Novel dual inhibitors of AChE and MAO derived from hydroxy aminoindan and phenethylamine as potential treatment for Alzheimer's disease

5260

J . Med. Chem. 2002, 45, 5260-5279

Novel Du a l In h ibitor s of ACh E a n d MAO Der ived fr om Hyd r oxy Am in oin d a n

a n d P h en eth yla m in e a s P oten tia l Tr ea tm en t for Alzh eim er ’s Disea se

J effrey Sterling,*^,†Yaacov Herzig,^†Tamar Goren,^†Nina Finkelstein,^†David Lerner,^†Willy Goldenberg,^†

Istvan Miskolczi,^‡Sandor Molnar,^‡Ferenc Rantal,^‡Tivadar Tamas,^‡Gyorgy Toth,^‡Adela Zagyva,^‡

Andras Zekany,^‡Gila Lavian,^§Aviva Gross,^§Rachel Friedman,^§Michal Razin,^|Wei Huang,^|Boris Krais,^|

Michael Chorev, Moussa B.Youdim,^§and Marta Weinstock^|

Research and Development Division, Teva Pharmaceutical Industries (Abic, P.O. Box 8077, Netanya 42504, Israel;

Teva, P. O. Box 1142, J erusalem 91010, Israel; and Biogal, Debrecen H-4042, Hungary); Department of Pharmacology,

The Rappaport Institute, School of Medicine, The Technion, Bat-Galim, Haifa 31096, Israel; Department of Pharmacology,

School of Pharmacy, The Hebrew University, Ein-Kerem, J erusalem 91120, Israel; and the Bone and Mineral Metabolism Unit,

Charles A. Dana and Thorndike Laboratories, Department of Medicine, Beth Israel Deaconess Medical Center and Harvard

Medical School, 330 Brookline Avenue, Boston, Massachusetts 02215

Received March 14, 2002

Carbamate derivatives of N-propargylaminoindans (Series I) and N-propargylphenethylamines

(Series II) were synthesized via multistep procedures from the corresponding hydroxy

precursors. The respective rasagiline- and selegiline-related series were designed to combine

inhibitory activities of both acetylcholine esterase (AChE) and monoamine oxidase (MAO) by

virtue of their carbamoyl and propargylamine pharmacophores. Each compound was tested

for these activities in vitro in order to find molecules with similar potencies against each enzyme.

Compounds with such dual AChE and MAO inhibitory activities are expected to have potential

for the treatment of Alzheimer’s disease. The observed SAR also offers insight into the

requirements of the active sites on these enzymes. A carbamate moiety was found to be essential

for AChE inhibition, which was absent in the corresponding hydroxy precursors. The propargyl

group caused 2-70-fold decrease in AChE inhibitory activity (depending on the position of the

carbamoyl group) of Series I, but had little or no effect in Series II. Thus, the 6- and

7-carbamyloxyphenyls in Series I were either equipotent to, or slightly (2- to 5-fold) less active

as AChE inhibitors than, the corresponding compounds in Series II, while the 4-carbamyloxy-

phenyls were more potent. The presence of the carbamate moiety in 6- and 7-carbamyloxyphen-

yls of Series I, considerably decreased MAO-A and -B inhibitory activity, compared to that of

the parent hydroxy analogues, while the opposite was true for Series II. Thus, the 6- and

7-carbamyloxyphenyls in Series I were 2-3 orders of magnitude weaker MAO inhibitors while

the 4- carbamyloxyphenyls were equipotent with the corresponding compounds in Series II. In

both series, N-methylation of the propargylamine enhanced the MAO (A and B equally)

inhibitory activities and decreased the AChE inhibitory activity. Two candidates belonging to

the indan and tetralin ring systems (24c, 27b) and one phenethylamine (53d ) were identified

as possible leads for further development based on the following criteria: (a) comparable AChE

and MAO-B inhibitory activities, (b) good to moderate AChE inhibitory activity, and (c) lack of

strong MAO-A selectivity. However, it is likely that these compounds will be metabolized to

the corresponding phenols, with inhibitory activities against AChE and/or MAO-A or -B,

different from those of the parent carbamates. Thus, the apparent enzyme inhibition will be a

result of the combined inhibition of all of these individual metabolites. The results of our ongoing

in vivo screening programs will be published elsewhere.

In tr od u ction

arising in the nucleus basalis of Meynert.¹In addition,

depressive symptoms occur in subjects with AD,²and

these may be associated with decreases in serotoninergic

and noradrenergic transmission in the limbic system.³

Although the cause of progressive neuronal degenera-

tion in AD is not known, evidence for the presence of

oxidative stress, mediated by increased levels of iron,

breakdown of peroxynitrite, and nitration of tyrosine

residues in cell membrane proteins has been reported.⁴

Monoamine oxidase B (MAO-B) activity also increases

in association with gliosis, which can result in higher

levels of H₂O₂and oxidative free radicals.⁵

Alzheimer’s disease (AD) is characterized by a pro-

gressive impairment in memory and intellectual ability,

accompanied by behavioral disturbances and a decreas-

ing ability to perform basic activities of daily living. The

degree of memory impairment correlates well with the

loss of cholinergic transmission in the temporal lobe and

other cortical brain regions innervated by neurons

* To whom correspondence should be addressed. Phone: + 972 2

5892985, Fax: + 972 2 5821163. E-mail: jeffrey.sterling@teva.co.il.

^†Teva Pharmaceutical Industries, Israel.

^‡Biogal Pharmaceutical Works, a Teva subsidiary, Hungary.

^§Department of Pharmacology, The Technion.

Currently, the only approved therapy for AD is based

on a reduction of the cognitive deficits by enhancing

^|School of Pharmacy, The Hebrew University.

Harvard Medical School.

Published on Web 10/23/2002

Novel Dual Inhibitors of AChE and MAO

J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5261

Sch em e 1^a

a

(a) (Boc)₂O, Et₃N, THF, rt; (b) R₃R₄NCXCl (X ) O, S), CH₃CN, NaH, rt or R₄NCO, Et₃N, CH₂Cl₂,rt; (c) HCl/dioxane, rt; (d) propargyl

bromide, K₂CO₃, CH₃CN or DMA, rt; (e) HCl/Et₂O, rt; (f) NaCNBH₃/(CH₂O)_n.

cholinergic transmission through inhibition of acetyl-

cholinesterase (AChE). These anti-AChE agents include

tacrine, galanthamine, donepezil, and rivastigmine (1b),

which have been shown to induce a modest improve-

ment in memory and cognitive function,⁶but do not

appear to prevent or slow the progressive neuro-

degeneration. On the other hand, selegiline, a selective

MAO-B inhibitor, has been reported to retard the

further deterioration of cognitive functions to more

advanced milestones in AD.⁷The propargylamine phar-

macophore of rasagiline (2), selegiline (3), and related

compounds also appears to have neuropotective activity

independent of MAO inhibition.^8-10Because of the

complexity of AD, it is unlikely that a single pharma-

cological action will provide a comprehensive and sat-

isfactory therapeutic solution for such patients. Such

therapy is more likely to be achieved by the use of

compounds that incorporate several pharmacological

traits into a single molecular entity and will work in a

synergistic manner. Attempts to combine anti-AChE

pounds were reversible MAO inhibitors in vitro. Some

of the imino 1,2,3,4-tetrahydrocyclopent[b]indole car-

bamates were potent dual inhibitors of AChE and MAO

in vitro, but exhibited low activity after oral administra-

tion, possibly because of poor brain penetration or low

bioavailability.^12,13N-Pyrimidine 4-acetylaniline deriva-

tives possessing AChE and reversible MAO-A inhibitory

activity in vitro have also been reported.¹⁴Several

7-aryloxycoumarin derivatives with known MAO inhibi-

tory activity were shown to act as noncompetitive AChE

inhibitors.¹⁵

The present study describes the preparation and

preliminary in vitro screening of two series of dual MAO

and AChE inhibitors that are based on introduction of

a carbamate moiety, to confer AChE inhibitory activity

on either rasagiline (Series I) or selegiline (Series II),

both of which are MAO-B inhibitors with neuroprotec-

tive activity in vitro and in vivo.^8,16-19We anticipated

that in addition to reduction in oxidative stress,²⁰drugs

that possess MAO-A inhibitory activity might also have

a direct effect on cognition and act as antidepressant

agents.

Our goal is identification of propargylamino carbam-

ates that are equipotent as MAO and AChE inhibitors

and therefore may be used for the treatment of AD and

other neurodegenerative diseases. By combining SAR

and modeling data, we will also gain insight into the

requirements of the active sites on these two enzymes.

and anti-MAO activities in one molecular entity have

previously been reported.^11-15Imino 1,2,3,4-tetrahydro-

cyclopent[b]indole carbamates, hybrids of physostig-

mine, an AChE inhibitor, and the MAO inhibitors

selegiline and tranylcypromine, were generated by

incorporating a carbamate and either a propargylamine

or a cyclopropylamine moiety into a single molecular

scaffold.¹²The N-alkylimine precursors of these com-

Ch em istr y

Ser ies I. In general, compounds of Series I were

synthesized via a four-step procedure from hydroxy

aminoindans 4 (Scheme 1) or analogous hydroxy ami-

notetralins, as follows. Hydroxy amines were N-Boc

protected (Boc₂O, Et₃N, THF) and carbamoylated by

carbamoyl/thiocarbamoyl chlorides or by alkyl isocyan-

5262 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24

Sterling et al.

Sch em e 2^a

a

(a) (CF₃CO)₂O, KOH, toluene, H₂O, rt; (b) ClCH₂COCl, AlCl₃,1,2-DCE, rt; (c) mCPBA,CH₂Cl₂,TFA,rt; (d) K₂CO₃, MeOH, H₂O, 70 °C.

Sch em e 3^a

a

(a) SO₂Cl₂, AcOH, rt; (b) 2,2-dimethyl-1,3-dioxane-4,6-dione (Meldrum’s acid), Et₃N, HCO₂H; (c) SOCl₂, reflux; (d) AlCl₃, CH₂Cl₂,

0 °C, then rt; (e) NH₂OH, NaOAc, H₂O, EtOH, reflux; (f) MoO₃, MeOH, DMF, rt; (g) HBr, AcOH, H₂O, reflux.

Sch em e 4. Preparation of 39u -w and 40u -w ^a

ates to give the dialkyl and monoalkyl carbamates (6),

respectively. The carbamates were selectively Boc-

deprotected (HCl, dioxane) and propargylated (propar-

gyl bromide, K₂CO₃, CH₃CN). The target compounds

were finally isolated as salts. This synthetic strategy

was chosen rather than the more economical approach

of preparing first hydroxy propargylaminoindans (39)

as common intermediates for the various carbamates,

because we also wanted to test the biological activities

of the non-propargyl carbamates. The order of propar-

gylation and carbamoylation was reversed in the 1-meth-

yl-2-propynyl analogues 26, and 6-hydroxy-1-aminoin-

dan was first propargylated by 3-mesyloxy-1-butyne.²¹

Compounds 31a and 32a were obtained by carbamoy-

lation of the corresponding hydroxy-N,N-dimethyl-1-

aminoindan 30. The N-methyl propargyl carbamates

19c, 22b , 22c, and 29b were prepared by N-mono-

methylation of the corresponding nor compounds.

