7.1,

2 H), 6.06 (bd, J = 7.1, NH), 7.28 (d, J = 8.8, 2 H), 7.65 (d, J = 8.8, 2 H),

8.03 (d, J = 2.4, 1 H), 8.52 (d, J = 2.4, 1 H). HRMS (m/z): [MH⁺] calcd

for C₂₃H₂₆F₃N₂O₄451.1839; found 451.1857.

6-(Cyclopropylmethoxy)-N-[(1R,2R)-2-hydroxycyclohexyl]-5-(4-

methoxyphenyl)-3-pyridinecarboxamide (15i). General procedure E,

with 13b (100 mg, 0.271 mmol) and B-(4-methoxyphenyl)-boronic acid

(45.3 mg, 0.298 mmol), was used to produce 15i (88 mg, 7482%) as

white solid. ¹H NMR (600 MHz, CDCl₃) δ 0.35 (m, 2 H), 0.58 (m, 2

H), 1.33 (m, 5 H), 1.77 (m, 2 H), 2.10 (m, 2 H), 3.43 (br s, OH), 3.44

(m, 1 H), 3.85 (m, 1 H), 3.86 (s, 3H), 4.26 (d, J = 7.1, 2 H), 6.04 (br d, J

= 7.1, NH), 6.97 (d, J = 8.3, 2 H), 7.57 (d, J = 8.3, 2 H), 7.99 (d, J = 2.5, 1

H), 8.48 (d, J = 2.5, 1 H). HRMS (m/z): [MH⁺] calcd for C₂₃H₂₉N₂O₄

397.2122; found 397.2133.

5-(4-Carbamoyl-phenyl)-6-(cyclopropylmethoxy)-N-[(1R,2R)-2-

hydroxycyclohexyl]-3-pyridinecarboxamide (15n). General proce-

dure E, with 13b (101.4 mg, 0.274 mmol) and B-(4-carbamoylphen-

yl)-boronic acid (67.8 mg, 0.411 mmol) in DME with Cs₂CO₃as base

and tetrakis(triphenylphosphine)palladium(0), was used to produce

15k (89.8 mg, 80%) as white solid. ¹H NMR (600 MHz, DMSO-d₆) δ

0.34 (m, 2 H), 0.53 (m, 2 H), 1.25 (m, 5 H), 1.65 (m, 2 H), 1.87 (m, 2

H), 3.40 (m, 1 H), 3.64 (m, 1 H), 4.25 (d, J = 7.1, 2 H), 4.65 (d, J = 5.0,

OH), 7.42 (br s, 1H), 7.75 (d, J = 8.5, 2 H), 7.97 (d, J = 8.5, 2 H), 8.04

(br s, 1 H), 8.20 (d, J = 8.2, NH), 8.25 (d, J = 2.3, 1 H), 8.63(d, J = 2.3, 1

H). HRMS (m/z): [MH⁺] calcd for C₂₃H₂₈N₃O₄410.2074; found

410.2082.

5-(4-Chloro-phenyl)-6-cyclopropylmethoxy-N-piperidin-1-yl-3-

pyridinecarboxamide (16a). General procedure D, with 10a (70 mg,

0.230 mmol) and 1-amino-piperidine (28 μL, 0.261 mmol), was used to

produce 16a (66 mg, 74%) as white solid. ¹H NMR (400 MHz, CDCl₃)

δ 0.34 (m, 2 H), 0.55 (m, 2 H), 1.27 (m, 1 H), 1.46 (br s, 2 H), 1.77 (br s,

4 H), 2.88 (br s, 4 H), 4.26 (d, J = 7, 1 H), 6.69 (br s, NH), 7.40 (d, J =

8.6, 2 H), 7.56 (d, J = 8.6, 2 H), 7.99 (br s, 1 H), 8.45 (br s, 1 H) major

rotamer. HRMS (m/z): [MH⁺] calcd for C₂₁H₂₅ClN₃O₂386.1635;

found 386.1634.

5-(4-Chlorophenyl)-N-[(2R)-2-cyclopropyl-2-hydroxypropyl]-6-

(cyclopropylmethoxy)-3-pyridinecarboxamide (16b). General proce-

dure D, with 10a (100 mg, 0.329 mmol) and (αR)-α-(aminomethyl)-α-

methyl-cyclopropanemethanol (42 mg, 0.365 mmol), was used to

1

produce 16b (124 mg, 94%) as white solid. H NMR (400 MHz,

CDCl₃) δ 0.34 (m, 3 H), 0.42 (m, 2 H), 0.50 (m, 1 H), 0.58 (m, 2 H),

0.93 (m, 1 H), 1.18 (s, 3 H), 1.28 (m, 1 H), 2.05 (s, OH), 3.50 (dd, J =

13.7, 5.4, 1 H), 3.64 (dd, J = 13.7, 6.4 1 H), 4.27 (d, J = 7, 2 H), 6.50 (br s,

NH), 7.41 (d, J = 8.6, 2 H), 7.56 (d, J = 8.6, 2 H), 8.04 (d, J = 2.4, 1 H),

8.53 (d, J = 2.4, 1 H). [α]²_D⁰= 1.1 (c = 0.558, MeOH). HRMS (m/z):

[MH⁺] calcd for C₂₂H₂₆ClN₂O₃401.1632; found 401.1631.

5-(4-Cyanophenyl)-6-(cyclopropylmethoxy)-N-[(1R,2R)-2-hydrox-

ycyclohexyl]-3-pyridinecarboxamide (15j). General procedure D, with

10g (200 mg, 0.679 mmol), was used to produce 15j (192 mg, 72%) as

white solid. ¹H NMR (400 MHz, CDCl₃) δ 0.33 (m, 2 H), 0.59 (m, 2

O

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

5-(4-Chlorophenyl)-6-(cyclopropylmethoxy)-N-[(1S,2S)-2-hydrox-

ycyclohexyl]-3-pyridinecarboxamide (16c). General procedure D,

with 10a (50 mg, 0.166 mmol) and (1S,2S)-2-amino-cyclohexanol

hydrochloride (27.6 mg, 0.182 mmol), was used to produce 16c (50 mg,

75%) as white solid. ¹H NMR (600 MHz, DMSO-d₆) δ 0.33 (m, 2 H),

0.53 (m, 2 H), 1.23 (m, 5 H), 1.63 (m, 2 H), 1.84 (m, 2 H), 3.39 (m, 1

H), 3.64 (m, 1H), 4.24 (d, J = 7.1, 2 H), 4.64 (d, J = 5, OH), 7.55 (d, J =

8.9, 2 H), 7.69 (d, J = 8.9, 2 H), 8.18 (bd, J = 8.3, NH), 8.21 (d, J = 2.4, 1

H), 8.62 (d, J = 2.4, 1 H). [α]_D²⁰= 26.3 (c = 1.0, MeOH). HRMS (m/z):

[MH⁺] calcd for C₂₂H₂₆ClN₂O₃401.1627; found 401.1631.

3.62 (m, 1 H), 4.60 (d, J = 5.1, OH), 7.50 (d, J = 8.7, 2 H), 7.54 (d, J =

8.7, 2 H), 7.91 (d, J = 2.4, 1 H), 8.00 (bd, J = 8.1, NH), 8.61(d, J = 2.4, 1

H). HRMS (m/z): [MH⁺] calcd for C₂₃H₂₉ClN₃O₂414.1948; found

414.1947.

6-Chloro-5-(4-chlorophenyl)-N-[(1R,2R)-2-hydroxycyclohexyl]-3-

pyridinecarboxamide (19). Following a procedure similar to the

preparation of 14g, 9a (1.4 g, 5.2 mmol) was used to produce 19 (1.4 g,

73%) as white solid. ¹H NMR (600 MHz, CDCl₃) 1.34 (m, 4 H), 1.78

(m, 2 H), 2.13 (m, 2 H), 2.71 (d, J = 5.7, OH), 3.46 (m, 1 H), 3.86 (m, 1

H), 6.31 (bd, J = 7.0, NH), 7.39 (d, J = 8.7, 2 H), 7.46 (d, J = 8.7, 2 H),

8.07 (d, J = 2.4, 1 H), 8.73 (d, J = 2.4, 1 H). HRMS (m/z): [MH⁺] calcd

for C₁₈H₁₉Cl₂N₂O₂365.0818; found 365.0825.

5-(4-Chlorophenyl)-6-(cyclopropylmethoxy)-N-[(1R,2S)-2-hydrox-

ycyclohexyl]-3-pyridinecarboxamide (16d). To a suspension of 10a

(1.5 g, 4.94 mmol) in DCM (1.5 mL) was added 1-chloro-N,N,2-

trimethyl-1-propen-1-amine (0.75 mL, 5.68 mmol) dropwise. The

mixture was stirred for 30 min at room temperature and added to a

solution of (1S,2R)-2-amino-cyclohexanol hydrochloride (0.94 g, 6.17

mmol) and DIEA (2.05 mL, 12.4 mmol) in DMF (2.0 mL). The

reaction was stirred for 1 h at room temperature and afterward

partitioned between ethyl acetate and citric acid solution (1 M). Organic

phases were combined, dried with magnesium sulfate, ﬁltered, and

concentrated. The resulting residue was puriﬁed by ﬂash chromato-

graphy, eluting with ethyl acetate/heptane (1:20 to 1:1) to aﬀord 16d

(1.88 g, 95%) as white solid. ¹H NMR (600 MHz, DMSO-d₆) δ 0.33 (m,

2H), 0.53 (m, 2H), 1.24 (m, 1 H), 1.31 (m, 2 H), 1.47 (m, 2 H), 1.54 (m,

1 H), 1.66 (m, 1 H), 1.74 (m, 2 H), 3.86 (m, 2 H), 4.24 (d, J = 7, 2H),

4.64 (d, J = 3.9, OH), 7.54 (d, J = 8.7, 2 H), 7.70 (d, J = 8.7, 2H), 8.04

(bd, J = 7.7, NH), 8.23 (d, J = 2.4, 1 H), 8.62 (d, J = 2.4, 1 H). HRMS (m/

5-(4-Chlorophenyl)-6-(2-cyclopropylethynyl)-N-[(1R,2R)-2-hy-

droxycyclohexyl]-3-pyridinecarboxamide (20). To a solution of 19

(0.47 g, 1.28 mmol) in DMF (8 mL) was added with stirring

bis(triphenylphosphine)palladium(II)dichloride (54 mg, 0.077 mmol),

copper(I)iodide (15 mg, 0.077 mmol), triphenylphosphine resin (174

mg, 1.48 mmol/g), ethynylcyclopropane (0.13 mL, 1.54 mmol), and

N,N-diethylamine (2 mL). The resulting mixture was microwaved twice

for 25 min at 120 °C, cooled, ﬁltered, and partitioned between water and

ethyl acetate. The organic portion was washed with water and sodium

bicarbonate solution, dried over magnesium sulfate, ﬁltered, and

concentrated. The crude product was puriﬁed by ﬂash chromatography,

eluting with ethyl acetate/heptane gradient to produce 20 (0.31 g, 61%)

as oﬀ-white solid. ¹H NMR (600 MHz, CDCl₃) δ 0.75 (m, 2H), 0.87 (m,

2 H), 1.35 (m, 5 H), 1.77 (m, 2 H), 2.12 (m, 2 H), 3.10 (d, J = 5.3, OH),

3.46 (m, 1 H), 3.87 (m, 1 H), 6.30 (bd, J = 7.3, NH), 7.42 (d, J = 8.7, 2

H), 7.51 (d, J = 8.7, 2 H), 8.05 (d, J = 2.1, 1 H), 8.84 (d, J = 2.1, 1 H).

HRMS (m/z): [MH⁺] calcd for C₂₃H₂₄ClN₂O₂395.1521; found

395.1528.

