The 3,7-diazabicyclo[3.3.1]nonane scaffold for subtype selective nicotinic acetylcholine receptor (nAChR) ligands. Part 1: The influence of different hydrogen bond acceptor systems on alkyl and (hetero)aryl substituents

Bioorganic & Medicinal Chemistry 21 (2013) 7283–7308

Contents lists available at ScienceDirect

Bioorganic & Medicinal Chemistry

journal homepage: www.elsevier.com/locate/bmc

The 3,7-diazabicyclo[3.3.1]nonane scaffold for subtype selective

nicotinic acetylcholine receptor (nAChR) ligands. Part 1:

The inﬂuence of different hydrogen bond acceptor systems

on alkyl and (hetero)aryl substituents

a,b

b

c

d

a,b,

⇑

Christoph Eibl , Isabelle Tomassoli , Lenka Munoz , Clare Stokes , Roger L. Papke , Daniela Gündisch

a

Pharmaceutical Institute, Pharmaceutical Chemistry I, University of Bonn, An der Immenburg 4, D-533121 Bonn, Germany

Department of Pharmaceutical Sciences, The Daniel K. Inouye College of Pharmacy, University of Hawai’i at Hilo, 34 Rainbow Drive, Hilo, HI 96720, USA

Department of Pharmacology, School of Medical Sciences, The University of Sydney, NSW 2006, Australia

Department of Pharmacology and Therapeutics, College of Medicine, University of Florida, Gainesville, FL 32610, USA

b

c

d

a r t i c l e i n f o

a b s t r a c t

Article history:

Available online 5 October 2013

3,7-Diazabicyclo[3.3.1]nonane is a naturally occurring scaffold interacting with nicotinic acetylcholine

receptors (nAChRs). When one nitrogen of the 3,7-diazabicyclo[3.3.1]nonane scaffold was implemented

in a carboxamide motif displaying a hydrogen bond acceptor (HBA) functionality, compounds with higher

⁄

Keywords:

afﬁnities and subtype selectivity for

sulfonamide, urea) had a strong impact on nAChR interaction. High afﬁnity ligands for

a

4b2 were obtained. The nature of the HBA system (carboxamide,

3

,7-Diazabicyclo[3.3.1]nonane

⁄

a

4b2 possessed

Bispidine

Nicotinic acetylcholine receptor

nAChR

small alkyl chains, small un-substituted hetero-aryl groups or para-substituted phenyl ring systems along

with a carboxamide group. Electrophysiological responses of selected 3,7-diazabicyclo[3.3.1]nonane

derivatives to Xenopus oocytes expressing various nAChR subtypes showed diverse activation proﬁles.

Compounds with strongest agonistic proﬁles were obtained with small alkyl groups whereas a shift to

partial agonism/antagonism was observed for aryl substituents.

Structure–activity relationship

1

. Introduction

membrane potential and the release of diverse neurotransmitters,

which is often subtype speciﬁc and can vary across areas of a spe-

1

–4

Nicotinic acetylcholine receptors (nAChRs) are ligand-gated

ciﬁc organ, for example, different brain areas.

NAChRs have been

excitatory cation channels and can form homomeric or hetero-

proposed as potential drug targets for the treatment of various cen-

1

–4

5–7

meric subtypes.

nation of ﬁve identical or different subunits (

and ) capable of forming a water-ﬁlled pore in the cell mem-

brane.

Each nAChR complex is composed of a combi-

tral nervous system (CNS) diseases.

nAChRs have a wide-spread expression in the brain and subunit

combination with 4, 5, 6, and/or b3 are implemented in

depression and addiction.

For example, b2-containing

a

1– 10, b1–b4, , d

a

c

e

a

1

–4

8–11

Stimulation of these receptors have an impact on

The majority of nAChR ligands have been derived from very po-

tent natural products displaying the important pharmacophoric

features found in the neurotransmitter acetylcholine (ACh) 1.⁶

Cation-p/HB forces along with HBA interactions corresponding to

the charged/protonable nitrogen and, for example, carbonyl or het-

3 3

Abbreviations: BBB, blood–brain barrier; CD OD, tetradeuteromethanol; CDCl ,

,7,12

deuterochloroform; CH

deuterium oxide; DCC, N,N -dicyclohexylcarbodiimide; DMAP, 4-(dimethyl-

2

Cl

2

, dichloromethane; CNS, central nervous system; D

2

O,

0

amino)pyridine; DME, 1,2-dimethoxyethane; DMF, N,N-dimethylformamide; Et

diethyl ether; Et N, triethylamine; EtOAc, ethyl acetate; HBA, hydrogen bond

acceptor; HCl, hydrogen chloride; HEPES, 4-(2-hydroxyethyl)piperazine-1-ethane-

sulfonic acid; HSS, HEPES-buffered salt solution; K CO , potassium carbonate; KBr,

potassium bromide; KMnO , potassium permanganate; KOH, potassium hydroxide;

MeCN, acetonitrile; MeI, iodomethane; MeOH, methanol; MgSO magnesium

sulfate; nAChR, nicotinic acetylcholine receptor; NaHCO sodium hydrogen

2

O,

3

eroaryl moieties, respectively, are involved in the interaction with

13–18

the receptors.

2

3

(

Di)azabicyclic octane and diazabicyclic nonane scaffold bear-

4

,

ing compounds are the majority representing the nAChR pharma-

6

,7

3

,

copeia in the recent past.

Among these bicyclic N-containing

carbonate; NaOH, sodium hydroxide; Pd/C, palladium on activated charcoal; PE,

petroleum ether; PEI, poly(ethyleneimine); Ro5, rule of ﬁve; THF, tetrahydrofuran;

TPSA, topological polar surface area; TRIS, tri(hydroxymethyl)aminomethane;

6,7

structural templates is 3,7-diazabicyclo[3.3.1]nonane. It is a nat-

urally occuring scaffold also known as bispidine found, for exam-

ple, in the nAChR ligand cytisine and sparteine, a sodium ion

2

ZnBr , zinc bromide.

⇑

Corresponding author. Tel.: +1 808 933 2943; fax: +1 808 933 2974.

E-mail address: danielag@hawaii.edu (D. Gündisch).

19

channel blocker. 3,7-Diazabicyclo[3.3.1]nonane derivatives have

0

http://dx.doi.org/10.1016/j.bmc.2013.09.059

7

284

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

been the subject of considerable interest in the ﬁeld of novel

nAChR compounds, but also as, for example, antiarrhythmic drugs,

opioid receptor ligands, and as multidentate ligands for organome-

but limited to a carboxamide HBA motif and limited substitution

pattern have been done by Targacept, Inc. which resulted in the

development of the

a4b2 agonist TC-6683/AZD1446 bearing the

2

0–37

27–29

tallic compounds.

The most prominent 3,7-diazabicy-

diazabicyclic system 3,7-diazabicyclo[3.3.0]octane.

This com-

clo[3.3.1]nonane compound interacting with nAChRs is the

pound showed positive effects on a rodent model for working

memory. Our extensive study around the bispidine scaffold started

with bispidine (3,7-diazabicyclo[3.3.1]nonane) itself which was

tested for afﬁnities at different nAChRs and functional properties.

Simple 3,7-diazabicyclo[3.3.1]nonane compounds with different

HBA systems, like carboxamides, sulfonamides and urea groups at-

tached to simple alkyl or (heter)aryl substituents were synthesized

(Fig. 2) to get more insight into structure activity relationships for

nAChRs. Compounds were tested for their afﬁnities, and selected

candidates displaying nAChR afﬁnity in the nanomolar range were

additionally evaluated for their functionality in Xenopus oocyte

experiments.

⁄

natural product cytisine 3 with high afﬁnity for

a

4b2 nAChR

3

8,39

(Fig. 1; Table 1).

Cytisine 3, its derivative 3-(pyridine-3-yl)-

cytisine (3PC) 4, recently developed by us, and varenicline 5

Ò

(

Chantix , used for smoking cessation) (Fig. 1), derived from cyti-

4

0–

sine, have antidepressant-like effects in mice (Fig. 1; Table 1).

All three compounds possess high afﬁnity and partial agonist

functionality for the

4

3

⁄

a

4b2 subtype to a various degree, but the

rigid compounds cytisine 3 and varenicline 5 have limited subtype

4

1–43

selectivity.

Since the intact cytisine template is limited in its synthetic ac-

cess, chemical space, ADMET properties, afﬁnity and functional

selectivity proﬁles, we revisited the 3,7-diazabicyclo[3.3.1]nonane

ring system (bispidine) as a nAChR ligand itself. This scaffold is

synthetically easily accessible via double Mannich reaction and

displays one pharmacophoric element. To explore the chemical

space around this scaffold further, 3,7-diazabicyclo[3.3.1]nonane

was attached to a variety of aromatic and non-aromatic substitu-

tents via different HBA motifs. In contrast to cytisine 3 where the

HBA system (pyridone) is fused to the 3,7-diazabicyclo[3.3.1]non-

ane scaffold, the HBA motifs of the synthesized derivatives are acy-

clic, thus compounds display higher ﬂexibility. A similar approach,

2

. Results and discussion

.1. Chemistry

The 3,7-diazabicyclo[3.3.1]nonane (bispidine) scaffold was syn-

2

4,25,46–48

thesized according to a previously published method.

N-

0

Benzyl-N -tboc-bispidinone 10 was obtained by a double Mannich

reaction from commercially available tert-butyl 4-oxopiperidine

carboxylate 9, benzylamine, and paraformaldehyde. It was puriﬁed

by ﬂash chromatography.

0

The reduction of the carbonyl functionality of N-benzyl-N -tboc-

bispidinone 10 was attempted by several published methods but in

O

our hands none of them produced more than 33% yield of the de-

N

O

N

CH₃

0

O

CH₃

sired N-benzyl-N -tboc-bispidine 11. We found the Huang–Minlon

N

variation of the Wolff–Kishner reduction suitable to produce 11 in

yields as high as 73%, if the reaction temperature would not exceed

1

2

NH

O

N

HBA linker

NH

5

N X

3

4

Substituent

Figure 1. Structures of acetylcholine 1, the natural products nicotine 2, and cytisine

, 3-(pyridine-3-yl)-cytisine (3PC) 4 and the FDA approved drug varenicline 5.

3

Figure 2. 3,7-Diazabicyclo[3.3.1]nonane based compounds.

Table 1

i

Afﬁnities (K values) and calculated physicochemical parameters of known nAChR ligands 3–7

⁄

ClogP^e

logBB^f

Compound

a4b2

a3b4

a7

(

a1)

2

b1

c

d

M

r

TPSA

K

i

[nM]

K

i

[nM]

K

i

[nM]

i

K [nM]

3

4

5

(Cyt)^a

0.122

0.91

0.06

19

119

240

250

1100

322

1300

>5000

3540

190.24

267.14

211.26

0.17

1.03

0.74

32.34

45.23

37.81

ꢀ0.25

ꢀ0.23

ꢀ0.06

b

(3PC)

(Var)^c

1

3

7

385

2000

19,000

194.27

ꢀ0.14

32.34

ꢀ0.10

4

2450

10,000

>50,000

208.30

0.26

23.55

ꢀ0.12

d

From Ref.: 44.

From Refs.: 41,44,45.

From Ref.: 42.

a,b

c

e

logP values have been calculated using the ACD/Labs Algorithm.

logP calculated with the ACD/Labs Algorithm was used to calculate logBB. TPSA: topological polar surface area.

f

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7285

2

4,25,49

0

1

40 °C.

The desired intermediate N-benzyl-N -tboc-bispidine

could easily be puriﬁed and cleaved from the N-tboc group in a

1

1 was obtained after puriﬁcation by ﬂash chromatography.

N-benzylbispidine 12 could be obtained by cleaving the N-tboc

ﬁnal step (Scheme 1).

0

The N-benzyl protecting group was cleaved from N-benzyl-N -

protection group under acidic conditions, for example, with hydro-

chloric acid in 1,4-dioxane. Exceeding 140 °C in the above men-

tioned Wolff-Kishner reaction (Huang–Minlon variation) also

lead to N-benzylbispidine 12 but side product formation hampered

the puriﬁcation process. N-benzylbispidine 12 could also be used

as an intermediate to further substitute the secondary amine of

tboc-bispidine 11 using palladium on activated charcoal (Pd/C)

5% as a catalyst under a hydrogen atmosphere. The resulting

N-tboc-bispidine 13 was obtained in quantitative yields and no

further puriﬁcation was necessary besides ﬁltering off the

catalyst.

N-tboc-Bispidine 13 was used as a starting material to introduce

a wide variety of moieties to the bispidine backbone (Scheme 1).

Different synthetic approaches were used to synthesize N-tboc

protected 3,7-diazabicyclo[3.3.1]nonane carboxamides, sulfona-

1

2, but the puriﬁcation of the N-benzyl protected intermediates

was often more difﬁcult and cumbersome than the puriﬁcation

of the N-tboc protected intermediates. Furthermore, the ﬁnal N-

benzyl deprotection often produced low yields depending on the

nature of the substituent. Therefore, the N-benzyl protection group

was cleaved ﬁrst leading to N-tboc protected intermediates, which

5

0–54

mides or urea compounds (Scheme 1).

The N-tboc protected

intermediates Boc-8, Boc-15-63, and Boc-67–69 were isolated

and puriﬁed, but not analyzed and described in detail.

Scheme 1. Synthesis of compounds. Reagents and conditions: (a) (CH

Dean Stark trap, 140 °C, 8 h; (c) HCl/1,4-dioxane (4 M), rt, 12 h; (d) Pd/C (5%), H

Et N, toluene, rt, 2 h; (g) carbamoyl chloride, Et N, toluene, rt, 2 h; (h) CDI, THF, reﬂux, 2 h = 14; then MeI, MeCN, THF, rt, 24 h, then R-COOH, Et

COOH, DCC, DMAP, CH Cl , 0 °C to rt, 12 h; for Boc-56: 4-hydroxybenzoic acid, DCC, THF, rt, 24 h; (j) 64, 65 or 66, MeI, MeCN, rt, 24 h; then 13, CH

,4-dioxane (4 M), rt, 4 h; (l) anhyd ZnBr , CH Cl , rt, 12–120 h; (m) CDI, THF, reﬂux, 16 h; (n) alkyl iodide, K CO , DMF, rt, 12 h.

2

O)

n

, BnNH

2

, AcOH, MeOH, [Ar] reﬂux, 6 h; (b) N

2

H

4

(80%), NaOH, diethylene glycol, 125 °C, 2 h, then

N, toluene, rt, 2 h; (f) sulfonyl chloride,

N, MeCN, rt, 12–120 h; (i) R-

Cl , Et N, rt, 24 h.; (k) HCl/

2

(2–4 bar), MeOH, rt, 4–24 h; (e) carboxyl chloride, Et

3

2

3

1

2

3

7

286

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

Boc-8 and Boc-15–33 intermediates were synthesized by ami-

3,7-diazabicyclo[3.3.1]nonane (bispidine) itself is interacting with

⁄

nolysis of N-tboc-bispidine and various carboxylic acid chlorides.

Due to the simplicity of this reaction and the fact that many car-

boxylic acids are commercially available in form of their acid chlo-

rides, we preferred this method. Puriﬁcation was accomplished by

ﬂash chromatography. Aminolysis was also used to synthesize N-

tboc protected sulfonamides Boc-34–40 and the urea compounds

nAChRs and has a K

12 showed afﬁnity for the

i

value of 600 nM for

a

4b2 . N-Benzyl-bispidine

⁄

a3b4 subtype in the high nanomolar

range (K

i

= 569.6 nM) but no afﬁnity for the other subtypes tested.

⁄

In contrast, N-tboc-bispidine 13 has high afﬁnity for the

a4b2 sub-

type in the low nanomolar range (K

i

= 45 nM) whereas afﬁnity for

= 1.3 M).

⁄

the

a3b4 subtype was in the low micromolar range (K

i

l

0

Boc-41–42. 1,1 -Carbonyldiimidazole (CDI) as coupling reagents

In general, all active compounds except for the intermediate N-

⁄

was used when certain carboxylic acid chlorides were not avail-

benzyl-bispidine 12 showed higher afﬁnity for the

a

4b2 nAChR

able. For the CDI method, N-tboc-bispidine 13 was allowed to react

with CDI, and N-carbonylimidazolyl-N -tboc-bispidine 14 was iso-

lated after chromatographic puriﬁcation. Then, 14 was activated

3

with an excess of MeI, and Et N and the carboxylic acid were added

subsequently. Flash chromatography provided the N-tboc pro-

tected bispidinecarboxamides Boc-44–55. DCC was used as an

alternative method, when CDI did not produce satisfying yields.

Using the DCC method, N-tboc-bispidine 13, the carboxylic acid,

DCC and N,N-dimethylaminopyridine (DMAP) were dissolved in

subtype than for the other subtypes tested.

0

As we reported before, acetylbispidine 8 can be considered as a

simpliﬁed cytisine 3 analog missing the pyridone moiety, but still

displaying the same pharmacophoric components as 3 (Fig. 1),

the protonable secondary nitrogen and the carbonyl functionality

2

4–26

which serves as a HBA system.

Compound 8 can also be con-

sidered as a more rigid and stable analog of ACh 1. In contrast to

the rigid cytisine 3 (Table 1), acetylbispidine 8 showed reduced

afﬁnity for all subtypes measured (Table 2), but still preference

⁄

2

CH Cl

2

at 0 °C and allowed to react at rt for 12 h. After ﬁltering

for the

a

4b2 subtype (K

i

= 5.6 nM) and higher afﬁnity than tetra-

off the dicyclohexyl urea by-product, the N-tboc protected 3,7-

diazabicyclo[3.3.1]nonane carboxamides were puriﬁed by ﬂash

chromatography. Occasionally, a second ﬁltration or a second chro-

matographic step was required to eliminate residual dicyclohexyl

urea by-product.

For the synthesis of Boc-67–69, the three aliphatic, cyclic

amines (pyrrolidine, morpholine and 4-benzylpiperidine) were al-

lowed to react with CDI. The carbonylimidazole adducts 64–66

were obtained after extraction, and activated with methyliodide,

subsequently. The activated adducts were coupled with N-tboc-

bispidine 13, and the N-tboc protected bispidine urea derivatives

Boc-67–69 were puriﬁed by ﬂash chromatography.

hydrocytisine 6 (K

Acyclic and small cyclic alkyl substituents: Methylene homologs

15 and 16 of acetylbispidine 8 provided compounds with similar

i

= 17 nM), an analog with intermediate rigidity.

⁄

high afﬁnity for the

a

4b2 receptor (K

i

for 8: 5.6 nM; 15: 4.5 nM;

16: 3.6 nM). The butan-1-one homolog 16 exhibited higher afﬁnity

⁄

for the

a

3b4 subtype compared to 8, but still displayed an about

⁄

74-fold higher afﬁnity for the

spidine 8 showed similar afﬁnity for

a

4b2 receptor. In contrast, acetylbi-

⁄

a3b4 and a7 receptors.

Compound 17, bearing an isopropyl chain, had lower afﬁnities for

⁄

the

i i

a4b2 (K = 50.3 nM) and the a3b4 subtypes (K = 828 nM)

than its unbranched isomer 16. A cyclopentyl moiety (compound

43) and its homolog bearing a cyclopentylmethyl moiety (com-

⁄

A small homologous series 69–71 (methyl, ethyl, propyl) of

acetylbispidine 8 was prepared to investigate the inﬂuence of the

pound 55) showed afﬁnities for the

molar range (K

a

4b2 subtype in the low nano-

i

for 43: 10.8 nM; 55: 43.5 nM) whereas its

secondary versus a tertiary amine group at the 3,7-diazabicy-

cyclohexyl homolog 18 was inactive at all nAChR subtypes tested.

These data indicate that small hydrocarbon moieties in close prox-

⁄

clo[3.3.1]nonane scaffold on the interaction with

a4b2 nAChRs.

The compounds were obtained by alkylating acetylbispidine 8 with

imity to the carbonyl functionality contribute to high afﬁnity for

⁄

alkyl iodides in the presence of K CO in DMF. The ﬁnal products

2

3

the

a

4b2 nAChR subtype. Compounds 8, 15–17, and 43 have also

⁄

could be obtained after ﬂash chromatography. N-Methyl-acetyl-

bispdine 70 has been synthesized before but was neither tested

been synthesized and evaluated by Targacept, Inc. (8: r

a

4b2 :

⁄

K

i

K

i

K

i

K

i

K

i

= 18.7 nM; h

= 12.2 nM; h

= 8.1 nM; h

= 101.6 nM; h

a

4b2 : K

i

= 20.2 nM; r

= 10.4 nM; r

a

7 : 3277.9 nM; 15: r

4b2 :

5

⁄

for nAChR afﬁnity nor have analytical data been published.

4b2 : K

i

7 : 795.1 nM; 16: r

4b2 :

⁄

The cleavage of the N-tboc protecting group from Boc-8, Boc-15–

3, and Boc-69–71 was accomplished by using acidic conditions at

4b2 : K

i

= 2.9 nM; r

= 44.4 nM; r

= 20.2 nM; r

a

7 : 230.3 nM; 17: r

4b2 :

⁄

6

a

4b2 : K

i

a

7 : 644.7 nM; 43: r

4b2 :

⁄

22,23,28

room temperature, either hydrogen chloride in 1,4-dioxane or anhy-

= 26.6 nM; h

a

4b2 : K

i

a

7 : 211.2 nM).

Com-

5

6

drous ZnBr

2

2 2

in CH Cl .

Both methods resulted in the desired

pound 18 has only been claimed in this patent but no biological

data have been published. The use of a different radioligand and

different tissues for the membrane preparation may account for

bispidine derivatives 8, 15–63, and 67–69 after alkaline workup.

Most compounds were transferred into their corresponding salt

forms, either fumaric or hydrochloric acid salts for in vitro testing.

This is indicated as ‘F’ or ‘H’ for the compounds in the Section 4 and

the compounds are also analyzed in their salt forms. The salt for-

mation step was advantageous for further purifying our ﬁnal

compounds from residual N-tboc-protected 3,7-diazabicyclo[

i

some, but minor differences in K values compared to our results.

It is known that a tertiary amine motif in cytisine, displayed in

the naturally occurring compound caulophylline, reduces the

⁄

37

a

4b2 nAChR afﬁnity. Similar observations have also been made

for tetrahydrocytisine 6 and its N-methyl analog 7. To study the

more ﬂexible compounds bearing the same diazabicyclic scaffold

attached to the same HBA linker, we evaluated three N-alkyl ana-

3

.3.1]nonane derivatives or other by-products. Compounds 35–

8, 40, 48, 62, 63, and 70–72 were used as free bases.

logs of acetylbispidine (cpds 70–72). There is a dramatic decrease

⁄

2

.2. Biological activity and structure–activity relationship (SAR)

of

a

4b2 afﬁnity from N-methyl (K

i

= 200 nM) to N-propyl (K

i

>

5000 nM; Table 3). Only the methyl group was tolerated but pro-

⁄

The afﬁnities of the 3,7-diazabicyclo[3.3.1]nonane derivatives 8,

2–13, 15–63, and 67–72 were determined in radioligand binding

duced a compound with a 35-fold lower afﬁnity for the

subtype.

a4b2

1

38,39,57

studies as previously described.

tive tissue from Sprague–Dawley rat forebrains (

Membrane fractions of na-

Simple aryl substituents: With the introduction of an aromatic

moiety at the 3,7-diazabicyclo[3.3.1]nonane carboxamide tem-

plate (19), a reduction of afﬁnity was observed compared to acet-

ylbispidine 8, but 19 displayed higher afﬁnity comparing with its

⁄

a

4b2 ,

a

7 ), calf

1) b1 d)

b1 d) and

⁄

adrenals (

a

3b4 ) or Torpedo californica electroplax ((

a

2

c

3

⁄

were prepared and [ H]epibatidine (

a

4b2 ,

7 ) were used as radioligands.

values of 3,7-diazabicyclo[3.3.1]nonane (bispidine), the

two intermediates 12 and 13, and of the ﬁnal compounds 8,

5–63, and 67–72 are summarized in Tables 2 and 3. The scaffold

a

3b4 , (

a

1)

2

c

3

⁄

[

H]MLA (

a

saturated cyclohexyl derivative 18. Benzoylbispidine 19 exhibited

⁄

The K

i

afﬁnity for the

(K

subtypes tested.

a

4b2 nAChR subtype in the nanomolar range

i

= 453.6 nM) and did not show any afﬁnity for the other receptor

1

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7287

Table 2

i

Afﬁnities (K values ± SEM) and calculated physicochemical parameters (logP, TPSA, logBB) of bispidine, the bispidine intermediates 12 and 13 and of the bispidine carboxamide

derivatives 8, 15–33, and 43–63

⁄

ClogP^b

logBB^c

0.01

Compd

R =

H

a

4b2

a

3b4

a

7

(

a

i

1)

2

b1

c

d

M

r

TPSA

K

i

[nM]

K

i

[nM]

K

i

[nM]

K [nM]

Bispidine

600.0 ± 23

>1000

126.20

216.32

0.06

24.06

1

2

3

>10,000

569.6 ± 150.0

>5000

>10,000

1.88

1.18

15.27

41.57

0.23

0.08

1

45.0 ± 5.3

1276.5 ± 9.3

>5000

226.31

8

1

5.6 ± 1.4

4.5 ± 2.3

663.8 ± 40.1

328.7 ± 70,2

636^a

n. d.