Racemic starting hydroxy amines 4 were prepared

according to known procedures.^22-27Optically pure

6-hydroxy-1-aminoindan was prepared either by optical

resolution of the racemic mixture²⁸or via a regioselec-

tive Friedel-Crafts chloroacetylation of the N-trifluo-

roacetyl derivative (33) of optically pure 1-aminoindan

(prepared from indene via N-benzyl-1-aminoindan²⁹),

followed by a Baeyer-Villiger oxidation and finally

hydrolysis (Scheme 2). This method was first reported³⁰

for the preparation of 6-hydroxyindole and was success-

fully applied to the synthesis of our aminoindan deriva-

tives. The chloro analogues 24 were synthesized from

5-chloro-6-hydroxy-1-aminoindan 38, which was pre-

pared from p-methoxybenzaldehyde (Scheme 3) as fol-

lows: reaction with sulfuryl chloride gave the 3-chloro

derivative,³¹which was reacted³²with Meldrum’s acid³³

(HCO₂H, Et₃N) to give 3-(3-chloro-4-methoxyphenyl)-

a

(a) Propargyl bromide; (b) HCO₂Et, then LAH; (c) NaCNBH₃,

(CH₂O)_n.

propionic acid 34, which after being converted to the

acid chloride was cyclized to the indanone 36. The latter

was converted to the amine 37³⁴via the oxime and

finally demethylated. Hydroxy aminotetralins were

prepared according to previously reported procedures.³⁵

Compounds 39u -w (potential metabolites of the target

carbamates) were prepared (Scheme 4) by reacting

hydroxy aminoindans 4(R5)H) with propargyl bromide

in N,N-dimethylacetamide (DMA) or acetonitrile with

potassium carbonate as base. The N-Me derivatives 40

were prepared either by N-methylation of 39 (NaC-

NBH₃/(CH₂O)_n) or by propargylation of N-Me hydroxy

aminoindans 41. NMR data of these compounds are

shown in Table 1.

Intermediates and target compounds are depicted in

Table 2. Full spectral (¹H NMR, IR, MS) and elemental

data are included in the Supporting Information. ¹H

NMR data for a few representative compounds are

shown in Table 1. The N-Boc-protected intermediates

5 (obtained in nearly quantitative yields) and their

carbamates 6 were shown to be pure by TLC and were

not further characterized.

Novel Dual Inhibitors of AChE and MAO

J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5263

Ta ble 1. ¹H NMR of Hydroxy N-Propargylaminoindans 39 and Selected Compounds of Series I and II

compd

δ in ppm, D₂O

39u

7.28 (d), 7.01 (d), 6.94 (dd), 4.88 (dd, 1H), 3.96 (m, 2H), 3.03 (m, 1H), 3.02 (s, 1H), 2.90 (m, 1H), 2.80 (s, 3H), 2.56 (m, 1H),

2.27 (m, 1H)

39w

39v

7.30 (t), 7.16 (d), 6.98 (d), 4.99 (dd, 1H), 4.02 (m, 2H), 3.06 (m, 1H), 3.06 (s, 1H), 2.95 (m, 1H), 2.60 (m, 1H), 2.32 (m, 1H)

7.35 (t), 6.98 (d), 6.83 (dd), 5.07 (dd, 1H), 4.01 (m, 2H), 3.16 (m, 1H), 3.01 (m, 1H), 3.0 (s, 1H), 2.57 (m, 1H), 2.30 (m, 1H)

7.47 (d, 1H, J ) 8.3 Hz), 7.31 (d, 1H, J ) 2.2 Hz), 7.20 (dd, 1H, J ) 8.3, 2.2 Hz), 5.0 (dd, 1H, J ) 7.7, 3.2 Hz), 4.34 (s, 1H,

CH_tartaric), 4.0 (m, 2H, CH₂CtCH), 3.56 (q, 1H, J ) 7.1, NCH₂CH₃), 3.40 (q, 1H, J ) 7.1, NCH₂CH₃), 3.18 (m, 1H,

CH₂), 3.15 (s, 1.5H, NCH₃), 3.02 (s, 1.5H, NCH₃), 3.02 (m, 1H, CH₂), 3.07 (t, 1H, J ) 2.5, CtCH), 2.64 (m, 1H, CH₂),

2.36 (m, 1H, CH₂), 1.28 (t, 1.5H, J ) 7.1 Hz, CH₃), 1.21 (t, 1.5H, J ) 7.1 Hz, CH₃)

7.56 (t, 1H, J ) 8.1 Hz), 7.39 (d, 1H, J ) 8.1 Hz), 7.12 (d, 1H, J ) 8.1 Hz), 5.30 (br d, 1H, J ) 8.2 Hz), 4.12 (m, 2H,

CH₂CtCH), 3.28 (m, 1H, CH₂), 3.20 (s, 1.5H, CONCH₃), 3.02 (s, 1.5H, CONCH₃), 3.08 (m, 1H, CH₂), 3.23 (t, 1H J ) 2.5,

CtCH), 2.78 (s, 3H, NCH₃), 2.55 (m, 2H, CH₂)

18b^a

22a

23a

7.51 (d, 1H, J ) 7.7 Hz), 7.47 (t, 1H, J ) 7.7 Hz), 7.23 (d, 1H, J ) 7.7 Hz), 5.07 (dd, 1H, J ) 7.7, 3.2 Hz), 4.05 (m, 2H,

CH₂CtCH), 3.29 (s, 1.5H, NCH₃), 3.08 (s, 1H, CH₂), 3.07 (t, 1H, J ) 2.6 Hz), 3.03 (s, 1.5H, NCH₃), 2.95 (m, 1H, CH₂),

3.23 (t, 1H J ) 2.5, CtCH), 2.65 (m, 1H, CH₂), 2.35 (m, 2H, CH₂)

7.50 (dd, 1H, J ) 8.7, 7.7 Hz), 7.29 (d, 1H, J ) 7.7 Hz), 7.14 (m, 2H), 4.15 (d, 2H, HCtCH₂), 4.02 (m, 1H, ArCH₂C(Me)H),

3.25 (dd, 1H, J ) 13.5, 5.5, ArCH₂), 3.18 (m, 1H, CtCH), 3.18 (s, 3H, CONCH₃), 3.02 (s, 3H, CONCH₃), 2.99 (s, 3H,

NCH₃), 2.97 (m, 1H, ArCH₂)

53a

(R)-52i

7.50 (t, 1H, J ) 7.7 Hz), 7.28 (d, 1H, J ) 7.7 Hz), 7.12 (m, 2H), 4.02 (m, 2H, HCtCCH₂), 3.82 (m, 1H, ArCH₂C(Me)H),

3.55 (t, 1H, J ) 7.7 Hz, CH₃CH₂CH₂CH₂N), 3.39 (t, 1H, J ) 7.7 Hz, CH₃CH₂CH₂CH₂N), 3.18 (m, 1H, ArCH₂), 3.03 (t, 1H,

J ) 2.5 Hz, CtCH), 2.98 (m, 1H, ArCH₂), 1.67 (m, 2H, CH₃CH₂CH₂CH₂N), 1.42 (m, 2H, CH₃CH₂CH₂CH₂N), 0.99 (t, 1.5H,

J ) 7.0 Hz), 0.97 (t, 1.5H, J ) 7.0 Hz)

a

Isolated as hemitartrate.

The dialkyl (but not mono) carbamates exhibit a

typical pattern, resulting from the presence in solution

of two equilibrating rotameric species. The rotational

energy barrier of the C-N bond in some representative

compounds (Table 3) lie in the range of 14.6-16.6 kcal/

mol³⁶and are in good agreement with literature data

for a series of dialkyl 3-(dimethylaminoethyl)phenyl

carbamates.³⁷The energy barrier (∆G^‡) for each com-

pound was calculated from the following approxima-

tion: ∆G^‡) 19.14 × T_coal(9.97 + log T_coal/δν) × 2.39 ×

10^-4kcal/mol; where: T_coalis the coalescence temper-

ature in K, and δν is the chemical shift difference, in

hertz, between the signals of the same alkyl substituent

at the syn and anti positions.

initial resolution of 3-(2-aminopropyl)phenol, was found

to be superior to the reported asymmetric synthesis of

58.^43,44

Compounds 55n -q (potential metabolites of the

target carbamates) were prepared by reacting amino-

ethyl phenols 42 with propargyl bromide in DMA with

potassium carbonate as base.

Intermediates and target compounds are depicted in

Table 4. Full spectral (¹H NMR, IR, MS) and elemental

data are included in the Supporting Information. ¹H

NMR data for a few representative compounds are

depicted in Table 1. The N-Boc-protected intermediates

43 (obtained in nearly quantitative yields) and the

corresponding carbamates 44 were shown to be pure by

TLC and were not further characterized.

Ser ies II. Racemic compounds were synthesized via

a four-step procedure from the corresponding 3-(2-

aminoethyl)phenols 42 (Scheme 5), and optical isomers

were prepared either from 57 or 58 (Scheme 6). Thus,

in analogy with Series I, hydroxy amines 42 were N-Boc-

protected (Boc₂O, Et₃N, THF) and carbamoylated by

carbamoyl chlorides or alkyl isocyanates. The carbam-

ates were selectively Boc-deprotected (HCl, dioxane) and

propargylated (propargyl bromide, K₂CO₃, ACN). The

target compounds were finally isolated as salts. The

order of propargylation and carbamoylation was re-

versed for 53 (compound 42, R1 ) Me, R6 ) Me, was

first propargylated to 55q which was then carbamy-

lated), thus avoiding the need for Boc protection and

deprotection (Scheme 5). Also, the 1-methyl-2-propynyl

analogues 54 were prepared by first propargylating

2-aminoethyl-3-phenol 42 with 3-mesyloxy-1- butyne,²¹

followed by Boc protection and carbamylation. Racemic

starting hydroxy amines were prepared by known

procedures.^22,24,38-40

In analogy to Series I, the dialkyl carbamates exhibit

a typical pattern, resulting from the presence in solution

of two equilibrating rotameric species with rotational

energy barriers in the range of 15.5-16.2 kcal/mol³⁶

(50a -c).

Resu lts a n d Discu ssion

Compounds were tested for their in vitro inhibitory

activity on purified AChE (from human erythrocytes)

and MAO-A and -B (from rat brain homogenates).

Selected compounds were also tested for in vitro BuChE

(from horse serum) inhibitory activity. Data are sum-

marized for series I and II in Tables 2 and 4, respec-

tively.

Ser ies I. ACh E. Several structural elements were

found to influence AChE inhibitory (AChEI) activity.

One of the most important is the nature of the carbam-

oyl nitrogen substituents. Analysis of the results of

subgroups of Me,R disubstituted carbamates in the

6-substituted series reveals that the aryl,Me carbamates

18f and 18m are the most, and the Et,Me 18b the least,

potent (Ar > Me > Bu = Pr > Et). A similar order of

potency of Me > Pr > Et was also found for the 4- and

7-substituted carbamates. An analogous relationship

between chemical structure and AChEI activity in a

series of mono- and dialkyl 3-(1-dimethylamino ethyl)-

phenyl carbamates has previously been reported by

Optical isomers of 58 were prepared from enantio-

merically pure 57 which was obtained from its racemic

mixture (56) by resolution with tartaric acid (Scheme

6). (S)-57 was isolated with L-tartaric acid,^41,42and the

(R) isomer was prepared either by using D-tartaric acid,

or by isolation from the mother liquors of the (S) isomer.