5-(4-Chlorophenyl)-6-(2-cyclopropylethyl)-N-[(1R,2R)-2-hydroxy-

cyclohexyl]-3-pyridinecarboxamide (21). To a solution of 20 (0.247 g,

0.63 mmol) in ethyl acetate (35 mL) was added palladium on carbon

(45 mg, 10%). The resulting mixture was hydrogenated at 0.4 bar and

room temperature, palladium was removed by ﬁltration over diatoma-

ceous earth, and the solution was concentrated to give 21 (0.19 g, 76%)

as yellow foam. ¹H NMR (400 MHz, CDCl₃) δ −0.10 (m, 2 H), 0.31 (m,

2H), 1.20 (m, 1H), 1.32 (m, 4 H), 1.53 (m, 2H), 1.76 (m, 2 H), 2.11 (m,

2 H), 2.88 (t, J = 7.6, 2H), 3.17 (d, J = 5.1, OH), 3.45 (m, 1 H), 3.85 (m,

1 H), 6.20 (bd, J = 7, NH), 7.25 (d, J = 8.3, 2 H), 7.43 (d, J = 8.3, 2 H),

7.88 (d, J = 2.4, 1 H), 8.90 (d, J = 2.4, 1 H). HRMS (m/z): [MH⁺] calcd

for C₂₃H₂₈ClN₂O₂399.1834; found 399.1837.

3-Bromo-2-(cyclopropylmethoxy)-5-methyl-pyridine (23a). To a

solution of cyclopropane−methanol (2.15 mL, 26.6 mmol) in DMF (50

mL) was added sodium hydride dispersion (1.16 g, ∼55% in oil, 26.6

mmol). The resulting suspension was stirred for 1 h at room

temperature, 3-bromo-2-chloro-5-methyl-pyridine (22, Combi-Blocks,

5.0 g, 24.2 mmol) was added, and the mixture was stirred for 6 h at 70

°C. After cooling, the reaction mixture was poured onto saturated

sodium bicarbonate solution (1200 mL) and partitioned into diethyl

ether and all organic phases were, after a ﬁnal wash with sodium

bicarbonate, combined, dried with Na₂SO₄, ﬁltered, and concentrated.

The residue was puriﬁed by ﬂash chromatography, eluting with

heptane/ethyl acetate (8:1) to aﬀord 23a (3.1 g, 53%) as reddish oil.

¹H NMR (400 MHz, CDCl₃) δ 0.38 (m, 2H), 0.59 (m, 2H), 1.31 (m,

z): [MH⁺] calcd for C₂₂H₂₆ClN₂O₃401.1632; found 401.1628. [α]_D²⁰

+34.0 (c = 0.992, MeOH).

=

5-(4-Chloro-phenyl)-6-cyclopropylmethoxy-N-((1S,2R)-2-hy-

droxy-cyclohexyl)-3-pyridinecarboxamide (16e). Following a proce-

dure similar to the preparation of 16d, 10a (1.50 g, 4.94 mmol) and

(1R,2S)-2-amino-cyclohexanol hydrochloride (0.94 g, 6.17 mmol) were

used to produce 16e (1.88 g, 95%) as white solid. ¹H NMR (600 MHz,

DMSO-d₆) δ 0.33 (m, 2H), 0.53 (m, 2H), 1.24 (m, 1 H), 1.31 (m, 2 H),

1.47 (m, 2 H), 1.55 (m, 1 H), 1.66 (m, 1 H), 1.75 (m, 2 H), 3.86 (m, 2

H), 4.24 (d, J = 7, 2H), 4.64 (d, J = 3.9, OH), 7.54 (d, J = 8.7, 2 H), 7.70

(d, J = 8.7, 2H), 8.04 (bd, J = 7.7, NH), 8.23 (d, J = 2.4, 1 H), 8.62 (d, J =

2.4, 1 H). HRMS (m/z): [MH⁺] calcd for C₂₂H₂₆ClN₂O₃401.1632;

found 401.1624. [α]_D²⁰= −32.3 (c = 0.988, MeOH).

5-Bromo-6-[(cyclopropylmethyl)methylamino]-3-pyridinecarbox-

ylic Acid. A mixture of 7 (1.00 g, 4.0 mmol), N-methyl-cyclo-

propanemethanamine hydrochloride (1:1) (1.21 g, 10 mmol), and

DBU (3.07 g, 20 mmol) was stirred for 3 h at 90 °C. Sodium hydroxide

solution (2 N, 4 mL) was added, and heating continued for 15 min. The

mixture was cooled, brought to a pH of ∼4 with hydrochloric acid (2 N,

4 mL) and citric acid (10%), and partitioned into ethyl acetate. The

organic portion was dried over magnesium sulfate, ﬁltered, and

concentrated. The crude product was puriﬁed by ﬂash chromatography,

eluting with a DCM/methanol gradient to produce the title compound

(0.83 g, 73%) as colorless solid. ¹H NMR (600 MHz, DMSO-d₆) δ 0.20

(m, 2 H), 0.48 (m, 2 H), 1.08 (m, 1 H), 3.10 (s, 3H), 3.40 (d, J = 6.8, 2

H), 8.18 (d, J = 2.0, 1 H), 8.63 (d, J = 2.0, 1 H), 12.80 (br s, 1 H). HRMS

(m/z): [MH⁺] calcd for C₁₁H₁₄BrN₂O₂285.0233, 287.0214; found

285.0239, 287.0220.

1H), 2.23 (s, 3 H), 4.18 (d, J = 7, 2H), 7.64 (d, J = 1.9, 1 H), 7.85 (d, J =

1.9, 1 H). HRMS (m/z): [MH⁺] calcd for C₁₀H₁₃BrNO 242.0175,

244.0155; found 242.0181, 244.0160.

3-Bromo-5-methyl-2-phenoxy-pyridine (23b). Following a proce-

dure similar to the preparation of 23a, phenol (1.33 g, 14.1 mmol) and 3-

bromo-2-chloro-5-methyl-pyridine (3.77 g, 18.2 mmol) were used to

produce 23b (1.42 g, 42%) as colorless oil. ¹H NMR (300 MHz, CDCl₃)

δ 2.27 (s, 3 H), 7.12 (d, J = 7.5, 2 H), 7.20 (t, J = 7.4, 1 H), 7.40 (t, J = 7.7,

2 H), 7.76 (s, 1 H), 7.88 (s, 1 H).

3-Bromo-2-cyclopentyloxy-5-methyl-pyridine (23c). Following a

procedure similar to the preparation of 23a, cyclopentanol (1.45 mL,

16.0 mmol) and 3-bromo-2-chloro-5-methyl-pyridine (3.0 g, 14.5

mmol) were used to produce 23c (1.75 g, 47%) as reddish oil. ¹H NMR

(400 MHz, CDCl₃) δ 1.62 (m, 2 H), 1.83 (m, 4 H), 1.94 (m, 2 H), 2.22

(s, 3 H), 5.40 (m, 1 H), 7.61 (d, J = 2.2, 1 H), 7.87 (d, J = 2.2, 1 H).

5-(4-Chlorophenyl)-6-[(cyclopropylmethyl)methylamino]-3-pyri-

dinecarboxylic A (17). Following a procedure similar to the preparation

of 10h, 5-bromo-6-[(cyclopropylmethyl)methylamino]-3-pyridinecar-

boxylic acid (0.40 g, 1.40 mmol) and B-4-chlorophenyl-boronic acid

(0.249 g, 1.54 mmol) were used to produce 17 (0.323 g, 73%) as

brownish foam. ¹H NMR (400 MHz, DMSO-d₆) δ 0.05 (m, 2 H), 0.38

(m, 2 H), 0.92 (m, 1 H), 2.74 (s, 3 H), 3.11 (m, 2 H), 7.48 (m, 4 H), 7.81

(d, J = 2.2, 1 H), 8.63 (d, J = 2.2, 1 H), 12.79 (br s, 1 H). MS (ESI) m/z =

315.3 [M − H⁻].

5-(4-Chlorophenyl)-6-[(cyclopropylmethyl)methylamino]-N-

[(1R,2R)-2-hydroxycyclohexyl]-3-pyridinecarboxamide (18). General

procedure D, with 17 (120 mg, 0.379 mmol), was used to produce 18

1

(133 mg, 85%) as colorless foam. H NMR (600 MHz, DMSO-d₆) δ

0.03 (m, 2 H), 0.37 (m, 2 H), 0.89 (m, 1 H), 1.21 (m, 4 H), 1.64 (m, 2

H), 1.86 (m, 2 H), 2.76 (s, 3 H), 3.05 (d, J = 6.8, 2 H), 3.38 (m, 1 H),

P

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

HRMS (m/z): [MH⁺] calcd for C₁₁H₁₅BrNO 256.0332, 258.0312;

found 256.0338, 258.0317.

with Na₂SO₄, ﬁltered, and concentrated. The solid residue was digested

with dichloromethane to aﬀord 26 (12.0 g, 65%) as oﬀ-white solid. ¹H

NMR (400 MHz, DMSO-d₆) δ 4.40 (d, J = 5.7, 2H), 5.31 (t, J = 5.8,

OH), 7.27 (d, J = 8.2, 1 H), 7.33 (d, J = 8.2, 1 H), 10.45 (br s, 1 H).

HRMS (m/z): [MH⁺] calcd for C₆H₇ClNO₂160.0160; found 160.0161.

6-Chloro-5-hydroxy-4-iodo-2-pyridinemethanol (27). To a solu-

tion of 26 (11.6 g, 72.7 mmol) in water (200 mL) was added sodium

bicarbonate (18.3 g, 218 mmol). The resulting suspension was stirred

for 15 min (foam formation) cooled to 0 °C, and iodide (19.4 g, 76.3

mmol) was added. The mixture was stirred at 0 °C for 18 h and afterward

allowed to reach room temperature. Sodium hydrogen sulﬁte solution

(2 N, 20 mL) was added slowly with cooling, and the pH was adjusted

carefully to 3 with hydrochloric acid (2 N). The product was partitioned

into ethyl acetate; all organic phases were combined, dried with MgSO₄,

ﬁltered, and concentrated. 27 (11.4 g; mp 142−144 °C) crystallized

from the concentrated solution and additional material (3.1 g, 70%

total) could be obtained by chromatography (heptane/ethyl acetate

2:1) of the residue obtained from the mother liquor as white solid. ¹H

NMR (400 MHz, DMSO-d₆) δ 4.40 (s, 2H), 5.42 (br s, OH), 7.75 (s, 1

H), 10.34 (br s, 1 H). HRMS (m/z): [MH⁺] calcd for C₆H₆ClINO₂

285.9126; found 285.9133.

6-Chloro-5-(cyclopropylmethoxy)-4-iodo-2-pyridinemethanol

(28a). To a solution of 27 (45.0 g, 158 mmol) in DMF (450 mL) was

added sodium hydride dispersion (6.3 g, ∼60% in oil, 158 mmol) in

portions. The resulting suspension was stirred for 30 min at room

temperature, (bromomethyl)cyclopropane (19.3 mL, 189 mmol,

dropwise) was added, and the mixture was stirred at 90 °C for 4 h.

Solvent was removed in high vacuum, ice−water (800 mL) was added,

and the mixture was extracted with ethyl acetate. Organic layers were

washed with water and brine, combined, dried over Na₂SO₄, ﬁltered, and

concentrated to a volume of ∼200 mL. n-Heptane (150 mL) was added

with stirring. The product precipitated, was ﬁltered, washed (ethyl

acetate/n-heptane 1:1), and dried to give 28a (46.1 g, 86%) as oﬀ-white

solid. ¹H NMR (600 MHz, CDCl₃) δ 0.41 (m, 2H), 0.67 (m, 2 H), 1.41

(m, 1 H), 2.66 (br s, OH), 3.90 (d, J = 7.3, 2 H), 4.67 (br s, 2H), 7.71 (s,

1 H). HRMS (m/z): [MH⁺] calcd for C₁₀H₁₂ClINO₂339.9601; found

339.9592.

3-Bromo-2-butoxy-5-methyl-pyridine (23d). Following a procedure

similar to the preparation of 23a, 1-butanol (2.44 mL, 26.6 mmol) and 3-

bromo-2-chloro-5-methyl-pyridine (5.0 g, 24.2 mmol) were used to

produce 23d (4.20 g, 71%) as reddish oil. ¹H NMR (400 MHz, CDCl₃)

δ 0.98 (t, J = 7.5, 3 H), 1.50 (m, 2 H), 1.78 (m, 2 H), 2.23 (s, 3 H), 4.32

(t, J = 6.5, 2 H), 7.63 (d, J = 2.2, 1 H), 7.87 (d, J = 2.2, 1 H). HRMS (m/

z): [MH⁺] calcd for C₁₀H₁₅BrNO 244.0332, 246.0312; found 244.0340,

246.0320.