168.24

182.26

-0.14

0.39

32.34

- 0.02

0.00

5

>2000

>1000

1

6

3.6 ± 0.7

265.9 ± 83.4

828.0^a

>1000

>5000

n. d.

196.29

0.92

0.73

32.34

0.04

7

50.3 ± 10.4

43

55

18

10.8 ± 0.8

43.5

n. d.

222.33

236.35

1.35

1.88

1.92

32.34

0.11

0.22

0.28

n. d.

>5000

>5,000

>10,000

19

44

20

21

45

22

46

23

453.6 ± 14.8

>10,000

>5000

>10,000

n. d.

>10,000

n. d.

>10,000

n. d.

230.30

244.33

264.75

275.30

248.30

264.75

298.30

275.30

0.56

1.02

1.67

0.06

0.82

1.36

1.62

0.47

32.34

81.17

32.34

81.17

ꢀ0.09

ꢀ0.13

ꢀ0.01

ꢀ0.52

ꢀ0.05

ꢀ0.11

0.03

>10,000

>5,000

n. d.

>10,000

n. d.

>10,000

n. d.

386.4 ± 42.7

>1,000

n. d.

147.1 ± 6.8

>2000

>3000

>10,000

n. d.

>10,000

ꢀ0.47

(

continued on next page)

7

288

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

Table 2 (continued)

⁄

ClogP^b

logBB^c

Compd

R =

a

4b2

[nM]

a

3b4

[nM]

a

7

(

a

i

1)

2

b1

c

d

M

r

TPSA

K

i

K

i

K

i

[nM]

K [nM]

2

4

5

221.0 ± 84.0

>10,000

>5,000

>10,000

244.33

286.41

1.02

32.34

ꢀ0.13

2

>10,000

2.25

2.21

32.34

0.12

⁄

4

7

39.9 ± 7.1

1089.5 ± 290.5

306.40

ꢀ0.65

26

27

28

56

197.3

>10,000

n. d.

>10,000

>1000

>10,000

n. d.

248.30

264.75

309.20

246.30

0.78

1.33

1.50

0.20

32.34

52.57

ꢀ0.06

-0.12

ꢀ0.25

130.2

138.0 ± 17.3

104.6

>10,000

n. d.

48

49

29

20.7 ± 8.9

15.9 ± 3.9

>1000

>10,000

4200

>10,000

1726

>5000

>10,000

255.31

275.30

290.36

0.22

0.52

1.24

56.13

81.17

50.80

ꢀ0.12

-0.46

>5000

ꢀ0.01

5

0

7

454.8 ± 89.4

73.2 ± 9.1

>10,000

231.29

232.28

ꢀ0.60

ꢀ1.00

45.23

58.12

ꢀ0.07

ꢀ0.11

5

1

8

291.6 ± 50.1

>5000

>1,000

>10,000

n. d.

>10,000

n. d.

310.19

265.74

0.62

0.24

45.23

-0.19

-0.42

5

194

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7289

Table 2 (continued)

⁄

ClogP^b

logBB^c

Compd

R =

a4b2

a3b4

a7

(a1)

2

b1c

d

M

r

TPSA

K

i

[nM]

K

i

[nM]

K

i

[nM]

i

K [nM]

30

31

59

21.4 ± 2.3

17.6 ± 0.8

661.6 ± 47.3

2242.5

>10,000

n. d.

>10.000

>10,000

n. d.

220.27

236.33

233.31

0.53

45.48

60.58

37.27

0.04

>10,000

n. d.

0.88

-0.11

ꢀ0.11

ꢀ0.81

60

61

62

>10,000

1800

>10,000

n. d.

>10,000

n. d.

>10,000

n. d.

269.34

0.12

0.49

48.13

ꢀ0.33

ꢀ0.39

ꢀ0.43

6

3

2

3

205.07

n. d.

270.33

280.36

0.17

1.79

61.02

32.34

ꢀ0.21

ꢀ0.37

ꢀ0.31

>10,000

n. d.

>10,000

447.2 ± 100.5

>10,000

5

2

3

>1000

n. d.

>10,000

281.35

0.84

0.66

45.23

ꢀ0.36

ꢀ0.39

162.1 ± 38.1

>5000

5

4

81.3 ± 20.6

>5000

>10,000

n. d.

281.35

0.48

45.23

-0.42

⁄

Values are generated from 2 to 10 independent experiments; = radioligand binding was increased; n. d. = not determined.

a

n = 2.

b

ClogP values have been calculated using the ACD/Labs Algorithm.

logP calculated with the ACD/Labs Algorithm was used to calculate logBB.

c

Additional substitution at the aromatic part provided important

22 (3-chloro) and 23 (3-nitro) were inactive at this receptor sub-

type. This seems to indicate that small electron withdrawing groups

(ﬂuoro 45, triﬂuoromethyl 46) are still tolerated in the meta posi-

tion, whereas bulkier electron withdrawing groups (chloro 22, nitro

23) are not.

⁄

insight into structural requirements for

a

4b2 nAChR ligands. The

steric input caused by substituents at the ortho position resulted

in a complete loss of activity. Substitution at the ortho position

compounds 44, 20, and 21) is likely to force the aromatic ring to

(

an out-of-plane arrangement which seems to be not tolerated by

the nAChRs evaluated. Compounds with a substitution at the meta

Compounds with substitution at the para position of the phenyl

ring 24-28, 47-49, and 56 exhibited higher afﬁnities for a4b2

⁄

position of the phenyl ring 45 (3-ﬂuoro) and 46 (3-triﬂuoromethyl)

nAChR in most cases compared with the meta-substituted

ligands except for the bulky 4-tert-butyl group (25, K values

>10,000 nM), or if an additional ortho substituent was present

⁄

showed afﬁnity in the nanomolar range for the

= 386.4 nM and K

a

4b2 subtype

i

(K

i

= 147.1 nM, respectively), but compounds

7

290

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

Table 3

Afﬁnities (K

i

values ± SEM) and calculated physicochemical parameters (logP, TPSA, logBB) of the bispidine sulfonamides 34–40, bispidine urea derivatives 41–42, 67–69, and

alkylated acetylbispidine derivatives 70–72

⁄

ClogP^b

logBB^c

Compd

R =

a

4b2

[nM]

a

3b4

[nM]

a

7

(

a

i

1)

2

b1

c

d

M

r

TPSA

K

i

K

i

K

i

[nM]

K [nM]

3

4

5

>5000

n. d.

>10,000

n. d.

204.29

266.36

ꢀ0.04

57.79

ꢀ0.03

ꢀ0.07

3

>10,000

1.64

3

6

7

970.4

>10,000

n. d.

291.37

280.39

1.58

2.10

81.58

57.79

0.10

0.13

3

858.3 ± 66.7

3

4

8

9

0

645.1

>10,000

n. d.

>10,000

n. d.

291.37

345.26

311.36

1.54

2.67

1.84

81.58

57.79

0.08

0.25

>5000

n. d.

135.2 ± 8.1

5700

>10,000

106.62

ꢀ0.20

4

1

2

499.3

>1,000

n. d.

197.28

225.33

0.32

1.38

35.58

ꢀ0.01

4

>3000

0.13

6

7

8

>5000

3800

>10,000

n. d.

>10,000

n. d.

>10,000

n. d.

223.31

239.31

0.76

35.58

44.81

ꢀ0.07

6

ꢀ0.22

6

9

>10,000

>5000

327.46

3.31

35.58

0.50

70

71

72

Methyl

Ethyl

n-Propyl

200

>5000

n. d.

182.26

196.29

210.31

0.25

0.78

1.31

23.55

0.03

0.09

0.16

Values are generated from 2 to 10 independent experiments; ^an = 2; n. d. = not determined.

b

ClogP values have been calculated using the ACD/Labs Algorithm.

logP calculated with the ACD/Labs Algorithm was used to calculate logBB.

c

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7291

⁄

(

2,4-dimethoxy compound 29). Halogen substituents (compound

2-yl derivative (59) showed less afﬁnity for the a4b2 subtype

⁄

2

6, 27, and 28) increased the afﬁnity for

= 453.6 nM). The 4-chloro 27 and 4-bro-

mo 28 analogs showed very similar K values (27: K = 130.2 nM;

= 138 nM) and slightly higher afﬁnities comparing with the

a4b2 receptors compared

compared to 30 and 31, which could be due to its N-methyl substi-

tuent mimicking an ortho-substituent. In contrast to 44 (2-methyl-

phenyl), which is lacking any afﬁnity, 59 displayed a K value of

i

661.6 nM for this subtype. Bispidine amide derivatives substituted

to benzoylbispidine 19 (K

i

2

8: K

i

4

K

-ﬂuoro derivative 26. (K

i

= 197.3 nM). A methyl group (24,

with small ﬁve-membered aromatic ring systems (30 and 31),

⁄

i

= 221 nM) also enhances afﬁnity for

a

4b2 receptors while keep-

including hetero atoms with hydrogen bond acceptor properties,

⁄

ing high subtype selectivity. Interestingly, the nitrile group (com-

pound 48) was capable to increase

are likely to enhance the afﬁnity for

a

4b2 nAChR.

⁄

a

4b2 afﬁnity by a factor of

Fused heteroaryl substituents: The inﬂuence of fused aryl systems

(naphthoyl derivatives 32 and 33) was already mentioned above,

but the introduction of an additional nitrogen atom at various posi-

tions created some additional important aspects. If the nitrogen is

in close proximity to the carbonyl function (e.g., 2-quinolinyl com-

pound 52, 2-indole compound 60, or 3-indole compound 61), the

afﬁnity drops enormously or even disappears, and this is regardless

of the HB nature of this nitrogen. Interestingly, the naphthoyl

ten compared to 24 (methyl group) along with high subtype selec-

tivity. This group, displaying a polarized triple bond, has a tiny ste-

ric demand and is about eight times smaller than a methyl group.

Because of its strong dipole character, polar interactions or hydro-

gen bonds are possible. Compared to the 4-cyanobenzoylbispidine

4

8, the 4-nitro group (compound 49) showed a similar K

i

value for

⁄

the

playing K

a

4b2 subtype (K

i

= 15.9 nM), but lower subtype selectivity dis-

⁄

i

values for

a

3b4 and

a7

subtypes in the micromolar

derivative 33, showed afﬁnity in the higher nanomolar range for

⁄

range. Compound 56 bearing a hydroxy group in the para position,

which can serve as a hydrogen bond donor, showed also high afﬁn-

ity (K

tron withdrawing groups exhibited increased afﬁnity for the

a

substituent in position 4 had the same effect on afﬁnity for

a

if the aromatic part directly attached to the carbonyl function is

considered as an arenologue spacer. Fused ring systems (com-

pound 32 and 33) showed no activity or reduced afﬁnity, respec-

a

4b2 (K

i

= 447.2 nM), whereas the nitrogen in 52 causes a dra-

>1000 nM). In contrast, the quinolin-6-yl analog

54 displayed an increase in afﬁnity (K = 81.3 nM) compared to its

non nitrogen containing naphthoyl derivative 33. The afﬁnity of

matic drop (K

i

⁄

i

= 104.6 nM) for the

a

4b2 nAChR subtype. In general, elec-

i

⁄

4b2 nAChR subtype. Also, the introduction of a

p

-electron rich

the naphthoyl derivative 32, which showed

a

K

i

value of

⁄

>10,000 nM for

a4b2 could be improved by the introduction of a

⁄

4b2 . In addition, it provides insight about spatial requirements

nitrogen observed in the quinolin-4-yl analog 53 displaying a K

i

va-

⁄

lue of 162.1 nM for

a4b2 . In summary, fused N-heteroaryl systems

with nitrogen atoms in close proximity to the HBA/carbonyl group

⁄

showed a reduced afﬁnity for the

a

4b2 subtype compared to com-

tively, compared to the biphenyl derivative 47 (K

i

= 39.9 nM). The

pounds bearing the nitrogen atom in a longer distance to this

1

-naphthoyl derivative 32 mimicking an ortho/meta substitution

group.

was lacking any afﬁnity for the subtypes tested, whereas 2-naph-

HBA linkers: To investigate the inﬂuence of the carboxamide lin-

ker as the HBA group, we replaced it by a sulfonamide or urea

group. As shown in Table 3 the two sulfonamides 34 and 35 do

not exhibit any afﬁnity at the tested receptor subtypes. Com-

thoyl derivative 33, which mimicks a meta/para substitution dis-

⁄

played

a

similar

K

i

value for

a4b2

i

(K = 447 nM) as

benzoylbispidine 19. Except for the 4-biphenyl derivative 47

⁄

(

l

a

M;

3b4 : K

i

= 1.1

l

M) and the 4-nitro derivative 49 (

l

a

3b4 : K

i

= 4.2 -

pounds 36 (3-cyano), 37 (4-methyl), and 38 (4-cyano) exhibited

⁄

a7 : K

i

= 1.7

M), no compound showed activity for the other

afﬁnity for the

a

4b2 subtype in the high nanomolar range (36:

tested receptor subtypes. Taken together, these data show that

derivatives with a substitution at the ortho position of the phenyl

ring are not tolerated by the receptor, regardless of the nature of

the substituent. Derivatives with a substituent at the meta position

are only tolerated if the substituents are small electron withdraw-

ing groups. Bulkier electron withdrawing substituents are not tol-

erated. And derivatives with a substitution at the para position

showed the broadest tolerance. Both, small electron donating as

K

i

= 970.4 nM; 37: K

i

= 858.3 nM; 38: K

i

= 645.1 nM), while show-

ing no afﬁnity for the other nAChR subtypes tested. Interestingly,

the small electron withdrawing 3-cyano group (compound 36)

exhibited a similar low afﬁnity than the small electron donating

4-methyl group (compound 37). The small electron withdrawing

4-cyano group (compound 38) displayed a slightly higher afﬁnity

for the

position of the phenyl ring (compound 40) resulted in a K

135.2 nM for the

nitro group by a bromo atom at the para position (compound

39), caused a non-active compound for the

⁄

a

4b2 nAChR. The introduction of a nitro group at the para

i

value of

⁄

well as electron withdrawing groups, resulted in compounds with

a

4b2 subtype. In contrast, the exchange of this

⁄

high afﬁnity for the

a

4b2 nAChR subtype.

⁄

Heteroaryl substitutents: Diverse heteroaryl containing deriva-

a

4b2 subtype. Within

tives (30–31, 50-54, and 57-63) were tested for their ability to

compete with [ H]epibatidine or [ H]MLA (Table 2). The 3-pyridyl

this subset of bispidine sulfonamides, only compound 40 showed a

3

⁄

weak afﬁnity for the

a

3b4 subtype (K

i

= 5.7

lM). It appears that

moiety (compound 50) has an almost identical afﬁnity for the

either small electron withdrawing groups in the meta or para posi-

⁄

a4b2 nAChR subtype compared to 19, also displaying high

tion or electron donating groups in the para position generate com-

⁄

subtype selectivity. An additional bromo substituent in the 5

pounds with afﬁnity for the

a

4b2 nAChR. However, it is not

position (compound 51) increases afﬁnity slightly, and the incor-

explainable, why the 4-bromo compound is inactive at the nAChR,

whereas the 4-nitro compound showed afﬁnity in the nanomolar

range. So, sulfonamide derivatives 34–35 and 37–40 compared

poration of an additional nitrogen (pyrazine derivative 57) had a

⁄

strong inﬂuence on the afﬁnity for

a

4b2 (K

i

= 73.2 nM) together

with a high subtype selectivity proﬁle. Other ring equivalents like

with their corresponding carboxamide analogs 8, 19, 24, 48, 28,

and 49 (see Table 2) exhibited lower or no afﬁnity for the a4b2

subtype. Thus, the replacement of the carboxamide linker by a sul-

⁄

2

-furanyl (30) and 2-thiophenyl (31) showed afﬁnity in the low

⁄

nanomolar range for the

1: K

a4b2 receptor subtype (30: K

i

= 21.4 nM;

3

i

= 17.6 nM). Whereas compound 31 displayed high subtype

fonamide group erases or decreases the afﬁnity of the compounds

⁄

selectivity, the furanyl containing compound 30 showed afﬁnity

for the

a

4b2 subtype. The sulfonamide linker has a weaker HB ba-

⁄

for the

a

3b4 subtype, but in the micromolar range (K

i

= 2.2

lM).

sicity than the carboxamide, but this difference is too small to ex-

plain the strong inﬂuence on afﬁnity. The impact of change in

linker geometry and therefore the orientation of the HBA system

in space seem to be of much higher importance.

Since both an acyclic amide and acyclic urea have about the

same HB basicity, compounds with urea linker were investigated

for afﬁnity. The N,N-dimethyl urea compound 41 showed an

Targacept, Inc. also synthesized and evaluated the 2-furanyl analog

⁄

3

0 as a nAChR ligand and the reported afﬁnities (r

a

4b2 : K

i

= 31 -

4b2 :

3

⁄

nM using [ H]nicotine on rat brain membrane fractions; h

= 9.9 nM using [ H]nicotine on SH-EP1 clonal cell membrane

a

3

K

i

⁄

3

fractions; r

tions) correspond well with our data.

a7 : 15 lM using [ H]MLA on rat brain membrane frac-

22,23,28

The N-methyl-pyrrol-

7

292

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

⁄

afﬁnity in the nanomolar range (K

i

= 499.3 nM) for the

a4b2 sub-

1

μM applied alone

type, but the homologous N,N-diethyl urea analog 42 lost its afﬁn-

ity. Interestingly, compound 41 displayed a 10-fold lower afﬁnity

0.08

⁄

for the

a

4b2 nAChR than the corresponding carboxamide analog

1

7. From the three cyclic urea compounds 67–69 only the morpho-

⁄

0.06

lino compound 68 had afﬁnity for the

only in the micromolar range (K = 3.8 micromolar). The carboxam-

ide analog 43 exhibited high afﬁnity in the low nanomolar range

= 10.8 nM), whereas its urea analog 67 was not active. Taken to-

a4b2 subtype, however,

i

0

.04

.02

(K

i

gether this emphasises that the urea group cannot replace the car-

boxamide linker at the bispidine backbone and that bulkier alkyl

groups are not tolerated by the receptor. Similar observations have

0

⁄

been obtained in the afﬁnity evaluation of

a

7 ligands bearing the

1

,4-diazabicyclo[3.2.2]-nonane scaffold which was directly linked

0.00

bispidine

13

48

via a sulfonamide or urea motif to a spectrum of phenyl substitu-

5

8

ents. The afﬁnity dropped enormously in this case in contrast

compounds

to a carbamate functionality used.⁵⁸

⁄

Selected compounds showing afﬁnity for

a

4b2 in the lower

1

0 μM applied alone

nanomolar range were evaluated electrophysiologically in an ini-

tial characterization for their functionality using human and

mouse (for muscle) nAChRs expressed in Xenopus oocytes

α4β2*

α3β4*

α7

5

9–61

(Fig. 3).

The observed responses should help to promote cer-

1.0

tain compounds for more detailed and complete functional evalu-

ation in the future. Most compounds showed a tendency towards a

⁄

partial agonistic/antagonistic proﬁle for

a4b2 except compound

α1β1εδ

1

a

6 bearing a simple alkyl chain. Possible agonistic effects for

3b4 were found for compounds 15, 16 and 43. All of them are

⁄

0.5

displaying small acyclic and cyclic substitutents. For compounds

, 16, and 43, belonging to the same structural category, agonism

at 7 was observed. There were only weak or no effects on the

muscle type. The scaffold 3,7-diazabicyclo[3.3.1]nonane (bispi-

dine) showed weak activation of 7, and even lower activity at

8

a

0

.0

a

8

15

16

43

55

47

56

31

other nAChR subtypes. When a spacer motif was introduced, for

compounds

example, a methylene motif for compound 43, obtaining com-

⁄

pound 55, the agonistic effect at

having essentially no effect at

activity at

showed the tendency towards agonism at

for compound 16), and 7, whereas aromatic substituents dis-

a

3b4 and

a7 of 43 changed to

⁄

1 μM co-applied with ACh

a

3b4 and apparent antagonistic

7 and muscle types. In general, small alkyl substituents

a

0

.6

⁄

a

4b2 , a3b4 (strongest

α4β2*

α3β4*

α7

a

played partial agonism/antagonism (e.g. compound 47). Several re-

search groups have used the bispidine scaffold for the development

of nAChR ligands like mentioned before, but most of them were fo-

0.4

α1β1εδ

0

.2

.0

cused on the development of a4b2 agonists and the chemical space

used was limited. Our ﬁrst part of an extensive SAR investigation

around the bispidine scaffold showed that a carboxamide HBA sys-

tem was preferred over other HBA systems investigated here. Para-

substitution in the phenyl group series was best tolerated and

0

could serve as a starting point for further evaluation of the chem-

-0.2

⁄

ical space for

a

4b2 ligands. Compounds 47 and 55 gave ﬁrst hints

1

3

8

15 16 43 55 47 56 48 31

compounds

that the introduction of a spacer motif has an important impact on

functionality leading to ligands with a possible antagonistic proﬁle

speculating about an additional binding site accessible through a

spacer motif. Therefore, a more detailed SAR examination on

spacer motifs will be presented in another study (‘‘part 2’’). Both

studies were the foundation for the development of in vivo active

compounds recently published.⁶²

Figure 3. Responses of oocytes expressing diverse nAChR subtypes to 1 or 10 lM of

selected compounds (numbering are following the sequence in the tables) relative

to ACh control responses. Responses of oocytes expressing diverse nAChRs to

compounds co-applied at 1 lM with ACh compared to responses to ACh alone. Bars

above zero indicate additive effects; bars below zero indicate reduced responses.

2

.3. Physicochemical properties and druglikeness

Supplementary data) for compounds showing nanomolar afﬁnity

for a4b2 (K <1000 nM). Most active compounds showed ‘good’

i

⁄

64

ACD/ADME Suite 5.0 (ACD/Labs) software has been used to cal-

or ‘acceptable’ values for most parameters calculated. There were

no correlations between afﬁnity and ClogP or TPSA values. The

logBB values can also be used to estimate the likelihood for

blood–brain barrier (BBB) penetration. Values below ꢀ0.5 would

reﬂect very poor or no BBB penetration and >0.7 very high pene-

trants. The cyano group is producing a borderline TPSA value,

and especially the nitro group shows the least ideal values for TPSA

culate physicochemical properties and druglikeness parameters

e.g., ClogP, TPSA, and logBB; Tables 1–3). The compounds do not

(

6

3

violate the Ro5. Some physicochemical properties associated

with drug-likeness parameters for CNS drugs are ideally between

2

50 and 350 for Mr, ClogP between 1-3, PSA <75, and more param-

eters important for CNS compounds have been calculated (Table 4;

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7293

and logBB. In addition, the nitro group is also a toxophoric group

and along with bromoaryl substituents which could also produce

toxic metabolites, both are here only used to get a fast SAR insight

regarding lipophilic input, steric aspects, and additional pharmaco-

phoric points.

an Agilent 1100 HPLC system (LC/ESI-MS) or on a Varian 500-MS

mass spectrometer (ESI-MS). The purity of the compounds was

determined by LC/ESI-MS or a Shimadzu Prominence HPLC system

at an appropriate wavelength. HRMS runs were performed on Agi-

lent Technologies 6530 Accurate-Mass Q-TOF LC/MS. All com-

pounds proved to possess P95% purity. Melting points were

determined in open capillary tubes with a melting point apparatus

3

. Conclusions

(

Weiss-Gallenkamp) or with a Melting Point B-540 (Büchi) and are

uncorrected. Infrared spectroscopy was performed with a Tensor-

7 FTIR infrared spectrometer (Bruker Optic) using KBr pellet or

with a Nicolet iS10 (Thermo Scientiﬁc). Elemental microanalyses

C, H, N) were performed with a VarioEL (Elementar Analysensys-

We prepared a simple series of 3,7-diazabicyclo[3.3.1]nonane

2

based compound series with three different HBA linkers and eval-

uated their afﬁnities and initial functionality for different nAChR

subtypes. The scaffold 3,7-diazabicyclo[3.3.1]nonane displaying

(

teme) or a Costech elemental combustion apparatus and the deter-

mined values are generally within ±0.4% of the theoretical values.