The N-Me group was then introduced by reductive

alkylation (HCO₂Et,²⁶, LAH ²⁷). This approach, with

5264 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24

Sterling et al.

Ta ble 2. AChE, BuChE, and MAO Inhibitory Activities (IC₅₀, µM)^aof Non-Propargyl and Propargyl Series I Compounds

mp

O- N-

pos pos

AChE/

BuChE

compd

1a

1b

7a

7b

(solvent)^b

R1 R2^cR3

R4

R5

X

AChE

BuChE

MAO-A

MAO-B

-

Me Me

Me Et

0.03

0.92

0.83

(0.81-0.86)

0.63

1.1

(0.74-1.2)

156-8 (A)

150-2 (A)

H

Me Me

Me Et

H

O

u

v

0.76

1.2

80

(30-230)

12.9

>1000

(0.45-1.8)

19.0

(0.34-1.03)

H

1.9

10.0

(8.99-49.4)

(0.76-4.2)

1.96

(1.63-2.45)

(9.4-17.7)

(R)-7b 197-8 (A)

H

6.14

5.6

(3.18-17.9)

7.3

(4.3-7.3)

32

7c

165-7 (A)

111-12 (B)

207-8 (A)

225-7 (D)

191-2 (C)

171-3 (C)

178-80 (A)

172-4 (A)

190-2 (A)

156-60 (A)

185-7 (A)

H

Me n-Pr

(5.5-10.2)

7d

7e

H

Me n-hexyl

Me cyclohexyl

Me p-MeOPh

0.53

(0.49-0.58)

3.96

H

(2.85-6.01)

7f

H

0.30

9.91

(7.65-13.2)

0.03

(0.28-0.32)

17.7

7g

7h

8a

H

Et

(7.33-61.5)

H

n-Pr

1.48

(1.27-1.80)

1.07

Me

Et

Me Me

Me Et

Me Me

0.95

(0.85-1.96)

1

50

(32-82)

22

(9-55)

500

>1000

(0.99-1.20)

8b

9a

38.6

(28.1-58.3)

24.5

(14.8-44.9)

(290-920)

10a

11a

11b

n-Pr Me Et

3.6

(2.5-5.8)

0.46

H

Me Me

Me Et

(0.44-0.49)

10.5

>1000

(6.02-21.4)

11c

12a

153-5 (A)

169-71 (B)

H

Me

Me n-Pr

Me Me

H

O

v

1000

0.51

(0.50-0.53)

0.009

13a

13b

16a

16b

17a

17b

18a

198-200 (A)

183-5 (A)

196-8 (A)

166-8 (A)

d

H

Me Me

Me Et

Me Me

Me Et

Me Me

Me Et

Me Me

Me Et

Me n-Pr

H

O

w

0.014

0.64

0.48

9.8

>1000

(0.008-0.012) (0.009-0.024)

(7.6-12.5)

H

0.026 0.054

7.9

(0.025-0.027) (0.038-0.064)

1.48

(1.27-1.86)

6.22

(3.87-11.9)

3.24

(5.6-11.1)

H

u

s

t

H

5.41

(3.19-4.99)

1.15

1.40

1000

750

H

(2.35-4.98)

d

H

t

79.6

(33.4-148)

2.9

180-2 (F)

Pg

2.07

(1.78-2.48)

530

(1.64-7.91)

(560-1000) (200-1400)

(R)-18a 139-41 (A)

(S)-18a 138-40 (A)

1.8

(1.57-2.08)

2.25

(1.67-3.57)

18b

194-6 (F)

47.0

1.5

(0.8-3.2)

1.98

31.3

16.1

15.9

1

250

(190-310)

300

>1000

410

(11.9-128)

31.8

(R)-18b 159-60 (A)

(S)-18b 160-2 (A)

(11.5-133)

(1.54-2.83)

(220-410)

34.9

2.2

550

(13.6-128)

14.6

(1.6-3.6)

14.8

(460-1400)

240

(170-340) (360-470)

18c

183-5 (F)

(6.4-45.9)

(6.3-52.4)

(R)-18c 126-8 (A)

10.8

14.9

0.72

(5.24-30.8)

(6.8-31.2)

Novel Dual Inhibitors of AChE and MAO

J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5265

Ta ble 2 (Continued)

mp

O- N-

R5 X pos pos

AChE/

BuChE

compd

(solvent)^bR1

R2^c

Pg

R3

R4

AChE

BuChE

MAO-A

MAO-B

(S)-18c 135-7 (A)

H

Me n-Pr

H

O

u

8.5

12.4

(4.8-73.9)

7.50

0.7

(2.0-15.2)

18d

18e

18f

18g

18h

106-8 (F)

174-5 (A)

172-4 (A)

175-7 (F)

165-7 (F)

Pg

Me n-hexyl

15.7

2.1

>1000

(7.81-39.2)

41.2

(5.02-12.2)

Me cyclohexyl H

5.56

7.4

170

(94-340)

700

10

(9-20)

500

(13.6-198)

(2.77-15.4)

6.4

(4.9-11.2)

Me p-MeOPh

H

0.86

0.13

(0.76-1.05)

13.7

(540-850) (260-1100)

H

Et

(4.50-28.6)

n-Pr

2.38

69

25

(1.85-3.26)

(47-101)

>1000

(14-43)

>1000

(S)-18h 124-6 (E)

18i 168-70 (A)

H

Pg

Me n-Pr

Me n-Bu

H

O

u

11.3

(5.96-26.7)

11.6

53.7

(29.6-110)

0.2

(R)-18i 86-8 (A)

(S)-18i 88-9 (A)

H

Pg

Me n-Bu

Et n-Bu

H

O

u

220

150

(7.5-19.5)

(150-320)

(30-230)

9.3

(4.6-25.8)

72.4

18j

148-50 (A)

178-80 (A)

188-90 (A)

182-4 (A)

3.03

(1.19-6.10)

23.9

(31.2-202)

18k

18l

Et cyclohexyl H

17.9

(6.9-40.6)

1.78

Me Bn

Me Ph

Me Me

H

1.26

1.4

40

(21-77)

80

120

(60-240)

100

(1.28-3.30)

(0.99-1.91)

18m

19a

19b

0.55

16.6

0.03

(0.54-0.56)

12.9

(8.68-38.6)

(55-108)

(70-140)

199-201 (F) Me Pg

36

47

(5.5-45.9)

(29-44)

(6-350)

196-8 (A)

(R)-19b 78-80 (A)

Me Pg

Me Et

H

O

u

>1300

1200

0.55

(0.21-1.26)

2182

4

12

(2-8)

5.6

(5-28)

9.2

19c

20a

21a

21b

21c

22a

22b

22c

23a

23b

24b

24c

25a

119-21 (A) Me Pg

Me n-Pr

Me Me

Me Et

H

O

u

v

w

u

.1000

(3.7-8.5)

>1000

(4.4-19)

>1000

212-3 (F)

219-20 (F)

208-9 (F)

185-6 (F)

Et Pg

17.9

(8.6-46.9)

2.5

H

Pg

2.8

(2.1-4.1)

0.31

0.9

>1000

(1.7-6.4)

525

1693

(224-1012)

45.8

(0.25-0.40)

Me n-Pr

Me Me

Me Et

(33.2-96.7)

169-71 (F) Me Pg

148-50 (H) Me Pg

7.0

330

(260-420)

65

730

(590-900)

100

(3.6-18.5)

439

(252-806)

.1000

20

22.0

113

(10.7-43.7)

(49-86)

(40-280)

65-7 (A)

Me Pg

Me n-Pr

Me Me

Me Et

71

63

(57-89)

0.26

(40-100)

86

196-8 (A)

183-5 (A)

161-3 (A)

164-6 (A)

152-4 (A)

H

Pg

0.053

(0.045-0.057)

2.15

(0.22-0.29)

(80-92)

150

0.019

0.83

(1.89-2.50) (0.016-0.021)

(0.72-0.97)

(90-260)

Me Et

5-Cl O

25.5

(11.3-56.8)

43.9

Me n-Pr

Me Me

Me Et

375

(315-465)

130

32

(42.5-45.9)

(18-59)

H

S

>500

98

(75-240)

(77-123)

25b

26a

193-5 (A)

195-7 (F)

H

S

O

u

>500

1-MePg Me Me

1.80

(1.52-2.21)

3.94

2.34

(1.44-5.39)

0.77

>1000

27a

27b

28a

28b

29b

31a

32a

207-9 (A)

201-3 (A)

206-8 (G)

208-9 (F)

72-4 (A)

H

Pg

Me Me

Me Et

Me Me

Me Et

Me Me

H

O

u

s

t

560

(460-680)

85

(3.18-5.07)

u

52.4

120

(70-2100

1000

(29.3-103)

4.1

(57-125)

u

0.77

(0.71-0.86)

0.29

5.5

2

(2.2-10.1)

(1.8-2.4)

2

(1.8-2.75)

0.027

u

t

204

703

18.9

120

(75-200)

4

(26.2-5898)

14.9

(0.28-0.30)

Me Pg

Me Me

w

u

0.79

(12.0-19.1)

(0.70-0.92)

(0.022-0.035) (3.0-5.3)

164-6 (A)

134-5 (A)

2.15

1000

>1000

(1.66-3.14)

0.013

(0.012-0.014)

w

10

>1000

(5.6-19.6)

5266 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24

Ta ble 2 (Continued)

Sterling et al.

mp

O-

pos

N-

pos

AChE/

BuChE

compd

(solvent)^b

R1

H

R2^cR3

R4

R5

H

X

AChE

BuChE

MAO-A

MAO-B

39u

172-4 (F)

174-6 (A)

175-7 (D)

166-8 (A)

196-8 (A)

Pg

-

u

v

w

u

v

w

0.67

(0.47-0.93)

0.3

0.6

(0.4-0.8)

0.23

(R)-39u

(S)-39u

39v

H

-

H

-

(0.28-0.34)

)0.1-0.5)

440

300

(370-520)

1.3

(120-570)

2.1

(1.0-1.8)

(1.6-2.9)

39w

0.9

(0.5-1.6)

0.015

(0.7-1.3)

0.03

40u

210-11 (F) Me

(0.014-0.016)

(0.02-0.06)

(R)-40u

(S)-40u

40v

71-2 (A)

82-4 (A)

83-5 (A)

160-2 (A)

Me

0.007

0.02

(0.005-0.010) (0.019-0.021)

12

(6-23)

0.07

23

(22-24)

0.05

(0.05-0.010)

(0.04-0.07)

0.07

40w

0.008

(0.005-0.012)

(0.06-0.09)

a

b

Numbers in parentheses represent 95% confidence interval. Blank box where IC₅₀was not determined. A, Et₂O; B, dioxane; C,

MeOH/EtOAc; D, ⁱPrOH; E, dioxane/Et₂O; F, ⁱPrOH/Et₂O; G, MeOH/Et₂O; H, MeOH; I, EtOAc, Et₂O; J , EtOAc. ^cPg ) propargyl, 1-MePg

) 1-methylprop-2-ynyl. Not determined.

d

Ta ble 3. C-N Rotational Energy Barrier Data (¹H NMR) of

the corresponding methyl disubstituted analogues. Here,

too, the propyl is more potent than the ethyl carbamate,

Selected Series I Propargyl Carbamates^a

δν (Hz), T_coal.