3-Bromo-2-cyclohexyloxy-5-methyl-pyridine (23e). Following a

procedure similar to the preparation of 23a, cyclohexanol (2.84 mL,

26.6 mmol) and 3-bromo-2-chloro-5-methyl-pyridine (5.0 g, 24.2

mmol) were used to produce 23e (3.1 g, 47%) as reddish oil. ¹H NMR

(400 MHz, CDCl₃) δ 1.40 (m, 1 H), 1.45 (m, 2 H), 1.55 (m, 1 H), 1.62

(m, 2 H), 1.80 (m, 2 H), 1.93 (m, 2 H), 2.22 (s, 3 H), 5.06 (m, 1 H), 7.62

(d, J = 2.2, 1 H), 7.85 (d, J = 2.2, 1 H). HRMS (m/z): [MH⁺] calcd for

C₁₂H₁₇BrNO 270.0488, 272.0468; found 270.0491, 272.0471.

3-(2-Chloro-5-(triﬂuoromethyl)phenyl)-2-cyclopropylmethoxy-5-

methyl-pyridine (24a). To a solution of 23a (1.0 g, 4.1 mmol) in

toluene (7 mL) was added with stirring the [1,1-bis-

(diphenylphosphino)ferrocene] dichloropalladium DCM complex

(0.169 g, 0.02 mmol), B-[2-chloro-5-(triﬂuoromethyl)phenyl]-boronic

acid (1.39 g, 6.2 mmol), and sodium carbonate solution (6.2 mL, 2M).

The resulting mixture was stirred at 90 °C overnight, cooled, and eluted

over Chem Elut (Varian) with ethyl acetate. The solution was

concentrated and puriﬁed by ﬂash chromatography, eluting with an

ethyl acetate/heptane gradient to produce 24a (1.21 g, 86%) as

yellowish oil. ¹H NMR (400 MHz, CDCl₃) δ 0.26 (m, 2 H), 0.49 (m, 2

H), 1.18 (m, 1 H), 2.30 (s, 3 H), 4.15 (d, J = 7.0, 1 H), 7.36 (d, J = 2.4, 1

H), 7.57 (m, 3 H), 8.01 (d, J = 2.4, 1 H). HRMS (m/z): [MH⁺] calcd for

C₁₇H₁₆ClF₃NO 342.0867; found 342.0870.

3-(4-Chloro-phenyl)-5-methyl-2-phenoxy-pyridine (24b). Follow-

ing a procedure similar to the preparation of 24a, 23b (0.79 g, 2.99

mmol) and B-[4-chloro-phenyl]-boronic acid (0.70 g, 4.49 mmol) were

used to produce 24b (0.61 g, 68%) as yellowish oil. ¹H NMR (300 MHz,

CDCl₃) δ 2.33 (s, 3 H), 7.07 (d, J = 8.0, 2 H), 7.15 (t, J = 8.0, 1 H), 7.35

(t, J = 8.0, 2 H), 7.40 (d, J = 8.6, 2 H), 7.55 (s, 1 H), 7.57 (d, J = 8.6, 2 H),

7.98 (s, 1 H). MS (ESI) m/z = 296.3 [MH⁺].

3-(4-Chloro-phenyl)-2-cyclopentyloxy-5-methyl-pyridine (24c).

Following a procedure similar to the preparation of 24a, 23c (0.85 g,

3.3 mmol) and B-[4-chloro-phenyl]-boronic acid (0.82 g, 5.0 mmol)

were used to produce 24c (0.74 g, 78%) as yellowish oil. ¹H NMR (300

MHz, CDCl₃) δ 1.6−1.8 (m, 6 H), 1.78 (m, 2 H), 2.28 (s, 1 H), 5.46 (m,

1 H), 7.36 (d, J = 8.4, 2 H), 7.39 (d, J = 2.2, 1 H), 7.47(d, J = 8.4, 2 H),

7.95 (d, J = 2.2, 1 H). MS (ESI) m/z = 288.1 [MH⁺].

6-Chloro-4-iodo-5-(2,2,2-triﬂuoroethoxy)-2-pyridinemethanol

(28b). To a solution of 27 (40.0 g, 140 mmol) in hexamethylphosphor-

amide (400 mL) was added sodium hydride dispersion (5.6 g, ∼60% in

oil, 140 mmol) in portions. The resulting suspension was stirred for 45

min at room temperature, 1,1,1-triﬂuoro-methanesulfonic acid 2,2,2-

triﬂuoroethyl ester (23.4 mL, 168 mmol, dropwise) was added, and the

mixture was stirred at 120 °C for 24 h. After cooling, the mixture was

poured into water (1500 mL), acidiﬁed with hydrochloric acid (2 N, 100

mL), and extracted with ethyl acetate. Organic layers were washed with

water, combined, dried over Na₂SO₄, ﬁltered, and concentrated. The

residue was puriﬁed by ﬂash chromatography, eluting with ethyl acetate/

2-Butoxy-3-(4-chloro-phenyl)-5-methyl-pyridine (24d). Following

a procedure similar to the preparation of 24a, 23d (1.0 g, 4.1 mmol) and

B-[4-chloro-phenyl]-boronic acid (0.96 g, 6.1 mmol) were used to

1

heptane (1:1) to produce 28b (47.4 g, 92%) as white solid. H NMR

1

produce 24d (0.86 g, 76%) as yellowish oil. H NMR (600 MHz,

(600 MHz, DMSO-d₆) δ 4.47 (d, J = 5.9, 2 H), 4.69 (q, J = 8.9, 2 H), 5.64

(t, J = 5.9, OH), 7.89 (s, 1 H). HRMS (m/z): [MH⁺] calcd for

C₈H₇ClF₃INO₂367.9157; found 367.9159.

DMSO-d₆) δ 0.89 (t, J = 7.4, 3 H), 1.37 (m, 2 H), 1.65 (m, 2 H), 2.79 (s,

3 H), 4.27 (t, J = 6.6, 2 H), 7.49 (d, J = 8.8, 2 H), 7.59 (d, J = 8.8, 2 H),

7.60 (d, J = 2.3, 1 H), 7.98 (d, J = 2.3, 1 H). HRMS (m/z): [MH⁺] calcd

for C₁₆H₁₉ClNO 276.1150; found 276.1155.

3-(4-Chloro-phenyl)-2-cyclohexyloxy-5-methyl-pyridine (24e).

Following a procedure similar to the preparation of 24a, 23e (1.00 g,

3.7 mmol) and B-[4-chloro-phenyl]-boronic acid (0.91 g, 5.6 mmol)

were used to produce 24e (0.91 g, 82%) as colorless oil. ¹H NMR (300

MHz, CDCl₃) δ 1.49 (m, 4 H), 1.69 (m, 2 H), 1.91 (m, 2 H), 2.28 (s, 3

H), 5.12 (m, 1 H), 7.36 (d, J = 8.6, 2 H), 7.39 (s, 1 H), 7.52 (d, J = 8.6,

2H), 7.92 (s, 1 H). MS (ESI) m/z = 302.1 [MH⁺].

6-Chloro-5-hydroxy-2-pyridinemethanol (26). To a suspension of

2-chloro-3-hydroxy-pyridine (25, 15.0 g, 116 mmol) in water (110 mL)

was added sodium bicarbonate (14.6 g, 174 mmol). The resulting

suspension was stirred for 15 min (foam formation) warmed to 90 °C,

and a formaldehyde solution (37% in water, 30.2 mL, 405 mmol) was

added in portions over 18 h. After cooling the reaction mixture, ice (75

g) was added to the mixture and the pH was adjusted to 1 with

hydrochloric acid (6 N, 30 mL, CO₂evolution). The product was

partitioned into ethyl acetate; all organic phases were combined, dried

6-Chloro-4-(4-chlorophenyl)-5-(cyclopropylmethoxy)-2-pyridine-

methanol (29a). To a solution of 28a (23.0 g, 67.7 mmol) in toluene

(250 mL) was added with stirring the [1,1-bis(diphenylphosphino)-

ferrocene] dichloropalladium DCM complex (2.75 g, 3.4 mmol), B-(4-

chlorophenyl)boronic acid (10.6 g, 67.7 mmol), and sodium carbonate

solution (67.7 mL, 2M). The resulting mixture was stirred at 90 °C for

1.5 h, cooled, and partitioned between water and ethyl acetate. The

organic portion was dried over Na₂SO₄, ﬁltered, and concentrated. The

crude product was puriﬁed by ﬂash chromatography, eluting with an

ethyl acetate/heptane gradient to produce 29a (21.2 g, 97%) as light-

brown oil. ¹H NMR (600 MHz, CDCl₃) δ 0.05 (m, 2H), 0.45 (m, 2 H),

1.00 (m, 1 H), 2.81 (t, J = 5.6, OH), 3.49 (d, J = 7.3, 2 H), 4.74 (d, J = 5.6,

2 H), 7.24 (s, 1 H), 7.45 (d, J = 8.7, 2 H), 7.60 (d, J = 8.7, 2 H). HRMS

(m/z): [MH⁺] calcd for C₁₆H₁₆Cl₂NO₂324.0558; found 324.0554.

6-Chloro-4-(4-chlorophenyl)-5-(2,2,2-triﬂuoroethoxy)-2-pyridine-

methanol (29b). Following a procedure similar to the preparation of

29a, 28b (23.0 g, 62.6 mmol) was used to produce 29b (22.0 g, quant) as

brown oil. ¹H NMR (600 MHz, CDCl₃) δ 2.77 (t, J = 5.6, OH), 4.03 (q, J

Q

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

= 8.2, 2 H), 4.77 (d, J = 5.6, 2 H), 7.30 (s, 1 H), 7.47 (d, J = 8.7, 2 H), 7.55

(d, J = 8.7, 2 H). HRMS (m/z): [MH⁺] calcd for C₁₄H₁₁Cl₂F₃NO₂

352.0119; found 352.0116.

phino)ferrocene] dichloropalladium DCM complex (4.5 g, 5.5 mmol),

B-4-chlorophenylboronic acid (43.5 g, 0.278 mol), sodium carbonate

(43.8 g, 413 mmol), and water (80 mL). The resulting mixture was

stirred at 90 °C for 5 h, cooled, and partitioned between water and ethyl

acetate. The organic portion was washed with brine, dried over

magnesium sulfate, ﬁltered, and concentrated. The residue was puriﬁed

by ﬂash chromatography, eluting with a heptane/ethyl acetate gradient

4-(4-Chlorophenyl)-5-(cyclopropylmethoxy)-2-pyridinemethanol

(30a). To a solution of 29a (22.9 g, 70.6 mmol) in acetic acid (95%, 70

mL) was added at 40 °C with stirring tetramethylammonium bromide

(0.111 g, 0.71 mmol) and in several portions zinc powder (13.85 g, 212

mmol). The resulting mixture was stirred for 18 h at 50 °C, more zinc

powder (12.0 g, 183 mmol) was added, and stirring was continued for

another 6 h. After cooling, the mixture was poured into water (1000

mL), conc NaOH solution (130 mL) was added, and the zinc hydroxide

precipitate was removed by ﬁltration over diatomaceous earth. The

precipitate was washed with ethyl acetate (10 × 200 mL), and the water

phase was extracted with ethyl acetate; all organic phases were

combined, dried over Na₂SO₄, ﬁltered, and concentrated in vacuo.

The residue was puriﬁed by ﬂash chromatography, eluting with an ethyl

acetate/methanol gradient to produce 30a (16.4 g, 80%) as oﬀ-white

solid. ¹H NMR (600 MHz, CDCl₃) δ 0.29 (m, 2H), 0.59 (m, 2 H), 1.20

(m, 1 H), 3.46 (bt, J = 5.1, OH), 3.90 (d, J = 6.8, 2 H), 4.79 (d, J = 5.0, 2

H), 7.20 (s, 1 H), 7.42 (d, J = 8.6, 2 H), 7.57 (d, J = 8.6, 2 H), 8.27 (s, 1

H). HRMS (m/z): [MH⁺] calcd for C₁₆H₁₇ClNO₂290.0948; found

290.0938.