Hydrogen for hydrogenations was produced by a Hogen GC hydro-

gen generator (Proton Energy Systems) or by a 60H hydrogen gen-

erator (Parker, domnick hunter). Lyophilizations were performed

with an Alpha 1-4 LSC freeze dryer (Martin Christ).

the essential pharmacophoric element for cation-

p

/HB interaction

⁄

has an afﬁnity for

nM) and weak

a

4b2 in the higher nanomolar range (K

i

= 600 -

a

7 agonism. Intermediates in the synthetic pathway

of this series like N-tboc-bispidine 13 and N-benzyl-bispidine 12

are active at nAChRs and display different subtype preferences.

Structure–activity relationship studies revealed that the carbox-

amide linker was preferred over a sulfonamide or urea motif. Ac-

tive carboxamide derivatives showed selectivity for the

Compounds in their salt form are named with ‘F’ or ‘H’ along

with their number, and they are also analyzed in their salt forms.

⁄

a4b2

nAChR except for small hydrocarbon substituents which exhibit

⁄

4

.1. General procedure A: synthesis of N-tboc protected

high afﬁnity for the

a

4b2 nAChR subtype but comparably low

⁄

bispidinecarboxamides, bispidinesulfonamides, or bispidine

urea derivatives

selectivity over

a3b4 and

a7

subtypes. These compounds dis-

played also more agonistic proﬁles for the neuronal subtypes.

Para-substituted benzoylbispidines with small electron withdraw-

⁄

The appropriate carboxyl chloride, sulfonyl chloride, or carbam-

oyl chloride (1 mmol), either neat or dissolved in dry toluene (1–

ing groups were well tolerated by the

a4b2 nAChR displaying

nanomolar afﬁnities. The impact of heteroaryl substituents (ﬁve-

⁄

2

mL), was added dropwise to a stirred solution of N-tboc-bispidine

membered, six-membered, fused) on

a

4b2 afﬁnity were depen-

1

3 (230 mg, 1 mmol) and Et N (101 mg, 1 mmol) in dry toluene

3

dent on size and the position of the heteroatom.

Further SAR studies will focus on the inﬂuence of an additional

spacer motif between the HBA system and the attached substituent

moiety.

(

5 mL) at rt. The volatiles were removed under reduced pressure

after 2 h and the residue was puriﬁed by ﬂash chromatography

silica gel, mixtures of CH Cl and MeOH—40:1, 20:1 or 9:1).

(

2

In summary, the 3,7-diazabicyclo[3.3.1]nonane scaffold can

serve as an important starting point for the development of nAChR

compounds with diverse and desired afﬁnity and functionality

pattern.

4

.2. General procedure B: synthesis of N-tboc protected

bispidinecarboxamides

Methyl iodide (570 mg, 4 mmol) was added to a stirred solution

of 14 (320 mg, 1 mmol) dissolved in dry MeCN (2 mL) and dry THF

(2 mL) at rt. The volatiles were removed under reduced pressure

after 24 h, the residue was dissolved in dry MeCN (4 mL), and

4

. Experimental section

All reagents and solvents were obtained from various suppliers

3

Et N (101 mg, 1 mmol) and the appropriate carboxylic acid

(

ABCR, Acros, Aldrich, Alfa Aesar, Fluka, Merck or Sigma) and used

(1 mmol) were added. The solution was allowed to stir at rt for

without further puriﬁcation unless otherwise noted. Dichloro-

methane was freshly distilled from calcium hydride. Methanol

was treated with sodium, distilled afterwards and stored under

nitrogen. Sodium wires were used to dry diethyl ether, petroleum

ether, tetrahydrofuran, and toluene. Water was taken from a water

puriﬁcation system PureLab Plus UV (ELGA Labwater) or Direct-

Q™ 5 (Millipore). Amines were puriﬁed prior to use with a Kugel-

rohr distillation apparatus (Büchi). Reactions were monitored by

thin-layer chromatography (TLC) using aluminum sheets coated

with silica gel 60 F254 (Merck). Compounds were visualized using

12–120 h before the volatiles were removed under reduced pres-

sure. The residue was puriﬁed by ﬂash chromatography (silica

gel, mixtures of CH Cl and MeOH—40:1, 20:1 or 9:1).

2

4.3. General procedure C: synthesis of N-tboc protected

bispidinecarboxamides

The appropriate carboxylic acid (1 mmol) and N-tboc-bispidine

13 (248 mg, 1.1 mmol) were dissolved in dry CH Cl₂(5 mL) and

2

cooled to 0 °C. DMAP (6.1 mg, 0.05 mmol) and DCC (206 mg,

UV light (254 or 365 nm) and using a KMnO

4

solution (1% in

1 mmol) were added and the mixture was allowed to warm up

and stir at rt. The precipitate was ﬁltered off after 12 h and washed

with cold CH Cl (2 mL). The solvent of the ﬁltrate was evaporated

water). Column chromatography was carried out on silica gel

(

2 2

0.035–0.060 nm) using different mixtures of CH Cl with MeOH

2

(

40:1, 20:1 or 9:1) or of PE with EtOAc (4:1 or 3:1) as mobile

under reduced pressure and the residue was puriﬁed by ﬂash chro-

matography (silica gel, mixtures of CH Cl and MeOH—40:1. 20:1

or 9:1).

1

13

phases. H NMR spectra (400 or 500 MHz) and C NMR spectra

100 or 125 MHz) were recorded on an Avance 400 or on an Avance

00 NMR spectrometer (Bruker). All NMR spectra were recorded at

2

(

5

rt. Chemical shifts (d) are given in parts per million (ppm) relative

to the remaining protons of the deuterated solvents used as inter-

nal standard. Coupling constants J are given in Hertz (Hz) and spin

multiplicities are given as s (singlet), d (doublet), dd (doublet of

doublets), t (triplet), q (quartet), m (multiplet) and br (broad). Mass

spectra were recorded on an API 2000 mass spectrometer with an

electron spray ionization source (Applied Biosystems) coupled to

4.4. General procedure D: cleavage of the N-tboc protecting

group from N-tboc protected bispidine derivatives

HCl in 1,4-dioxane (4 M, 4-5 mL) was added to a stirred solution

of the N-tboc protected bispidine derivative (0.2–1.0 mmol), dis-

solved in 4–5 mL of 1,4-dioxane, and the mixture was allowed to stir

at rt for 2–12 h. The volatiles were removed under reduced pressure

7

294

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

before the residue was dissolved in KOH solution (0.25 M, 20 mL)

and extracted with CH Cl

(3–5 ꢁ 20 mL). The combined organic

layers were washed with saturated NaHCO solution (10 mL), water

10 mL), dried with MgSO and ﬁltered. The product was obtained

after evaporating the solvent under reduced pressure.

compound was obtained as a white solid in 94% yield; mp 272–

1

2

279 °C (dec). H NMR (400 MHz, D

2H); 3.43–3.57 (br m, 8H). C NMR (100 MHz, D O) d 24.0, 25.2,

2

O) d 2.01 (br s, 2H), 2.53 (br s,

1

3

2

(

4

46.0. HRMS for

z = 127.1126. Anal. (C

C

7

H

14

N

2

:

calcd m/z = 127.123, found m/

⁄

7

H

14

N

2

2.0HCl 0.5H

2

O) C, H, N.

4

.5. General procedure E: cleavage of the N-tboc protecting

4.10. (1R,5S)-1-(3,7-Diazabicyclo[3.3.1]nonan-3-yl)ethanone

group from N-tboc protected bispidine derivatives

fumaric acid salt (8F)

The N-tboc protected bispidine derivative (0.2–1.0 mmol) was

The N-tboc protected compound was obtained by using general

dissolved in dry CH

2

Cl

2

(5 mL), anhydrous ZnBr

2

(2–3 equiv) was

procedure A with acetyl chloride (79 mg, 1 mmol). A mixture of

added, and the mixture was allowed to stir at rt for 12–120 h. After

the removal of the volatiles under reduced pressure the residue

was dissolved in KOH solution (0.25 M, 20 mL) and extracted with

2 2

CH Cl and MeOH (9:1) was used for the chromatographic puriﬁca-

tion. After removal of the solvents a white solid (268 mg, 98%) was

obtained. The N-tboc protection group of this solid (250 mg,

0.93 mmol) was cleaved using general procedure E with anhydrous

CH

with saturated NaHCO

MgSO , and ﬁltered. The product was obtained after evaporating

the solvent under reduced pressure.

2

Cl

2

(5 ꢁ 20 mL). The combined organic layers were washed

3

solution (10 mL), water (10 mL), dried over

2

ZnBr (420 mg, 1.86 mmol) for 48 h and an off white solid 8

4

(146 mg, 93%) was obtained after extraction. This solid 8 (48 mg,

0.29 mmol) was transferred to its fumaric acid salt 8F by using

the general procedure G with fumaric acid (33 mg, 0.29 mmol).

Compound 8F (66 mg, 69%) was obtained as a white solid in 63%

4

.6. General procedure F: formation of fumaric acid salts of

1

bispidine derivatives

yield over three steps; mp 152–156 °C (dec). H NMR (500 MHz,

2

D O) d 1.93 (br m, 1H), 1.99 (br m, 1H), 2.16 (s, 3H), 2.32 (br m,

The amine (0.2–1.0 mmol) was dissolved in a mixture of Et

2

O

2H), 3.06 (br d, J = 14.1 Hz, 1H), 3.32 (br m, 2H), 3.44 (br d,

J = 13.1 Hz, 1H), 3.50, (br d, J = 13.4 Hz, 1H), 3.54 (br d,

J = 13.4 Hz, 1H), 4.05 (br d, J = 13.2 Hz, 1H), 4.35 (br d, J = 14.0 Hz,

and MeOH (9:1, 2–5 mL). A saturated solution of fumaric acid in

the same mixture of solvents was added dropwise to a stirred solu-

tion of the amine until no further precipitation was observed. The

solution was kept at 4-8 °C overnight before the precipitate was ﬁl-

tered off and washed with the same mixture of solvents (2 ꢁ 5 mL)

1

3

1H), 6.68 (s, 2.5H). C NMR (125 MHz, D

2

O) d 24.3, 28.0, 28.4,

30.1, 48.7, 50.3, 50.6, 52.8, 137.4, 173.6, 178.9. LC/ESI-MS: positive

+

ꢀ1

mode m/z = 169.1 ([M+H] ). Purity (>99.9%). IR (KBr, cm ) 3455,

⁄ ⁄

9 16 2 4 4 4 2

H N O 1.25C H O 1.15H O) C,

2

and dry Et O (5 mL). The solid was dissolved in water (20-30 mL),

1696, 1624, 984, 975. Anal. (C

H, N.

and freeze-dried.

4

.7. General procedure G: formation of fumaric acid salts of

4.11. (1R,5S)-tert-Butyl 7-benzyl-9-oxo-3,7-

bispidine derivatives

diazabicyclo[3.3.1]nonane-3-carboxylate (10)

The amine (0.2–1.0 mmol) was dissolved in isopropanol (3–

mL), ﬁltered, and heated to 70–80 °C. The same molar amount

of fumaric acid was dissolved in isopropanol (3 mL) and also

Compound 10 was obtained following the synthetic procedures

4

6

5

published by Stead et al. A solution of tert-butyl 4-oxopiperidine-

1-carboxylate (10.0 g, 50.2 mmol), acetic acid (2.9 mL,

9

heated to 70–80 °C. The two solutions were combined and allowed

50.9 mmol), and benzylamine (5.5 mL, 50.4 mmol) in methanol

(40 mL) was added dropwise to a stirred suspension of paraformal-

dehyde (3.32 g, 110.6 mmol). The resulting mixture was heated

under reﬂux for 1 h before another portion of paraformaldehyde

(3.32 g, 110.6 mmol) was added. This mixture was heated at reﬂux

for additional 5 h before the reaction was allowed to cool to rt and

the solvent was evaporated under reduced pressure. The residue

to cool to rt. Dry Et

2

O (10 mL) were added and the mixture was

kept at 4–8 °C overnight. The solid was ﬁltered off, washed with

2

dry Et O (3 ꢁ 5 mL), dissolved in water (20–30 mL), and freeze-

dried.

4

.8. General procedure H: synthesis of alkyl substituted

acetylbispidine derivatives

was dissolved in Et

solution (1 M, 2 ꢁ 80 mL). The combined aqueous layers were ex-

tracted with Et

O (3 ꢁ 50 mL). The combined organic layers were

dried with MgSO , ﬁltered, and the solvent was evaporated under

2

O (150 mL) and washed with aqueous KOH

Acetylbispidine 8 (110 mg, 0.65 mmol) was dissolved in 4 mL of

2

dry DMF and K

2

CO

3

(90.4 mg, 0.65 mmol) and the appropriate al-

4

kyl iodide (0.65 mmol) were added. The reaction was allowed to

stir at rt for 12 h before the volatiles were removed under reduced

pressure. The residue was dissolved in aqueous KOH solution (1 M,

reduced pressure. Flash column chromatography (silica gel, PE:E-

tOAc 3:1) of the residue afforded a white solid 10 (13.0 g, 78%) after

1

extensive evaporation of the solvents; mp 83 °C.

(500 MHz, CDCl

H NMR

5

mL) and extracted with CH

layers were dried with MgSO

moved under reduced pressure. The residue was puriﬁed by ﬂash

chromatography on silica gel using a mixture of CH Cl and MeOH

20:1 or 9:1) as eluents. After evaporating the solvents the ﬁnal

products were obtained as clear oils.

2

Cl

2

(3 ꢁ 5 mL). The combined organic

3

) d 1.53 (s, 9H), 2.42 (br m, 2H), 2.70 (br m, 2H),

4

, ﬁltered, and the solvent was re-

3.17 (br m, 2H), 3.27 (br d, J = 12.5 Hz, 1H), 3.35 (br d, J = 12.5 Hz,

1H), 3.52 (br m, 2H), 4.41 (br d, J = 12.9 Hz, 1H), 4.57 (br d,

1

3

2

3

J = 12.9 Hz, 1H), 7.24-7.38 (m, 5H). C NMR (125 MHz, CDCl ) d

(

28.7, 47.7, 50.0, 50.6, 58.8, 59.1, 62.0, 80.2, 127.4, 128.5, 128.9,

137.5, 154.9, 213.6. LC/ESI-MS: positive mode m/z = 331.3

+

ꢀ1

(

[M+H] ). Purity (>98.5%). IR (KBr, cm ) 1731, 1695. Anal.

4

.9. 3,7-Diazabicyclo[3.3.1]nonane dihydrochloric acid salt

(C19H N O ) C, H, N.

26 2 3

(

bispidine dihydrochloric acid salt)

4

.12. (1R,5S)-tert-Butyl 7-benzyl-3,7-diazabicyclo[3.3.1]nonane-

N-tboc-bispidine 13 (150 mg, 0.66 mmol) was dissolved in 1,4-

3-carboxylate (11)

dioxane (3 mL) and a HCl/1,4-dioxane mixture (4 M) was added.

The volatiles were removed under reduced pressure after 6 h (stir-

A solution of 10 (18.9 g, 57.2 mmol), NaOH (10.0 g, 250 mmol),

and hydrazine hydrate (80%, 10.0 mL, 160 mmol) in 150 mL of

diethylene glycol was heated at 125 °C under reﬂux conditions.

ring at rt). The residue was dissolved in 5 ml Et

2

O and ﬁltered. The

O. The ﬁnal

precipitate was washed at least three times with Et

2

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7295

After 2 h the reﬂux condenser was exchanged for a Dean-Stark

apparatus and the mixture was heated at 140 °C for additional

reﬂuxed for 2 h before the solvent was evaporated under reduced

pressure. The residue was puriﬁed by ﬂash chromatography (silica

8

h. After cooling to rt 250 mL of water were added and the result-

ing mixture was extracted with toluene (4 ꢁ 150 mL). The com-

bined organic layers were washed with saturated NaHCO

and ﬁl-

gel, mixture of CH

2 2

Cl and MeOH - 20:1). Evaporation of the sol-

vents afforded a white solid 14 (1.98 g, 6.2 mmol, 93%); mp

1

3

143 °C (dec). H NMR (500 MHz, CDCl ) d 1.43 (s, 9H), 1.87 (s,

solution (1 ꢁ 30 mL), water (2 ꢁ 30 mL), dried with MgSO

4

2H), 1.96 (br s, 2H), 3.01 (br m, 1H), 3.09 (br m, 1H), 3.26 (br s,

tered. The solvent was evaporated under reduced pressure and the

2H), 3.97 (br s, 1H), 4.25 (br m, 3H), 7.07 (s, 1H), 7.33 (s, 1H),

1

3

residue was puriﬁed by ﬂash chromatography (silica gel, PE:EtOAc

3

7.89 (s, 1H). C NMR (125 MHz, CDCl ) d 27.8, 28.5, 31.2, 47.4,

1

4

:1) to afford 11 as a white solid (13.2 g, 73%); mp 66 °C. H NMR

48.8, 50.2, 51.8, 80.3, 118.0, 129.4, 137.0, 151.8, 155.0. LC/ESI-

MS: positive mode m/z = 321.1 ([M+H] ), negative mode m/

+

(

500 MHz, CDCl

3

) d 1.52 (s, 9H), 1.61 (m, 1H), 1.66 (m, 1H), 1.79 (br

-

ꢀ1

s, 1H), 1.87 (br s, 1H), 2.16 (br d, J = 10.9 Hz, 1H), 2.22 (br d,

J = 10.9 Hz, 1H), 2.89 (br d, J = 10.8 Hz, 1H), 2.99 (br d, J = 10.9 Hz,

z = 319.9 ([M – H] ). Purity (>99.8%). IR (KBr, cm ) 1675. Anal.

) C, H, N.

24 4 3

(C16H N O

1

H), 3.05 (ddd, J = 13.1, 3.9, 1.7 Hz, 1H), 3.10 (ddd, J = 13.1, 3.9,

.7 Hz, 1H), 3.30 (d, J = 13.5 Hz, 1H), 3.44 (d, J = 13.5 Hz, 1H), 3.99

4.16. (1R,5S)-1-(3,7-Diazabicyclo[3.3.1]nonan-3-yl)propan-1-

(

5

br d, J = 13.1 Hz, 1H), 4.16 (br d, J = 13.1 Hz, 1H), 7.19-7.34 (m,

one fumaric acid salt (15F)

1

3

H). C NMR (125 MHz, CDCl ) d 28.9, 29.2, 31.3, 47.7, 48.6,

5

8.9, 59.2, 63.7, 78.9, 126.8, 128.2, 128.7, 139.1, 155.2. LC/ESI-

The N-tboc protected compound was obtained by using general

procedure A with propionyl chloride (93 mg, 1 mmol). A mixture of

CH Cl and MeOH (20:1) was used for the chromatographic puriﬁ-

2 2

+

MS: positive mode m/z = 317.1 ([M+H] ). Purity (>99.5%). IR (KBr,

cm ) 1681. Anal. (C19

ꢀ1

H

28

N

2

O

2

) C, H, N.

cation. After removal of the solvents a white solid (285 mg, 99%)

was obtained. The N-tboc protection group of this solid (210 mg,

0.74 mmol) was cleaved using the general procedure D for 12 h

and an off white solid 15 (132 mg, 97%) was obtained after extrac-

tion. This solid 15 (35 mg, 0.19 mmol) was transferred to its fuma-

ric acid salt 15F by using the general procedure G with fumaric acid

(22 mg, 0.19 mmol). Compound 15F (32 mg, 54%) was obtained as

4

.13. (1R,5S)-3-Benzyl-3,7-diazabicyclo[3.3.1]nonane fumaric

acid salt (12F)

HCl dissolved in 1,4-dioxane (4 M, 6 mL) was added to a solu-

tion of 11 (192 mg, 0.61 mmol) in 1,4-dioxane (6 mL). The mixture

was stirred at rt for 12 h before the volatiles were evaporated un-

der reduced pressure. The residue was dissolved in aqueous KOH

1

a white solid in 52% yield over three steps; mp 158-161 °C (dec). H

solution (1 M, 20 mL) and extracted with Et

combined organic layers were dried with MgSO

solvent was evaporated under reduced pressure. General proce-

2

O (3 ꢁ 20 mL). The

2

NMR (500 MHz, D O) d 1.07 (t, J = 7.5 Hz, 3H), 1.95 (br m, 1H), 2.00

(br m, 1H), 2.32 (br s, 2H), 2.50 (dt, J = 17.5, 7.5 Hz, 1H), 3.06 (br d,

J = 13.1 Hz, 1H), 3.33 (br m, 2H), 3.46 (br m, 2H), 3.53 (br d,

4

, ﬁltered, and the

dure F afforded 12F as an off-white solid (139 mg, 0.42 mmol,

J = 13.2 Hz, 1H), 4.11 (br d, J = 13.2 Hz, 1H), 4.39 (br d, J = 13.8 Hz,

1H), 6.70 (s, 2.0H). C NMR (125 MHz, D O) d 11.1, 28.1, 28.4,

2

1

13

6

8% over two steps); mp 167-170 °C (dec). H NMR (500 MHz,

MeOD) d 1.77 (br m, 1H), 1.93 (br m, 1H), 2.10 (br s, 2H), 2.46

dt, J = 11.9, 2.4 Hz, 2H), 3.13 (br m, 2H), 3.25 (dt, J = 12.8, 2.8 Hz,

29.9, 30.2, 48.9, 50.3, 50.6, 52.0, 137.5, 174.3, 182.0. LC/ESI-MS: po-

+

ꢀ1

(

sitive mode m/z = 183.4 ([M+H] ). Purity (>99.6%). IR (KBr, cm )

⁄ ⁄

18 2 4 4 4 2

H N O 1.0C H O 0.6H O)

2

5

1

H), 3.43 (br m, 2H), 3.53 (s, 2H), 6.69 (s, 2.0H), 7.26-7.38 (m,

H). ¹C NMR (125 MHz, MeOD) d 28.6, 31.1, 50.5, 59.0, 64.0,

28.6, 129.6, 130.5, 136.2, 138.2, 171.5. LC/ESI-MS: positive mode

3440, 1679, 1638, 984, 972. Anal. (C10

C, H, N.

3

+

ꢀ1

m/z = 217.4 ([M+H] ). Purity (>99.8%). IR (KBr, cm ) 3433, 1708,

4.17. (1R,5S)-1-(3,7-Diazabicyclo[3.3.1]nonan-3-yl)butan-1-one

fumaric acid salt (16F)

⁄

1

636, 986, 970. Anal. (C19

H

28

N

2

O

2

1.0C H

4 4

O

4

) C, H, N.