∆G^‡,

kcal/mol

as previously reported for other carbamates by Wein-

stock et al.⁴⁵

compd coal. peak (*)

RT

(°C)

solvent

18a

18b

CH₃(*)

CH₂(*)CH₃

CH₃(*)

71.4

72.0

67.9

75.5

76.0

68.4

79.6

79.4

63.2

71.7

55.9

67-68 buffer

16.56

16.66

16.04

15.97

16.16

16.49

16.39

15.78

14.69

15.50

The steric influence of the size of the alkyl substituent

on inhibitory potency against BuChE is quite different.

In this enzyme, the two phenylalanines at the active

site of AChE have been replaced by leucine and valine.

These amino acid substitutions enlarge the size of the

active site on BuChE relative to that of AChE, resulting

in a different order of potency among the alkyl substit-

uents: Me ) Et > Bu > Pr ) Ar for the inhibition of

this enzyme by Series I compounds. The different

inhibitory potencies of these compounds against AChE

and BuChE may also result from variations in other

amino acids, such as tryptophan, at the entrance to the

gorge of AChE, which is replaced by alanine in BuChE.⁴⁶

Thus, the Et,Me carbamates were found to show the

highest selectivity for BuChE inhibition and the Me,Ph

were the most selective for AChE. Since AChE is lost

in the brains of subjects with AD as the cholinergic

neurons degenerate,⁴⁸but BuChE, which is found in

glial cells remains unchanged, or is even increased,⁴⁹

drugs that also block BuChE could increase acetylcho-

line levels more effectively than AChE-selective inhibi-

tors and help to maintain ACh levels for interaction with

its receptors.

69

56

60

65

50

30

43

D₂O

buffer

D₂O

buffer

D₂O

buffer

D₂O

18c

CH₃(*)

CH₂(*)CH₂CH₃

CH₃(*)

CH₂(*)CH₃

18e

18m

18k

D₂O/buffer

D₂O

a

Measurements were performed on Varian 500 Inova. Coales-

cence temperature is (1°C.

Weinstock et al.^37,45The authors showed a close cor-

relation between the amount of energy needed to

overcome the restriction of rotation about the amide

bond (∆G^q) and the affinity of the inhibitor for the

enzyme. However, additional examples in Series I, such

as the Et,cyclohexyl analogue 18k , which has a ∆G^q

value lower than that of 18b (Table 3), but about three

times the potency of the latter (IC₅₀17.9 vs 47 µM),

suggests that there may be other explanations for the

reduction in potency induced by the ethyl substituent.

Furthermore, the most potent compound of the prop-

argylated 6-carbamates of Series I, the Me,Ph derivative

18m , exhibits the lowest ∆G^qvalue of all the compounds

for which T_cwas determined. The high potency of the

aryl carbamates 18f and 18m may be explained by the

π-π interaction with phenylalanines 288 and 290

located at the bottom of the gorge, adjacent to the active

site.⁴⁶The anomalous “ethyl” effect was also observed

by Lieske and co-workers⁴⁷who found the diethyl

carbamoyl derivative to be the least active in a series

of 5-(1,3,3-trimethylindolinyl) carbamates, ∼7400 less

potent (k_i) than the dimethylcarbamoyl analogue. The

two monosubstituted carbamates tested for AChE in-

hibitory activity (18g and 18h ) are more potent than

Substituting sulfur for oxygen (thiocarbamates) re-

sults in a decrease of AChEI activity (25a vs 18a ). We

also studied the effect of N-indanylamine substitution

on AChEI activity. The N-propargyl-6-carbamoyl ami-

noindans (18a ,b,c) are 2-5 times less potent in AChEI

than the parent primary amines (7a ,b,c). For the 4- and

7-substituted carbamoyl aminoindans, N-propargylation

results in 5-7-fold loss of potency for the Me,Me

carbamates (21a and 23a ) and 50-83-fold loss of

potency for the Et,Me carbamates (21b and 23b) in

comparison to the parent primary amines (11a , 13a and

11b,13b, respectively). The effect of other N-alkylations

on AChEI activity is less clear-cut. In the 6- and

7-carbamoyl series, N-methyl substitution at the prop-

Novel Dual Inhibitors of AChE and MAO

J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5267

Ta ble 4. AChE, BuChE, and MAO Inhibitory Activities (IC₅₀, µM)^aof Non-Propargyl and Propargyl Series II Compounds

compd

mp (solvent)^bR1

R2^c

R3

R4

R6

H

AChE

BuChE

AChE/BuChE

MAO-A

MAO-B

>1000

45a

125-7 (A)

103-5 (A)

83-5 (A)

H

Me Me

0.23

(0.18-0.44)

35.5

95

(60-150)

180

45b

Me Et

H

>1000

(22.2-56.8)

(100-310)

45c

Me n-Pr

Me Me

Me Et

H

7.8

240

(4.7-15.6)

0.28

(110-520)

22

(9-52)

20

(11-38)

51

(43-61)

2.7

46a

126-8 (A)

d

Me

H

(0.12-0.41)

46b

H

20.7

(12.7-36.8)

0.16

48a

127-9 (F)

138-9 (F)

113-15 (F)

108-10 (F)

150-2 (F)

152-4 (F)

166-70 (A)

119-121 (A)

170-2 (A)

100-2 (A)

87-9 (A)

Me Me

Me Et

H

(0.12-0.32)

50a

H

Pg

H

0.22

0.21

1.05

55.9

8.1

24

(11-52)

7.8

(0.18-0.32) (0.16-0.41)

(1.3-5.6)

2.6

50b

H

30.2 0.54

(9.46-68.1) (0.22-1.54)

(1.7-3.9)

(5.8-10.5)

50c

Me n-Pr

Me Me

Me Et

H

15.4

(6.0-41.2)

0.85

1.9

(1.6-2.4)

1.8

0.29

(1.5-2.2)

0.3

(0.19-0.44)

2.8

51a

Me Pg

H

(0.79-0.93)

(0.1-0.9)

(1.5-5.2)

51b

H

16.6

0.8

(7.3-50.4)

0.54

(0.32-0.98) (1.00-1.20)

33.9 0.93

(12.6-74.0) (0.90-0.95)

19.1 2.07

(0.5-1.4)

6.9

(0.4-1.3)

5.7

(4.8-6.9)

12.8

(9.5-17.2)

0.17

52a

H

Pg

Me Me

Me Et

Me

1.09

0.50

36.5

9.2

(6.3-7.7)

52b

2.1

(1.8-2.5)

14.6

(12.1-17.6) (0.08-0.38)

18

(14-24)

15

(13-17)

0.12

(0.09-0.16)

0.12

(0.09-0.16) (0.07-0.59)

52c

Me n-Pr

(13.6-27.3) (1.19-8.56)

52e

Me cyclohexyl Me

3.6

(2.2-6.9)

12.2

0.23

(0.18-0.30)

0.4

52i

Me n-Bu

Me Me

Me

2.75

4.4

2.2

(5.4-38.2) (1.89-4.67)

1.64 0.75

(1.32-2.23) (0.73-0.77)

234 1.5

(91.8-732) (1.38-1.65)

33.1

(9.8-157)

3.06

(0.2-0.8)

53a

158-160 (F) Me Pg

0.8

(0.3-1.2)

0.1

53b

144-6 (F)

d

Me Pg

Me Et

153

53c

Me n-Pr

Me n-hexyl

22.8

(13.7-27.8)

0.72

1.45

4.25

53d

95-7 (A)

92-4 (I)

95-6 (J )

105-7 (A)

118-9 (I)

119-20 (I)

151-3 (F)

135-7 (F)

140-2 (F)

111-2 (F)

20

(8-43)

1.5

(0.8-2.9)

(2.36-4.22) (0.64-0.89)

(R)-53d

(S)-53d

53e

2.0

(1.7-2.5)

3.4

(2.2-6.4)

8.67

Me cyclohexyl Me

H

0.67

12.9

1.7

(0.8-3.7)

0.9

0.24

(0.11-0.57)

0.22

(3.69-33.1) (0.45-1.08)

(R)-53e

(S)-53e

54a

7.9

(5.6-11.9)

(0.8-1.0)

(0.19-0.26)

6.3

1.1

0.28

(2.8-27.5)

(0.8-1.5)

440

(230-820)

(0.23-0.34)

> 1000

H

1-MePg Me Me

0.17

0.24

0.7

(0.16-0.18) (0.23-0.26)

13.9 0.24

(7.99-27.5) (0.20-0.36)

54b

1-MePg Me Et

H

57.9

55n

Pg

-

67

107

(22-530)

6.4

(14-340)

55o

Me Pg

1.3

(0.5-3.7)

(3.6-11.3)

55p

55q

111-3 (A)

88-90 (A)

H

Pg

-

Me

830

230

(600-1100) (150-360)

8.9

(4.7-16.8)

Me Pg

4.8

(2.8-8.3)

a

b

Numbers in parentheses represent 95% confidence interval. Blank box where IC₅₀was not determined. A, Et₂O; B, dioxane; C,

MeOH/EtOAc; D, ⁱPrOH; E, dioxane/Et₂O; F, ⁱPrOH/Et₂O; G, MeOH/Et₂O; H, MeOH; I, EtOAc, Et₂O; J , EtOAc. ^cPg ) propargyl, 1-MePg

) 1-methylprop-2-ynyl. Not determined.

d

5268 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24

Sterling et al.

Sch em e 5^a

methylenes of 31a that prevents it from aligning itself

to the active site.⁵¹Compound 13a (the primary amine

analogue of 32a ) is the most active non-propargyl

carbamate of both series, with an IC₅₀of 9 nM.

Such an analysis may also explain the observed

influence of the position of the ring substitution of the

carbamate moiety on AChE inhibitory activity of the

propargylamines, with potency decreasing as a function

of its position from 4 > 6 > 7. In fact, the 4-carbamate

23a is the most potent AChE inhibitor of all the

propargyl carbamates of both series, with an IC₅₀of 53

nM. A similar SAR was reported by Bolognesi et al. for

a series of carbamate regioisomers.⁵⁰The influence of

the position of the carbamate on AChEI activity in

Series I seems to be due to the overall shape of the

molecule and the effect this has on its fit in the active

pocket (Figure 1). The lower potencies of the N-methyl

propargylamines may also relate to this steric interfer-

ence in the active site and reflect subtle differences in

the binding orientation of the Me,Me carbamates in

comparison to that of the Et,Me analogues.

a

(a) (Boc)₂O, Et₃N, THF, rt; (b) R₃R₄NCOCl, CH₃CN,NaH, rt;

(c) HCl, dioxane, rt; (d) propargyl bromide, K₂CO₃, CH₃CN or

DMA, rt; (e) HCl/Et₂O, rt.