4-(4-Chlorophenyl)-5-(2,2,2-triﬂuoroethoxy)-2-pyridinemethanol

(30b). Following a procedure similar to the preparation of 30a, 29b

(22.0 g, 62.5 mmol) was used to produce 30b (16.5 g, 83%) as oﬀ-white

solid. ¹H NMR (600 MHz, CDCl₃) δ 3.33 (bt, J = 5.2, OH), 4.34 (q, J =

8.0, 2 H), 4.78 (d, J = 5.2, 2 H), 7.28 (s, 1 H), 7.45 (d, J = 8.7, 2 H), 7.52

(d, J = 8.7, 2 H) 8.31 (s, 1 H). HRMS (m/z): [MH⁺] calcd for

C₁₄H₁₂ClF₃NO₂318.0509; found 318.0502.

4-(4-Chlorophenyl)-5-(cyclopropylmethoxy)-2-pyridinecarboxylic

Acid (31a). To a solution of 30a (16.3 g, 56.3 mmol) in pyridine (250

mL) was added a solution of tetrabutylammonium permanganate (40.66

g, 113 mmol) in pyridine (120 mL). The combined solution was stirred

for 1 h at 80 °C. After cooling, the mixture was poured into water (1500

mL), saturated sodium hydrogensulﬁte solution (110 mL) was added,

and after acidiﬁcation with concentrated hydrochloric acid (400 mL) the

resulting suspension was stirred for 15 min at room temperature. Solids

were removed by ﬁltration, washed well with water, and the ﬁltrate was

concentrated in vacuo to produce 31a (16.4 g, 96%) as oﬀ-white solid.

¹H NMR (600 MHz, DMSO-d₆) δ 0.34 (m, 2H), 0.56 (m, 2 H), 1.22 (m,

1 H), 4.12 (d, J = 4.0, 2 H), 7.20 (s, 1 H), 7.56 (d, J = 8.7, 2 H), 7.71 (d, J

= 8.7, 2 H), 7.98 (s, 1 H), 8.53 (s, 1 H), 12.98 (br s, 1H). HRMS (m/z):

[MH⁺] calcd for C₁₆H₁₅ClNO₃304.0741; found 304.0742.

4-(4-Chlorophenyl)-5-(2,2,2-triﬂuoroethoxy)-2-pyridinecarboxylic

Acid (31b). Following a procedure similar to the preparation of 31a, 30b

(0.50 g, 1.57 mmol) was used to produce 31b (0.53 g, quant) as white

solid. ¹H NMR (600 MHz, DMSO-d₆) δ 5.09 (q, J = 8.8, 2 H), 7.58 (d, J

= 8.8, 2 H), 7.66 (d, J = 8.8, 2 H), 8.02 (s, 1 H), 8.66 (s, 1 H), 13.16 (br s,

1 H). HRMS (m/z): [MH⁺] calcd for C₁₄H₁₀ClF₃NO₃332.0296; found

332.0299.

4-(4-Chlorophenyl)-5-(cyclopropylmethoxy)-N-[(1R,2R)-2-hydrox-

ycyclohexyl]-2-pyridinecarboxamide (32a). General procedure D,

with 31a (100 mg, 0.329 mmol), was used to produce 32a (105 mg,

80%) as colorless foam. ¹H NMR (400 MHz, CDCl₃) δ 0.33 (m, 2 H),

0.63 (m, 2 H), 1.25 (m, 1 H), 1.40 (m, 4 H), 1.78 (m, 2 H), 2.10 (m, 2

H), 3.49 (m, 1 H), 3.57 (d, J = 4.0, OH), 3.81 (m, 1 H), 3.99 (d, J = 6.7, 2

H), 7.42 (d, J = 8.6, 2 H), 7.61 (d, J = 8.6, 2 H), 7.89 (bd, J = 7.3, NH),

8.18 (s, 1 H), 8.22 (s, 1 H). HRMS (m/z): [MH⁺] calcd for

C₂₂H₂₆ClN₂O₃401.1362; found 401.1633.

1

to produce 34 (37.2 g, 65%) as oﬀ-white solid. H NMR (600 MHz,

CDCl₃) δ 7.28 (m, 1 H), 7.46 (d, J = 8.8, 2 H), 7.50 (m, 1 H), 7.95 (d, J =

8.8, 2 H), 8.52 (d, J = 4.6, 1 H). MS (ESI) m/z = 208.03 [MH⁺].

2-(4-Chlorophenyl)-3-(cyclopropylmethoxy)-pyridine. To a solu-

tion of 34 (28.5 g, 137 mmol) in DMSO (160 mL) was added sodium

hydride dispersion (7.8 g, ∼55% in oil, 179 mmol) in portions. The

resulting suspension was stirred for 30 min at room temperature,

cyclopropanemethanol (14.0 mL, 172 mmol, dropwise) was added, and

the mixture was stirred at room temperature for 3 h. Afterward, water

(1000 mL) was added carefully and the mixture was extracted with

DCM. Organic layers were washed with brine, combined, dried over

MgSO₄, ﬁltered, and concentrated. The residue was puriﬁed by ﬂash

chromatography, eluting with an ethyl acetate/heptane gradient to

produce the title compound (30.1 g, 84%) as yellow oil. ¹H NMR (600

MHz, CDCl₃) δ 0.33 (m, 2 H), 0.63 (m, 2H), 1.25 (m, 1 H), 3.87 (d, J =

6.8, 2 H), 7.20 (m, 1 H), 7.25 (d, J = 6.8, 1 H,) 7.41 (d, J = 8.4, 2 H), 7.98

(d, J = 8.4, 2 H), 8.30 (d, J = 5.7, 1 H). HRMS (m/z): [MH⁺] calcd for

C₁₅H₁₅ClNO 260.0842; found 260.0840.

2-(4-Chlorophenyl)-3-(2,2,2-triﬂuoroethoxy)-pyridine. To a solu-

tion of 34 (0.425 g, 2.04 mmol) in DMSO (5 mL) was added cesium

carbonate and (1.33 g, 4.08 mmol) and 2,2,2-triﬂuoroethanol (1.5 mL,

20 mmol), and the mixture was stirred at room temperature for 40 min.

The resulting mixture was partitioned between water and ethyl acetate.

Organic layers were washed with brine, combined, dried over MgSO₄,

ﬁltered, and concentrated. The residue was puriﬁed by ﬂash

chromatography, eluting with an ethyl acetate/heptane gradient to

produce the title compound (0.38 g, 65%) as white solid. ¹H NMR (600

MHz, CDCl₃) δ 4.34 (q, J = 8.0, 2 H), 7.28 (m, 2 H), 7.43 (d, J = 8.7, 2

H), 7.89 (d, J = 8.7, 2 H), 8.43 (m, 1 H). HRMS (m/z): [MH⁺] calcd for

C₁₃H₁₀ClF₃NO 288.0403; found 288.0397.

2-(4-Chlorophenyl)-3-cyclopropylmethoxy-pyridine 1-oxide

(35a). To a solution of 2-(4-chlorophenyl)-3-(cyclopropylmethoxy)-

pyridine (30.0 g, 116 mmol) in acetic acid (300 mL) was added

hydrogen peroxide solution (606 mL 30% in water). The resulting

solution was stirred for 5 h at 70 °C. After cooling, the mixture was

poured into saturated bicarbonate solution and the mixture was

extracted with DCM. Organic layers were washed with water and brine,

combined, dried over MgSO₄and charcoal, ﬁltered, and concentrated.

The residue was puriﬁed by ﬂash chromatography, eluting with an ethyl

acetate/methanol gradient to produce 35a (25.4 g, 80%) as oﬀ-white

solid. ¹H NMR (600 MHz, CDCl₃) δ 0.23 (m, 2 H), 0.56 (m, 2H), 1.11

(m, 1 H), 3.82 (d, J = 6.8, 2 H), 6.89 (m, 1 H), 7.14 (m, 1 H), 7.44 (d, J =

8.8, 2 H), 7.52 (d, J = 8.8, 2 H), 8.03 (m, 1 H). HRMS (m/z): [MH⁺]

calcd for C₁₅H₁₅ClNO₂276.0791; found 276.0788.

2-(4-Chlorophenyl)-3-(2,2,2-triﬂuoroethoxy)-pyridine 1-oxide

(35b). Following a procedure similar to the preparation of 35a, 2-(4-

chlorophenyl)-3-(2,2,2-triﬂuoroethoxy)-pyridine (1.0 g, 3.48 mmol)

was used to produce 35b (0.71 g, 67%) as oﬀ-white solid. ¹H NMR (600

MHz, CDCl₃) δ 4.28 (q, J = 7.9, 2 H), 6.92 (d, J = 8.7, 1 H), 7.21 (m, 1

H), 7.46 (d, J = 8.7, 2 H), 7.50 (d, J = 8.7, 2 H), 8.14 (d, J = 6.6, 1 H).

HRMS (m/z): [MH⁺] calcd for C₁₃H₁₀ClF₃NO₂304.0352; found

304.0337.

4-(4-Chlorophenyl)-5-(2,2,2-triﬂuoroethoxy)-N-[(1R,2R)-2-hydrox-

ycyclohexyl]-2-pyridinecarboxamide (32b). General procedure D,

with 31b (200 mg, 0.603 mmol), was used to produce 32b (226 mg,

87%) as white solid. ¹H NMR (600 MHz, DMSO-d₆) δ 1.21−1.34 (m, 4

H), 1.64 (m, 2 H), 1.91 (m, 2 H), 3.47 (m, 1 H), 3.59 (m, 1 H), 4.69 (d, J

= 5.5, OH), 5.07 (q, J = 8.7, 2 H), 7.58 (d, J = 8.7, 2 H), 7.65 (d, J = 8.7, 2

H), 7.99 (s, 1 H), 8.31 (bd, J = 8.1, NH), 8.57 (s, 1 H). HRMS (m/z):

[MH⁺] calcd for C₂₀H₂₀ClF₃N₂O₃429.1193; found 429.1192.

2-(4-Chlorophenyl)-3-ﬂuoro-pyridine (34). To a solution of 2-

chloro-3-ﬂuoro-pyridine (33, 36.2 g, 0.275 mol) in toluene (380 mL)

and DMF (40 mL) was added with stirring the [1,1-bis(diphenylphos-

6-(4-Chlorophenyl)-5-(cyclopropylmethoxy)-2-pyridinecarboni-

trile (36a). To a solution of 35a (25.2 g, 91 mmol) in acetonitrile (500

mL) was added triethylamine (26.7 mL, 192 mmol) and N,N-

dimethylcarbamic chloride (9.2 mL, 101 mmol) with stirring. Five

minutes later, trimethylsilylcyanide (34.4 mL, 274 mmol) was added

and the resulting solution was stirred for 15 h at 90 °C. After cooling, the

mixture was partitioned between water and ethyl acetate; organic layers

were washed with brine, combined, dried over MgSO₄, ﬁltered, and

concentrated. The residue was puriﬁed by ﬂash chromatography, eluting

with an heptane/ethyl acetate gradient to produce 36a (14.2 g, 55%) as

light−yellow solid. ¹H NMR (600 MHz, CDCl₃) δ 0.38 (m, 2 H), 0.70

R

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

(m, 2H), 1.30 (m, 1 H), 3.96 (d, J = 7.0, 2 H), 7.26 (d, J = 8.5, 1 H), 7.43

(d, J = 8.7, 2 H), 7.61 (d, J = 8.5, 1 H), 8.00 (d, J = 8.7, 2 H). HRMS (m/

z): [MH⁺] calcd for C₁₆H₁₄ClN₂O 285.0795; found 285.0793.

6-(4-Chlorophenyl)-5-(2,2,2-triﬂuoroethoxy)-2-pyridinecarboni-

trile (36b). Following a procedure similar to the preparation of 36a, 35b

(89 mg, 0.29 mmol) was used to produce 36b (56 mg, 61%) as light-

yellow solid. ¹H NMR (600 MHz, CDCl₃) δ 4.46 (q, J = 7.8, 2 H), 7.33

(d, J = 8.5, 1 H), 7.46 (d, J = 8.7, 2 H), 7.68 (d, J = 8.5, 1 H), 7.91 (d, J =

8.7, 2 H). HRMS (m/z): [MHCO₂⁻] calcd for C₁₅H₉ClF₃N₂O₃

357.0254; found 357.0270.

potassium carbonate (15.8 g, 114 mmol). The resulting mixture was

stirred at reﬂux temperature for 4 h, cooled, and partitioned between

water and ethyl acetate. The organic portion was dried over magnesium

sulfate, ﬁltered, and concentrated. The crude product was puriﬁed by

ﬂash chromatography, eluting with ethyl acetate in heptane (1:3) to

produce 39 (12.99 g, 80%) as yellow solid. ¹H NMR (600 MHz, CDCl₃)

δ 4.76 (br s, NH₂), 7.48 (d, J = 8.6, 2 H), 7.69 (d, J = 8.6, 2 H), 8.09 (s, 1

H). HRMS (m/z): [MH⁺] calcd for C₁₀H₈BrClN₃283.9585, 285.9562;

found 283.9587, 285.9564.