4

.14. (1R,5S)-tert-Butyl 3,7-diazabicyclo[3.3.1]nonane-3-

The N-tboc protected compound was obtained by using the gen-

carboxylate fumaric acid salt (13F)

eral procedure A with butyryl chloride (107 mg, 1 mmol). A mix-

ture of CH

2

Cl

2

and MeOH (20:1) was used for the

2

00 mg of Pd/C (5%) was added to a solution of 11 (1.5 g,

chromatographic puriﬁcation. After removal of the solvents a

white solid (292 mg, 97%) was obtained. The N-tboc protection

group of this solid (200 mg, 0.67 mmol) was cleaved using the gen-

eral procedure D for 12 h and a yellowish oil 16 (128 mg, 97%) was

obtained after extraction. This oil 16 (45 mg, 0.23 mmol) was

transferred to its fumaric acid salt 16F by using the general proce-

dure G with fumaric acid (27 mg, 0.23 mmol). Compound 16F

4

.74 mmol) in MeOH (7 mL) and the mixture was allowed to react

under an atmosphere of hydrogen (10 psi) at rt for 4 h. After the

mixture was ﬁltered and washed thoroughly with MeOH the sol-

vent was evaporated under reduced pressure. The product was ob-

tained as a clear oil (1.05 g, 4.64 mmol) in 98% yield. The free

amine 13 was used for further syntheses. General procedure F

was used to transfer this clear oil 13 (161 mg, 0.71 mmol) into a

(46 mg, 73%) was obtained as a white solid in 59% yield over three

1

white solid, its fumaric acid salt 13F (200 mg, 82%); mp 170 °C

dec). H NMR (500 MHz, D

steps; mp 159–161 °C (dec). H NMR (500 MHz, D

2

O) d 0.94 (t,

1

(

2

O) d 1.47 (s, 9H), 1.86 (br d,

J = 7.4 Hz, 3H), 1.58 (br m, 2H), 1.95 (br m, 1H), 2.00 (br m, 1H),

J = 13.5 Hz, 1H), 1.96 (br d, J = 13.5 Hz, 1H), 2.25 (br s, 2H), 3.17

2.32 (br s, 2H), 2.42 (br m, 1H), 2.53 (br m, 1H), 3.07 (br d,

(

br d, J = 13.2 Hz, 2H), 3.31 (br d, J = 13.2 Hz, 2H), 3.48 (br d,

J = 13.9 Hz, 1H), 3.33 (br m, 2H), 3.42-3.55 (br m, 3H), 4.14 (br d,

1

3

13

J = 13.2 Hz, 2H), 4.05 (br d, J = 13.2 Hz, 2H), 6.80 (s, 2.0H).

NMR (125 MHz, MeOD) d 28.2, 30.3, 30.5, 50.7, 50.9, 85.3, 137.6,

1

ity (>99.9%). IR (KBr, cm ) 3438, 1691, 1657, 985. Anal. (C12

O

C

J = 13.4 Hz, 1H), 4.39 (br d, J = 14.1 Hz, 1H), 6.69 (s, 2.0H).

NMR (125 MHz, D

50.2, 50.6, 52.2, 137.6, 174.5, 181.3. LC/ESI-MS: positive mode m/

C

2

O) d 15.9, 20.7, 28.1, 28.4, 30.2, 38.5, 48.7,

+

61.0, 174.4. LC/ESI-MS: positive mode m/z = 226.9 ([M+H] ). Pur-

ꢀ1

+

ꢀ1

H

22

N

2-

z = 197.4 ([M+H] ). Purity (>99.8%). IR (KBr, cm ) 3466, 1700,

⁄

2

1.0C H

4 4

O

4

0.1H

2

O) C, H, N

1650, 1619, 994, 980. Anal. (C11H N O 1.0C H O 0.3H O) C, H, N.

20 2 4 4 2

4

.15. (1R,5S)-tert-Butyl 7-(1H-imidazole-1-carbonyl)-3,7-

4.18. (1R,5S)-1-(3,7-Diazabicyclo[3.3.1]nonan-3-yl)-2-

diazabicyclo[3.3.1]nonan-3-carboxylate (14)

methylpropan-1-one fumaric acid salt (17F)

N-tboc-bispidine 13 (1.5 g, 6.63 mmol) was dissolved in dry THF

The N-tboc protected compound was obtained by using the gen-

(

20 mL) and CDI (1.18 g, 7.29 mmol) was added. The solution was

eral procedure A with isobutyryl chloride (107 mg, 1 mmol). A

7

296

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

mixture of CH

graphic puriﬁcation. After removal of the solvents a clear oil

281 mg, 95%) was obtained. The N-tboc protection group of this

2

Cl

2

and MeOH (20:1) was used for the chromato-

4.21. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(2-

chlorophenyl)methanone fumaric acid salt (20F)

(

oil (210 mg, 0.71 mmol) was cleaved using the general procedure

D for 18 h and a clear oil 17 (130 mg, 93%) was obtained after

extraction. This oil 17 (64 mg, 0.33 mmol) was transferred to its fu-

maric acid salt 17F by using the general procedure G with fumaric

acid (38 mg, 0.33 mmol). Compound 17F (60 mg, 57%) was ob-

The N-tboc protected compound was obtained by using the gen-

eral procedure A with 2-chlorobenzoyl chloride (175 mg, 1 mmol).

A mixture of CH Cl and MeOH (20:1) was used for the chromato-

2 2

graphic puriﬁcation. After removal of the solvents a white solid

(323 mg, 87%) was obtained. The N-tboc protection group of this

solid (250 mg, 0.69 mmol) was cleaved using the general proce-

dure D for 12 h and a clear oil 20 (180 mg, 99%) was obtained after

extraction. This solid 20 (96 mg, 0.36 mmol) was transferred to its

fumaric acid salt 20F by using the general procedure G with

fumaric acid (42 mg, 0.36 mmol). Compound 20F (109 mg, 79%)

tained as a white solid in 51% yield over three steps; mp 165-

1

68 °C (dec). H NMR (500 MHz, D

2

O) d 1.04 (br m, 3H), 1.12 (br

m, 3H), 1.95 (br m, 1H), 2.00 (br m, 1H), 2.33 (br s, 2H), 3.06 (br

m, 2H), 3.28-3.39 (br m, 2H), 3.44 (br m, 1H), 3.52 (br m, 2H),

4

2

3

.24 (br d, J = 12.7 Hz, 1H), 4.40 (br d, J = 13.3 Hz, 1H), 6.69 (s,

1

3

.0H). C NMR (125 MHz, D

2

O) d 20.4, 21.1, 28.1, 28.5, 30.3,

was obtained as a white solid in 68% yield over three steps; mp

1

3.5, 49.1, 50.1, 50.7, 52.1, 137.6, 174.4, 184.9. LC/ESI-MS: positive

183-184 °C (dec). H NMR (500 MHz, D

2

O, rotamers present) d

+

ꢀ1

mode m/z = 197.3 ([M+H] ). Purity (>99.7%). IR (KBr, cm ) 3448,

1.95–2.07 (br m, 2H), 2.17–2.24 (br m, 1H), 2.43–2.50 (br m, 1H),

⁄ ⁄

20 2 4 4 4 2

H N O 1.0C H O 0.45H O)

1

700, 1650, 1638, 984, 975. Anal. (C11

3.31–3.45 (br m, 4H), 3.49–3.65 (br m, 3H), 4.51–4.62 (br m, 1H),

1

3

C, H, N.

2

6.68 (s, 1.8H), 7.35–7.61 (m, 4H). C NMR (125 MHz, D O, rota-

mers present) d 27.8, 28.2, 29.9, 48.8, 49.5, 50.5, 53.0, 129.8,

4

.19. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-

130.7, 131.0, 132.6, 132.7, 134.2, 137.0, 137.6, 174.4, 175.5. LC/

+

yl(cyclohexyl)methanone fumaric acid salt (18F)

ESI-MS: positive mode m/z = 265.4 ([M+H] ). Purity (>99.8%). IR

ꢀ

1

⁄

(

H

KBr, cm ) 3433, 1705, 1648, 984, 977. Anal. (C14

H17ClN

2

O 0.9C4-

⁄

The N-tboc protected compound was obtained by using the gen-

4

O

4

0.5H

2

O) C, H, N.

eral procedure A with cyclohexanecarbonyl chloride (147 mg,

1

mmol). A mixture of CH

2

Cl

2

and MeOH (20:1) was used for the

4.22. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(2-

chromatographic puriﬁcation. After removal of the solvents a

white solid (332 mg, 97%) was obtained. The N-tboc protection

group of this solid (220 mg, 0.65 mmol) was cleaved using the gen-

eral procedure D for 18 h and a white solid 18 (153 mg, 99%) was

obtained after extraction. This solid 18 (93 mg, 0.39 mmol) was

transferred to its fumaric acid salt 18F by using the general proce-

dure G with fumaric acid (46 mg, 0.39 mmol). Compound 18F

nitrophenyl)methanone fumaric acid salt (21F)

The N-tboc protected compound was obtained by using the gen-

eral procedure A with 2-nitrobenzoyl chloride (186 mg, 1 mmol). A

mixture of CH Cl and MeOH (20:1) was used for the chromato-

2 2

graphic puriﬁcation. After removal of the solvents a yellowish solid

(371 mg, 97%) was obtained. The N-tboc protection group of this oil

(290 mg, 0.77 mmol) was cleaved using the general procedure D

for 18 h and a yellowish oil 21 (205 mg, 96%) was obtained after

extraction. This oil 21 (103 mg, 0.37 mmol) was transferred to its

fumaric acid salt 21F by using the general procedure G with fuma-

ric acid (43 mg, 0.37 mmol). Compound 21F (95 mg, 65%) was ob-

(

107 mg, 73%) was obtained as a white solid in 70% yield over three

1

steps; mp 166–167 °C (dec). H NMR (500 MHz, D

H), 1.31 (br m, 2H), 1.47 (br m, 1H), 1.65-1.82 (br m, 5H), 1.95 (br

m, 1H), 2.00 (br m, 1H), 2.32 (br s, 2H), 2.75 (tt, J = 11.6, 2.9 Hz, 1H),

.05 (br d, J = 13.3 Hz, 1H), 3.28-3.39 (br m, 2H), 3.43 (br d,

J = 12.7 Hz, 1H), 3.51 (br m, 2H), 4.24 (br d, J = 12.9 Hz, 1H), 4.39

br d, J = 13.6 Hz, 1H), 6.69 (s, 2.0H). ¹³C NMR (125 MHz, D

O) d

7.9, 28.07, 28.11, 28.3, 28.5, 30.3, 30.9, 31.7, 43.8, 50.0, 50.1,

0.7, 52.1, 137.6, 174.4, 183.9. LC/ESI-MS: positive mode m/

2

O) d 1.21 (br m,

2

3

tained as a yellow solid in 61% yield over three steps; mp 184-

1

(

2

5

2

185 °C (dec). H NMR (500 MHz, D

2

O, rotamers present) d 1.97–

2.04 (br m, 2H), 2.16-2.24 (br m, 1H), 2.48 (br s, 1H), 2.34–3.73

(br m, 7H), 4.49–4.57 (br m, 0.5H), 4.68–4.76 (br m, 0.5H), 6.68

(s, 1.8H), 7.54 (br s, 0.5H), 7.70–7.76 (br m, 0.5H), 7.78 (m, 1H),

7.93 (t, J = 7.5 Hz, 1H), 8.35 (d, J = 8.3 Hz, 1H).

+

ꢀ1

z = 237.4 ([M+H] ). Purity (>99.7%). IR (KBr, cm ) 3443, 1709,

⁄

13

1

633, 985, 975. Anal. (C14

H

24

N

2

O 1.0C H

4 4

O

4

2

1.05H O) C, H, N.

C NMR

(

125 MHz, D O, rotamers present) d 27.8, 28.1, 30.2, 48.7, 49.6,

2

4

.20. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-

50.8, 53.3, 128.3, 130.3, 133.6, 134.0, 137.6, 138.9, 147.2, 174.4,

+

yl(phenyl)methanone fumaric acid salt (19F)

175.4. LC/ESI-MS: positive mode m/z = 276.4 ([M+H] ). Purity

ꢀ

1

(

>99.9%). IR (KBr, cm ) 3442, 1706, 1647, 1531, 1348, 970. Anal.

⁄

The N-tboc protected compound was obtained by using the gen-

(C14

H

17

N

3

O

3

0.9C

4

H

4

O

4

2

0.6H O) C, H, N.

eral procedure A with benzoyl chloride (141 mg, 1 mmol). A mix-

ture of CH

Cl

2 2

and MeOH (20:1) was used for the

4.23. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(3-

chromatographic puriﬁcation. After removal of the solvents a

white solid (325 mg, 97%) was obtained. The N-tboc protection

group of this solid (200 mg, 0.61 mmol) was cleaved using the gen-

eral procedure D for 12 h and an off white solid 19 (138 mg, 99%)

was obtained after extraction. This solid 19 (67 mg, 0.29 mmol)

was transferred to its fumaric acid salt 19F by using the general

procedure G with fumaric acid (34 mg, 0.29 mmol). Compound

chlorophenyl)methanone fumaric acid salt (22F)

The N-tboc protected compound was obtained by using the gen-

eral procedure A with 3-chlorobenzoyl chloride (175 mg, 1 mmol).

A mixture of CH Cl and MeOH (20:1) was used for the chromato-

2 2

graphic puriﬁcation. After removal of the solvents a clear oil

(362 mg, 98%) was obtained. The N-tboc protection group of this

oil (280 mg, 0.77 mmol) was cleaved using the general procedure

D for 12 h and a white solid 22 (200 mg, 98%) was obtained after

extraction. This solid 22 (132 mg, 0.50 mmol) was transferred to

its fumaric acid salt 22F by using the general procedure G with fu-

maric acid (58 mg, 0.50 mmol). Compound 22F (126 mg, 58%) was

1

9F (46 mg, 44%) was obtained as a white solid in 42% yield over

1

three steps; mp 162–165 °C (dec). H NMR (500 MHz, D

2

O) d

.99 (br m, 1H), 2.07 (br m, 1H), 2.27 (br s, 2H), 3.35-3.37 (br m,

H), 3.37-3.40 (br m, 2H), 3.46 (br d, J = 13.1 Hz, 1H), 3.98 (br m,

1

2

1

13

H), 4.56 (br s, 1H), 6.73 (s, 1.8H), 7.53 (m, 5H).

O) d 27.3, 29.1, 48.3, 48.5, 128.3, 129.7, 131.2,

36.2, 136.8, 171.3, 175.4. LC/ESI-MS: positive mode m/z = 231.4

C NMR

(

125 MHz, D

2

obtained as a yellow solid in 56% yield over three steps; mp 163–

1

165 °C (dec). H NMR (500 MHz, D

2

O) d 2.00 (br m, 2H), 2.22 (br s,

+

ꢀ1

(

[M+H] ). Purity (>99.9%). IR (KBr, cm ) 3420, 1701, 1613, 987,

1H), 2.43 (br s, 1H), 3.30–3.40 (br m, 3H), 3.42–3.54 (br m, 3H),

3.84 (br d, J = 13.2 Hz, 1H), 4.50 (br d, J = 13.6 Hz, 1H), 6.73 (s,

⁄

9

69. Anal. (C14

H

18

N

2

O 0.9C H

4 4

O

4

1.5H

2

O) C, H, N.

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7297

2

D

1

.65H), 7.39 (m, 1H), 7.50 (m, 2H), 7.57 (m 1H). ¹³C NMR (125 MHz,

O) d 28.0, 28.4, 30.1, 49.3, 49.6, 50.6, 54.6, 127.9, 129.5, 133.26,

33.29, 137.0, 137.4, 139.0, 173.6, 176.4. LC/ESI-MS: positive mode

(100 mg, 60%) was obtained as a white solid in 56% yield over three

steps; mp 151-153 °C (dec). H NMR (500 MHz, D O) d 1.34 (s, 9H),

2

2.00 (br m, 2H), 2.22 (br s, 1H), 2.43 (br s, 1H), 3.30–3.40 (br m, 3H),

1

2

+

ꢀ1

m/z = 265.5 ([M+H] ). Purity (>99.9%). IR (KBr, cm ) 3445, 1699,

1

3.43–3.54 (br m, 3H), 3.91 (br m, 1H), 4.51 (br m, 1H), 6.70 (s,

⁄

13

616, 983, 969. Anal. (C14

H17ClN

2

O 1.32C

4

H O

4

2

0.85H O) C, H, N.

1.9H), 7.44 (d, J = 8.5 Hz, 2H), 7.62 (d, J = 8.6 Hz, 2H). C NMR

(

125 MHz, D

2

O) d 28.0, 28.5, 30.3, 33.2, 37.2, 49.4, 49.6, 50.6,

4

.24. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(3-

54.8, 128.6, 129.8, 134.4, 137.6, 157.3, 174.2, 178.1. LC/ESI-MS: po-

+

ꢀ1

nitrophenyl)methanone fumaric acid salt (23F)

sitive mode m/z = 287.0 ([M+H] ). Purity (>99.9%). IR (KBr, cm )

⁄ ⁄

26 2 4 4 4 2

H N O 0.95C H O 1.6H O) C,

3

491, 1705, 1611, 975. Anal. (C18

The N-tboc protected compound was obtained by using the gen-

H, N.

eral procedure A with 3-nitrobenzoyl chloride (186 mg, 1 mmol). A

mixture of CH

graphic puriﬁcation. After removal of the solvents a yellowish oil

369 mg, 97%) was obtained. The N-tboc protection group of this

2

Cl

2

and MeOH (20:1) was used for the chromato-

4.27. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(4-

ﬂuorophenyl)methanone fumaric acid salt (26F)

(

oil (290 mg, 0.77 mmol) was cleaved using the general procedure

D for 18 h and a yellowish oil 23 (206 mg, 97%) was obtained after

extraction. This oil 23 (87 mg, 0.32 mmol) was transferred to its fu-

maric acid salt 23F by using the general procedure G with fumaric

acid (37 mg, 0.32 mmol). Compound 23F (101 mg, 79%) was ob-

The N-tboc protected compound was obtained by using the gen-

eral procedure A with 4-ﬂuorobenzoyl chloride (159 mg, 1 mmol).

A mixture of CH Cl and MeOH (20:1) was used for the chromato-

2 2

graphic puriﬁcation. After removal of the solvents a clear oil

(347 mg, 98%) was obtained. The N-tboc protection group of this

oil (260 mg, 0.75 mmol) was cleaved using the general procedure

D for 12 h and a clear oil 26 (178 mg, 96%) was obtained after

extraction. This oil 26 (87 mg, 0.35 mmol) was transferred to its fu-

maric acid salt 26F by using the general procedure G with fumaric

acid (41 mg, 0.35 mmol). Compound 26F (126 mg, 95%) was ob-

tained as a yellow solid in 74% yield over three steps; mp 166-

1

69 °C (dec). H NMR (500 MHz, D

2

O) d 2.02 (br m, 2H), 2.23 (br

s, 1H), 2.46 (br s, 1H), 3.34–3.46 (br m, 4H), 3.52–3.61 (br m,

H), 3.82 (br d, J = 12.9 Hz, 1H), 4.55 (br d, J = 13.6 Hz, 1H), 6.68

s, 2.0H), 7.77 (m, 1H), 7.90 (m, 1H), 8.38 (m, 1H), 8.40 (m 1H).

2

(

1

3

C NMR (125 MHz, D

2

O) d 27.9, 28.4, 30.1, 49.4, 49.6, 50.5, 54.6,

tained as a white solid in 89% yield over three steps; mp 157-

1

25.0, 128.0, 133.2, 136.1, 137.6, 138.8, 150.7, 174.5, 175.4. LC/

158 °C (dec). H NMR (500 MHz, D

2

O) d 2.00 (br m, 2H), 2.22 (br

+

ESI-MS: positive mode m/z = 276.4 ([M+H] ). Purity (>99.7%). IR

s, 1H), 2.43 (br s, 1H), 3.31–3.39 (br m, 3H), 3.44–3.54 (br m,

ꢀ

1

(

KBr, cm ) 3443, 1707, 1643, 1535, 1355, 978, 970. Anal. (C14

H

17-

3H), 3.91 (br m, 1H), 4.51 (br m, 1H), 6.69 (s, 1.8H), 7.62 (m, 2H),

⁄

13

N

O

3 3

1.0C

4

H

4

O

4

0.75H

2

O) C, H, N.

2

7.52 (m, 2H). C NMR (125 MHz, D O) d 28.0, 28.4, 30.2, 49.4,

4

9.6, 50.6, 54.8, 118.6 (d, JC,F = 22.1 Hz), 132.2 (d, JC,F = 8.9 Hz),

4

.25. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(p-

133.4 (d, JC,F = 3.3 Hz), 137.5, 166.4 (d, JC,F = 248.2 Hz), 174.2,

+

tolyl)methanone fumaric acid salt (24F)

177.2. LC/ESI-MS: positive mode m/z = 249.6 ([M+H] ). Purity

ꢀ

1

(

FN

>99.5%). IR (KBr, cm ) 3427, 1701, 1616, 984, 969. Anal. (C14H

17-

⁄

The N-tboc protected compound was obtained by using the gen-

O 0.9C H

2 4 4

O

4

1.5H

2

O) C, H, N.

eral procedure A with p-toluoyl chloride (155 mg, 1 mmol). A mix-

2

ture of CH Cl

2

and MeOH (20:1) was used for the chromatographic

4.28. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(4-

puriﬁcation. After removal of the solvents a clear oil (322 mg, 92%)

chlorophenyl)methanone fumaric acid salt (27F)

was obtained. The N-tboc protection group of this oil (200 mg,

0

.58 mmol) was cleaved using the general procedure D for 12 h

The N-tboc protected compound was obtained by using the gen-

and an off white solid 24 (130 mg, 92%) was obtained after extrac-

tion. This solid 24 (49 mg, 0.20 mmol) was transferred to its fuma-

ric acid salt 24F by using the general procedure G with fumaric acid

eral procedure A with 4-chlorobenzoyl chloride (175 mg, 1 mmol).

A mixture of CH Cl and MeOH (20:1) was used for the chromato-

2 2

graphic puriﬁcation. After removal of the solvents a clear oil

(355 mg, 96%) was obtained. The N-tboc protection group of this

oil (300 mg, 0.82 mmol) was cleaved using the general procedure

D for 12 h and a clear oil 27 (211 mg, 97%) was obtained after

extraction. This oil 27 (83 mg, 0.31 mmol) was transferred to its fu-

maric acid salt 27F by using the general procedure G with fumaric

acid (36 mg, 0.31 mmol). Compound 27F (78 mg, 63%) was ob-

(

23 mg, 0.20 mmol). Compound 24F (49 mg, 56%) was obtained as

a white solid in 47% yield over three steps; mp 166–167 °C (dec).

1

2

H NMR (500 MHz, D O) d 1.99 (br m, 2H), 2.22 (br s, 1H), 2.40

(

s, 3H), 2.4, (br s, 1H), 3.30–3.40 (br m, 3H), 3.43–3.53 (br m,

3

H), 3.93 (br s, 1H), 4.50 (br s, 1H), 6.74 (s, 3.0H), 7.38 (m, 4H).

1

3

C NMR (125 MHz, D

2

O) d 23.4, 28.1, 28.4, 30.3, 49.4, 49.7, 50.6,

5

4.8, 129.8, 132.1, 134.2, 137.3, 144.2, 173.4, 178.3. LC/ESI-MS: po-

tained as a yellow solid in 59% yield over three steps; mp 150-

+

ꢀ1

1

sitive mode m/z = 245.4 ([M+H] ). Purity (>99.8%). IR (KBr, cm

3

)

154 °C (dec). H NMR (500 MHz, D

2

O) d 2.00 (br m, 2H), 2.22 (br

⁄

429, 1702, 1610, 982. Anal. (C15

H

20

N

2

O 1.5C

4 4

H O

4

1.0H

2

O) C, H,

s, 1H), 2.43 (br s, 1H), 3.30–3.40 (br m, 3H), 3.42–3.54 (br m,

N.

3H), 3.87 (br m, 1H), 4.50 (br m, 1H), 6.70 (s, 2.0H), 7.46 (d,

J = 8.7 Hz, 2H), 7.55 (d, J = 8.7 Hz, 2H). C NMR (125 MHz, D O) d

2

1

3

4

.26. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(4-tert-

28.0, 28.5, 30.2, 49.4, 49.6, 50.6, 54.7, 131.4, 131.8, 135.8, 137.5,

butylphenyl)methanone fumaric acid salt (25F)

138.8, 174.1, 177.0. LC/ESI-MS: positive mode m/z = 265.5

+

ꢀ1

(

[M+H] ). Purity (>99.8%). IR (KBr, cm ) 3433, 1706, 1637, 981,

⁄

The N-tboc protected compound was obtained by using the gen-

eral procedure with 4-tert-butylbenzoyl chloride (197 mg,

mmol). A mixture of CH Cl and MeOH (20:1) was used for the

970. Anal. (C14

H17ClN O 1.0C H

2 4 4

O

4

0.75H

2

O) C, H, N.

A

1

2

4.29. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(4-

chromatographic puriﬁcation. After removal of the solvents a

white solid (384 mg, 98%) was obtained. The N-tboc protection

group of this solid (340 mg, 0.88 mmol) was cleaved using the gen-

eral procedure D for 12 h and a white solid 25 (242 mg, 96%) was

obtained after extraction. This solid 25 (113 mg, 0.39 mmol) was

transferred to its fumaric acid salt 25F by using the general proce-

dure G with fumaric acid (46 mg, 0.39 mmol). Compound 25F

bromophenyl)methanone fumaric acid salt (28F)

The N-tboc protected compound was obtained by using the gen-

eral procedure A with 4-bromobenzoyl chloride (219 mg, 1 mmol).