Sch em e 6^a

MAO. We also addressed the SAR implications of our

results for MAO inhibition and A/B selectivity. Intro-

duction of a hydroxy group in any position on the parent

propargylaminoindan nucleus results in a decrease of

1-2 orders of magnitude in MAO-B inhibitory activity,

thereby causing a loss of A/B selectivity. The addition

of the carbamate moiety (which is essential for AChE

inhibition) in the case of the 6- and 7-hydroxy propargyl-

aminoindans 39u and 39v, respectively, results in a

further 100-1000-fold decrease of MAO-A and -B

inhibitory potency in 18a ,b,c and 21a ,b. By contrast,

carbamylation of 4-hydroxy-N-propargyl-1-aminoindan

39w results in only a 70-fold decrease in MAO-B and a

4-fold increase in MAO A inhibitory potency, making

23a and 23b MAO-A selective inhibitors (virtually the

only substituted propargylaminoindans (PAI’s) with

appreciable A/B selectivity, Tables 2 and 5).

The nature of the N-carbamoyl substituents had some

effect on MAOI potency and selectivity. In the 6-sub-

stituted series (compounds 18), which was more exten-

sively studied, the more bulky substituents tend to

increase inhibitory potency for both MAO-A and -B (cf

for example 18e,l,m ).

As shown for AChE, the position of the carbamate has

a profound effect on MAO inhibitory activity. The order

of potency is the same as for AChEI, and decreases as

a function of position from 4 > 6 > 7. Thus, in this

series, only the 4-substituted carbamates have signifi-

cant (<1 µM) MAOI potency in vitro. This structure-

activity relationship may be useful in gaining some

insight into the putative active site of MAO (vide infra).

a

(a) (1) L-Tartaric acid, MeOH, reflux. (2) 25 % NH₄OH, rt; (b)

HCO₂Et, reflux; (c) LAH, THF, 5 °C, then rt; (d) HCl, Et₂O, rt.

argylamine nitrogen virtually eliminates the AChEI

activity (IC₅₀> 1000 µM) of the Et,Me and Me,Pr

analogues (19b , 19c, 22c). The effect is considerably

smaller in the 4-series, where 29b is only ∼7-fold less

potent than 23b. Similarly, among the Me,Me carbam-

ates, the N-methyl propargylamines (19a , 22a ) have

activities slightly lower (3-4-fold) than those of the

corresponding secondary propargylamines (18a , 21a ).

By contrast, N-methylation increases both BuChEI

activity and selectivity of the 6-carbamates, while

having the opposite effect on the 4- and 7-analogues.

The chirality of the indanylamine does not affect

AChEI, in analogy with the lack of enantioselectivity

reported for a series of hexahydrochromeno[4,3-b]pyr-

role carbamates.⁵⁰

The Me,Me carbamates of the 1-aminotetralin (27a )

and 2-aminotetralin (28a ) analogues, as well as the

Et,Me derivative of 1-aminotetralin (27b), are compa-

rable to the corresponding 1-aminoindan derivatives

(18a ,b) in AChEI activity. However, the 2-aminotetralin

Et,Me carbamate 28b is about 4 times less active than

18b.

Aminoindans 31a and 32a may be considered as

conformationally constrained analogues of 1a , the di-

methyl analogue of rivastigmine. The comparable AChEI

activities of 32a and 1a suggest that the bioactive

conformation of (the flexible) 1a is similar to that of 32a .

While in both aminoindans the carbamate moiety is

oriented “meta” to the “aminobenzyl” group, the two

structures differ significantly. Thus, assuming that the

amine orients the carbamate in the active site, there

seems to be some steric interference with the ring

We also modified the orientation of the two pharma-

cophores by preparing aminotetralins. The dimethyl-

carbamoyl derivative of 1-propargylaminotetralin (27a )

shares low MAO-A and -B inhibitory potency with the

analogous indan derivative 18a , while the Et,Me ana-

logue 27b is comparatively more active than 18b.

However, the 2-aminotetralin derivatives 28a and 28b

have remarkably high MAO-A inhibitory activity as

compared to 18a and 18b. In fact, they approach the

4-carbamoyl propargylaminoindans (compounds 23) in

MAOI potency. Compounds 28 and 23 may be consid-

ered rigid conformers of the phenethylamines 52 (Table

Novel Dual Inhibitors of AChE and MAO

J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5269

F igu r e 2. Superposition of 1- and 2-aminotetralin derivatives

on 52b. Red, 52b; blue, 28b; green, 27b.

N-Methyl substitution at the propargylamine of pro-

pargylamino hydroxy and carbamate derivatives results

in an increase in MAOI (both A and B) activity of all

positional isomers (4-, 6-, and 7-), so that they are also

not A/B selective (Table 5). In contrast, N-methyla-

tion of R-PAI itself (rasagiline)⁵³resulted in a slight

decrease in MAO-B inhibitory activity. Thus, MPAI is

a potent MAOI with some A selectivity in comparison

to rasagiline, which is a potent B-selective MAOI. The

potency of S-PAI as an inhibitor of both A and B was

significantly increased by N-methylation. The effect of

the N-Me group on MAOI activity may be the net

outcome resulting from several parameters, such as

hydrophobicity, steric volume, hydrogen bonding, all of

which may have a significant bearing on binding to the

enzyme:⁵⁴

• The propargylamine nitrogen may act either as an

hydrogen bonding (HB) donor or acceptor, and both

these features are incorporated in the putative phar-

macophore modeled (vide infra). However, the consis-

tently higher potency of all the N-Me analogues may

imply that the HB donating effect of the nitrogen is not

crucial for binding, since in this case it can only act as

an HB acceptor.

• The steric constraint imposed by the Me group may

stabilize a specific conformation which binds better to

the enzyme.

• There may be favorable hydrophobic interactions

between the Me group and a specific lipophilic feature

in the binding pocket.

The A/B selectivities of PAI and its N-Me derivative

(both racemic and R) have been investigated by several

groups. Kalir, Sabbagh, and Youdim⁵⁵found that race-

mic PAI (AGN-1135) is more B-selective than its N-

methyl analogue (AGN-1133). Riederer et al.⁵⁶explained

the different selectivities of AGN-1135 and 1133 toward

the two MAO isoforms (1135: B-selective, 1133: non-

selective) by the steric hindrance conferred by the

methyl group in the active site on MAO-B. However,

the lack of selectivity actually results from the increased

potency of the N-Me compound toward MAO-A (the

reduced B activity is secondary). In fact, rasagiline ((R)-

AGN-1135) may be the only PAI with an N-Me deriva-

tive, which is less potent as a MAO-B inhibitor. We have

now shown with great consistency, that N-Me deriva-

tives of all ring substituted PAI’s are more potent

F igu r e 1. Overall shape of minimum energy conformations

of representative propargyl carbamates (Series I). (a) Blue,

18b; (b) red, 21b; (c) magenta, 23b.

4) and are superimposable using Catalyst molecular

modeling software (Molecular Simulations Inc., San

Diego, CA). Compounds 27 have a higher displacement

RMS value and thus are less superimposable than the

others (Figure 2). However, compounds 28 behave in

analogy with other compounds in Series I with relatively

low potency for MAO-B inhibition, thus having more

A-selectivity in comparison to the nonselective 2-prop-

argylaminotetralin (with IC₅₀’s for A and B inhibition

of of 0.05 and 0.04 µM, respectively⁵²).

5270 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24

Sterling et al.

Ta ble 5. MAO Inhibitory Activity of Rasagiline, Its N-Me Analogue, and Their Hydroxy and Carbamyloxy Congeners

IC₅₀(µM)^a

NH/NMe^b

A B

R

compd/ stereochem

rasagiline

subst posit

R1

H

A

B

A/B

93

0.41

(0.28-0.54)

0.003

0.0044

136

0.44

(0.0035-0.053)

H

R-MPAI

S-PAI

Me

H

0.01

17

0.3

1.3

1.4

22

314

45

340

S-MPAI

Me

0.07

0.05

39u

6

7

4

H

0.67

(0.47-0.93)

0.015

0.6

(0.4-0.8)

0.03

1.1

20

40u

Me

H

0.5

(0.014-0.016)

(0.02-0.06)

(R)-39u

(R)-40u

(S)-39u

(S)-40u

39v

0.3

0.23

1.3

43

37

11.5

13

(0.28-0.34)

0.007

)0.1-0.5)

0.02

Me

H

0.35

1.47

0.52

0.62

1.4

(0.005-0.010)

(0.019-0.021)

OH

440

300

(370-520)

12

(6-23)

1.3

(1.0-1.8)

0.07

(0.05-0.010)

0.9

(120-570)

23

(22-24)

2.1

(1.6-2.9)

0.05

Me

H

19

42

40v

Me

H

(0.04-0.07)

39w

0.9

1.0

113

13

(0.5-1.6)

0.008

(0.7-1.3)

0.07

40w

Me

0.11

(0.005-0.012)

(0.06-0.09)

(R)-18b

(R)-19b

18c

6

H

300

(220-410)

4

(2-8)

240

>1000

0.3

75

43

83

45

Me

H

12

(5-28)

410

0.33

0.59

0.61

(170-340)

(360-470)

c

-OCON(Me)R₄

19c

Me

5.6

9.2

(3.7-8.5)

>1000

65

(4.4-19)

>1000

100

21b

22b

7

H

Me

∼1

>15

>10

0.65

(49-86)

(40-280)

23b

29b

4

H

0.83

150

0.005

0.007

31

38

(0.72-0.97)

0.027

(90-260)

4

Me

(0.022-0.035)

(3.0-5.3)

a

b

Numbers in parentheses represent 95% confidence interval. Ratio between IC₅₀’s of unsubstituted and N-Me substituted

propargylamines. ^cb, R₄) Et; c, R₄) n-Pr.

inhibitors of both MAO-A and -B than the parent

secondary PAI’s.

N-methylation is strongly dependent on the indole

substituent, ranging from an enhancement of 3 orders

of magnitude to a decrease in potency by a factor of 10

with concomitant differences between MAO-A and -B.⁵⁹

These observations are more consistent with those of

Polymeropoulos,⁵⁷who explained the greater MAO-B

inhibitory potency of (R)-N-Me PAI relative to (R)-PAI

(he does not address the question of selectivity), by the

higher lipophilicity of the tertiary amine allowing better

binding to the lipophilic pocket of MAO-B and offering

more effective orientation to the flavin complex

However, the uncertainty of extrapolating such data

is apparent from results recently reported by Moron et

al.⁵⁸for MAO inhibition of a series of substituted

propargylaminoindoles (59). In these, the effect of

In the one case where MAOI was determined for each

member of a chiral pair (18b), there was no real

difference between the enantiomers in potency or se-

lectivity. This is further indication of the very special

properties of rasagiline, in which there is a difference

of four-orders-of-magnitude between the enantiomers in

MAO-B inhibitory potency.