2,5-Dibromo-3-(4-chlorophenyl)-pyrazine (40). To a solution of 39

(2.36 g, 8.3 mmol) in dibromomethane (15 mL) was added with stirring

isoamylnitrite (1.3 mL g, 9.4 mmol). To this mixture was added a

solution of trimethylbromosilane (1.5 g, 9.8 mmol) in dibromomethane

(5 mL). The resulting mixture was stirred at room temperature for 1 h

and poured into sodium bicarbonate solution (30 mL, 10%). The

organic portion was puriﬁed by ﬂash chromatography, eluting with ethyl

acetate in heptane (1:9) to produce 40 (2.13 g, 74%) as light-yellow

solid. ¹H NMR (600 MHz, CDCl₃) δ 7.48 (d, J = 8.8, 2 H), 7.75 (d, J =

8.8, 2 H), 8.43 (s, 1 H). MS (ESI) m/z = 346.0, 348.0, 350.0 [M].

5-Bromo-3-(4-chlorophenyl)-2-(cyclopropylmethoxy)-pyrazine

(41). To a solution of cyclopropanemethanol (79.3 mg, 1.1 mmol) in

DMSO (2 mL) was added sodium hydride dispersion (96 mg, ∼55% in

oil, 2.2 mmol). The resulting suspension was stirred for 45 min at room

temperature, 40 (348 mg, 1.0 mmol) was added, and the mixture was

stirred at room temperature for 3 h. The mixture was portioned between

ethyl acetate and water; organic layers were washed with water and

brine, combined, dried over Na₂SO₄, ﬁltered, and concentrated. The

residue was puriﬁed by ﬂash chromatography, eluting with an heptane/

DCM gradient to produce 41 (190 mg, 59%) as colorless crystals; mp

86−87 °C. ¹H NMR (400 MHz, CDCl₃) δ 0.36 (m, 2 H), 0.64 (m, 2 H),

1.29 (m, 1 H), 4.17 (d, J = 7.3, 2 H), 7.45 (d, J = 8.7, 2 H), 7.73 (d, J = 8.7,

1H), 8.01 (s, 1 H). MS (ESI) m/z = 339.2, 341.2 [MH⁺].

6-(4-Chlorophenyl)-5-(cyclopropylmethoxy)-2-pyrazinecarboxylic

Acid Methyl Ester (42). To a solution of 41 (195 mg, 0.574 mmol) in

methanol (3.5 mL) and ethyl acetate (1.5 mL) was added the [1,1-

bis(diphenylphos-phino)ferrocene] dichloropalladium DCM complex

(21.2 mg, 0.0259 mmol) and triethylamine (0.161 mL, 1.15 mmol). The

resulting solution was stirred in a CO atmosphere (70 bar) at 110 °C for

18 h. The mixture was concentrated and puriﬁed by ﬂash

chromatography, eluting with an heptane/DCM gradient to produce

42 (168 mg, 92%) as light-brown solid. ¹H NMR (400 MHz, CDCl₃) δ

0.41 (m, 2 H), 0.66 (m, 2 H), 1.33 (m, 1 H), 4.00 (s, 3 H), 4.35 (d, J =

7.3, 2 H), 7.45 (d, J = 8.7, 2 H), 8.13 (d, J = 8.7, 2 H), 8.79 (s, 1 H). MS

(ESI) m/z = 319.1 [MH⁺].

6-(4-Chlorophenyl)-5-(cyclopropylmethoxy)-2-pyrazinecarboxylic

Acid (43). To a solution of 42 (0.16 g, 0.50 mmol) in THF (2 mL),

methanol (0.5 mL) and water (0.5 mL) was added lithium hydroxide

solution (2 mL, 1 M in water), and the mixture was stirred for 2 h at

room temperature. Afterward, citric acid solution (10 mL, 10%) was

added, and the organic phase was separated, dried over MgSO₄, ﬁltered,

and concentrated to produce 43 (0.152 g, 94%) as oﬀ-white solid. ¹H

NMR (600 MHz, DMSO-d₆) δ 0.40 (m, 2 H), 0.59 (m, 2 H), 1.33 (m, 1

H), 4.34 (d, J = 7.3, 2 H), 7.61 (d, J = 8.8, 2 H), 8.15 (d, J = 8.8, 1 H), 8.77

(s, 1 H), 13.33 (br s, 1 H). HRMS (m/z): [MH⁺] calcd for

C₁₅H₁₄ClN₂O₃305.0687; found 305.0691.

6-(4-Chlorophenyl)-5-(cyclopropylmethoxy)-N-[(1R,2R)-2-hy-

droxy-cyclohexyl]-pyrazine-2-carboxamide (44). General procedure

D, with 43 (50 mg, 0.164 mmol), was used to produce 44 (36 mg, 55%)

as white solid. ¹H NMR (400 MHz, CDCl₃) δ 0.40 (m, 2 H), 0.65 (m, 2

H), 1.2−1.5 (m, 5 H), 1.78 (m, 2 H), 2.11 (m, 2 H), 3.36 (d, J = 4.4,

OH), 3.50 (m, 1 H), 3.85 (m, 1 H), 4.35 (d, J = 7.3, 2 H), 7.48 (d, J = 8.9,

2 H), 7.64 (bd, J = 7.7, NH), 8.05 (d, J = 8.9, 2 H), 8.87 (s, 1 H). [MH⁺]

calcd for C₂₁H₂₅ClN₃O₃402.1579; found 402.1595.

6-(4-Chlorophenyl)-5-(cyclopropylmethoxy)-2-pyridinecarboxylic

Acid. Acetyl chloride (2.6 mL, 35 mmol) was added to ethanol (40 mL)

at 0 °C. After stirring for 5 min at 0 °C, 36a (104 mg, 7.0 mmol) was

added and the mixture was stirred for 20 h at 90 °C. After cooling, the

mixture was concentrated, partitioned between water and ethyl acetate;

organic layers were washed with brine, combined, dried over MgSO₄,

ﬁltered, and concentrated. The residue was redissolved in THF (15 mL)

and water (7 mL), lithium hydroxide monohydrate (0.59 g, 14 mmol)

was added, and the mixture was stirred for 5 h at 80 °C. After cooling, the

mixture was brought to pH 6 with acetic acid and extracted with DCM.

Organic layers were washed with water and brine, combined, dried over

MgSO₄, ﬁltered, and concentrated. The residue was puriﬁed by ﬂash

chromatography, eluting with an heptane/ethyl acetate gradient to

1

produce the title compound (1.01 g, 47%) as light-yellow solid. H

NMR (600 MHz, CDCl₃) δ 0.39 (m, 2 H), 0.70 (m, 2 H), 1.30 (m, 1 H),

3.99 (d, J = 7.0, 2 H), 7.41 (d, J = 8.6, 1 H), 7.46 (d, J = 8.7, 2 H), 7.94 (d,

J = 8.7, 2 H), 8.16 (d, J = 8.6, 1 H), COOH not visible (br s at 1.60).

HRMS (m/z): [MH⁺] calcd for C₁₆H₁₅ClNO₃304.0735; found

304.0736.

6-(4-Chlorophenyl)-5-(2,2,2-triﬂuoroethoxy)-2-pyridinecarboxylic

Acid. Acetyl chloride (0.12 mL, 1.66 mmol) was added to ethanol (2

mL) at 0 °C. After stirring for 5 min at 0 °C, 36b (104 mg, 0.33 mmol)

was added and the mixture was stirred for 20 h at 80 °C. After cooling,

the mixture was concentrated, partitioned between water and ethyl

acetate; organic layers were washed with brine, combined, dried over

MgSO₄, ﬁltered, and concentrated. The residue was redissolved in THF

(2 mL), sodium hydroxide solution (2 N, 1 mL) was added, and the

mixture was stirred for 30 h at 80 °C. After cooling the mixture was

brought to pH 6 with acetic acid and extracted with DCM. Organic

layers were washed with water and brine, combined, dried over MgSO₄,

ﬁltered, and concentrated. The residue was puriﬁed by ﬂash

chromatography, eluting with an heptane/ethyl acetate gradient to

produce the title compound (54 mg, 49%) as oﬀ-white solid. ¹H NMR

(600 MHz, CDCl₃) δ 4.49 (q, J = 7.7, 2 H), 7.47 (d, J = 8.5, 1 H), 7.49 (d,

J = 8.7, 2 H), 7.85 (d, J = 8.7, 2 H), 8.23 (d, J = 8.5, 1 H), 10.69 (br s, 1

H). HRMS (m/z): [MH⁺] calcd for C₁₄H₁₀ClF₃NO₃332.0301; found

332.0298.

6-(4-Chlorophenyl)-5-(cyclopropylmethoxy)-N-[(1R,2R)-2-hydrox-

ycyclohexyl]-2-pyridinecarboxamide (37a). General procedure D,

with 6-(4-chlorophenyl)-5-(cyclopropylmethoxy)-2-pyridinecarboxylic

acid (180 mg, 0.593 mmol), was used to produce 37a (174 mg, 73%) as

oﬀ-white foam. ¹H NMR (600 MHz, DMSO-d₆) δ 0.36 (m, 2 H), 0.59

(m, 2 H), 1.25 (m, 4 H), 1.31 (m, 1 H), 1.64 (m, 2 H), 1.91 (m, 2 H),

3.47 (m, 1 H), 3.59 (m, 1 H), 4.03 (d, J = 7.0, 2 H), 4.71 (d, J = 5.5, OH),

7.55 (d, J = 8.8, 2 H), 7.68 (d, J = 8.7, 1 H), 7.98 (d, J = 8.7, 1 H), 8.11 (d,

J = 8.8, 2 H), 8.12 (bd, J = 7.0, NH). HRMS (m/z): [MH⁺] calcd for

C₂₂H₂₆ClN₂O₃401.1632; found 401.1630.

6-(4-Chlorophenyl)-5-(2,2,2-triﬂuoroethoxy)-N-[(1R,2R)-2-hydrox-

ycyclohexyl]-2-pyridinecarboxamide (37b). General procedure D,

with 6-(4-chlorophenyl)-5-(2,2,2-triﬂuoroethoxy)-2-pyridinecarboxylic

acid (150 mg, 0.452 mmol), was used to produce 37b (117 mg, 60%) as

oﬀ-white solid. ¹H NMR (600 MHz, CDCl₃) δ 4.49 (q, J = 7.7, 2 H),

7.47 (d, J = 8.6, 1 H), 7.49 (2, J = 8.7, 2 H), 7.85 (d, J = 8.7, 2 H), 8.23 (d,

J = 8.6, 1 H), 10.69 (br s, 1 H). HRMS (m/z): [MH⁺] calcd for

C₁₄H₁₀ClF₃NO₃332.0301; found 332.0298.

5-Bromo-3-(4-chlorophenyl)-2-pyrazinamine (39). To a solution of

3,5-dibromo-2-pyrazinamine (38, 14.4 g, 57 mmol) in THF (360 mL)

and water (360 mL) was added with stirring tetrakis-

(triphenylphosphine)palladium(0) (3.29 g, 29.9 mmol). To this mixture

was added B-4-chlorophenyl-boronic acid (9.36 g, 9.66 mmol) and

3-Chloro-6-(cyclopropylmethoxy)-pyridazine (46). To a solution of

cyclopropanemethanol (17.95 mL, 222 mmol) in DMSO (150 mL) was

added sodium hydride dispersion (8.86 g, ∼60% in oil, 222 mmol, in

portions). The resulting suspension was stirred for 30 min at room

temperature, and the resulting solution was added dropwise at room

temperature to a solution of 3,6-dichloro-pyridazine (45, 30.0 g, 201

mmol) in DMSO (300 mL). Stirring at room temperature was

S

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

continued for another hour; afterward, the mixture was partitioned

between ethyl acetate and water, and organic layers were washed with

water and brine, combined, dried over MgSO₄, ﬁltered, and

concentrated. The residue was puriﬁed by ﬂash chromatography,

eluting with an heptane/ethyl acetate gradient to produce 46 (27.2 g,

73%) as white solid. ¹H NMR (000 MHz, CDCl₃) δ 0.38 (m, 2 H), 0.64

(m, 2 H), 1.32 (m, 1 H), 4.30 (d, 2 H), 6.96 (d, 1 H), 7.33 (d, 1 H).