A mixture of CH Cl and MeOH (20:1) was used for the chromato-

2 2

graphic puriﬁcation. After removal of the solvents a white solid

(406 mg, 99%) was obtained. The N-tboc protection group of this

7

298

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

solid (320 mg, 0.78 mmol) was cleaved using the general proce-

dure D for 18 h and a clear oil 28 (237 mg, 98%) was obtained after

extraction. This oil 28 (134 mg, 0.43 mmol) was transferred to its

fumaric acid salt 28F by using the general procedure G with

fumaric acid (50 mg, 0.43 mmol). Compound 28F (143 mg, 78%)

4.32. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(thiophen-2-

yl)methanone fumaric acid salt (31F)

The N-tboc protected compound was obtained by using the gen-

eral procedure A with 2-thiophenecarbonyl chloride (147 mg,

was obtained as a yellow solid in 76% yield over three steps; mp

2 2

1 mmol). A mixture of CH Cl and MeOH (20:1) was used for the

1

47–149 °C (dec). H NMR (500 MHz, D

2

O) d 2.00 (br m, 2H),

chromatographic puriﬁcation. After removal of the solvents a

white solid (275 mg, 80%) was obtained. The N-tboc protection

group of this solid (240 mg, 0.71 mmol) was cleaved using the gen-

eral procedure D for 12 h and a yellowish solid 31 (150 mg, 89%)

was obtained after extraction. This solid 31 (67 mg, 0.28 mmol)

was transferred to its fumaric acid salt 31F by using the general

procedure G with fumaric acid (33 mg, 0.28 mmol). Compound

2

.22 (br s, 1H), 2.42 (br s, 1H), 3.29–3.39 (br m, 3H), 3.42–3.54

(

br m, 3H), 3.86 (br m, 1H), 4.50 (br m, 1H), 6.68 (s, 1.7H), 7.39

d, J = 8.4 Hz, 2H), 7.70 (d, J = 8.4 Hz, 2H). ¹³C NMR (125 MHz,

D

2

O) d 28.0, 28.4, 30.2, 49.4, 49.6, 50.6, 54.7, 127.1, 131.5, 134.7,

1

36.2, 137.6, 174.4, 177.0. LC/ESI-MS: positive mode m/z = 309.4

+

ꢀ1

and 311.4 ([M+H] ). Purity (>99.8%). IR (KBr, cm ) 3433, 1705,

1

⁄

636, 970. Anal. (C14

H17BrN

2

O 0.85C

4

H O

4

2

0.7H O) C, H, N.

31F (73 mg, 70%) was obtained as a white solid in 50% yield over

1

three steps; mp 158–162 °C (dec). H NMR (500 MHz, D

2

O) d

4

.30. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(2,4-

2.03 (br m, 2H), 2.36 (br s, 2H), 3.35–3.40 (br m, 2H), 3.44–3.56

(br m, 4H), 4.46 (br d, J = 13.8 Hz, 2H), 6.69 (s, 1.9H), 7.19 (dd,

J = 5.0, 3.7 Hz, 1H), 7.50 (dd, J = 3.7, 1.0 Hz, 1H), 7.72 (dd, J = 5.0,

dimethoxyphenyl)methanone fumaric acid salt (29F)

1

3

The N-tboc protected compound was obtained by using the gen-

0.8 Hz, 1H). C NMR (125 MHz, D

2

O) d 28.4, 30.4, 50.2, 130.2,

eral procedure A with 2,4-dimethoxybenzoyl chloride (201 mg,

133.3, 133.8, 137.6, 138.1, 171.2, 174.4. LC/ESI-MS: positive mode

+

ꢀ1

1

mmol). A mixture of CH

2

Cl

2

and MeOH (20:1) was used for the

m/z = 237.1 ([M+H] ). Purity (>99.8%). IR (KBr, cm ) 3379, 1702,

⁄

chromatographic puriﬁcation. After removal of the solvents a

white solid (325 mg, 82%) was obtained. The N-tboc protection

group of this solid (320 mg, 0.82 mmol) was cleaved using the gen-

1610, 989, 969. Anal. (C12H N OS 0.95C H O 1.25H O) C, H, N.

16 2 4 4 4 2

4.33. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(naphthalene-1-

eral procedure E with anhydrous ZnBr

2

(369 mg, 1.6 mmol) for

yl)methanone fumaric acid salt (32F)

1

2 h and a yellowish oil 29 (231 mg, 97%) was obtained after

extraction. This oil 29 (96 mg, 0.33 mmol) was transferred to its fu-

maric acid salt 29F by using the general procedure G with fumaric

acid (38 mg, 0.33 mmol). Compound 29F (106 mg, 78%) was ob-

The N-tboc protected compound was obtained by using the gen-

eral procedure A with 1-naphthoyl chloride (191 mg, 1 mmol). A

mixture of CH Cl and MeOH (20:1) was used for the chromato-

2 2

tained as an off white solid in 62% yield over three steps; mp

graphic puriﬁcation. After removal of the solvents a white solid

(353 mg, 91%) was obtained. The N-tboc protection group of this

solid (240 mg, 0.63 mmol) was cleaved using the general proce-

dure D for 12 h and a white solid 32 (172 mg, 97%) was obtained

after extraction. This solid 32 (62 mg, 0.22 mmol) was transferred

to the fumaric acid salt 32F by using the general procedure G with

fumaric acid (26 mg, 0.22 mmol). Compound 32F (73 mg, 80%) was

1

3

75 °C (dec). H NMR (500 MHz, D

H), 2.04 (br d, J = 13.7 Hz, 1H), 2.16 (br s, 1H), 2.40 (br s, 1H),

.25 (br d, J = 13.6 Hz, 1H), 3.33–3.42 (br m, 3H), 3.48 (br d,

2

O) d 1.98 (br d, J = 13.4 Hz,

J = 12.8 Hz, 1H), 3.61 (br d, J = 12.6 Hz, 1H), 3.72 (br d, J = 14.4 Hz,

1

H), 3.88 (s, 3H), 3.90 (s, 3H), 4.55 (br d, J = 13.8 Hz, 1H), 6.68 (s,

.8H), 6.74 (m, 2H), 7.23 (d, J = 8.8 Hz, 1H). ¹³C NMR (125 MHz,

D

2

O) d 28.4, 31.1, 49.2, 50.8, 51.0, 54.2, 58.3, 58.5, 101.5, 109.5,

obtained as a white solid in 70% yield over three steps; mp 152–

1

19.4, 132.2, 137.5, 158.0, 165.0, 174.3, 175.2. LC/ESI-MS:

154 °C (dec). H NMR (500 MHz, D

2

O) d 1.96 (br m, 2H), 2.00 (br

+

positive mode m/z = 291.4 ([M+H] ). Purity (>99.7%). IR (KBr,

s, 1H), 2.47 (br s, 1H), 3.27–3.33 (br m, 3H), 3.35–3.40 (br m,

1H), 3.48–3.53 (br m, 3H), 3.55 (br d, J = 13.0 Hz, 1H), 4.66 (br d,

J = 13.9 Hz, 1H), 6.66 (s, 2.0H), 7.56–7.66 (m, 5H), 8.00–8.06 (m,

ꢀ

1

⁄

cm ) 3442, 1704, 1628, 985, 970. Anal. (C16

H

22

N

2

O

3

0.9C H

4 4

O

4

-

⁄

1

2

.0H O) C, H, N.

1

3

2

5

1

(

9

2

H). C NMR (125 MHz, D O) d 27.9, 28.2, 29.9, 48.8, 49.3, 50.6,

3.6, 126.6, 126.8, 128.4, 129.8, 130.6, 131.5, 131.6, 132.7, 135.4,

36.0, 137.5, 174.3, 177.8. LC/ESI-MS: positive mode m/z = 281.5

[M+H] ). Purity (>98%). IR (KBr, cm ) 3433, 1707, 1635, 983,

70. Anal. (C18

4

.31. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(furan-2-

yl)methanone fumaric acid salt (30F)

+

ꢀ1

⁄ ⁄

20 2 4 4 4 2

H N O 1.0C H O 1.5H O) C, H, N.

The N-tboc protected compound was obtained by using the gen-

eral procedure A with 2-furoyl chloride (131 mg, 1 mmol). A mix-

4

.34. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(naphthalene-2-

ture of CH

Cl

2 2

and MeOH (20:1) was used for the

yl)methanone fumaric acid salt (33F)

chromatographic puriﬁcation. After removal of the solvents a yel-

lowish solid (267 mg, 82%) was obtained. The N-tboc protection

group of this solid (240 mg, 0.75 mmol) was cleaved using the gen-

eral procedure D for 12 h and a yellowish oil 30 (164 mg, 99%) was

obtained after extraction. This oil 30 (79 mg, 0.36 mmol) was

transferred to its fumaric acid salt 30F by using the general

procedure G with fumaric acid (42 mg, 0.36 mmol). Compound

The N-tboc protected compound was obtained by using the gen-

eral procedure A with 2-naphtoyl chloride (191 mg, 1 mmol). A

mixture of CH

graphic puriﬁcation. After removal of the solvents a white solid

342 mg, 88%) was obtained. The N-tboc protection group of this

2 2

Cl and MeOH (20:1) was used for the chromato-

(

solid (250 mg, 0.66 mmol) was cleaved using the general proce-

dure D for 12 h and a white solid 33 (181 mg, 98%) was obtained

after extraction. This solid 33 (103 mg, 0.37 mmol) was transferred

to the fumaric acid salt 33F by using the general procedure G with

fumaric acid (43 mg, 0.37 mmol). Compound 33F (101 mg, 68%)

3

0F (48 mg, 38%) was obtained as an off white solid in 31% yield

1

2

over three steps; mp 157–162 °C (dec). H NMR (500 MHz, D O)

d 2.05 (br m, 2H), 2.37 (br s, 2H), 3.35–3.45 (br m, 4H), 3.57 (br

d, J = 13.0 Hz, 2H), 4.51 (br d, J = 13.2 Hz, 2H), 6.65 (dd, J = 3.6,

1

.8 Hz, 1H), 6.69 (s, 2.0H), 7.13 (dd, J = 3.6, 0.6 Hz, 1H), 7.73 (dd,

was obtained as a white solid in 58% yield over three steps; mp

1

3

J = 1.8, 0.6 Hz, 1H). C NMR (125 MHz, D

2

O) d 28.4, 30.7, 50.2,

1

52–154 °C (dec). H NMR (500 MHz, D

2

O) d 1.97 (br m, 2H),

1

14.6, 120.8, 137.6, 148.6, 148.7, 166.7, 174.5. LC/ESI-MS:

2

.11 (br s, 1H), 2.42 (br s, 1H), 3.30–3.55 (br m, 6H), 3.82 (br d,

+

positive mode m/z = 221.1 ([M+H] ). Purity (>99.4%). IR (KBr,

J = 11.9 Hz, 1H), 4.54 (br d, J = 12.6 Hz, 1H), 6.64 (s, 2.0H), 7.51

dd, J = 8.7, 1.7 Hz, 1H), 7.64 (m, 2H), 7.94.7.99 (m, 3H), 8.01 (d,

ꢀ

1

⁄

cm ) 3426, 1701, 1613, 983, 968. Anal. (C12

H

16

N

2

O

2

1.0C H

4 4

O

4

-

(

⁄

1

2

.0H O) C, H, N.

13

2

J = 8.5 Hz, 1H). C NMR (125 MHz, D O) d 28.0, 28.4, 30.2, 49.4,

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7299

+

ꢀ1

4

1

9.6, 50.6, 54.7, 126.5, 129.7, 130.1, 130.69, 130.70, 131.3, 131.5,

34.7, 135.1, 136.4, 137.5, 174.2, 177.8. LC/ESI-MS: positive mode

z = 292.3 ([M+H] ). Purity (95%). IR (KBr, cm ) 3364, 2232, 1343,

⁄

1159. Anal. (C14

H

17

N

3

O

2

S 0.75H

2

O) C, H, N.

+

ꢀ1

m/z = 281.3 ([M+H] ). Purity (>99.9%). IR (KBr, cm ) 3432, 1704,

1

⁄

635, 983, 970. Anal. (C18

H

20

N

2

O 1.0C

4

H O

4

0.6H

2

O) C, H, N.

4.38. (1R,5S)-3-Tosyl-3,7-diazabicyclo[3.3.1]nonane (37)

The N-tboc protected compound was obtained by using the gen-

4

.35. (1R,5S)-3-(Methylsulfonyl)-3,7-diazabicyclo[3.3.1]nonane

fumaric acid salt (34F)

eral procedure

A

with p-toluenesulfonyl chloride (190 mg,

Cl and MeOH (9:1) was used for the

1

mmol). A mixture of CH

2

The N-tboc protected compound was obtained by using the gen-

eral procedure A with methylsulfonyl chloride (115 mg, 1 mmol). A

chromatographic puriﬁcation. After removal of the solvents a

white solid (377 mg, 97%) was obtained. The N-tboc protection

group of this solid (200 mg, 0.53 mmol) was cleaved using the gen-

eral procedure E with anhydrous ZnBr (237 mg, 1.1 mmol) for

2

48 h and compound 37 was obtained as an off white solid

mixture of CH

graphic puriﬁcation. After removal of the solvents a white solid

280 mg, 91%) was obtained. The N-tboc protection group of this

2 2

Cl and MeOH (20:1) was used for the chromato-

(

solid (230 mg, 0.76 mmol) was cleaved using the general proce-

dure D for 12 h and an white solid 34 (146 mg, 95%) was obtained

after extraction. This solid 34 (110 mg, 0.54 mmol) was transferred

to its fumaric acid salt 34F by using the general procedure G with

fumaric acid (63 mg, 0.54 mmol). Compound 34F (163 mg, 94%)

(145 mg, 98%) after extraction, in 95% yield over two steps; mp

1

3

111 °C. H NMR (500 MHz, CDCl ) d 1.50 (br m, 1H), 1.71 (br m,

1H), 1.85 (br s, 2H), 2.44 (s, 3H), 2.58 (br m, 2H), 2.73 (br s, 1H),

3.02 (br m, 2H), 3.21 (br d, J = 14.1 Hz, 2H), 3.89 (br d, J = 10.8 Hz,

2H), 7.34 (d, J = 8.0 Hz, 2H), 7.62 (d, J = 8.3 Hz, 2H). ¹³C NMR

was obtained as a white solid in 81% yield over three steps; mp

(125 MHz, CDCl

3

) d 21.7, 28.3, 31.3, 50.9, 51.3, 128.1, 129.7,

1

+

1

79 °C (dec). H NMR (500 MHz, D

2

O) d 1.90 (br m, 1H), 2.00 (br

131.4, 143.8. LC/ESI-MS: positive mode m/z = 281.4 ([M+H] ). Pur-

ꢀ1

m, 1H), 2.38 (br s, 2H), 2.99 (s, 3H), 3.18 (br m, 2H), 3.36 (br d,

ity (99.5%). IR (KBr, cm ) 3343, 1341, 1166. Anal. (C14

H

20

N

2

O

2-

⁄

J = 13.3 Hz, 2H), 3.59 (br d, J = 13.3 Hz, 2H), 3.84 (br m, 2H), 6.69

S 0.5H

2

O) C, H, N.

1

3

(

1

s, 1.8H). C NMR (125 MHz, D

2

O) d 28.4, 30.4, 34.3, 50.2, 53.2,

+

37.6, 174.5. LC/ESI-MS: positive mode m/z = 205.1 ([M+H] ). Pur-

4.39. 4-((1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-

ylsulfonyl)benzonitrile (38)

ꢀ1

ity (99%). IR (KBr, cm ) 3457, 1705, 1654, 1337, 1164, 986, 962.

⁄ ⁄

8 16 2 2 4 4 4 2

Anal. (C H N O S 0.9C H O 0.7H O) C, H, N.

The N-tboc protected compound was obtained by using the gen-

eral procedure A with p-cyanophenylsulfonyl chloride (202 mg,

4

.36. (1R,5S)-3-(Phenylsulfonyl)-3,7-diazabicyclo[3.3.1]nonane

1

2 2

mmol). A mixture of CH Cl and MeOH (9:1) was used for the

(

35)

chromatographic puriﬁcation. After removal of the solvents a

white solid (295 mg, 75%) was obtained. The N-tboc protection

group of this solid (200 mg, 0.51 mmol) was cleaved using the gen-

The N-tboc protected compound was obtained by using the gen-

eral procedure A with benzenesulfonyl chloride (177 mg, 1 mmol).

A mixture of CH Cl and MeOH (20:1) was used for the chromato-

graphic puriﬁcation. After removal of the solvents a white solid

324 mg, 87%) was obtained. The N-tboc protection group of this

eral procedure E with anhydrous ZnBr

8 h and compound 38 was obtained as an off white solid

105 mg, 71%) after extraction, in 53% yield over two steps; mp

2

(230 mg, 1.0 mmol) for

2

4

(

1

2

3

69 °C. H NMR (500 MHz, CDCl ) d 1.53 (br m, 1H), 1.75 (br s,

solid (90 mg, 0.25 mmol) was cleaved using the general procedure

D for 4 h and compound 35 was obtained as an off white solid

H), 1.89 (br m, 1H), 2.44 (br s, 1H), 2.62 (br m, 2H), 3.05 (br m,

H), 3.22 (br d, J = 14.0 Hz, 2H), 3.94 (br d, J = 11.4 Hz, 2H), 7.86

(

63 mg, 96%) after extraction, in 84% yield over two steps; mp

1

3

1

3

(m, 4H). C NMR (125 MHz, CDCl ) d 28.2, 31.2, 51.0, 51.3,

1

2

01 °C. H NMR (500 MHz, CDCl

H), 2.57–2.61 (br m, 4H), 3.05 (br m, 2H), 3.23 (br d, J = 14.0 Hz,

H), 3.92 (br d, J = 11.5 Hz, 2H), 7.55 (m, 2H), 7.62 (m, 1H), 7.75

3

) d 1.51 (br m, 1H), 1.86 (br m,

1

16.8, 117.4, 128.6, 133.0, 139.1. LC/ESI-MS: positive mode m/

+

ꢀ1

z = 292.1 ([M+H] ). Purity (>99.8%). IR (KBr, cm ) 3405, 2237,

1344, 1164. Anal. (C₁₄H₁₇N O S 0.75H O) C, H, N.

⁄

1

3

2

(

m, 2H). C NMR (125 MHz, CDCl

3

) d 28.1, 31.2, 50.7, 51.3,

1

28.1, 129.2, 133.1, 134.4. LC/ESI-MS: positive mode m/z = 267.0

+

ꢀ1

4.40. (1R,5S)-3-(4-Bromophenylsulfonyl)-3,7-

diazabicyclo[3.3.1]nonane hydrochloric acid salt (39H)

(

[M+H] ). Purity (>99%). IR (KBr, cm ) 3417, 1345, 1172. Anal.

⁄

(

C

13

H

18

N

2

O

2

S 0.7H

2

O) C, H, N.

The N-tboc protected compound was obtained by using the gen-

eral procedure A with p-bromophenylsulfonyl chloride (255.5 mg,

mmol). A mixture of CH Cl and MeOH (20:1) was used for the

2 2

4

.37. 3-((1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-

ylsulfonyl)benzonitrile (36)

1

chromatographic puriﬁcation. After removal of the solvents a

white solid (407 mg, 91%) was obtained. This solid (264 mg,

The N-tboc protected compound was obtained by using the gen-

eral procedure A with m-cyanophenylsulfonyl chloride (202 mg,

mmol). A mixture of CH Cl and MeOH (9:1) was used for the

2 2

chromatographic puriﬁcation. After removal of the solvents a

white solid (385 mg, 98%) was obtained. The N-tboc protection

group of this solid (200 mg, 0.51 mmol) was cleaved using the gen-

0

2 2

.59 mmol) was dissolved in a mixture of CH Cl (2 mL) and 1,4-

1

dioxane (2 mL), HCl in 1,4-dioxane (4 M, 2 mL) was added for the

cleavage of the N-tboc protection group, and the reaction mixture

was allowed to stir at rt for 2 h. Then dry Et

the precipitate was ﬁltered off, and washed with dry Et

3 ꢁ 2 mL). Compound 39H (224 mg, 99%) was obtained as a white

2

O (2 mL) was added,

2

O

eral procedure E with anhydrous ZnBr

8 h and compound 36 was obtained as an off white solid

138 mg, 93%) after extraction, in 91% yield over two steps; mp

2

(230 mg, 1.0 mmol) for

(

4

(

1

2

1

solid in 90% yield over two steps; mp 266–269 °C (dec). H NMR

400 MHz, O) 1.57 (br d, J = 13.5 Hz, 1H), 1.89 (br d,

J = 13.6 Hz, 1H), 2.29 (br s, 2H), 2.63 (br d, J = 11.6 Hz, 2H), 3.34

br d, J = 13.3 Hz, 2H), 3.61 (br d, J = 13.2 Hz, 2H), 3.89 (br d,

J = 11.4 Hz, 2H), 7.70 (d, J = 8.6 Hz, 2H), 7.78 (d, J = 8.6 Hz, 2H).

(

D

2

d

1

3

46 °C. H NMR (500 MHz, CDCl ) d 1.53 (br m, 1H), 1.75 (br s,

H), 1.89 (br m, 1H), 2.52 (br s, 1H), 2.62 (br m, 2H), 3.04 (br m,

H), 3.21 (br d, J = 14.1 Hz, 2H), 3.93 (br d, J = 11.4 Hz, 2H), 7.70

ddd, J = 7.9, 7.9, 0.5 Hz, 1H), 7.89 (ddd, J = 7.8, 1.6, 1.2 Hz, 1H),

.97 (ddd, J = 8.0, 1.8, 1.2, 1H), 8.04 (ddd, J = 1.7, 1.7, 0.5 Hz, 1H).

(

13

C NMR (100 MHz, D

2

O) d 26.1, 28.0, 47.8, 51.2, 129.6, 130.4,

7

1

31.1, 133.4. ESI-MS: positive mode m/z = 345.3 and 347.2

1

3

C NMR (125 MHz, CDCl ) d 28.1, 31.1, 50.8, 51.3, 113.9, 117.3,

+

ꢀ1

(

[M+H] ). Purity (>99%). IR (cm ) 2848, 2556, 1570, 1352, 908,

1

30.2, 131.4, 131.9, 136.0, 136.6. LC/ESI-MS: positive mode m/

⁄

7

47. Anal. (C13

H17BrN

2

O

2

S 1.0HCl) C, H, N.

7

300

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

4

.41. (1R,5S)-3-(4-Nitrophenylsulfonyl)-3,7-

4.44. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-

diazabicyclo[3.3.1]nonane (40)

yl(cyclopentyl)methanone fumaric acid salt (43F)

The N-tboc protected compound was obtained by using the gen-

eral procedure A with p-nitrophenylsulfonyl chloride (222 mg,

eral procedure

1 mmol) for 72 h. A mixture of CH

B

with cyclopentanecarboxylic acid (114 mg,

Cl and MeOH (40:1) was used

1

mmol). A mixture of CH

2

Cl

2

and MeOH (40:1) was used for the

2

chromatographic puriﬁcation. After removal of the solvents a yel-

lowish solid (272 mg, 65%) was obtained. The N-tboc protection

group of this solid (253 mg, 0.61 mmol) was cleaved using the gen-

for the chromatographic puriﬁcation. After removal of the solvents

a white solid (228 mg, 71%) was obtained. The N-tboc protection

group of this solid (198 mg, 0.61 mmol) was cleaved using the gen-

eral procedure D for 12 h and a white solid 43 (134 mg, 98%) was

obtained after extraction. This solid 43 (134 mg, 0.60 mmol) was

transferred to its fumaric acid salt 43F by using the general proce-

dure G with fumaric acid (70 mg, 0.60 mmol). Compound 43F

eral procedure E with anhydrous ZnBr

2 h and compound 40 was obtained as

177 mg, 92%) after extraction, in 60% yield over two steps; mp

2

(277 mg, 1.23 mmol) for

1

a

yellowish solid

(

1

3

85 °C (dec). H NMR (500 MHz, CDCl ) d 1.52 (br m, 1H), 1.74

(

br s, 2H), 1.89 (br m, 1H), 2.16 (br s, 1H), 2.64 (br m, 2H), 3.03

br m, 2H), 3.21 (br d, J = 14.1 Hz, 2H), 3.96 (br d, J = 10.9 Hz, 2H),

(164 mg, 79%) was obtained as a white solid in 55% yield over three

1

steps; mp 164–166 °C (dec). H NMR (500 MHz, D

2

O) d 1.40–1.48

7

2

1

.94 (ddd, J = 8.9, 2.2, 2.0 Hz, 2H), 8.40 (ddd, J = 8.9, 2.2, 2.0 Hz,

(br m, 1H), 1.57–1.72 (br m, 4H), 1.77–1.87 (br m, 2H), 1.93–2.03

(br m, 3H), 2.32 (br s, 2H), 3.06 (br d, J = 13.7 Hz, 1H), 3.18 (quint,

J = 8.0 Hz, 1H), 3.29-3.39 (br m, 2H), 3.45 (br m, 2H), 3.52 (br d,

1

3

H). C NMR (125 MHz, CDCl ) d 28.2, 31.2, 50.9, 51.3, 124.4,

29.1, 140.7, 150.4. LC/ESI-MS: positive mode m/z = 312.0

+

ꢀ1

(

[M+H] ). Purity (>99%). IR (KBr, cm ) 3424, 1530, 1349, 1168.