Replacement of the propargyl group with 1-methyl-

2-propynyl results in a further decrease in MAO inhibi-

tory activity (26a vs 18a ), in analogy to other propar-

gylamines.²¹For instance, the pargyline derivative is

several orders of magnitude less potent than the parent

(from 9 × 10^-7to >10^-3).⁶⁰

Compound 24c, with an additional ring substituent

(5-chloro), has an interesting profile, with less AChE

Novel Dual Inhibitors of AChE and MAO

J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5271

Ta ble 6. MAO Inhibitory Activity of Selegiline, Desmethylselegiline (DMS), and Their Hydroxy and Carbamyloxy Congeners

IC₅₀(µM)^a

A/B

NH/NMe^b

R

compd

DMS

R1

H

R6

A

B

A

B

H

Me

195

(100-340)

1.2

0.32

610

171

0.63

162

46

(0.27-0.36)

selegiline

Me

0.007

(1.1-1.3)

(0.004-0.014)

55n

55o

55p

55q

H

67

(14-340)

1.3

107

(22-530)

6.4

51

17

OH

Me

H

0.20

3.6

(0.5-3.7)

(3.6-11.3)

Me

830

230

>93

>48

(600-1100)

8.9

(150-360)

4.8

Me

1.85

(4.7-16.8)

(2.8-8.3)

50a

51a

50b

51b

52a

53a

52b

53b

H

2.7

(1.3-5.6)

0.3

24

(11-52)

2.8

0.11

0.33

1

9

8.6

Me

H

(0.1-0.9)

(1.5-5.2)

H

2.6

7.8

3.3

9.8

7

(1.7-3.9)

0.8

(5.8-10.5)

0.8

c

OCON(Me)R₄

Me

H

(0.5-1.4)

(0.4-1.3)

Me

6.9

5.7

1.21

0.15

0.16

1.2

58

(6.3-7.7)

0.12

(4.8-6.9)

0.8

(0.3-1.2)

12.8

(9.5-17.2)

0.1

Me

H

(0.09-0.16)

2.1

17.5

128

(1.8-2.5)

0.12

Me

(0.09-0.16)

(0.07-0.59)

a

b

Numbers in parentheses represent 95% confidence interval. Ratio between IC₅₀’s of unsubstituted and N-Me substituted

propargylamines. ^ca , R₄) Me; b, R₄) Et.

and MAO-A inhibitory potency than 18c, but 8-fold

more MAO-B inhibitory potency. As a result, this is the

only MAO-B-selective compound in Series I with similar

IC₅₀values for MAO-B and AChE inhibition.

supporting the nonsignificance of steric requirements

in this area.

Amine N-substitution (propargyl and/or additional

methyl) has a tendency to lower AChEI potency, most

markedly in 52b and 53b, but this effect is not fully

consistent.

Ser ies II. ACh E. As in Series I, AChEI activity is

highly dependent on the nature of the carbamoyl

substituents, exhibiting the same order of reactivity,

namely Me ∼ Bu, hexyl > Pr > Et. The low potency of

the Et substituent is most pronounced in the R1)R6)Me

case (53b vs 51b, 52b, 54b). This anomalous behavior

of the Et,Me carbamates has already been discussed for

Series I.

MAO. Introduction of a hydroxy group into the parent

compounds of Series II (Table 6) has a more subtle effect

on activity than in Series I. The MAO-B inhibitory

potencies of the N-methyl derivatives (55o and 55q) are

500 times lower than that of selegiline (2), although they

are about the same for MAO-A inhibition. As a result,

there is little or no A/B selectivity in these compounds.

The nor analogues (55n and 55p ) too are considerably

less potent against MAO-B than is N-desmethylsel-

egiline (DMS).

In both Series I and II, the change in the size of the

alkyl substituents on the carbamoyl nitrogen has a

much less pronounced effect on BuChE inhibition than

on that of AChE. Thus, the Et,Me carbamates have

similar or greater potency as BuChE inhibitors than the

other carbamates, and tend to be the most selective

against BuChE (50b, 52b, 53b, and 54b). In Series II,

we have prepared derivatives with methyl substitution

at (i) phenethyl C2 (R6 ) Me), (ii) the 2-propynylamine

nitrogen (R1 ) Me), and (iii) C1 of 2-propynyl, and

combinations thereof. The AChEI potency of the 2-Me

phenethyl derivatives is little affected by the presence

of the Me group (cf 52a ,b,c vs 50a ,b,c). In contrast, the

AChE potency decreases in most of the N-Me analogues

(cf 53b vs 50b ). The 1-methyl-2-propynyl derivatives

54a ,b are equipotent to the propargyl analogues 50a ,b,

As in Series I, the N-Me propargyl compounds are

more potent than the secondary amines (51 and 53 vs

50 and 52, respectively). Likewise, selegiline has been

reported to be 60 times more potent an inhibitor of

MAO-B than its desmethyl analogue⁶¹(Table 6). How-

ever, in contrast to Series I, the presence of the

carbamate moiety confers a greater MAO inhibitory

activity than that of the parent hydroxy analogues,

sometimes very significantly (50a ,b vs 55n , or 52a ,b

vs 55p ). No clear trend for the effect of carbamate

N-alkyl substituents on MAO inhibitory activity could

be discerned.

5272 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24

Sterling et al.

27b, and (R)-18b) have a slower rate of inhibition of

AChE than the open-chain amines (rivastigmine 1b and

53d ). The presence of a propargyl group and the identity

of the carbamoyl substituents exert a smaller effect. The

slow onset of enzyme activity by the ring-closed com-

pounds is expected to enhance their therapeutic utility

by reducing the intensity of adverse effects due to the

slower build-up of excess cholinergic activity.

In Vitr o/in Vivo Cor r ela tion a n d Meta bolism .

Preliminary in vivo studies suggest that the propargyl

carbamates of both series will undergo analogous me-

tabolism including stepwise loss of either or both the

carbamoyl and propargyl moieties (X, Y, Z, Scheme 7,

as well as other metabolites). Thus, the in vivo phar-

macological profile will be determined not only by the

enzyme inhibitory potency of the parent compounds but

also by that of their metabolites. Thus, in Series I, most

of the in vivo MAOI will be due to metabolites X while

the principal AChE inhibitors (metabolites Y or the

parent) will depend on the carbamate substitution

position. For instance, the metabolites for compound

18b include compound 39u (X) which is at least 4 orders

of magnitude more potent in MAOI (A and B), and

compound 7b (Y) which is 5 times more potent in

AChEI.

By contrast, in Series II, the parent compound is more

active than, or at least equipotent to, either metabolite

X or Y for both enzymes. Metabolites Z have negligible

inhibitory activity in both series. In any case, the

ultimate MAOI/AChEI ratio in vivo will be determined

by the pharmacokinetics of the various metabolites and

their bioavailability.

F igu r e 3. Putative MAO pharmacophore. Light blue, hydro-

phobes; yellow, carbamate centroid; magenta, HB donor or

acceptor.

The absolute configuration of the 2-methylphenethyl

has no effect on MAO-inhibitory activity, as shown by

the almost equal potency of the two optical isomers of

53e. Thus, in both series the bulky ring substitution

decreases the importance of chirality and of A/B selec-

tivity in comparison to rasagiline or selegiline. In

common with all other examples (vide supra), the

1-methyl-2-propynylamine derivatives have a very low

MAOI activity,⁶⁰e.g., 54a vs 50a . It is noteworthy that

both selegiline and rasagiline are unique in their series

in MAO-B inhibitory potency and selectivity, relative

to analogues in each series, despite the structural

differences between them.

Im p lica tion s for a P u ta tive MAO Active Site. The

in vitro results described above have important implica-

tions for understanding the steric requirements of the

active sites of the various enzymes.⁵⁴

In an attempt to correlate MAOI activity to structure

in general, and the carbamate position in particular, we

modeled five representative compounds (18b, 21b, 23b,

28b, 52b, Figure 3) using Catalyst molecular modeling

software. As the absolute configuration of the propar-

gylamine-bearing carbon does not significantly affect

MAOI activity in these compounds, only one optical

isomer (S) of each compound was modeled. A putative

pharmacophore, based on the three active analogues

(28b, 23b, 52b), was identified, comprising the following

elements (Figure 3): (1) a carbamate group, featured

by its geometrical center (carbamate centroid) and by a

hydrophobic element (Et,Me substituents), (2) an aro-

matic hydrophobe, (3) an HB donor/acceptor (propar-

gylamine nitrogen), and (4) an HB donor (acetylenic

CH). If we assume that the carbamate moiety interacts

with a feature of the active site and thus determines

the orientation of the propargylamine, the three active

analogues correlate well. However, for 18b and 21b

(Figure 4) the propargylamine moiety (essential for

MAOI potency) is markedly misoriented and presum-

ably cannot interact with the FAD at the active site.⁶²

In fact, the 6-carbamoyl-N-propargylaminoindans (18)

are even less potent MAO-A inhibitors than their parent

6-carbamoylaminoindans (7).

Con clu sion s

We have described the preparation and in vitro

activity of aminoindan (Series I) and phenethylamine

(Series II) derivatives incorporating two pharmaco-

phores: a carbamate and a propargyl group.

In Series I, MAO and AChE inhibitory potency of the

carbamates decreases as a function of position 4 > 6 >

7. Series II can be viewed as being somewhat isosteric

to either 4- or 6-substituted Series I, with pharmaco-

logical activity also intermediate between the two. In

general, 6- and 7-carbamoyloxy PAI’s (Series I) are

either equipotent to, or slightly (2-5-fold) less active

(as AChE inhibitors) than, the corresponding pheneth-

ylamine derivatives (PPE’s, Series II), while the 4-an-

alogues are more potent. The same trend is observed

for MAO inhibition, but in this case the 6- and 7-car-

bamoyl PAI’s are 2-3 orders of magnitude less potent

inhibitors of this enzyme, while the 4-analogues are

equipotent to the PPE’s (MAO-A).

In Series I, the 6- and 7-hydroxy compounds are more

potent MAO inhibitors than the corresponding carbam-

ates while the 4-hydroxy (A only) and Series II hydroxy

compounds are less potent than their corresponding

carbamates. However, in all examples the hydroxy

derivatives are less potent than the unsubstituted

parent compounds (rasagiline and selegiline). In both

series, and for all regioisomers, AChEI activity depends

primarily on the carbamoyl nitrogen substituents, with

Et,Me being the least potent.

The higher potency of the N-Me derivatives for both

MAO-A and -B suggests that methyl substitution exerts

a counter-balancing effect.

Kin etics of ACh E In h ibition . The bimolecular rate

constants for carbamoylation of AChE (k_i) by four of the

compounds were determined and compared to that of

rivastigmine 1b (Table 7). The results of the kinetics

experiment suggest that the cyclic compounds (e.g., 24c,

The N-propargyl group decreases AChEI activity (as

compared to the non-propargyl analogues) in Series I,

Novel Dual Inhibitors of AChE and MAO

J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5273

F igu r e 4. Correlation between active and nonactive compounds and the putative MAO pharmacophore. (a) Actives: red, 52b;

green, 28b; blue, 23b. (b) Nonactives: yellow, 21b; cyan, 18b.

Ta ble 7. Bimolecular Rate Constants for Carbamylation by Selected Compounds

pseudo-first-order rate constant k_obs(min^-1) ( SEM

compd

2.5 µM

5 µM

10 µM

20 µM

40 µM

50 µM

k_i^a

1b

53d

19 ( 3

14 ( 1

-

15 ( 1

33 ( 3

23 ( 2

44 ( 3

30 ( 2

-

55 ( 1

-

63 ( 1

1432 ( 191

1035 ( 60

50 µM

100 µM

200 µM

300 µM

400 µM

24c

27b

(R)-18b

-

17 ( 1

21 ( 1

16 ( 1

22 ( 1

30 ( 1

21 ( 1

-

39 ( 1

32 ( 1

34 ( 1

48 ( 2

42 ( 2

38 ( 2

68 ( 1

52 ( 1

59 ( 5

101 ( 12

102 ( 5

a

Bimolecular rate constant (M^-1min^-1) ( SEM.

while having little or no effect in Series II. N-methyla-

tion at the propargylamine nitrogen has a dual effect

in both series: it increases MAOI (A and B) and

decreases AChEI activities.

of all of these individual metabolites. The results of our

ongoing in vivo screening program will be published

elsewhere.