HRMS (m/z): [MH⁺] calcd for C₈H₁₀ClN₂O 185.0476; found

185.0481.

Cell Culture. CHO-K1 β-arrestin cells (DiscoveRx Inc., Fremont,

CA) expressing human CB1 and human CB2 were cultured in F-12

Nutrient Mixture (HAM) supplemented with 10% FBS, 300 μg/mL

hygromycin, and 800 μg/mL Geneticin (G418). Cells were incubated in

a humidiﬁed atmosphere at 37 °C with 5% CO₂.

Radioligand Binding Assay. For membrane preparation, stably

transfected cells were treated for 24 h with 5 mM butyrate in growth

medium before harvesting, followed by homogenization in 15 mM

Hepes, 0.3 mM EDTA, 1 mM EGTA, 2 mM MgCl₂, and complete

EDTA-free protease inhibitor (Roche Applied Science, Rotkreuz,

Switzerland), pH 7.4, using a glass potter and centrifugation at

47800g at 4 °C for 30 min. The pellet was then rehomogenized twice in

the same buﬀer and centrifuged (47800g, 4 °C, 30 min). The ﬁnal pellet

was then resuspended in 75 mM Tris, 0.3 mM EDTA, 1 mM EGTA,

12.5 mM MgCl₂, and 250 mM sucrose, pH 7.4, at a protein

concentration of 1−3 mg per mL, aliquoted, frozen on dry ice, and

stored at −80 °C. Saturation binding was performed with 0.05−2.6 nM

[³H]CP55940 (Perkin-Elmer) and 1.0 μg of membrane protein.

CP55940 (10 μM) was used to deﬁne nonspeciﬁc binding. More than

95% of the total binding signal was speciﬁc. Assay buﬀer consisted of 50

mM Tris-HCl, 5 mM MgCl₂, 2.5 mM EGTA, and 0.1% fatty acid-free

BSA, pH 7.4. Assays were initiated by addition of membranes in a ﬁnal

volume of 250 μL per well. Assays were incubated for 3 h at room

temperature and then vacuum ﬁltered and rinsed with wash buﬀer (50

mM Tris-HCl, 5 mM MgCl₂, 2.5 mM EGTA, and 0.5% fatty acid-free

BSA, pH 7.4) on a Filtermate cell harvester through Packard GF/B

ﬁlters presoaked in 0.3% polyethylenimine.

For competition binding, membrane preparations were incubated

with 0.3 nM [³H]CP55940 in the presence or absence of increasing

concentrations of ligands for 60 min at 30 °C in a ﬁnal volume of 0.2 mL

of 50 mM Tris-HCl, 5 mM MgCl₂, 2.5 mM EGTA, 0.1% fatty acid-free

BSA, and 1% DMSO, pH 7.4, buﬀer, gently shaking. Nonspeciﬁc

binding was deﬁned with 10 μM CP55940. All binding reactions were

terminated by vacuum ﬁltration onto 0.5% polyethylenimine presoaked

GF/B ﬁlter plates (Packard) followed by seven brief washes with 2 mL of

ice-cold binding buﬀer containing 0.5% fatty acid-free BSA. Plates were

dried at 50 °C for 1 h, and liquid scintillation counting was used to

determine bound radiolabel. IC₅₀values and Hill slopes were

determined by a four-parameter logistic model using ActivityBase (ID

Business Solution, Ltd.). K_ivalues were determined by the generalized

Cheng−Prusoﬀ equation.⁴⁰

3-Chloro-6-(cyclopropylmethoxy)-5-iodo-pyridazine (47). To a

solution of 2,2,6,6-tetramethylpiperidine (79.67 mL, 459 mmol) in

THF (800 mL) was added dropwise at room temperature 1.6 N n-

butyllithium in hexane (285 mL, 456 mmol). Stirring was continued for

20 min, and afterward the solution was cooled to −78 °C. A precooled

solution of 46 (24.2 g, 131 mmol) in THF (500 mL) was added rapidly

(−69 °C maximum), and the mixture was stirred for 5 min at −75 °C

before a precooled solution of iodine (39.9 g, 157 mmol) in THF (500

mL) was added rapidly (−54 °C maximum). Stirring continued at −75

°C for 15 min, the mixture was quenched by addition of saturated

ammonium chloride solution, and afterward the mixture was partitioned

between ethyl acetate and water; organic layers were washed with water,

combined, dried over MgSO₄, ﬁltered, and concentrated. The residue

was puriﬁed by ﬂash chromatography, eluting with an heptane/DCM

1

gradient to produce 47 (10.3 g, 25%) as white solid. H NMR (400

MHz, CDCl₃) δ 0.42 (m, 2 H), 0.65 (m, 2H), 1.37 (m, 1H), 4.37 (d, J =

7.1, 2 H), 7.90 (s, 1 H). MS (ESI) m/z = 310.9 [MH⁺].

3-Chloro-6-(cyclopropylmethoxy)-5-(4-chlorophenyl)-pyridazine

(48). To a solution of 47 (14.7 g, 47.2 mmol) in THF (380 mL) and

water (380 mL) was added with stirring tetrakis(triphenylphosphine)-

palladium(0) (2.7 g, 2.36 mmol). To this mixture was added B-4-

chlorophenyl-boronic acid (7.38 g, 47.2 mmol) and potassium

carbonate (13.1 g, 94.4 mmol). The resulting mixture was stirred at

reﬂux temperature for 25 h, cooled, and partitioned between water and

ethyl acetate. The organic portion was dried over magnesium sulfate,

ﬁltered, and concentrated. The crude product was puriﬁed by ﬂash

chromatography, eluting with an heptane/ethyl acetate gradient to

produce 48 (4.40 g, 32%) as white solid. ¹H NMR (400 MHz, CDCl₃) δ

0.37 (m, 2 H), 0.62 (m, 2 H), 1.34 (m, 1 H), 4.41 (d, J = 7.1, 2 H), 7.39

(s, 1 H), 7.47 (d, J = 8.6, 2 H), 7.62 (d, J = 8.6, 2 H). MS (ESI) m/z =

295.1, 297.1 [MH⁺].

5-(4-Chlorophenyl)-6-(cyclopropylmethoxy)-3-pyridazinecarbox-

ylic Acid Methyl Ester (49). To a solution of 48 (4.4 g, 14.9 mmol) in

methanol (53 mL) was added the [1,1-bis(diphenylphos-phino)-

ferrocene] dichloropalladium−DCM complex (531 mg, 0.65 mmol)

and triethylamine (4.16 mL, 29.8 mmol). The resulting solution was

stirred in a CO atmosphere (70 bar) at 120 °C for 20 h. The mixture was

ﬁltered, concentrated, and puriﬁed by ﬂash chromatography, eluting

with an heptane/ethyl acetate gradient to produce 49 (2.93 g, 62%) as

white solid. ¹H NMR (400 MHz, CDCl₃) δ 0.41 (m, 2H), 0.63 (m, 2 H),

1.39 (m, 1 H), 4.05 (s, 3 H), 4.54 (d, J = 7.1, 2 H), 7.48 (d, J = 8.6, 2 H),

7.68 (d, J = 8.6, 2 H), 8.11 (s, 1 H). MS (ESI) m/z = 319.2 [MH⁺].

5-(4-Chlorophenyl)-6-(cyclopropylmethoxy)-3-pyridazinecarbox-

ylic Acid (50). To a solution of 49 (2.85 g, 8.94 mmol) in THF (30 mL)

was added lithium hydroxide solution (11.6 mL, 1 M in water), and the

mixture was stirred for 30 min at room temperature. Afterward, the

mixture was acidiﬁed with hydrochloric acid (1 N), and the precipitate

was ﬁltered and dried to give 50 (2.56 g, 94%) as white solid. ¹H NMR

(600 MHz, DMSO-d₆) δ 0.40 (m, 2 H), 0.58 (m, 2 H), 1.33 (m, 1 H),

4.47 (d, J = 7.3, 2 H), 7.60 (d, J = 8.7, 1 H), 7.82(d, J = 8.7, 2 H), 8.09 (s, 1

H), 13.67 (br s, 1 H). HRMS (m/z): [MH⁺] calcd for C₁₅H₁₄ClN₂O₃

305.0687; found 305.0688.

5-(4-Chlorophenyl)-6-(cyclopropylmethoxy)-N-[(1R,2R)-2-hydrox-

ycyclohexyl]-pyridazine-3-carboxamide (51). Following a procedure

similar to the preparation of 16c, 50 (100 mg, 0.33 mmol) and (1R,2R)-

2-amino-cyclohexanol hydrochloride (47 mg, 0.41 mmol) were used to

produce 51 (90 mg, 68%) as white solid. ¹H NMR (400 MHz, CDCl₃) δ

0.40 (m, 2 H), 0.65 (m, 2 H), 1.2−1.45 (m, 5 H), 1.77 (m, 2 H), 2.10 (m,

2 H), 3.18 (d, J = 4.2, OH), 3.49 (m, 1 H), 3.88 (m, 1 H), 4.49 (d, J = 7.2,

2 H), 7.47 (d, J = 8.7, 2 H), 7.69 (d, J = 8.7, 2 H), 8.00 (bd, NH), 8.23 (s,

1 H). [MH⁺] calcd for C₂₁H₂₅ClN₃O₃402.1580; found 402.1579.

GTPγ[³⁵S] Binding Assay. Membranes were generated as

described.²⁵Fractions were washed and diluted in assay buﬀer (10

mM HEPES, 32 mM MgCl₂, 100 mM NaCl, 20 μM GDP, pH 7.4),

homogenized brieﬂy using a Polytron and incubated for 10 min at 30 °C.

Following preincubation, assay-mixtures were prepared in 96-well

microtiter plates. The composition of the assay mixtures in a ﬁnal

volume of 200 μL of assay buﬀer per well was as follows: 25 μg of

membrane protein (pretreated as described above), 0.25 nM GTPγ-

[³⁵S], and the test compounds at the appropriate concentrations.

Nonspeciﬁc binding was measured in the presence of unlabeled

GTPγ[S] in excess (10 μM). All binding reactions were terminated by

vacuum ﬁltration onto GF/B ﬁlter plates (Packard) presoaked with

wash buﬀer followed by three brief washes with 2 mL of ice-cold wash

buﬀer (Tris 50 mM, 2.5 mM EDTA, 5 mM MgCl₂, and 0.5% fatty acid

free BSA). Plates were dried at 50 °C for 1 h, and liquid scintillation

counting was used to determine bound radiolabel in a Packard

TopCount (Canberra Packard SA, Zurich, Switzerland).

̈

Microsomal Stability Testing. Incubations were performed in 96-

well deep-well plates with a ﬁnal incubation volume of 600 μL.

Incubations contained (ﬁnally) 2 μM test compound, 0.5 mg/mL rat

liver microsomes, and NADPH regenerating system. Then 50 μL

aliquots were removed after 1, 3, 6, 9, 15, 25, 35, and 45 min and

quenched in 150 μL of acetonitrile containing internal standard.

Samples were then cooled and centrifuged before analysis by LC-MS/

MS.

Log peak area ratio (test compound peak area/internal standard peak

area) was plotted against incubation time and a linear ﬁt made to the

data with emphasis upon the initial rate of compound disappearance.

The slope of the ﬁt is then used to calculate the intrinsic clearance:

T

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

were harvested (epididymal fat pads, liver, and brain) for further

analysis.

Clmic(μL/min /mg)

= −slope(min⁻¹) × 1000/[protein concentration(mg/mL)]

ASSOCIATED CONTENT

■

Results were not corrected for protein binding in the microsomal

incubation.