J = 13.0 Hz, 1H), 4.27 (br d, J = 13.1 Hz, 1H), 4.41 (br d, J = 13.8 Hz,

⁄

13

Anal. (C13

H

17

N

3

O

4

S 0.5H

2

O) C, H, N.

2

1H), 6.69 (s, 2.0H). C NMR (125 MHz, D O) d 28.2, 28.4, 28.5,

2

8.6, 30.4, 31.9, 32.2, 44.5, 49.2, 50.2, 50.7, 52.3, 137.6, 174.4,

+

4

.42. (1R,5S)-N,N-Dimethyl-3,7-diazabicyclo[3.3.1]nonane-3-

184.0. LC/ESI-MS: positive mode m/z = 223.4 ([M+H] ). Purity

(>99.8%). IR (KBr, cm ) 3441, 1707, 1654, 979. Anal. (C13H N

22 2-

O 1.0C H O 0.35H O) C, H, N.

ꢀ1

carboxamide fumaric acid salt (41F)

⁄

4 4 4 2

The N-tboc protected compound was obtained by using the gen-

eral procedure

A

with dimethylcarbamoyl chloride (108 mg,

Cl and MeOH (20:1) was used for the

4.45. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(o-

tolyl)methanone fumaric acid salt (44F)

1

mmol). A mixture of CH

2

chromatographic puriﬁcation. After removal of the solvents a

white solid (280 mg, 94%) was obtained. The N-tboc protection

group of this solid (240 mg, 0.81 mmol) was cleaved using the gen-

eral procedure D for 4 h and a white solid 41 (152 mg, 95%) was ob-

tained after extraction. This solid 41 (140 mg, 0.71 mmol) was

transferred to the fumaric acid salt 41F by using the general proce-

dure G with fumaric acid (82 mg, 0.71 mmol). Compound 41F

The N-tboc protected compound was obtained by using the gen-

eral procedure B with 2-methylbenzoic acid (136 mg, 1 mmol) for

48 h. A mixture of CH Cl and MeOH (20:1) was used for the chro-

2 2

matographic puriﬁcation. After removal of the solvents a clear oil

(186 mg, 54%) was obtained. The N-tboc protection group of this

oil (165 mg, 0.48 mmol) was cleaved using the general procedure

(

180 mg, 81%) was obtained as a white solid in 72% yield over three

2

E with anhydrous ZnBr (216 mg, 0.96 mmol) for 12 h and a white

1

steps; mp 144–148 °C (dec). H NMR (400 MHz, D

H), 2.23 (br s, 2H), 2.90 (s, 6H), 3.18 (br d, J = 13.1 Hz, 2H), 3.35 (br

d, J = 13.0 Hz, 2H), 3.56 (d, J = 13.2 Hz, 2H), 3.72 (d, J = 13.0 Hz, 2H),

2

O) d 1.94 (br m,

solid 44 (92 mg, 79%) was obtained after extraction. This solid 44

(77 mg, 0.32 mmol) was transferred to its fumaric acid salt 44F

by using the general procedure G with fumaric acid (37 mg,

0.32 mmol). Compound 44F (61 mg, 52%) was obtained as a white

2

1

3

6

5

.69 (s, 1.9H). C NMR (100 MHz, D

2

O) d 25.7, 28.5, 37.5, 47.4,

1

0.8, 134.4, 165.8, 171.1. ESI-MS: positive mode m/z = 198.3

solid in 22% yield over three steps; mp 155–157 °C (dec). H NMR

+

ꢀ1

(

[M+H] ). Purity (>99%). IR (cm ) 2951, 1713, 1604, 1409, 984,

2

(500 MHz, D O) d 2.00 (br m, 2H), 2.17 (br s, 1H), 2.20 (s, 3H), 2.45

⁄

7

56, 637. Anal. (C10

19

H N

3

O 1.0C

H

4 4

O

4

0.7H

2

O) C, H, N.

(br s, 1H), 3.33–3.44 (br m, 5H), 3.52 (br d, J = 13.2 Hz, 1H), 3.66 (br

d, J = 13.4 Hz, 1H), 4.54 (br d, J = 14.2 Hz, 1H), 6.68 (s, 2.0H), 7.33–

1

3

4

.43. (1R,5S)-N,N-Diethyl-3,7-diazabicyclo[3.3.1]nonane-3-

7.39 (m, 3H), 7.41–7.45 (m, 1H). C NMR (125 MHz, D

20.9, 28.3, 30.0, 48.6, 49.5, 50.6, 53.3, 127.9, 129.1, 132.6, 133.6,

37.4, 137.5, 137.6, 174.4, 178.5. LC/ESI-MS: positive mode m/

2

O) d 20.8,

carboxamide fumaric acid salt (42F)

1

+

ꢀ1

The N-tboc protected compound was obtained by using the gen-

z = 245.4 ([M+H] ). Purity (>99.7%). IR (KBr, cm ) 3485, 1704,

⁄ ⁄

20 2 4 4 4 2

1615, 984, 967. Anal. (C15H N O 1.0C H O 0.55H O) C, H, N.

eral procedure

1

A

with diethylcarbamoyl chloride (135.6 mg,

Cl and MeOH (20:1) was used for the

mmol). A mixture of CH

2

chromatographic puriﬁcation. After removal of the solvents a

white solid (353 mg, 91%) was obtained. The N-tboc protection

group of this solid (240 mg, 0.63 mmol) was cleaved using the gen-

eral procedure D for 12 h and a white solid 42 (172 mg, 97%) was

obtained after extraction. This solid 42 (62 mg, 0.22 mmol) was

transferred to the fumaric acid salt 42F by using the general proce-

dure G with fumaric acid (26 mg, 0.22 mmol). Compound 42F

4.46. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(3-

ﬂuorophenyl)methanone fumaric acid salt (45F)

The N-tboc protected compound was obtained by using the gen-

eral procedure B with 3-ﬂuorobenzoic acid (140 mg, 1 mmol) for

24 h. A mixture of CH Cl and MeOH (20:1) was used for the chro-

2 2

matographic puriﬁcation. After removal of the solvents a clear oil

(230 mg, 66%) was obtained. The N-tboc protection group of this

oil (210 mg, 0.60 mmol) was cleaved using the general procedure

D for 12 h and a clear oil 45 (107 mg, 71%) was obtained after

extraction. This oil 45 (96 mg, 0.39 mmol) was transferred to its fu-

maric acid salt 45F by using the general procedure G with fumaric

acid (45 mg, 0.39 mmol). Compound 45F (84 mg, 58%) was ob-

(

73 mg, 80%) was obtained as a white solid in 70% yield over three

1

2

steps; mp 111–115 °C (dec). H NMR (400 MHz, D O) d 1.14 (t,

J = 7.1 Hz, 6H); 1.93 (br m, 2H); 2.22 (br s, 2H); 3.20 (br d,

J = 13.2 Hz, 2H); 3.27 (q, J = 7.2 Hz, 4H); 3.34 (br d, J = 13.1 Hz,

2

H); 3.53 (br d, J = 13.1 Hz, 2H); 3.64 (br d, J = 13.3 Hz, 2H); 6.68

1

3

(

5

s, 2.0H). C NMR (100 MHz, D

2.1, 135.3, 166.5, 172.1. ESI-MS: positive mode m/z = 226.3

2

O) d 13.0, 26.7, 29.4, 42.5, 48.4,

tained as a yellow solid in 27% yield over three steps; mp 157–

+

1

(

[M+H] ). HRMS for C12

H

23

N

3

O: calc m/z = 226.1914, found m/

158 °C (dec). H NMR (500 MHz, D

2

O) d 2.00 (br m, 2H), 2.22 (br

ꢀ1

z = 226.1899. Purity (>99%). IR (cm ) 2969, 2850, 1703, 1634,

s, 1H), 2.43 (br s, 1H), 3.30–3.41 (br m, 3H), 3.43–3.56 (br m,

3H), 3.86 (br d, J = 12.9 Hz, 1H), 4.50 (br d, J = 13.9 Hz, 1H), 6.70

⁄

1

251, 1121, 968, 641. Anal. (C12

H

23

N

3

O 1.0C

4

H O

4

2

1.3H O) C, H, N.

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7301

(

s, 2.0H), 7.28 (m, 3H), 7.54 (dt, JH,H = 8.0 Hz, JH,F = 5.8 Hz, 1H). ¹³C

NMR (125 MHz, D O) d 27.9, 28.4, 30.1, 49.3, 49.6, 50.6, 54.6,

16.7 (d, JC,F = 23.6 Hz), 120.2 (d, JC,F = 21.2 Hz), 125.5 (d, JC,F = 3.0 -

Hz), 133.8 (d, JC,F = 8.3 Hz), 137.5, 139.2 (d, JC,F = 7.4 Hz), 165.1 (d,

C,F = 245.5 Hz), 174.0, 176.5. LC/ESI-MS: positive mode m/

(69 mg, 96%) after extraction in 30% yield over two steps; mp

157 °C (dec). H NMR (500 MHz, CDCl ) d 1.68 (br s, 1H), 1.88 (br

3

m, 3H), 2.12 (br m, 1H), 3.00 (br s, 1H), 3.05 (br d, J = 12.2 Hz,

2H), 3.14 (br d, J = 12.1 Hz, 2H), 3.27 (br d, J = 11.3 Hz, 1H), 3.36

(br d, J = 10.1 Hz, 1H), 3.66 (br d, J = 10.8 Hz, 1H), 4.81 (br d,

1

2

1

J

+

ꢀ1

z = 249.4 ([M+H] ). Purity (>99.8%). IR (KBr, cm ) 3448, 1701,

1

J = 11.7 Hz, 1H), 7.51 (d, J = 8.0 Hz, 2H), 7.70 (d, J = 8.0 Hz, 2H).

⁄

13

648, 982, 971. Anal. (C14

H17FN

2

O 1.0C

4 4

H O

4

0.7H

2

O) C, H, N.

3

C NMR (125 MHz, CDCl ) d 28.4, 28.8, 32.2, 47.1, 51.0, 51.3,

5

2.7, 113.1, 118.4, 127.6, 132.6, 141.7, 168.9. LC/ESI-MS: positive

+

ꢀ1

4

.47. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(3-

mode m/z = 256.1 ([M+H] ). Purity (>99.5%). IR (KBr, cm ) 3365,

2234, 1633. Anal. (C15

⁄

(

triﬂuoromethyl)phenyl)methanone fumaric acid salt (46F)

17

H N

3

O 0.25H

2

O) C, H, N.

The N-tboc protected compound was obtained by using the gen-

4.50. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(4-

eral procedure B with 3-triﬂuoromethylbenzoic acid (190 mg,

mmol) for 72 h. A mixture of CH Cl and MeOH (40:1) was used

nitrophenyl)methanone fumaric acid salt (49F)

1

2

for the chromatographic puriﬁcation. After removal of the solvents

a clear oil (190 mg, 48%) was obtained. The N-tboc protection

group of this oil (180 mg, 0.45 mmol) was cleaved using the gen-

eral procedure D for 12 h and a clear oil 46 (126 mg, 93%) was ob-

tained after extraction. This oil 46 (126 mg, 0.42 mmol) was

transferred to its fumaric acid salt 46F by using the general proce-

dure G with fumaric acid (49 mg, 0.42 mmol). Compound 46F

The N-tboc protected compound was obtained by using the gen-

eral procedure B with 4-nitrobenzoic acid (167 mg, 1 mmol) for

12 h. A mixture of CH Cl and MeOH (40:1) was used for the chro-

2 2

matographic puriﬁcation. After removal of the solvents a yellowish

solid (184 mg, 49%) was obtained. The N-tboc protection group of

this solid (170 mg, 0.45 mmol) was cleaved using the general pro-

2

cedure E with anhydrous ZnBr (204 mg, 0.90 mmol) for 12 h and a

(

70 mg, 39%) was obtained as a yellow solid in 18% yield over three

yellowish solid 49 (76 mg, 61%) was obtained after extraction. This

solid 49 (42 mg, 0.15 mmol) was transferred to its fumaric acid salt

49F by using the general procedure G with fumaric acid (18 mg,

0.15 mmol). Compound 49F (37 mg, 54%) was obtained as a yellow

1

steps; mp 154–156 °C (dec). H NMR (500 MHz, D

H), 2.22 (br s, 1H), 2.44 (br s, 1H), 3.32-3.56 (br m, 6H), 3.81 (br d,

J = 13.3 Hz, 1H), 4.54 (br d, J = 13.4 Hz, 1H), 6.65 (s, 1.6H), 7.68-7.90

2

O) d 2.01 (br m,

2

1

3

1

(

m, 4H). C NMR (125 MHz, D

2

O) d 27.9, 28.4, 30.1, 49.4, 49.6,

solid in 20% yield over three steps; mp 144-146 °C (dec). H NMR

5

1

0.5, 54.7, 126.61 (q, JC,F = 271.8 Hz), 126.63 (q, JC,F = 3.8 Hz),

2

(500 MHz, D O) d 1.96–2.07 (br m, 2H), 2.23 (br s, 1H), 2.46 (br s,

1H), 3.34–3.49 (br m, 6H), 3.77 (br d, J = 13.4 Hz, 1H), 4.50 (br d,

30.0 (q, JC,F = 3.6 Hz), 132.5, 133.2, 133.3 (q, JC,F = 32.6 Hz), 137.7,

38.1, 175.0, 176.4. LC/ESI-MS: positive mode m/z = 299.3

J = 14.0 Hz, 1H), 6.73 (s, 2.0H), 7.72 (d, J = 8.9 Hz, 2H), 8.37 (d,

+

ꢀ1

13

(

[M+H] ). Purity (>99.7%). IR (KBr, cm ) 3440, 1680, 1637, 985,

2

J = 8.9 Hz, 2H). C NMR (125 MHz, D O) d 27.9, 28.4, 30.1, 49.3,

⁄

9

75. Anal. (C15

H

17

F

3

N

2

O 0.8C

H

4 4

O

4

1.6H

2

O) C, H, N.

49.5, 50.5, 54.5, 127.0, 130.9, 137.6, 143.7, 151.3, 174.5, 175.8.

+

LC/ESI-MS: positive mode m/z = 276.3 ([M+H] ). Purity (>99%). IR

ꢀ1

4

.48. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(biphenyl-4-

(KBr, cm ) 3433, 1704, 1634, 1521, 1353, 983, 969. Anal. (C14H

17-

⁄ ⁄

3 3 4 4 4 2

N O 1.0C H O 3.0H O) C, H, N.

yl)methanone fumaric acid salt (47F)

The N-tboc protected compound was obtained by using the gen-

4.51. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(pyridine-3-

eral procedure B with 4-biphenylcarboxylic acid (198 mg, 1 mmol)

yl)methanone fumaric acid salt (50F)

2 2

for 12 h. A mixture of CH Cl and MeOH (40:1) was used for the

chromatographic puriﬁcation. After removal of the solvents a clear

oil (293 mg, 72%) was obtained. The N-tboc protection group of this

oil (250 mg, 0.61 mmol) was cleaved using the general procedure D

for 12 h and a clear oil 47 (141 mg, 75%) was obtained after extrac-

tion. This oil 47 (60 mg, 0.20 mmol) was transferred to its fumaric

acid salt 47F by using the general procedure G with fumaric acid

The N-tboc protected compound was obtained by using the gen-

eral procedure B with nicotinic acid (185 mg, 1 mmol) for 12 h. A

mixture of CH Cl and MeOH (9:1) was used for the chromato-

2 2

graphic puriﬁcation. After removal of the solvents a clear oil

(197 mg, 59%) was obtained. The N-tboc protection group of this

oil (175 mg, 0.53 mmol) was cleaved using the general procedure

(

23 mg, 0.20 mmol). Compound 47F (54 mg, 63%) was obtained

2

E with anhydrous ZnBr (358 mg, 1.6 mmol) for 72 h and a clear

as a white solid in 34% yield over three steps; mp 148–150 °C

oil 50 (105 mg, 86%) was obtained after extraction. This oil 50

(48 mg, 0.21 mmol) was transferred to its fumaric acid salt 50F

by using the general procedure G with fumaric acid (24 mg,

0.21 mmol). Compound 50F (47 mg, 63%) was obtained as a white

solid in 32% yield over three steps; mp 144–148 °C (dec). H NMR

2

(500 MHz, D O) d 2.01 (br m, 2H), 2.25 (br s, 1H), 2.45 (br s, 1H),

1

(

2

(

4

dec). H NMR (500 MHz, D

.43 (br s, 1H), 3.30–3.41 (br m, 3H), 3.42–3.54 (br m, 3H), 3.91

br m, 1H), 4.53 (br m, 1H), 6.67 (s, 2.0H), 7.48 (m, 1H), 7.56 (m,

H), 7.74 (d, J = 7.9 Hz, 2H), 7.79 (d, J = 8.3 Hz, 2H). C NMR

O) d 28.0, 28.5, 30.3, 49.4, 49.6, 50.6, 54.8, 129.9,

2

O) d 1.99 (br m, 2H), 2.20 (br s, 1H),

1

3

1

(

125 MHz, D

2

1

30.0, 130.5, 131.2, 132.1, 136.2, 137.6, 142.4, 145.5, 174.6,

3.33–3.40 (br m, 3H), 3.46 (br d, J = 13.3 Hz, 1H), 3.52 (br d,

J = 12.7 Hz, 1H), 3.59 (br d, J = 13.0 Hz, 1H), 3.84 (br d, J = 12.8 Hz,

+

77.8. LC/ESI-MS: positive mode m/z = 307.5 ([M+H] ). Purity

ꢀ1

(

>99.9%). IR (KBr, cm ) 3427, 1705, 1624, 982, 970. Anal. (C20

H

22-

1H), 4.54 (br d, J = 13.6 Hz, 1H), 6.65 (s, 2.0H), 7.68 (dd, J = 8.0,

⁄

13

N

2

O 1.0C

4

H O

4

0.9H

2

O) C, H, N.

5.2 Hz, 1H), 8.10 (ddd, J = 8.0, 2.1, 1.6 Hz, 1H), 8.71 (m, 2H).

NMR (125 MHz, D

C

2

O) d 27.9, 28.4, 30.1, 49.4, 49.6, 50.5, 54.5,

4

.49. 4-((1R,5S)-3,7-Diazabicyclo[3.3.1]nonane-3-

127.6, 134.6, 137.7, 140.4, 148.6, 151.9, 174.3, 174.9. LC/ESI-MS:

+

carbonyl)benzonitrile (48)

positive mode m/z = 232.3 ([M+H] ). Purity (>99.4%). IR (KBr,

ꢀ

1

⁄

cm ) 3433, 1701, 1635, 984, 969. Anal. (C13

H

17

N

3

O 1.0C

4 4

H O

4

-

⁄

The N-tboc protected compound was obtained by using the gen-

2

0.6H O) C, H, N.

eral procedure B with 4-cyanobenzoic acid (147 mg, 1 mmol) for

1

2 h. A mixture of CH

2

Cl

2

and MeOH (9:1) was used for the chro-

4.52. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(5-

bromopyridin-3-yl)methanone fumaric acid salt (51F)

matographic puriﬁcation. After removal of the solvents a white so-

lid (109 mg, 31%) was obtained. The N-tboc protection group of this

solid (100 mg, 0.28 mmol) was cleaved using the general proce-

dure D for 2 h and compound 48 was obtained as a white solid

The N-tboc protected compound was obtained by using the gen-

eral procedure B with 5-bromopyridine-3-carboxylic acid (303 mg,

7

302

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

1

.5 mmol) for 12 h. A mixture of CH

2

Cl

2

and MeOH (40:1) was used

1H), 9.01 (d, J = 4.7 Hz, 1H). ¹³C NMR (125 MHz, D

O) d 27.7,

2

for the chromatographic puriﬁcation. After removal of the solvents

a clear oil (166 mg, 41%) was obtained. The N-tboc protection

group of this oil (210 mg, 0.51 mmol) was cleaved using the gen-

28.1, 29.8, 48.7, 49.2, 50.5, 53.3, 121.1, 126.7, 127.5, 129.8, 132.1,

135.0, 137.6, 146.8, 147.9, 152.0, 174.1, 174.9. LC/ESI-MS: positive

+

ꢀ1

mode m/z = 282.3 ([M+H] ). Purity (>99.7%). IR (KBr, cm ) 3424,

⁄ ⁄

19 3 4 4 4 2

H N O 1.0C H O 1.45H O) C, H,

eral procedure E with anhydrous ZnBr

2

(231 mg, 1.0 mmol) for

1700, 1646, 985, 967. Anal. (C17

N.

4

8 h and a yellowish oil 51 (142 mg, 89%) was obtained after

extraction. This oil 51 (81 mg, 0.26 mmol) was transferred to its fu-

maric acid salt 51F by using the general procedure G with fumaric

acid (30 mg, 0.26 mmol). Compound 51F (84 mg, 75%) was ob-

4.55. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(quinolin-6-

yl)methanone fumaric acid salt (54F)

tained as a white solid in 27% yield over three steps; mp 186–

1

87 °C (dec). H NMR (500 MHz, D

2

O) d 2.01 (br m, 2H), 2.25 (br

The N-tboc protected compound was obtained by using the gen-

s, 1H), 2.44 (br s, 1H), 3.36 (br m, 3H), 3.45 (br d, J = 13.4 Hz, 1H),

.52 (br d, J = 12.6 Hz, 1H), 3.59 (br d, J = 13.2 Hz, 1H), 3.83 (br d,

J = 13.0 Hz, 1H), 4.52 (br d, J = 13.9 Hz, 1H), 6.68 (s, 1.84H), 8.18

eral procedure

1 mmol) for 72 h. A mixture of CH

for the chromatographic puriﬁcation. After removal of the solvents

a clear oil (303 mg, 79%) was obtained. The N-tboc protection

group of this oil (270 mg, 0.71 mmol) was cleaved using the gen-

B

with 6-quinolinecarboxylic acid (173 mg,

Cl and MeOH (20:1) was used

3

2

(

dd, J = 2.1, 1,9 Hz, 1H), 8.61 (d, J = 1.7 Hz, 1H), 8.79 (d, J = 2.1 Hz,

1

3

1

H). C NMR (125 MHz, D

2

O) d 27.9, 28.4, 30.0, 49.4, 49.5, 50.4,

5

4.5, 123.4, 135.3, 137.6, 141.2, 148.0, 154.4, 173.2, 174.5. LC/

2

eral procedure E with anhydrous ZnBr (319 mg, 1.4 mmol) for

+

ESI-MS: positive mode m/z = 310.3 and 312.3 ([M+H] ). Purity

72 h and a clear oil 54 (191 mg, 96%) was obtained after extraction.

This oil 54 (45 mg, 0.16 mmol) was transferred to its fumaric acid

salt 54F by using the general procedure G with fumaric acid

ꢀ1

(

>99.5%). IR (KBr, cm ) 3440, 1697, 1639, 984, 969. Anal. (C13H

16-

⁄

BrN

3

O 0.92C

H

4 4

O

4

2

0.6H O) C, H, N.