Exp er im en ta l Section

Three candidates (24c, 27b, 53d ) were identified as

possible leads for further development, based on the

following criteria: (a) comparable AChE and MAO-B

inhibitory activities, (b) good to moderate AChE inhibi-

tory activity, and (c) lack of strong A selectivity.

However, it is likely that these compounds will be

metabolized to derivatives with inhibitory activities

against AChE and/or MAO-A or -B, which are different

from those of the parent compounds. Thus, the final

enzyme inhibition will be a result of the combined action

Ch em istr y. Gen er a l. All commercial chemicals and sol-

vents were reagent grade and were used without further

purification unless otherwise specified. Melting points were

determined on a Buchi B-540 apparatus and are uncorrected.

Elemental analyses were carried out at the Hebrew University

of J erusalem, and the results are within (0.4% of the theoreti-

cal values. Merck silica gel 60 F254 plated were used for

analytical TLC (visualized with UV light and iodine vapors);

flash column chromatography was performed on Merck silica

gel 60 (230-400 mesh). ¹H NMR spectra were recorded in

DMSO-d₆, CDCl₃, or D₂O by means of a Varian Gemini-300

5274 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24

Sterling et al.

3. Boc-Dep r otection : 6-(N-Me,N-Et ca r ba m yloxy)-1-

a m in oin d a n ‚HCl (7b‚HCl). Compound 6, R1)H (7.8 g, 23.3

mmol) was dissolved in dioxane (80 mL), and a 20% HCl/

dioxane (80 mL) was added. After 2 h stirring at room

temperature, the solvent was evaporated in vacuo and the

residue was treated with dry ether (200 mL). The mixture was

stirred at room temperature for 4 h and filtered, to give 6.15

g (22.7 mmol, 97%).

Sch em e 7. Putative Metabolic Pathways for Series I

and II

4. P r op a r gyla tion : 6-(N-Me,N-Et ca r ba m yloxy)-N-p r o-

p a r gyl-1-a m in oin d a n ‚HCl (18b‚HCl). To a stirred mixture

of 7b‚HCl (5.2 g, 19.2 mmol) and potassium carbonate (5.31

g, 38.4 mmol) in acetonitrile (250 mL) was added a solution of

propargyl bromide (2.06 g, 17.28 mmol) in acetonitrile (10 mL).

The reaction mixture was stirred at room temperature under

nitrogen for 25 h and filtered. The filtrate was evaporated to

dryness in vacuo, and the residue was purified by column

chromatography (EtOAc) to give 3.6 g (13.2 mmol, 69%) of the

free base as a yellow oil. The free base was dissolved in dry

ether (150 mL), and HCl/ether (15 mL) was added. The

mixture was stirred at room temperature for 1 h and filtered,

i

and the solid was recrystallized from PrOH/ether to give 3.5

g (11.3 mmol, 59%) of the title compound as a white solid.

6-(N,N-Dim eth ylcar bam yloxy)-N-pr opar gyl-1-am in oin -

d a n Mesyla te (18a Mesyla te). To a stirred mixture of 7a ‚

HCl (1.88 g, 7.33 mmol), K₂CO₃(2.03 g, 14.66 mmol), and

acetonitrile (70 mL) was added a solution of propargyl bromide

(0.79 g, 6.6 mmol) in CH₃CN (5 mL) dropwise over 5 min,

under nitrogen. The mixture was stirred under N₂for 24 h

and filtered, and the solvent was removed at reduced pressure.

The residue was taken up into water (150 mL) and toluene

(150 mL). This mixture was stirred while adjusting the pH of

the aqueous layer to 3.75 by the addition of 20% aq HCl. The

aqueous layer was separated and extracted with toluene (2 ×

100 mL) and brought carefully to pH 7.5 by the addition of

10% aq NaOH solution. It was then extracted with toluene (5

× 70 mL). The combined toluene layers were dried (Na₂SO₄)

and filtered, and the solvent was removed under reduced

pressure to give 1.06 g (62%) of a yellow oil. To a stirred

solution of the free base (1.65 g, 6.4 mmol) in anhyd ether (60

mL) was added dropwise a solution of methanesulfonic acid

(0.7 g, 7.29 mmol) in ether (10 mL). The resulting suspension

was stirred at 25° C for 30 min and then allowed to settle for

an additional 30 min. The ether was then decanted off, and

the residue was dried in vacuo. It was then recrystallized from

ⁱPrOH/ether to give 2.05 g of a white solid (90.3%).

7-(N-Me,N-Et ca r ba m yloxy)-N-m eth yl,N-p r op a r gyl-1-

a m in oin d a n ‚HCl (22b‚HCl). A mixture of 21b (330 mg, 1.21

mmol), paraformaldehyde (165 mg), and NaCNBH₃(95 mg,

1.51 mmol) in absol MeOH (25 mL) was refluxed under

nitrogen for 4 h. The reaction mixture was evaporated to

dryness under reduced pressure, and the viscous oily residue

was purified by chromatography (hexane:EtOAc 70:30) to give

270 mg (78%) of the free base as a light viscous oil. The latter

was converted to the HCl salt by ethereal HCl. The crude salt

was then dissolved in dry MeOH (30 mL), to give, after

evaporation of the solvent, 245 mg (81%) of a white solid, mp

147-50 °C.

spectrometer. Chemical shifts were expressed in δ (ppm)

relative to TMS (in DMSO-d₆or CDCl₃) as internal standard,

or to HOD (4.80 ppm, in D₂O), and coupling constants (J ) are

in hertz. Mass spectra were recorded on a Finnigan 4021

spectrometer. Reaction conditions were not optimized.

Dialkylcarbamoyl chlorides were prepared by reacting di-

alkylamines with either phosgene in toluene⁶³or carbon

dioxide and thionyl chloride.⁶⁴Dialkyl thiocarbamoyl chlorides

were prepared from thiophosgene.⁶⁵

Full experimental details are given for representative

examples of the various synthetic steps. Full spectral and

analytical data may be found in the Supporting Information.

A note on nomenclature: To avoid confusion (due to the

different ranking of hydroxy and carbamoyl functionalities)

and maintain uniformity through the entire text, we have

decided to use the more straightforward (albeit not always

formal) numbering system for the compounds of Series I. Thus,

both hydroxy compounds and their carbamates are designated

4-, 6-, and 7- substituted derivatives of 1-aminoindan (e.g., 4-,

6-, and 7-hydroxy-1-aminoindan instead of 1-amino-indan-4-

ol, 3-amino-indan-5-ol, and 3-amino-indan-4-ol, respectively).

Ser ies I. 1. Boc-P r otection : 6-Hyd r oxy-N-Boc 1-a m i-

n oin d a n (5, R1)H). A solution of 6-hydroxy 1-aminoindan

(4,R1)H) (16 g, 107 mmol), di-tert-butyl dicarbonate (23.8 g,

109.2 mmol), and Et₃N (16.74 mL, 120 mmol) in THF (375

mL) was stirred at room temperature for 20 h. The reaction

mixture was evaporated to dryness under reduced pressure,

and the residue was dissolved in CH₂Cl₂(200 mL), washed

with water (200 mL), dried over Na₂SO₄, and evaporated to

dryness under reduced pressure. The crude product was

purified by column chromatography (hexane/EtOAc 2:1) to give

23 g of a solid (86%), mp: 104-5 °C.

2. Ca r ba m yla tion : 6-(N-Me,N-Et ca r ba m yloxy)-N-Boc

1-a m in oin d a n (6, R1)H). To a stirred and ice-cooled solution

of 5, R1)H (7.5 g, 30 mmol) in acetonitrile (75 mL) was added

N-Me,N-Et carbamoyl chloride (4.4 g, 36 mmol), followed by a

dropwise addition of NaH (60% in oil, 1.56 g, 39 mmol). The

reaction mixture was stirred for 2 h at room temperature

under argon. After evaporation of the solvent in vacuo, water

(100 mL) was added and extracted with ether (3 × 100 mL).

The organic phase was washed with dilute NaOH (pH 10-

11), dried, and evaporated to dryness in vacuo. Purification

by column chromatography (hexane:EtOAc 2:1) afforded 7.8

g (77%) of a solid, mp: 97-8 °C.

P r ep a r a tion of 5-Ch lor o-6-h yd r oxy-1-a m in oin d a n (38,

Sch em e 3): (1) 3-Ch lor o-4-m eth oxyben za ld eh yd e. Sulfu-

ryl chloride (270 g, 2 mol) was added to a solution of

4-methoxybenzaldehyde (136 g, 1 mol) in acetic acid (300 mL)

at 20-30 °C for 2 h while the reaction mixture was well stirred

and cooled. It was then allowed to stand at room temperature

for 1 day. The yellow solution was poured onto a mixture (1.2

kg) of water and ice. The solid was collected by filtration,

washed three times with ice-cold water and once with n-

hexane, dried under vacuum over CaCl₂, then over NaOH, to

give 170.6 g (100%), mp 40-46 °C. It was used in the next

step without any purification. A small sample was crystallized

from n-hexane, mp 54-56 °C (54-56 °C^31a, 62 °C^31b).

6-(N-Alkyl car bam yloxy)-N-Boc 1-am in oin dan (6, R1)H,

R3)H). The alkyl isocyanate (1 mmol) and a few drops of Et₃N

were added to a solution of 5,R1)H (250 mg, 1 mmol) in CH₂-

Cl₂(5 mL). The mixture was stirred at room temperature for

20 h and evaporated to dryness and the residual solid

triturated with n-hexane. The crude free base was converted

to the HCl salt by 6.7 M HCl/dioxane, and the crude salt was

crystallized from MeOH/EtOAc.

(2) 3-(3-Ch lor o-4-m eth oxyp h en yl)p r op ion ic Acid (34).

A mixture of crude 3-chloro-4-methoxybenzaldehyde (170.6 g,

1 mol), Meldrum’s acid³³(144.1 g, 1 mol), triethylammonium

formate (250 mL, prepared from triethylamine (168 mL, 1.2

Novel Dual Inhibitors of AChE and MAO

J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5275

mol) and formic acid (113 mL, 3 mol)), and DMF (200 mL) was

stirred at room temperature for 6 h and set aside at room

temperature for 18 h. The reaction mixture was then heated

to 100 °C within 2 h, stirred at 95-100 °C for 2 h, cooled to 40

°C and poured onto a mixture of ice-water (1200 g) and concd

HCl (120 mL). The solid was collected by filtration, washed

twice with cold water, dissolved in acetone (300 mL) at room

temperature, and treated with charcoal (10 g). Ice (450 g) was

added to the filtrate (450 mL), and the mixture was allowed

to stand at 5 °C for 3 days. The crystals were collected by

filtration, washed with 50% EtOH, and dried in a desiccator.