S

* Supporting Information

Hepatocytes Stability Testing: Short Overview. Freshly isolated

rat hepatocytes were incubated in suspension in a Williams E medium

supplemented with 10% FCS. The hepatocyte suspension cultures were

freshly prepared by liver perfusion (as described by Seglen⁴¹).

Incubations of a test compound at 2 μM test concentration in

suspension cultures of 1 Mio cells/mL (∼1 mg/mL protein

concentration) were performed in 96-well plates and shaken at 900

rpm for up to 3 h in a 5% CO₂atm and 37 °C. After 2, 10, 20, 40, 60, 120,

and 180 min, 100 μL of cell suspension in each well was quenched with

200 μL of methanol containing an internal standard. Samples were then

cooled and centrifuged before analysis by LC-MS/MS.

CEREP data for 14g and 14h, as well as glucose excursion data

from the high fat diet rat study and PK data for 15h. This material

is available free of charge via the Internet at http://pubs.acs.org.

AUTHOR INFORMATION

■

Corresponding Author

*Phone: (+41) 61-688-5906. Fax: (+41) 61-688-6459. E-mail:

stephan.roever@roche.com.

Author Contributions

Log peak area ratio (test compound peak area/internal standard peak

area) was plotted against incubation time and a linear ﬁt made to the

data with emphasis upon the initial rate of compound disappearance.

The slope of the ﬁt was then used to calculate the intrinsic clearance:

The manuscript was written through contributions of all authors.

All authors have given approval to the ﬁnal version of the

manuscript.

Notes

Clint(μL/min /1 × 10⁶cells)

The authors declare the following competing ﬁnancial

interest(s): The authors are or have been employees of F.

Hoﬀmann-La Roche AG.

= −slope(min⁻¹) × 1000/[protein concentration(mg/mL)]

PK Study after 10 Days Food Admix Administration. 14g and

14h were administered by feed admix to male Sprague−Dawley rats (n =

3/test compound) at 30 mg/kg/d each over 10 days. The feed admix

chow was replaced by drug-free high fat diet (HFD) chow at the time of

ﬁrst blood sampling. The rats were implanted with a jugular vein

catheter 4 days before start of sample collection. After change to the

drug-free HFD chow, serial blood samples were collected from the

indwelling catheter (K₃EDTA as anticoagulant) over a period of 24 h.

Drug levels in plasma were analyzed by LC-MS.

ACKNOWLEDGMENTS

■

We thank Beat Frei, Noemi Raschetti, Jitka Weber, Bernd

Pullmann, Gabriela Schnider, Guy Wassner, Etienne Rauber, Joel

Ruegger, Adriane Gischig, and Indira Fabre for technical

assistance, J. Schneider for NMR spectroscopy, C. Bartelmus

for mass spectroscopy, Jacqueline Higelin and Elisabeth Zirwes

for conducting the biochemical assay, Anja Osterwald for

̈

High Fat Diet Rat Study. A group of 80 male Sprague−Dawley rats

(Iﬀa Credo/Charles River France; 10 weeks-old) was fed a high fat diet

(from SSNIFF 43% of energy, 19% fat, and equivalent to KLIBA 2157)

during 3 weeks (min average BW∼450 g). Ambient temperature was

approximately 21 °C, and relative humidity 55−65%. A 12 h light−dark

cycle was maintained in the room with test being performed during the

light phase. Access to tap water was ad libitum. At the end of the feeding

period, obese rats (high responders) were selected using the following

criteria: the diﬀerence in average BW of high responders compared with

that of rats fed standard chow diet was at least 2 times the standard

deviation (SD) of the average BW of the standard chow diet group:

́

executing cell-based assays, Marie Stella Gruyer, Veronique

Dall’Asen, Christelle Rapp-Mary, Christophe Flament, Stephen

Fowler, and Pascal Schenk for obtaining PK and ADME data and

Philippe Verry and Andree Roeckel for conducting PD/eﬃcacy

studies. The research described in the manuscript was funded by

F. Hoﬀmann-La Roche AG.

ABBREVIATIONS USED

■

AIBN, 2,2′-azobis(2-methylpropannitrile); 2-AG, 2-arachido-

noylglycerol; ANOVA, analysis of variance; AUC, area under the

curve; CB2, cannabinoid receptor type 2; DIO, diet-induced

obesity; DBU, 2,3,4,6,7,8,9,10-octahydropyrimido[1,2-a]-

azepine; DCM, dichloromethane; DEAD, 1,2-diazenedicarbox-

ylic acid 1,2-diethyl ester; DIEA, N,N-diisopropylethylamine;

DME, dimethoxyethane; DMF, dimethylformamide; DMSO,

dimethyl sulfoxide; GTP, guanosine triphosphate; HEK, human

embryonic kidney (cells); KO, knock out; LC-MS, liquid

chromatography−mass spectrometry; MS, mass spectrometry;

MW, microwave; NBS, 1-bromo-2,5-pyrrolidinedione; NMR,

nuclear magnetic resonance; OGTT, oral glucose tolerance test;

Pgp, P-glycoprotein; PK, pharmacokinetic; RT, room temper-

ature; SAR, structure−activity relationship; SD, standard

deviation; SDPK, single dose PK; SEM, standard error of the

mean; TBTU, O-(benzotriazol-1-yl)-N,N,N′,N′-tetramethyluro-

nium tetraﬂuoroborate; THF, tetrahydrofuran; TMP, 2,2,6,6-

tetramethylpiperidine; TMSBr, bromotrimethylsilane; TMSCN,

trimethylsilanecarbonitrile; TPP, triphenylphosphine

[BW(obese) − BW(chow) ≥ 2 × SD(chow)]

The rest of the diet-resistant rats (low responders) were kept for

separate investigation, such as pharmacokinetic characterization in age-

matched rats. Seven homogeneous groups of obese animals (n = 9)

according to BW, the body fat content (measured by MRS at the end of

the growing period) were constituted and rats were transferred to single

cages.

Treatment was given as food admix. Doses were 10 mg/kg/day for

rimonabant and 30 mg/kg/day for 14g and 14h administered as food

admix. During the treatment, body weight (BW) and food intake (FI)

are daily measured. Total body fat and lean mass were measured

noninvasively (by MRS). Liver triglyceride content was measured by

MRS from isolated liver harvested and kept frozen in dry ice. Satellite

groups of three rats each treated with either 14g or 14h were reserved for

PK determination after 10 days of treatment.

After 24 days of treatment, body fat content (fast mass) was measured

noninvasively by MRS method; blood was collected for plasma

parameters measurement and leptin plasma levels were measured by

ELISA assay (BioVendor, Heidelberg, Germany). OGTT was

performed after overnight fasting. Glucose challenge (2 g/kg BW)

was given as bolus per gavage and blood collected at several time-points

post glucose load (+15/+30 + 60′ and +120′) to measure blood glucose

(by GlucoTrend) and plasma insulin (by ELISA). Afterward, organs

REFERENCES

■

(1) Touwn, M. The religious and medicinal uses of Cannabis in China,

India and Tibet. J. Psychoactive Drugs 1981, 13, 23−34.

U

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

(2) Gaoni, Y.; Mechoulam, R. Isolation, Structure, and Partial

Synthesis of an Active Constituent of Hashish. J. Am. Chem. Soc.

1964, 86, 1646−1647.

models of obesity. J. Clin. Invest. 2010, 120, 3735−3735. (c) Tarzia, G.;

LoVerme, J.; Duranti, A.; Tontini, A.; Spadoni, G.; Mor, M.; Rivara, S.;

Stella, N.; Xu, C.; Piomelli, D. Synthesis and characterization of a

peripherally restricted CB(1) cannabinoid antagonist, URB447, that

reduces feeding and body-weight gain in mice. Bioorg. Med. Chem. Lett.

2009, 19, 639−643. (d) Barth, F.; Hortala, L.; Rinaldi-Carmona, M.;

Congy, C.; Boulu, L.; Sadoun, F.; Fabre, G.; Finance, O. Rational design

of a novel peripherally restricted, orally active CB(1) cannabinoid

antagonist containing a 2,3-diarylpyrrole motif. Bioorg. Med. Chem. Lett.

2010, 20, 4573−4577. (e) Hogberg, T.; Receveur, J. M.; Murray, A.;

Linget, J. M.; Norregaard, P. K.; Cooper, M.; Bjurling, E.; Nielsen, P. A.

Conversion of 4-cyanomethyl-pyrazole-3-carboxamides into CB1

antagonists with lowered propensity to pass the blood−brain barrier.

Bioorg. Med. Chem. Lett. 2010, 20, 453−457. (f) Hogberg, T.; Cooper,

M.; Receveur, J. M.; Bjurling, E.; Norregaard, P. K.; Nielsen, P. A.; Skold,

N. Exploring SAR features in diverse library of 4-cyanomethylpyrazole-

3-carboxamides suitable for further elaborations as CB1 antagonists.

Bioorg. Med. Chem. Lett. 2010, 20, 26−30. (g) Sasmal, P. K.; Reddy, D.

S.; Talwar, R.; Venkatesham, B.; Balasubrahmanyam, D.; Kannan, M.;

Srinivas, P.; Kumar, K. S.; Devi, B. N.; Jadhav, V. P.; Khan, S. K.; Mohan,

P.; Chaudhury, H.; Bhuniya, D.; Iqbal, J.; Chakrabarti, R. Novel

pyrazole-3-carboxamide derivatives as cannabinoid-1 (CB1) antago-

nists: journey from non-polar to polar amides. Bioorg. Med. Chem. Lett.

2011, 21, 562−568.

(20) Dow, L. D.; Carpino, P. A.; Gautreau, D.; Hadcock, J. R.; Iredale,

P. A.; Kelly-Sullivan, D.; Lizano, J. S.; O’Connor, R. E.; Schneider, S. R.;

Scott, D. A.; Ward, K. A. Design of a potent CB1 receptor antagonist

series: potential scaffold for peripherally-targeted agents. ACS Med.

Chem. Lett. 2012, 3, 397−401.

(21) Mayweg, A.; Marty, H. P.; Mueller, W.; Narquizian, R.; Neidhart,

W.; Pﬂieger, P.; Roever, S. Novel CB1 receptor inverse agonists. WO

2004060870 A1, 2004.

(22) (a) Kuo, G.-H.; Prouty, C.; Wang, A.; Emanuel, S.; DeAngelis, A.;

Zhang, Y.; Song, F.; Beall, L.; Connolly, P. J.; Karnachi, P.; Chen, X.;

Gruninger, R. H.; Sechler, J.; Fuentes-Pesquera, A.; Middleton, S. A.;

Jolliffe, L.; Murray, W. V. Synthesis and structure−activity relationships

of pyrazine−pyridine biheteroaryls as novel, potent, and selective

vascular endothelial growth factor receptor-2 inhibitors. J. Med. Chem.

2005, 48, 4892−4909. (b) Andjelkovic, M.; Benardeau, A.; Chaput, E.;

Hebeisen, P.; Nettekoven, M.; Obst Sander, U.; Panousis, C. G.; Roever,

S. 3-Pyridinecarboxamide and 2-pyrazinecarboxamide derivatives as

HDL-cholesterol raising agents. WO 2008040651 A1, 2008.

(3) Devane, W. A.; Dysarz, F. A., III; Johnson, M. R.; Melvin, L. S.;

Howlett, A. C. Determination and characterization of a cannabinoid

receptor in rat brain. Mol. Pharmacol. 1988, 34, 605−613.

(4) Matsuda, L. A.; Lolait, S. J.; Brownstein, M. J.; Young, A. C.;

Bonner, T. I. Structure of a cannabinoid receptor and functional

expression of the cloned cDNA. Nature 1990, 346, 561−564.

(5) Gerard, C.; Mollereau, C.; Vassart, G.; Parmentier, M. Nucleotide

sequence of a human cannabinoid receptor cDNA. Nucleic Acids Res.

1990, 18, 7142.

(6) Munro, S.; Thomas, K. L.; Abu-Shaar, M. Molecular character-

ization of a peripheral receptor for cannabinoids. Nature 1993, 365, 61−

65.

(7) Devane, W. A.; Hanus, L.; Breuer, A.; Pertwee, R. G.; Stevenson, L.

A.; Griffin, G.; Gibson, D.; Mandelbaum, A.; Etinger, A.; Mechoulam, R.