(

19 mg, 0.16 mmol). Compound 54F (27 mg, 40%) was obtained

4

.53. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(quinolin-2-

as a white solid in 30% yield over three steps; mp 162–165 °C

1

yl)methanone fumaric acid salt (52F)

2

(dec). H NMR (500 MHz, D O) d 2.02 (br m, 2H), 2.21 (br s, 1H),

2

.47 (br s, 1H), 3.34–3.48 (br m, 4H), 3.53–3.61 (br m, 2H), 3.86

The N-tboc protected compound was obtained by using the gen-

eral procedure with 2-quinolinecarboxylic acid (173 mg,

mmol) for 72 h. A mixture of CH Cl and MeOH (20:1) was used

for the chromatographic puriﬁcation. After removal of the solvents

a clear oil (210 mg, 55%) was obtained. The N-tboc protection

group of this oil (180 mg, 0.47 mmol) was cleaved using the gen-

(br d, J = 12.7 Hz, 1H), 4.59 (br d, J = 13.9 Hz, 1H), 6.58 (s, 2.0H),

7.79 (dd, J = 8.4, 4.7 Hz, 1H), 7.95 (dd, J = 8.7, 1.9 Hz, 1H), 8.18 (d,

J = 1.8 Hz, 1H), 8.19 (d, J = 8.9, 1H), 8.68 (d, J = 8.1 Hz, 1H), 9.00

B

1

2

1

3

2

(dd, J = 4.7, 1.4 Hz, 1H). C NMR (125 MHz, D O) d 28.0, 28.4,

30.1, 49.4, 49.6, 50.6, 54.7, 125.4, 129.0, 130.4, 130.8, 132.3,

136.7, 137.8, 144.2, 146.8, 152.7, 175.7, 176.4. LC/ESI-MS: positive

+

ꢀ1

eral procedure E with anhydrous ZnBr

2

(213 mg, 0.94 mmol) for

mode m/z = 282.5 ([M+H] ). Purity (>99.9%). IR (KBr, cm ) 3426,

⁄ ⁄

19 3 4 4 4 2

1701, 1633, 969. Anal. (C17H N O 1.0C H O 1.5H O) C, H, N.

7

2 h and a yellowish oil 52 (130 mg, 98%) was obtained after

extraction. This oil 52 (60 mg, 0.21 mmol) was transferred to its fu-

maric acid salt 52F by using the general procedure G with fumaric

acid (25 mg, 0.21 mmol). Compound 52F (49 mg, 56%) was ob-

4.56. 1-((1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl)-2-

cyclopentylethanone fumaric acid salt (55F)

tained as a white solid in 30% yield over three steps; mp 145-

1

50 °C (dec). H NMR (500 MHz, D

2

O) d 2.06 (br m, 2H), 2.13 (br

The N-tboc protected compound was obtained by using the gen-

s, 1H), 2.46 (br s, 1H), 3.40–3.49 (br m, 4H), 3.65 (br m, 2H), 3.83

br d, J = 13.4 Hz, 1H), 4.56 (br d, J = 12.9 Hz, 1H), 6.62 (s, 2.0H),

.70 (d, J = 8.5 Hz, 1H), 7.72 (ddd, J = 8.1, 7.0, 1.0 Hz, 1H), 7.87

ddd, J = 8.5, 6.9, 1.4 Hz, 1H), 7.98 (d, J = 8.5, 1H), 8.01 (d,

eral procedure C with cyclopentylacetic acid (128 mg, 1 mmol) for

12 h. A mixture of CH Cl and MeOH (20:1) was used for the chro-

2 2

(

7

(

matographic puriﬁcation. After removal of the solvents a white so-

lid (258 mg, 77%) was obtained. The N-tboc protection group of this

solid (200 mg, 0.59 mmol) was cleaved using the general proce-

dure D for 2 h and a white solid 55 (122 mg, 89%) was obtained

after extraction. This solid 55 (122 mg, 0.52 mmol) was transferred

to its fumaric acid salt 55F by using the general procedure G with

fumaric acid (60 mg, 0.52 mmol). Compound 55F (152 mg, 84%)

13

J = 8.1 Hz, 1H), 8.52 (d, J = 8.4 Hz, 1H). C NMR (125 MHz, D

2

O) d

8.4, 28.6, 30.9, 50.0, 50.1, 50.3, 53.3, 122.5, 130.6, 131.2, 131.3,

34.2, 137.5, 142.3, 148.7, 155.1, 174.4, 174.6. LC/ESI-MS: positive

2

1

+

ꢀ1

mode m/z = 282.5 ([M+H] ). Purity (>99.7%). IR (KBr, cm ) 3433,

⁄ ⁄

19 3 4 4 4 2

H N O 1.0C H O 0.75H O) C, H, N.

1

702, 1641, 973. Anal. (C17

was obtained as a white solid in 58% yield over three steps; mp

1

4

.54. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(quinolin-4-

162–164 °C (dec). H NMR (400 MHz, D

2

O) d 1.06–1.24 (br m,

yl)methanone fumaric acid salt (53F)

2H), 1.46–1.68 (br m, 4H), 1.68–1.84 (br m, 2H), 1.93 (br d,

J = 13.5 Hz, 1H), 1.99 (br d, J = 13.7 Hz, 1H), 2.13 (br sept,

J = 7.6 Hz, 1H), 2.31 (br s, 2H), 2.33–2.42 (br m, 1H), 2.63–2.72

(br m, 1H), 3.05 (br d, J = 13.3 Hz, 1H), 3.26–3.54 (br m, 5H), 4.15

(br d, J = 13.0 Hz, 1H), 4.41 (br d, J = 13.7 Hz, 1H), 6.68 (s, 1.88H).

The N-tboc protected compound was obtained by using the gen-

eral procedure

B

with 4-quinolinecarboxylic acid (173 mg,

Cl and MeOH (20:1) was used

1

mmol) for 72 h. A mixture of CH

2

1

3

for the chromatographic puriﬁcation. After removal of the solvents

a clear oil (177 mg, 46%) was obtained. The N-tboc protection

group of this oil (150 mg, 0.39 mmol) was cleaved using the gen-

C NMR (100 MHz, D

2

O) d 25.0, 25.8, 26.2, 28.0, 32.5, 33.0, 36.4,

40.2, 46.4, 47.9, 48.3, 50.2, 135.3, 172.0, 178.7. ESI-MS: positive

+

mode m/z = 237.3 ([M+H] ). HRMS for

14 24 2

C H N O: calc m/

ꢀ1

eral procedure E with anhydrous ZnBr

2

(177 mg, 0.79 mmol) for

z = 237.1961, found m/z = 237.1979. Purity (>99%). IR (KBr, cm

2947, 2914, 2864, 1654, 1400, 1216, 977. Anal. (C14

)

⁄

7

2 h and a yellowish oil 53 (98 mg, 89%) was obtained after extrac-

H

24

N

2

O 0.9C4-

⁄

tion. This oil 53 (45 mg, 0.16 mmol) was transferred to its fumaric

acid salt 53F by using the general procedure G with fumaric acid

4

H O

4

2

0.5H O) C, H, N.

(

19 mg, 0.16 mmol). Compound 53F (49 mg, 67%) was obtained

4.57. (1R,5S)-3,7-diazabicyclo[3.3.1]nonan-3-yl(4-

as a white solid in 30% yield over three steps; mp 168-170 °C

hydroxyphenyl)methanone hydrochloric acid salt (56H)

1

(

dec). H NMR (500 MHz, D

2

O) d 2.00 (br m, 2H), 2.09 (br s, 1H),

2

.50 (br s, 1H), 3.30–3.42 (br m, 4H), 3.45–3.60 (br m, 3H), 4.65

The N-tboc protected compound was obtained by using the gen-

eral procedure C with 4-hydroxybenzoic acid (138 mg, 1 mmol) for

24 h. DMAP was not used for this reaction and the solvent was

(

br d, J = 13.9 Hz, 1H), 6.63 (s, 2.0H), 7.73 (d, J = 8.2 Hz, 1H), 7.79

m, 1H), 7.83 (d, J = 4.7 Hz, 1H), 7.96 (m, 1H), 8.17 (d, J = 8.5 Hz,

(

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7303

changed to THF. After the reaction was ﬁnished the precipitate was

4.60. (1R5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(1-methyl-1H-

ﬁltered off, washed with THF (3 ꢁ 5 mL), and dried in a desiccator.

A white solid (330 mg, 95%) was obtained. The N-tboc protection

group of this solid (300 mg, 0.87 mmol) was cleaved using the gen-

eral procedure D for 2 h and compound 56H was obtained as a

white solid (210 mg, 86%) ﬁltering and washing the precipitate

pyrrol-2yl)methanone fumaric acid salt (59F)

The N-tboc protected compound was obtained by using the gen-

eral procedure C with 1-methyl-2-pyrrolecarboxylic acid (125 mg,

1 mmol) for 12 h. A mixture of CH Cl and MeOH (40:1) was used

2 2

with dry Et

steps; mp >320 °C (dec). H NMR (400 MHz, D

H); 2.33 (br s, 2H); 3.31-3.52 (br m, 6H); 4.24 (br s, 2H); 6.98

d, J = 8.2 Hz, 2H); 7.41 (d, J = 8.3 Hz, 2H). ¹³C NMR (100 MHz,

O) d 26.1, 28.1, 47.9, 115.9, 126.8, 129.9, 158.5, 175.5. ESI-MS:

2

O (3 ꢁ 2 mL) as a white solid in 81% yield over two

for the chromatographic puriﬁcation. After removal of the solvents

an off white solid (325 mg, 98%) was obtained. The N-tboc protec-

tion group of this solid (297 mg, 0.89 mmol) was cleaved using the

1

2

O) d 2.01 (br m,

2

(

D

2

general procedure E with anhydrous ZnBr (602 mg, 2.7 mmol) for

2

12 h and a yellowish oil 59 (201 mg, 97%) was obtained after

extraction. This oil 59 (201 mg, 0.86 mmol) was transferred to its

fumaric acid salt 59F by using the general procedure G with fuma-

ric acid (100 mg, 0.86 mmol). Compound 59F (226 mg, 67%) was

+

positive mode m/z = 247.3 ([M+H] ). HRMS for C14

m/z = 247.1441, found m/z = 247.1436. Purity (>99.5%). IR (cm

18 2 2

H N O : calc

ꢀ

1

)

3

H

084, 2665, 2570, 1607, 1574, 1430, 1255, 1095, 847. Anal. (C14-

⁄

18

N

2

O

2

1.0HCl 1.0H

2

O) C, H, N.

obtained as an off white solid in 64% yield over three steps; mp

1

36-137 °C (dec). H NMR (500 MHz, D

2

O) d 2.01 (br m, 2H), 2.35

(

6

1

br s, 2H), 3.36-3.44 (br m, 4H), 3.54 (br d, J = 13.4 Hz, 2H), 3.72

s, 3H), 4.47 (br d, J = 14.0 Hz, 2H), 6.22 (dd, J = 3.9, 2.6 Hz, 1H),

4

.58. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(pyrazin-2-

yl)methanone fumaric acid salt (57F)

.55 (dd, J = 3.9, 1.6 Hz, 1H), 6.72 (s, 2.4H), 6.95 (dd, J = 2.2,

1

3

.7 Hz, 1H). C NMR (125 MHz, D

2

O) d 28.4, 30.7, 38.0, 50.3,

The N-tboc protected compound was obtained by using the gen-

10.0, 117.4, 127.1, 137.5, 170.5, 173.9. LC/ESI-MS: positive mode

eral procedure C with pyrazinecarboxylic acid (124 mg, 1 mmol)

for 12 h. A mixture of CH

chromatographic puriﬁcation. After removal of the solvents a

white solid (285 mg, 86%) was obtained. The N-tboc protection

group of this solid (231 mg, 0.69 mmol) was cleaved using the gen-

+

ꢀ1

m/z = 234.4 ([M+H] ). Purity (99%). IR (KBr, cm ) 3434, 1709,

1

2

Cl

2

and MeOH (40:1) was used for the

⁄ ⁄

19 3 4 4 4 2

H N O 1.2C H O 0.95H O) C, H, N.

617, 972. Anal. (C13

4

.61. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(1H-indol-2-

yl)methanone fumaric acid salt (60F)

eral procedure D with anhydrous ZnBr

2

(469 mg, 2.1 mmol) for

1

2 h and a yellowish oil 57 (160 mg, 99%) was obtained after

The N-tboc protected compound was obtained by using the gen-

eral procedure C with indole-2-carboxylic acid (161 mg, 1 mmol)

for 12 h. A mixture of CH Cl and MeOH (40:1) was used for the

2 2

chromatographic puriﬁcation. After removal of the solvents a

white solid (353 mg, 96%) was obtained. The N-tboc protection

group of this solid (340 mg, 0.92 mmol) was cleaved using the gen-

eral procedure D for 4 h and a clear oil 60 (244 mg, 98%) was ob-

tained after extraction. This oil 60 (244 mg, 0.91 mmol) was

transferred to its fumaric acid salt 60F by using the general proce-

dure G with fumaric acid (105 mg, 0.91 mmol). Compound 60F

extraction. This oil 57 (160 mg, 0.69 mmol) was transferred to its

fumaric acid salt 57F by using the general procedure G with fuma-

ric acid (80 mg, 0.69 mmol). Compound 57F (187 mg, 76%) was ob-

tained as an orange solid in 65% yield over three steps; mp 154–

1

56 °C (dec). H NMR (500 MHz, D

2

O) d 2.06 (br m, 2H), 2.22 (br

s, 1H), 2.45 (br s, 1H), 3.37–3.47 (br m, 4H), 3.55 (br d,

J = 12.2 Hz, 1H), 3.72 (br d, J = 12.3 Hz, 1H), 4.00 (br d, J = 13.1 Hz,

1

(

H), 4.50 (br d, J = 13.2 Hz, 1H), 6.68 (s, 2.0H), 7.48 (m, 1H), 8.77

m, 1H), 8.91 (m, 1H). ¹³C NMR (125 MHz, D

O) d 28.4, 28.6, 30.9,

2

4

1

9.8, 50.2, 50.3, 54.0, 137.6, 146.8, 147.2, 148.8, 150.8, 172.4,

74.4. LC/ESI-MS: positive mode m/z = 233.4 ([M+H] ). Purity

(

228 mg, 67%) was obtained as a white solid in 63% yield over three

+

1

steps; mp 198–200 °C (dec). H NMR (500 MHz, D

2

O) d 1.97 (br m,

ꢀ

1

(

>99.7%). IR (KBr, cm ) 3446, 1701, 1639, 985, 967. Anal. (C12H

16-

2

4

7

1

H), 2.28 (br s, 2H), 3.25–3.34 (br m, 3H), 3.39–3.47 (br m, 3H),

⁄ ⁄

4 4 4 4 2

O 1.0C H O 0.5H O) C, H, N.

N

.51 (br d, J = 16.3 Hz, 2H), 6.62 (s, 1.5H), 6.91 (d, J = 0.7 Hz, 1H),

.20 (ddd, J = 8.0, 7.1, 0.9 Hz, 1H), 7.36 (ddd, J = 8.2, 7.0, 1.1 Hz,

H), 7.55 (dd, J = 8.3, 0.8, 1H), 7.75 (d, J = 8.1 Hz, 1H). ¹³C NMR

4

.59. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(6-

chloropyridin-3-yl)methanone fumaric acid salt (58F)

(

125 MHz, D

2

O) d 28.3, 30.4, 50.2, 109.8, 115.0, 123.5, 125.0,

1

27.7, 129.6, 131.8, 137.7, 138.9, 169.4, 175.0. LC/ESI-MS: positive

+

ꢀ1

The N-tboc protected compound was obtained by using the gen-

eral procedure C with 6-chloropyridine-3-carboxylic acid (158 mg,

mmol) for 12 h. A mixture of CH Cl and MeOH (9:1) was used

2 2

mode m/z = 270.3 ([M+H] ). Purity (>99.2%). IR (KBr, cm ) 3422,

1

⁄ ⁄

19 3 4 4 4 2

H N O 0.75C H O 1.0H O) C, H, N.

701, 1617, 970. Anal. (C16

1

for the chromatographic puriﬁcation. After removal of the solvents

a white solid was obtained. The N-tboc protection group of this so-

lid (340 mg, 0.92 mmol) was cleaved using the general procedure D

4

.62. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(1H-indol-3-

yl)methanone fumaric acid salt (61F)

4

h and a clear oil 58 (244 mg, 98%) was obtained after extraction.

The N-tboc protected compound was obtained by using the gen-

eral procedure C with indole-3-carboxylic acid (161 mg, 1 mmol)

for 12 h. A mixture of CH Cl and MeOH (20:1) was used for the

2 2

chromatographic puriﬁcation. After removal of the solvents a

white solid (85 mg, 23%) was obtained. The N-tboc protection

group of this solid (135 mg, 0.37 mmol) was cleaved using the gen-

eral procedure D for 2 h and a yellowish oil 61 (97 mg, 99%) was

obtained after extraction. This oil 61 (96 mg, 0.36 mmol) was

transferred to its fumaric acid salt 61F by using the general proce-

dure G with fumaric acid (41 mg, 0.36 mmol). Compound 61F

This oil 58 (244 mg, 0.91 mmol) was transferred to its fumaric acid

salt 58F by using the general procedure G with fumaric acid

(

105 mg, 0.91 mmol). Compound 58F (228 mg, 67%) was obtained

as a white solid in 63% yield over three steps; mp 154–157 °C (dec).

1

2

H NMR (400 MHz, D O) d 2.01 (br m, 2H), 2.25 (br s, 1H), 2.44 (br

s, 1H), 3.30–3.40 (br m, 3H), 3.42–3.64 (br m, 3H), 3.85 (br d,

J = 12.9 Hz, 1H), 4.53 (br d, J = 13.9 Hz, 1H), 6.67 (s, 1.86H), 7.63

(

d, J = 8.3 Hz, 1H), 7.97 (dd, J = 2.4, 8.3 Hz, 1H), 8.50 (d, J = 2.2 Hz,

H). ¹C NMR (100 MHz, D

3

1

5

2

O) d 24.7, 25.2, 26.9, 46.2, 46.4, 47.3,

1.4, 124.6, 129.8, 134.4, 138.6, 147.3, 151.8, 170.8, 171.4. ESI-

(

76 mg, 56%) was obtained as a white solid in 13% yield over three

+

1

MS: positive mode m/z = 266.3 ([M+H] ). HRMS for C13

calc m/z = 266.1055, found m/z = 266.1045. Purity (>99.5%). IR

3

H16ClN O:

steps; mp 185–190 °C (dec). H NMR (500 MHz, D

H), 2.28 (br s, 2H), 3.30–3.35 (br m, 2H), 3.40-3.48 (br m, 4H), 4.46

br d, J = 13.5 Hz, 2H), 6.62 (s, 1.6H), 7.27 (ddd, J = 8.1, 7.1, 1.1 Hz,

H), 7.32 (ddd, J = 8.2, 7.1, 1.2 Hz, 1H), 7.58 (d, J = 7.9 Hz, 1H),

2

O) d 1.98 (br m,

2

(

1

ꢀ1

(

cm ) 2854, 1640, 1581, 1405, 1357, 1105, 970, 640. Anal. (C13-

⁄ ⁄

3 4 4 4 2

16ClN O 0.75C H O 1.2H O) C, H, N.

H

7

304

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7

3

1

.70 (s, 1H), 7.79 (d, J = 7.9, 1H). ¹³C NMR (125 MHz, D

0.5, 50.3, 52.0, 111.0, 115.2, 123.2, 124.1, 125.8, 128.9, 132.0,

2

O) d 28.4,

evaporating the solvent under reduced pressure the residue was

dissolved in 30 mL of CH Cl

and washed with water (2 ꢁ 10 mL).

The organic layer was dried with MgSO and the solvent was re-

2

37.7, 138.5, 173.9, 175.1. LC/ESI-MS: positive mode m/z = 270.3

4

+

ꢀ1

(

[M+H] ). Purity (>99.8%). IR (KBr, cm ) 3398, 1701, 1600, 985.

moved under reduced pressure. Compound 65 was obtained as a

⁄

Anal. (C16

H

19

N

3

O 0.8C

4 4

H O

4

2

1.1H O) C, H, N.

white solid in 65% yield without further puriﬁcation; mp 89–

1

9

0 °C. H NMR (500 MHz, CDCl

3

) d 3.62 (m, 4H), 3.75 (m, 4H),

13

4

.63. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(1H-indol-5-

7.10 (m, 1H), 7.19 (t, J = 1.4 Hz, 1H), 7.87 (s, 1H).

(125 MHz, CDCl ) d 46.9, 66.6, 117.9, 130.1, 137.0, 151.0. LC/ESI-

MS: positive mode m/z = 181.9 ([M+H] ). Purity (>99%). IR (KBr,

C NMR

yl)methanone (62)

3

+

ꢀ1

The N-tboc protected compound was obtained by using the gen-

8 11 3 2

cm ) 3138, 1681, 1433, 1305. Anal. (C H N O ) C, H, N.

eral procedure C with indole-5-carboxylic acid (161 mg, 1 mmol)

for 12 h. A mixture of CH

2

Cl

2

and MeOH (20:1) was used for the

4.67. (4-Benzylpiperidin-1-yl)(1H-imidazol-1-yl)methanone

(66)

chromatographic puriﬁcation. After removal of the solvents an

off-white solid (364 mg, 98%) was obtained. The N-tboc protection

group of this solid (200 mg, 0.54 mmol) was cleaved using the gen-

eral procedure D for 4 h and compound 62 was obtained as a white

4-Benzylpiperidine (1.93 g, 11 mmol) and CDI (1.95 g, 12 mmol)

were dissolved in 20 mL of THF and stirred under reﬂux for 16 h.

After evaporating the solvent under reduced pressure the residue

solid (138 mg, 95%) after extraction, in 93% yield over two steps;

1

mp 225–226 °C. H NMR (400 MHz, CDCl

3

) d 1.88–1.96 (br m,

was dissolved in 30 mL of CH

2

Cl

2

and washed with water

and the sol-

4

1

H), 2.95–3.40 (br m, 6H), 4.14 (br s, 1H), 4.73 (br s, 1H), 6.55 (s,

H), 7.18 (dd, J = 1.4, 8.3 Hz, 1H), 7.22 (t, J = 2.7 Hz, 1H), 7.33 (d,

(2 ꢁ 10 mL). The organic layer was dried with MgSO

4

vent was removed under reduced pressure. Compound 66 was ob-

13

J = 8.3 Hz, 1H), 7.67 (s, 1H), 8.99 (br s, 1H). C NMR (100 MHz,

CDCl ) d 28.8, 32.4, 51.3, 103.1, 111.4, 119.9, 121.0, 125.7, 127.6,

27.9, 136.4, 172.7. ESI-MS: positive mode m/z = 271.3 ([M+H] ).

tained as a white solid in 87% yield without further puriﬁcation;

1

3

mp 64 °C. H NMR (500 MHz, CDCl

3

) d 1.30 (m, 2H), 1.77 (m, 2H),

+

1

1.83 (m, 1H), 2.59 (d, J = 7.1 Hz, 1H), 2.96 (m, 2H), 4.09 (br d,

J = 13.1 Hz, 2H), 7.07 (dd, J = 1.4, 0.9 Hz, 1H), 7.14 (m, 2H), 7.18

(t, J = 1.4 Hz, 1H), 7.22 (m, 1H), 7.29 (m, 2H), 7.84 (t, J = 1.0 Hz,

ꢀ

1

Purity (>99%). IR (cm ) 3316, 2859, 1614, 1580, 1425, 1233,

9

30, 886.

1

3

1

H). C NMR (125 MHz, CDCl

3

) d 32.0, 38.1, 42.9, 47.0, 118.1,

4

.64. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(1H-indazol-6-

126.4, 128.5, 129.2, 129.7, 137.0, 139.6, 151.0. LC/ESI-MS: positive

+

-

yl)methanone (63)

mode m/z = 270.1 ([M+H] ), negative mode m/z = 268.0 ([M – H] ).

ꢀ1

Purity (>99.5%). IR (KBr, cm ) 3154, 2951, 1680, 1464, 1430. Anal.

19 3

(C16H N O) C, H, N

The N-tboc protected compound was obtained by using the gen-

eral procedure C with 1H-indazole-6-carboxylic acid (162 mg,

1

mmol) for 12 h. A mixture of CH

2

Cl

2

and MeOH (20:1) was used

4.68. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(pyrrolidin-1-

for the chromatographic puriﬁcation. After removal of the solvents

a white solid (280 mg, 76%) was obtained. The N-tboc protection

group of this solid (160 mg, 0.43 mmol) was cleaved using the gen-

eral procedure D for 4 h and compound 63 was obtained as an off-

yl)methanone fumaric acid salt (67F)

Compound 64 (182 mg, 1.1 mmol) was dissolved in 2 mL of

MeCN and methyl iodide (625 mg, 4.4 mmol) was added. The solu-

tion was stirred at rt for 24 h before the volatiles were removed un-

white solid (110 mg, 94%) after extraction, in 71% yield over two

1

steps; mp 227–229 °C. H NMR (400 MHz, DMSO-d

6

) d 1.45–2.05

der reduced pressure. The residue was dissolved in 10 mL of CH

2 2

Cl

(

br m, 4H), 2.90–3.40 (br m, 6H), 3.64 (br s, 1H), 4.57 (br s, 1H),

and Et N (101 mg, 1 mmol) and N-tboc-bispidine 13 (226 mg,

3

7

(

.09 (d, J = 8.2 Hz, 1H), 7.51 (s, 1H), 7.80 (d, J = 8.2 Hz, 1H), 8.11

) d 27.9,

1 mmol) were added. The solution was stirred at rt for another

24 h before it was washed twice with 5 mL of hydrochloric acid

s, 1H), 13.19 (br s, 1H). ¹³C NMR (100 MHz, DMSO-d

6

3

1

1.6, 46.4, 50.7, 52.0, 108.3, 118.9, 120.6, 122.6, 133.4, 135.1,

39.3, 169.5. ESI-MS: positive mode m/z = 271.3 ([M+H] ). HRMS

(1 M) and once with 5 mL of H

MgSO , ﬁltered and the solvent is removed under reduced pres-

sure. The residue was puriﬁed by ﬂash chromatography using a

mixture of CH Cl and MeOH (40:1) and after removal of the sol-

2

O. The organic layer was dried with

+

4

19 3

for C16H N O: calc m/z = 270.1601, found m/z = 270.1594. Purity

ꢀ

1

(

>99%). IR (cm ) 3316, 2859, 1614, 1580, 1425, 1233, 930, 886.

2

vents a white solid (223 mg, 69%) was obtained. The N-tboc protec-

tion group of this solid (180 mg, 0.56 mmol) was cleaved using the

general procedure D for 3 h and a white solid 67 (120 mg, 97%) was

obtained after extraction. This solid 67 (105 mg, 0.47 mmol) was

transferred to its fumaric acid salt 67F by using the general proce-

dure F. Compound 67F (109 mg, 67%) was obtained as a white solid

4

.65. (1H-Imidazol-1-yl)(pyrrolidin-1-yl)methanone (64)

Pyrrolidine (0.78 g, 11 mmol) and CDI (1.95 g, 12 mmol) were

dissolved in 20 mL of THF and stirred under reﬂux for 16 h. After

evaporating the solvent under reduced pressure the residue was

dissolved in 30 mL of CH

The organic layer was dried with MgSO

moved under reduced pressure. Compound 64 was obtained as a

1

2

Cl

2

and washed with water (2 ꢁ 10 mL).

in 45% yield over three steps; mp 166–169 °C (dec). H NMR

4

and the solvent was re-

(500 MHz, MeOD) d 1.88–1.93 (br m, 5H), 1.97–2.02 (br m, 1H),

2.17 (br m, 1H), 3.09-3.13 (br m, 2H), 3.29-3.34 (br m, 2H), 3.43-

3.47 (br m, 4H), 3.50 (br d, J = 12.8 Hz, 2H), 3.81 (br d, J = 13.3 Hz,

white solid in 90% yield without further puriﬁcation; mp 54–

1

13

5

7

5 °C. H NMR (500 MHz, CDCl

3

) d 1.98 (m, 4H), 3.62 (m, 4H),

2H), 6.68 (s, 2.0H). C NMR (125 MHz, MeOD) d 26.4, 27.9, 30.5,

.06 (dd, J = 1.4, 0.9 Hz, 1H), 7.34 (t, J = 1.4 Hz, 1H), 8.01 (s, 1H).

48.9, 49.4, 52.1, 136.2, 164.8, 171.5. LC/ESI-MS: positive mode m/

1

3

+

ꢀ1

C NMR (125 MHz, CDCl

49.8. LC/ESI-MS: positive mode m/z = 166.1 ([M+H] ). Purity

3

) d 25.7, 48.0, 117.7, 129.6, 136.8,

z = 224.1 ([M+H] ). Purity (>99.4%). IR (KBr, cm ) 3441, 1710,

+

⁄ ⁄

21 3 4 4 4 2

H N O 1.0C H O 0.25H O) C, H, N.

1

1635, 972. Anal. (C12

ꢀ

1

(

>99.5%). IR (KBr, cm

O) C, H, N.

)

3138, 1672, 1423, 1339. Anal.

8 11 3

C H N

4.69. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-

yl(morpholino)methanone fumaric acid salt (68F)

4

.66. (1H-Imidazol-1-yl)(morpholino)methanone (65)

Compound 65 (182 mg, 1.1 mmol) was dissolved in 2 mL of

MeCN and methyl iodide (625 mg, 4.4 mmol) was added. The

solution was stirred at rt for 24 h before the volatiles were

Morpholine (0.96 g, 11 mmol) and CDI (1.95 g, 12 mmol) were

dissolved in 20 mL of THF and stirred under reﬂux for 16 h. After

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7305

removed under reduced pressure. The residue was dissolved in

2H), 2.03 (s, 3H), 2.09 (s, 3H), 2.12 (br d, J = 11.2 Hz, 1H), 2.20 (br

d, J = 11.0 Hz, 1H), 2.87 (br m, 3H), 3.34 (d, J = 13.1 Hz, 1H), 3.79

1

2 2 3

0 mL of CH Cl and Et N (101 mg, 1 mmol) and N-tboc-bispidine

13

1

3 (226 mg, 1 mmol) were added. The solution was stirred at rt

(br d, J = 13.1 Hz, 1H), 4.55 (br d, J = 13.2 Hz, 1H).

(100 MHz, CDCl ) d 22.2, 29.2, 29.6, 31.2, 46.5, 46.9, 51.2, 60.5,

61.0, 81.4, 170.3. ESI-MS: positive mode m/z = 183.2 ([M+H] ),

C NMR

for another 24 h before it was washed twice with 5 mL of hydro-

chloric acid (1 M) and once with 5 mL of H O. The organic layer

was dried with MgSO , ﬁltered and the solvent is removed under

reduced pressure. The residue was puriﬁed by ﬂash chromatogra-

phy using a mixture of CH Cl and MeOH (9:1) and after removal

3

+

2

+

ꢀ1

4

205.2 ([M+Na] ). Purity (>95%). IR (cm ) 2923, 2862, 2775,

1614, 1443, 1259, 997.

2

of the solvents a white solid (336 mg, 99%) was obtained. The N-

tboc protection group of this solid (130 mg, 0.38 mmol) was

cleaved using the general procedure D for 2 h and a clear oil 68

4.72. 1-((1R,5S)-7-Ethyl-3,7-diazabicyclo[3.3.1]nonan-3-

yl)ethanone (71)

(

0

87 mg, 95%) was obtained after extraction. This oil 68 (87 mg,

.36 mmol) was transferred to its fumaric acid salt 68F by using

Compound 71 was obtained by using general procedure H with

iodoethane (101.4 mg). Puriﬁcation was achieved by ﬂash chroma-

the general procedure G with fumaric acid (42 mg, 0.36 mmol).

2 2

tography using a mixture of CH Cl and MeOH (9:1) and after re-

Compound 68F (57 mg, 44%) was obtained as an off white solid

in 42% yield over three steps; mp 155–158 °C (dec). H NMR

moval of the solvents under reduced pressure the ﬁnal

compound 71 was obtained as a clear oil (74.0 mg, 58%). H NMR

1

(

2

4

(

4

500 MHz, D

2

O) d 1.93 (br m, 1H), 1.98 (br m, 1H), 2.24 (br s,

H), 3.27 (br m, 2H), 3.35 (br m, 2H), 3.38 (dd, J = 4.9, 4.7 Hz,

H), 3.54 (br d, J = 13.2 Hz, 2H), 3.76 (dd, J = 4.9, 4.7 Hz, 4H), 3.78

O) d 28.8, 31.4,

9.3, 50.4, 53.9, 69.1, 137.5, 168.0, 173.9. LC/ESI-MS: positive mode

3

(400 MHz, CDCl ) d 0.95 (t, J = 7.0 Hz, 3H), 1.61 (br m, 1H), 1.70

(br m, 1H), 1.84 (br s, 2H), 2.01 (s, 3H), 2.04-2.26 (br m, 5H), 2.81

(br d, J = 13.2 Hz, 1H), 2.96 (br d, J = 10.7 Hz, 2H), 3.33 (br d,

J = 12.7 Hz, 1H), 3.78 (br d, J = 13.4 Hz, 1H), 4.60 (br d, J = 13.4 Hz,

br m, 2H), 6.73 (s, 1.5H). ¹³C NMR (125 MHz, D

2

1

3

1H). C NMR (100 MHz, CDCl

3

) d 12.4, 22.2, 29.2, 29.7, 32.1,

+

ꢀ1

m/z = 240.4 ([M+H] ). Purity (>99.7%). IR (KBr, cm ) 3440, 1679,

1

46.4, 51.2, 52.7; 58.3; 58.4, 169.8. ESI-MS: positive mode m/

⁄

+

ꢀ1

637, 985, 967. Anal. (C12

H

21

N

3

O

2

0.75C

4 4

H O

4

2

1.5H O) C, H, N.

z = 197.3 ([M+H] ), 219.2 ([M+Na] ). Purity (>95%). IR (cm )

2

916, 2864, 2748, 1631, 1445, 1233, 994, 586.

4

.70. (1R,5S)-3,7-Diazabicyclo[3.3.1]nonan-3-yl(4-

benzylpiperidin-1-yl)methanone fumaric acid salt (69F)

4

.73. 1-((1R,5S)-7-Propyl-3,7-diazabicyclo[3.3.1]nonan-3-

yl)ethanone (72)

Compound 66 (296 mg, 1.1 mmol) was dissolved in 2 mL of

MeCN and methyl iodide (625 mg, 4.4 mmol) was added. The solu-

tion was stirred at rt for 24 h before the volatiles were removed

under reduced pressure. The residue was dissolved in 10 mL of

Compound 72 was obtained by using general procedure H with

1

-iodopropane (110.5 mg). Puriﬁcation was achieved by ﬂash chro-

2 2

matography using a mixture of CH Cl and MeOH (20:1) and after

CH

226 mg, 1 mmol) was added. The solution was stirred at rt for an-

other 24 h before it was washed twice with 5 mL of hydrochloric

acid (1 M) and once with 5 mL of H O. The organic layer was dried

with MgSO , ﬁltered and the solvent was removed under reduced

pressure. The residue was puriﬁed by ﬂash chromatography using

a mixture of CH Cl and MeOH (40:1) and after removal of the sol-

2 2 3

Cl and Et N (101 mg, 1 mmol) and N-tboc-bispidine 13

removal of the solvents under reduced pressure the ﬁnal com-

(

_p_o_u_n_d₇₂_w_a_s_o_b_t_a_i_n_e_d_a_s_a_c_l_e_a_r_o_i_l₍₉₁_.₆_m_g_,₆₇_%₎_.1H NMR

(

400 MHz, CDCl3) d 0.82 (t, J = 7.4 Hz, 3H), 1.32–1.49 (br m, 2H),

2

1

2

.69 (br s, 2H), 1.88 (br s, 2H), 2.03 (s, 3H), 2.06–2.32 (br m, 4H),

.83 (br d, J = 13.2 Hz, 1H), 2.95 (br d, J = 8.0 Hz, 2H), 3.32 (br d,

4

J = 13.4 Hz, 1H), 3.78 (br d, J = 13.0 Hz, 1H), 4.58 (br d, J = 13.5 Hz,

2

13

1

H). C NMR (100 MHz, CDCl3) d 11.8, 20.0, 22.3, 29.0, 29.5,

1.5, 46.5, 51.2, 61.2, 80.7, 170.0. ESI-MS: positive mode m/

vents a white solid (355 mg, 83%) was obtained. The N-tboc pro-

tection group of this solid (300 mg, 0.70 mmol) was cleaved

using the general procedure D for 3 h and a clear oil 69 (191 mg,

3

+

ꢀ1

z = 211.3 ([M+H] ). Purity (>95%). IR (cm ) 2921, 2858, 2204,

603, 1431, 1205, 758, 688.

1

8

0

3%) was obtained after extraction. This oil 69 (124 mg,

.38 mmol) was transferred to its fumaric acid salt 69F by using

the general procedure G with fumaric acid (44 mg, 0.38 mmol).

4.74. Membrane preparation: preparation of rat brains

Compound 69F (124 mg, 72%) was obtained as a white solid in

1

5

0% yield over three steps; mp 172–175 °C (dec). H NMR

Frozen rat brains (Pel-Freez Biologicals, AR) (–80 °C) were

thawed slowly before the preparation of the P2 rat brain mem-

brane fraction (30–60 min. on ice, afterwards at rt). A single cut

just behind the inferior colliculi was done to exclude the cerebel-

lum and the medulla. After the determination of the wet weight

(1.32 g on average), the brains were pressed into a pulp using a syr-

inge and homogenized in saccharose buffer with a glass teﬂon

homogenizer (potter, 10 sec.). The tissue was then centrifuged

(1000g, 20 min., 4 °C), the supernatant (S1) aspirated with a Pas-

teur pipette and stored on ice. The P1 pellet was re-suspended in

saccharose buffer and the centrifugation was repeated. The super-

(

500 MHz, D

2

O) d 1.17–1.27 (br m, 2H), 1.69 (br m, 2H), 1.82 (br

m, 1H), 1.91 (br d, J = 13.6 Hz, 1H), 1.97 (br d, J = 13.6 Hz, 1H),

.21 (br s, 2H), 2.61 (d, J = 7.1 Hz, 2H), 2.87 (dt, J = 12.4, 1.9 Hz,

H), 3.22 (br d, J = 13.1 Hz, 2H), 3.34 (br d, J = 13.1 Hz, 2H), 3.53

br d, J = 13.2 Hz, 2H), 3.70 (br m, 4H), 6.69 (s, 2.0H), 7.27-7.41

2

O) d 27.5, 30.2, 32.8, 38.8, 43.4,

8.1, 49.1, 52.9, 127.6, 129.9, 130.8, 136.2, 142.3, 166.8, 173.0.

LC/ESI-MS: positive mode m/z = 328.4 ([M+H] ). Purity (>99.8%).

2

(

1

3

m, 5H). C NMR (125 MHz, D

4

+

ꢀ1

⁄

IR (KBr, cm ) 3441, 1704, 1628, 981, 965. Anal. (C20

H

29

N

3

O 1.0C4-

⁄

4

O

4

0.5H

2

O) C, H, N.

0

natant S1 was collected and added to the supernatant S1. The

4

.71. 1-((1R,5S)-7-Methyl-3,7-diazabicyclo[3.3.1]nonan-3-

combined supernatants were centrifuged (25,000g, 20 min, 4 °C),

the supernatant S2 was removed and the pellet P2 collected and di-

luted with HSS buffer. The buffer volume added was calculated on

the basis of the wet weight on a ratio 1:2.

yl)ethanone (70)

Compound 70 was obtained by using general procedure H with

iodomethane (92.3 mg). Puriﬁcation was achieved by ﬂash chro-

matography using a mixture of CH Cl and MeOH (9:1) and after

2 2

The ﬁnal pellet was stored in aliquots at ꢀ80 °C. On the day of

the experiment, the P2 membrane fraction was thawed, diluted

with HSS buffer (30-fold volume), homogenized and centrifuged

(35,000g, 10 min, 4 °C). The collected pellet was suspended in

HSS buffer and used in the radioligand binding experiment.

removal of the solvents under reduced pressure the ﬁnal com-

1

pound 70 was obtained as a clear oil (47.4 mg, 40%). H NMR

(

3

400 MHz, CDCl ) d 1.59 (br m, 1H), 1.68 (br m, 1H), 1.87 (br s,

7

306

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

4

.75. Membrane preparation: preparation of calf adrenals

punched out and transferred into 4 mL scintillation vials. The scin-

tillation vials were ﬁlled with scintillation cocktail (2 mL) and the

radioactivity was measured using a liquid scintillation counter. As-

says were carried out in duplicates, triplicates or quadruplicates.

Frozen calf adrenals (Pel-Freez Biologicals, AR) (ꢀ80 °C) were

placed on ice (30-60 min.) and allowed to thaw slowly before they

were cut into small pieces. After determination of the wet weight

(

7

4

4–6 g), the tissue was homogenized in HSS buffer (Ultraturrax at

50 rpm). The homogenate was centrifuged (30,000g, 10 min,

°C), the pellets collected and washed. This procedure was re-

4.79. Radioligand binding assay: competition assay using (±)-

3

[ H]epibatidine ([ H]Epi) and calf adrenals membrane fraction

⁄

(a

3b4 nAChR)

peated ﬁve times. The buffer volume to re-suspend the pellet

was calculated on the basis of the wet weight in a ratio of 1:6.5.

The prepared tissues were stored in aliquots at ꢀ80 °C. One

hour before the experiments the tissues were slowly thawed,

homogenized in HSS buffer and centrifuged (25,000g, 20 min,

A dilution row of 6–9 concentrations of the test compound was

prepared. Each assay sample, with a total volume of 500

tained 200 L of the test compound, 100 L of (±)-[ H]Epi

(0.5 nM), 100 L of the calf adrenal membrane fraction (60–

70 g) and 100 L HSS buffer. Non-speciﬁc binding was deter-

mined in the presence of 300

M (ꢀ)-nicotine tartrate salt. The

lL con-

3

l

4

°C). The pellets were re-suspended in fresh HSS buffer and used

l

for radioligand binding assays.

l

samples were homogenized and incubated (90 min at 22 °C). The

incubation was terminated by vacuum ﬁltration through glass ﬁber

ﬁlters presoaked in 1% PEI solution. The ﬁlters were rinsed three

times with TRIS buffer, punched out and transferred into 4 mL scin-

tillation vials. The scintillation vials were ﬁlled with scintillation

cocktail (2 mL) and the radioactivity was measured using a liquid

scintillation counter. Assays were carried out in duplicates, tripli-

cates or quadruplicates.

4

.76. Membrane preparation: preparation of Torpedo

californica electroplax

Frozen samples of Torpedo californica electric organ (Dr.

Charles Winkler, CA) (ꢀ80 °C) were placed on ice (30–60 min.)

and allowed to thaw slowly before the membrane preparation.

The tissue was homogenized in ice-cold HSS buffer (Ultraturrax

at 750 rpm) and centrifuged (30,000g, 10 min, 4 °C). The pellets

were collected, washed four times with HSS buffer through re-

homogenization and centrifugation at the same settings. The

remaining pellets were collected, re-suspended in HSS buffer and

stored at aliquots at ꢀ80 °C.

4.80. Radioligand binding assay: competition assay using (±)-

3

[ H]epibatidine ([ H]Epi) and Torpedo californica electroplax

(( 1)2b1 d nAChR)

a

c

One hour before the experiments the tissues were slowly

thawed, homogenized in HSS buffer and centrifuged (25,000g,

A dilution row of 6–9 concentrations of the test compound was

prepared. Each assay sample, with a total volume of 500 L con-

L of (±)-[ H]Epi, 100

g) and 100 L HSS buf-

fer. Non-speciﬁc binding was determined in the presence of

300

M (ꢀ)-nicotine tartrate salt. The samples were homogenized

l

3

2

0 min, 4 °C). The pellets were re-suspended in fresh HSS buffer

tained 200

l

L of the test compound, 100

l

lL

and used for radioligand binding assays.

of Torpedo californica electroplax (60–70

l

4

.77. Radioligand binding assay: competition assay using (±)-

l

3

⁄

[

H]epibatidine ([ H]Epi) and rat brain P2 fraction (

a4b2

and incubated (90 min at 22 °C). The incubation was terminated by

vacuum ﬁltration through glass ﬁber ﬁlters presoaked in 1% PEI

solution. The ﬁlters were rinsed three times with TRIS buffer,

punched out and transferred into 4 mL scintillation vials. The scin-

tillation vials were ﬁlled with scintillation cocktail (2 mL) and the

radioactivity was measured using a liquid scintillation counter. As-

says were carried out in duplicates, triplicates or quadruplicates.

nAChR)

A dilution row of 6–9 concentrations of the test compound was

prepared. Each assay sample, with a total volume of 500 L con-

tained 100 L of the membrane protein (60 g), 100 L of (±)-

H]Epi (PerkinElmer, Inc., MA) (0.5 nM), 100 L of HSS buffer

L of the test compound. Non-speciﬁc binding was deter-

M (ꢀ)-nicotine tartrate salt. The

l

3

[

l

and 200

l

mined in the presence of 300

l

4.81. Heterologous expression of nAChRs in Xenopus laevis

oocytes

samples were homogenized and incubated (90 min at 22 °C). The

incubation was terminated by vacuum ﬁltration (Brandel Har-

vester, Adi Hassel, Munich) through glass ﬁber ﬁlters presoaked

in 1% PEI solution. The ﬁlters were rinsed three times with TRIS

buffer, punched out and transferred into 4 mL scintillation vials.

The scintillation vials were ﬁlled with scintillation cocktail (2 mL)

and the radioactivity was measured using a liquid scintillation

counter. Assays were carried out in duplicates, triplicates or

quadruplicates.

Mouse muscle nAChR

a, b, and d clones used for receptor

expression in Xenopus laevis oocytes were obtained from Dr. J.

Boulter (University of California, Los Angeles, CA), and the mouse

clone was provided by Dr. P. Gardner (University of Massachusetts

Medical School, Worcester, MA). Human nAChR clones were ob-

tained from Dr. J. Lindstrom (University of Pennsylvania, Philadel-

phia, PA). The human Resistance-to-cholinesterase 3 (RIC-3) clone,

obtained from Dr. M. Treinin (Hebrew University, Jerusalem, Is-

4

[

.78. Radioligand binding assay: competition assay using

H]methyllycaconitine ([ H]MLA) and rat brain P2 fraction (

rael), was co-injected with

a7 to improve the level and speed of

3

⁄

a7

a

7 receptor expression without affecting the pharmacological

5

7

nAChR)

properties of the receptors. Subsequent to linearization and puri-

ﬁcation of the plasmid cDNAs, cRNAs were prepared using the

mMessage mMachine in vitro RNA transfection kit (Ambion, Aus-

tin, TX).

A dilution row of 6–9 concentrations of the test compound was

prepared. Each assay sample, with a total volume of 250 L con-

L of [ H]MLA (Tocris,

UK/Biotrend, Germany; ARC, Inc., MO), 100 L of P2 membrane

protein fraction (60–70 g). Non-speciﬁc binding was determined

in the presence of 1 M MLA. The samples were homogenized

l

3

tained 50

l

L of the test compound, 100

l

Oocytes were surgically removed from mature female Xenopus

laevis frogs (Nasco, Ft. Atkinson, WI) and injected appropriate

l

6

0

l

nAChR subunit cRNAs as described previously. Frogs were main-

tained in the Animal Care Service facility of the University of Flor-

ida, and all procedures were approved by the University of Florida

Institutional Animal Care and Use Committee. In brief, the frog was

ﬁrst anesthetized for 15–20 min in 1.5 L frog tank water containing

l

and incubated (120 min at 22 °C). The incubation was terminated

by vacuum ﬁltration through glass ﬁber ﬁlters presoaked in 1%

PEI solution. The ﬁlters were rinsed three times with TRIS buffer,

C. Eibl et al. / Bioorg. Med. Chem. 21 (2013) 7283–7308

7307

1

g of 3-aminobenzoate methanesulfonate (MS-222) buffered with

References and notes

sodium bicarbonate. The harvested oocytes were treated with

1

2

3

.

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.25 mg/ml collagenase (Worthington Biochemicals, Freehold, NJ)

for 2 h at room temperature in a calcium-free Barth’s solution

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(

1

3

4

,

4

5

6

.

licular layer. Stage V oocytes were subsequently isolated and in-

jected with 50 nl of 5–20 ng nAChR subunit cRNA. Recordings

were carried out 1–7 days after injection.

7. Gündisch, D.; Eibl, C. Exp. Opin. Ther. Pat. 2011 , 21 , 1867 .

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Article Doi

The 3,7-diazabicyclo[3.3.1]nonane scaffold for subtype selective nicotinic acetylcholine receptor (nAChR) ligands. Part 1: The influence of different hydrogen bond acceptor systems on alkyl and (hetero)aryl substituents

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