Isolation and structure of a brain constituent that binds to the

cannabinoid receptor. Science 1992, 258, 1946−1949.

(8) Sugiura, T.; Kondo, S.; Sukagawa, A.; Nakane, S.; Shinoda, A.; Itoh,

K.; Yamashita, A.; Waku, K. 2-Arachidonoylglycerol: a possible

endogenous cannabinoid receptor ligand in brain. Biochem. Biophys.

Res. Commun. 1995, 215, 89−97.

́

(9) Pacher, P.; Batkai, S.; Kunos, G. The endocannabinoid system as an

emerging target of pharmacotherapy. Pharmacol. Rev. 2006, 58, 389−

462.

(10) Guindon, J.; Hohmann, A. G. The endocannabinoid system and

cancer: therapeutic implication. Br. J. Pharmacol. 2011, 163, 1447−1463.

(11) Mallat, A.; Teixeira-Clerc, F.; Deveaux, V.; Manin, S.; Lotersztajn,

S. The endocannabinoid system as a key mediator during liver diseases:

new insights and therapeutic openings. Br. J. Pharmacol. 2011, 163,

1432−1440.

(12) Montecucco, F.; Di Marzo, V. At the heart of the matter: the

endocannabinoid system in cardiovascular function and dysfunction.

Trends Pharmacol. Sci. 2012, 33, 331−340.

(13) Abood, M.; Barth, F.; Bonner, T. I.; Cabral, G.; Casellas, P.;

Cravatt, B. F.; Devane, W. A.; Elphick, M. R.; Felder, C. C.; Herkenham,

M.; Howlett, A. C.; Kunos, G.; Mackie, K.; Martin, B. R.; Mechoulam,

R.; Pertwee, R. G. Cannabinoid receptors: CB1 receptor. IUPHAR

database (IUPHAR-DB); International Union of Basic and Clinical

Pharmacology: Kansas City, Kansas, 2013; http://www.iuphar-db.org/

DATABASE/ObjectDisplayForward?objectId=56 and references cited

therein.

(23) The stability of a mixture of 0.3 g potassium hydroxide powder in

3 mL of DMSO was tested in an accelerating rate calorimeter under

pseudoadiabatic conditions. Decomposition occurred above 145 °C and

led to a fast rise in temperature (64.6 °C/min) and pressure. From the

obtained data, we extrapolate time to maximum rate of 8 h at 136 °C and

24 h at 119 °C. Reaction temperatures of 100 °C may be acceptable if

proper safety procedures are followed.

(14) Janero, D. R.; Makriyannis, A. Cannabinoid receptor antagonists:

pharmacological opportunities, clinical experience, and translational

prognosis. Expert Opin. Emerging Drugs 2009, 14, 43−65.

(15) Sam, A. H.; Salem, V.; Ghatei, M. A. Rimonabant: From RIO to

Ban. J. Obes. 2011, Article ID 432607.

(16) FDA Brieﬁng Document NDA 21-888, Zimulti (rimonabant)

Tablets, 20 mg, Sanoﬁ Aventis, Advisory Committee; U.S. Food and Drug

Administration: Silver Spring, MD, June 13, 2007; http://www.fda.gov/

ohrms/dockets/ac/07/brie ﬁ ng/2007-4306b1-fda-backgrounder.pdf

(17) Proietto, J.; Rissanen, A.; Harp, J. B.; Erondu, N.; Yu, Q.;

Suryawanshi, S.; Jones, M. E.; Johnson-Levonas, A. O.; Heymsfield, S.

B.; Kaufman, K. D.; Amatruda, J. M. A clinical trial assessing the safety

and efficacy of the CB1R inverse agonist taranabant in obese and

overweight patients: low-dose study. Int. J. Obes. 2010, 34, 1243−1254.

(18) Aronne, L. J.; Finer, N.; Hollander, P. A.; England, R. D.; Klioze, S.

S.; Chew, R. D.; Fountaine, R. J.; Powell, C. M.; Obourn, J. D. Efficacy

and safety of CP-945,598, a selective cannabinoid CB1 receptor

antagonist, on weight loss and maintenance. Obesity 2011, 19, 1404−

1414.

(19) (a) Fulp, A.; Bortoff, K.; Seltzman, H.; Zhang, Y.; Mathews, J.;

Snyder, R.; Fenell, T.; Maitra, R. Design and synthesis of cannabinoid

receptor 1 antagonists for peripheral selectivity. J. Med. Chem. 2012, 55,

2820−2834. (b) Tam, J.; Vemuri, V. K.; Liu, J.; Batkai, S.;

Mukhopadhyay, B.; Godlewski, G.; Osei-Hyiaman, D.; Ohnuma, S.;

Ambudkar, S. V.; Pickel, J.; Makriyannis, A.; Kunos, G. Peripheral CB1

cannabinoid receptor blockade improves cardiometabolic risk in mouse

(24) Wishka, D. G.; Graber, D. R.; Seest, E. P.; Dolak, L. A.; Han, F.;

Watt, W.; Morris, J. Stereoselective Synthesis of Furo[2,3-c]pyridine

Pyrimidine Thioethers, A New Class of Potent HIV-1 Non-nucleoside

Reverse Transcriptase Inhibitors. J. Org. Chem. 1998, 63, 7851−7859.

(25) Alig, L.; Alsenz, J.; Andjelkovic, M.; Bendels, S.; Ben

Bleicher, K.; Bourson, A.; David-Pierson, P.; Guba, W.; Hildbrand, S.;

Kube, D.; Lubbers, T.; Mayweg, A. V.; Narquizian, R.; Neidhart, W.;

́

ardeau, A.;

̈

Nettekoven, M.; Plancher, J.-M.; Rocha, R.; Rogers-Evans, M.; Rover, S.;

̈

Schneider, G.; Taylor, S.; Waldmeier, P. Benzodioxoles: Novel

Cannabinoid-1 Receptor Inverse Agonists for the Treatment of Obesity.

J. Med. Chem. 2008, 51, 2115−2127.

(26) Kuhn, B.; Mohr, P.; Stahl, M. Intramolecular Hydrogen Bonding

in Medicinal Chemistry. J. Med. Chem. 2010, 53, 2601−2611.

(27) Data for rimonabant can be found in the registration dossiers, e.g.:

Scientiﬁc Discussion; European Medicines Agency: London, 2006;

http://www.ema.europa.eu/docs/en_GB/document_library/

EPAR_-_Scienti ﬁ c_Discussion/human/000666/WC500021284.pdf

(28) A summary of the CEREP data can be found in the Supporting

Information.

V

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

(29) Flamment, M.; Gueguen, N.; Wetterwald, C.; Simard, G.;

̀

Malthiery, Y.; Ducluzeau, P. H. Effects of the cannabinoid CB1

antagonist rimonabant on hepatic mitochondrial function in rats fed a

high-fat diet. Am. J. Physiol. Endocrinol. Metab. 2009, 297, E1162−

E1170.

(30) Ahima, R. S.; Flier, J. S. Adipose Tissue as an Endocrine Organ.

Trends Endocrinol. Metab. 2000, 11, 327−332.

(31) Poirier, B.; Bidouard, J.-P.; Cadrouvele, C.; Marniquet, X.; Staels,

B.; O’Connor, S. E.; Janiak, P.; Herbert, J.-M. The anti-obesity effect of

rimonabant is associated with an improved serum lipid profile. Diabetes,

Obes. Metab. 2005, 7, 65−72.

(32) Macedo, I. C.; Medeiros, L. F.; Oliveira, C.; Oliveira, C. M.;

Rozisky, J. R.; Scarabelot, V. L.; Souza, A.; Silva, F. R.; Santos, V. S.;

Cioato, S. G.; Caumo, W.; Torres, I. L. Cafeteria diet-induced obesity

plus chronic stress alter serum leptin levels. Peptides 2012, 38, 189−196.

(33) Chen, R. Z.; Frassetto, A.; Lao, J. Z.; Huang, R. R.; Xiao, J. C.;

Clements, M. J.; Walsh, T. F.; Hale, J. J.; Wang, J.; Tong, X.; Fong, T. M.

Pharmacological evaluation of LH-21, a newly discovered molecule that

binds to cannabinoid CB1 receptor. Eur. J. Pharmacol. 2008, 584, 338−

342.

(34) Cluny, N. L.; Vemuri, V. K.; Chambers, A. P.; Limebeer, C. L.;

Bedard, H.; Wood, J. T.; Lutz, B.; Zimmer, A.; Parker, L. A.;

Makriyannis, A.; Sharkey, K. A. A novel peripherally restricted

cannabinoid receptor antagonist, AM6545, reduces food intake and

body weight, but does not cause malaise, in rodents. Br. J. Pharmacol.

2010, 161, 629−642.

(35) Tam, J.; Cinar, R.; Liu, J.; Godlewski, G.; Wesley, D.; Jourdan, T.;

Szanda, G.; Mukhopadhyay, B.; Chedester, L.; Liow, J. S.; Innis, R. B.;

Cheng, K.; Rice, K. C.; Deschamps, J. R.; Chorvat, R. J.; McElroy, J. F.;

Kunos, G. Peripheral cannabinoid-1 receptor inverse agonism reduces

obesity by reversing leptin resistance. Cell Metab. 2012, 16, 167−179.

(36) Pang, Z.; Wu, N. N.; Zhao, W.; Chain, D. C.; Schaffer, E.; Zhang,

X.; Yamdagni, P.; Palejwala, V. A.; Fan, C.; Favara, S. G.; Dressler, H. M.;

Economides, K. D.; Weinstock, D.; Cavallo, J. S.; Naimi, S.; Galzin, A.

M.; Guillot, E.; Pruniaux, M. P.; Tocci, M. J.; Polites, H. G. The central

cannabinoid CB1 receptor is required for diet-induced obesity and

rimonabant’s antiobesity effects in mice. Obesity 2011, 19, 1923−1934.

(37) Quarta, C.; Bellocchio, L.; Mancini, G.; Mazza, R.; Cervino, C.;

Braulke, L. J.; Fekete, C.; Latorre, R.; Nanni, C.; Bucci, M.; Clemens, L.

E.; Heldmaier, G.; Watanabe, W.; Leste-Lassere, T.; Maitre, M.;

Tedesco, L.; Fanelli, F.; Reuss, S.; Klaus, S.; Srivastava, R. K.; Monory,

K.; Valerio, A.; Grandis, A.; De Giorgio, R.; Pasquali, R.; Nisoli, E.; Cota,

D.; Lutz, B.; Marsicano, G.; Pagotto, U. CB1 Signaling in Forebrain and

Sympathetic Neurons Is a Key Determinant of Endocannabinoid

Actions on Energy Balance. Cell Metab. 2010, 11, 273−285.

(38) Mukhopadhyay, P.; Rajesh, M.; Bat

Liaudet, L.; Evgenov, O. V.; Mackie, K.; Hasko,

cannabinoid receptors promote oxidative stress and cell death in murine

models of doxorubicin-induced cardiomyopathy and in human

cardiomyocytes. Cardiovasc. Res. 2010, 85, 773−784.

́

kai, S.; Patel, V.; Kashiwaya, Y.;

G.; Pacher, P. CB1

́

(39) Cusack, K. P.; Breinlinger, E. C.; Fix-Stenzel, S.-R.; Stoﬀel, R. H.;

Woller, K. R. Novel oxadiazole compounds. WO 2011071570 A1, 2011.

(40) Cheng, Y.; Prusoff, W. H. Relationship between the inhibition

constant (K1) and the concentration of inhibitor which causes 50%

inhibition (I50) of an enzymatic reaction. Biochem. Pharmacol. 1973, 22,

3099−3108.

(41) Seglen, P. O. Preparation of isolated rat liver cells. Methods Cell

Biol. 1976, 13, 29−83.

W

dx.doi.org/10.1021/jm4010708 | J. Med. Chem. XXXX, XXX, XXX−XXX

Article Doi

6-alkoxy-5-aryl-3-pyridinecarboxamides, a new series of bioavailable cannabinoid receptor type 1 (CB1) antagonists including peripherally selective compounds

DOI: 10.1021/jm4010708

Source and publish data:

Authors:

Article abstract of DOI:10.1021/jm4010708

Full text of DOI:10.1021/jm4010708

Products guided by the article

R&D Labs maybe for 912454-29-6

Relevant to this article

Hot Product