The mother liquor was evaporated to dryness, and the oily

residue was triturated with cold 50% EtOH. The solid was then

collected by filtration, washed, and dried. The combined solids

(107 g) were dissolved in EtOH (450 mL) at room temperature

and treated with charcoal (5 g). Ice cold water (450 mL) was

added to the ethanolic filtrate, and the mixture was cooled and

filtered. The solid was washed with 50% EtOH and dried (88

g, 41%, based on 4-methoxybenzaldehyde), mp 113-116 °C.

(3) 3-(3-Ch lor o-4-m eth oxyp h en yl)p r op ion yl Ch lor id e

(35). A mixture of thionyl chloride (87 mL) and 34 (129 g, 0.6

mol) was stirred at room temperature for 0.5 h and heated to

100 °C within 1 h. This temperature was maintained for an

additional 1 h. The excess of SOCl₂was distilled off under

reduced pressure. Toluene was added and then distilled off,

to give 139.8 g (0.6 mol, 100%) of a crude product which was

used in the next step without further purification.

water, dried (17.6 g, 64%), and crystallized from MeOH (15.2

g, 55%), mp 207-209 °C (dec).

P r ep a r a tion of Op tica l Isom er s of 6-Hyd r oxy-1-a m i-

n oin d a n (R/S 4,R1)H,R5)H, Sch em e 2): (1) N-Tr iflu o-

r oa cetyl-(R/S)-1-a m in oin d a n (33). To an ice- cooled solution

of trifluoroacetic anhydride (194.6 g, 0.926 mol) in toluene (680

mL) was added dropwise a solution of 98% optically pure

(chiral HPLC)⁶⁶(R/ S)-1-aminoindan (113.32 g 0.85 mol) in

toluene (50 mL) and stirred under ice-cooling for 3.5 h. A

solution of KOH (67.25 g, 1.2 mol) in water (1000 mL) was

then added, and the reaction mixture stirred for further 2 h

at room temperature. The solid was collected by filtration,

washed with water (680 mL), and dried to give 152 g (78%) of

a white solid, mp: 153-154 °C. The solution was evaporated,

and the crystals were collected by filtration, washed with

water, and dried. This second crop (25 g) was crystallized from

a mixture of hexane and ethyl acetate to give 18 g (9%) of a

white solid, mp:153-154°C. The total yield was 170 g (87%).

6-Ch lor oa ce t yl-N -t r iflu or oa ce t yl-(R /S )-1-a m in oin -

d a n . To a suspension of AlCl₃(89.2 g, 0.67 mol) in 1,2-

dichloroethane (600 mL) was added chloroacetyl chloride (55.7

mL, 78.9 g, 0.7 mol) dropwise at 0-5 °C under nitrogen for 20

min and left to warm to room temperature. To this mixture

was added 33 (34.4 g, 0.15 mol) over a period of 3 h at room

temperature. The resulting mixture was then stirred for an

additional 30 min and poured onto a mixture of ice-cold water

(1.5 L) and 1,2-dichloroethane (1 L). The mixture was stirred

for 5 min, and the layers were separated. The aqueous layer

was extracted with 1,2-dichloroethane (2 × 750 mL). The

combined organic layers were washed with water (2 × 900 mL)

and 5% aqueous NaHCO₃(3 × 900 mL). The organic layer was

dried and the solvent evaporated to give a solid, which was

crystallized from ethanol to give 31.2 g (68%) of a white solid

mp: 166-167°C.

(3) 6-Ch lor oa cet oxyl-N-t r iflu or oa cet yl-(R/S)-1-a m i-

n oin d a n . 6-Choroacetyl-N-trifluoroacetyl-(R/ S)-1-aminoindan

(30.57 g, 0.1 mol) was dissolved in anhydrous dichloromethane

(210 mL) and 3-chloroperoxybenzoic acid (70%, 44.87 g, 0.26

mol) was added in one portion. The suspension was cooled to

0°C, and trifluoroacetic acid (11.4 g, 0.1 mol) was added

dropwise over 5-10 min. The reaction flask was protected from

light, and the mixture was stirred for 3-5 days at room

temperature, poured onto water (300 mL), and neutralized

with ammonium hydroxide solution. The layers were sepa-

rated, and the aqueous layer was extracted with dichlo-

romethane (200 mL). The combined organic layers were dried,

and the solvent was evaporated to give a solid, which was

crystallized from ethanol to give 15 g (48%) of a white solid

mp: 169-170 °C.

(4) 6-Hyd r oxy-(R/S)-1-a m in oin d a n ((R/S)-4,R1)H,-

R5)H). A suspension of 6-chloroacetoxy-N-trifluoroacetyl-(R/

S)-1-aminoindan (25.4 g, 0.11 mol) and K₂CO₃(38.0 g, 0.275

mol) in a mixture of methanol (275 mL) and water (175 mL)

was stirred at 70 °C for 1.5 h. Methanol was removed in vacuo,

and the aqueous phase was neutralized with 10% HCl. The

mixture was filtered and the solid was washed with water.

The mother liquor was concentrated to a small volume, and

the resulting suspension was neutralized and filtered. The

brown solid was crystallized twice from methanol to give 7.0

g (43%) of a white solid, mp 200-203 °C.

P r epar ation of 6-Hydr oxy-1-(1-m eth ylpr opar gyl)am in o-

in d a n ‚HCl (4,R1)1-m eth ylp r op a r gyl,R5)H). (1) 3-Mes-

yloxy-1-bu tyn e.²¹To a solution of 10 g (0.147 mol) of (()

3-butyn-2-ol and 29.8 mL (0.215 mol) of triethylamine in 250

mL of methylene chloride cooled at -50 °C was added 14.4

mL (0.185 mol) of mesyl chloride within 1.5 h under stirring.

The solution was allowed to warm to room temperature and

stirred for 1 h. The solution was washed twice with water and

then evaporated to dryness to give the crude product (21.3 g),

which was used for the next step without purification.

(4) 5-Ch lor o-6-m eth oxy-1-in d a n on e (36). Crude 35 (35

g, 27.5 mL) was added to a stirred suspension of AlCl₃(22 g)

in CH₂Cl₂(1350 mL) at 0-5 °C, and the suspension was stirred

at this temperature for 1 h. A second aliquot of crude 35 (27.5

mL) and AlCl₃(22 g) was added with additional stirring at

0-5 °C for 1 h. The reaction mixture was allowed to warm to

room temperature, stirred for an additional 1 h, and poured

onto a mixture of ice, water (1.5 kg), and concd HCl (72 mL).

CH₂Cl₂(600 mL) was added to the mixture under efficient

stirring. The phases were separated, and the organic layer was

dried and treated with charcoal. The filtrate was concentrated

to half of volume, stored at -15 °C, filtered, washed with cold

CH₂Cl₂and MeOH, and dried (102.7 g, 87%), mp 165-166 °C.

(5) 5-Ch lor o-6-m eth oxy-1-in d a n on e Oxim e. A mixture

of 36 (98.3 g, 0.5 mol), hydroxylamine hydorchloride (69.5 g, 1

mol), sodium acetate (82 g, 1 mol), ethanol (450 mL), and water

(250 mL) was refluxed for 1 h. The reaction mixture was

allowed to cool to room temperature, cooled to 5 °C, and

filtered. The solid was washed with water and cold EtOH and

dried (102.6 g, 97%), mp 208-210 °C.

(6) 5-Ch lor o-6-m eth oxy-1-a m in oin d a n ‚HCl (37‚HCl). A

solution of NaBH₄(37.8 g, 1 mol) in DMF (300 mL) was added

to a stirred mixture of MoO₃(54.0 g, 0.25 mol) and 5-chloro-

6-methoxy-1-indanone oxime (52.9 g, 0.25 mol) in MeOH (1000

mL) at room temperature under N₂for 2 h. Water (200 mL)

was added to the reaction mixture over a period of 10 min.

MeOH was distilled off under reduced pressure, and concd HCl

(150 mL) and crushed ice (70 g) were added to the residue.

The solid was removed by filtration, and ice (50 g) and 10 N

KOH (200 mL) were added to the filtrate. The suspension was

filtered and the cake washed with water and CH₂Cl₂. The

filtrate was extracted with CH₂Cl₂(3 × 600 mL), and concd

HCl (21 mL) was added to the combined organic layers under

stirring. The mixture was well shaken and stored at 5 °C for

3 days. The crystals were then collected by filtration, washed

three times with CH₂Cl₂, and dried (29.1 g). A second crop of

8.9 g was obtained from the mother liquors (total yield 65%),

mp 257-260 °C (dec).

(7) 5-Ch lor o-6-h yd r oxy-1-a m in oin d a n (38). A mixture

of 37‚HCl (35.1 g, 0.15 mol), 45% aq HBr (300 mL), and 33%

HBr/AcOH (150 mL) was refluxed for 1 h and evaporated to

dryness under reduced pressure, and the residue was dissolved

in water (350 mL). The dark solution was treated with charcoal

(1.5 g) at room temperature for 0.5 h, the charcoal was filtered

off, and 10 N KOH was added to the clear solution (pH 8.5).

The resulting solid was collected by filtration, washed with

¹H NMR (δ, DMSO-d₆): 1.52, 1.58 (3H, s, CH₃), 3.2 (3H, s,

CH₃), 3.8 (1H, s, CH), 5.35 (1H, m, CH) ppm.

(2) 6-Hyd r oxy-1-(1-m eth ylp r op a r gyl)a m in oin d a n ‚HCl

(4,R1)1-m eth ylp r op a r gyl,R5)H). To a solution of 17.4 g

5276 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24

Sterling et al.

of 6-hydroxy-1-aminoindan in 500 mL DMF, 16.2 mL triethyl-

amine and 17.1 g 3-mesyloxy-1-butyne was added; the solution

was allowed to stand at room temperature for 82 h, poured

into 1000 mL water and extracted with 3 × 200 mL ethyl

acetate. The organic phase was washed with 3 × 100 mL

water, dried on MgSO₄and the solvent removed under reduced

pressure. The residue was purified by column chromatography

(ethyl acetate); the free base (R_f) 0.765) was dissolved in ether

and treated with gaseous HCl to give the HCl salt (5.25 g,

mp: 206-209 °C). Anal. Calcd. for C₁₃H₁₅NO HCl (237.72):

C, 65.67; H, 6.78; N, 5.89. Found: C ,63.51; H , 6.71; N , 5.66.

purified by column chromatography (hexane:EtOAc 2:1), af-

fording 6.0 g (86%) of the title compound as a yellow oil.

The monoalkyl carbamates 45g and 45h were prepared by

the procedure described above for monoalkyl carbamates of

Series I.

3. Boc-Dep r ot ect ion : 3-(N-Me,N-n P r ca r b a m yloxy)

N-Met h ylp h en et h yla m in e ‚H Cl (46c‚H Cl) 44,R 1)Me,-

R6)H (6.0 g, 17.14 mmol) was dissolved in dioxane (60 mL),

and 20% HCl/ether (60 mL) was added. The mixture was

stirred at room temperature for 4 h and evaporated to dryness

in vacuo, and the residual oil was treated with ether (2 × 150

mL), to give, after stirring and ice-cooling, 4.6 g (93.5%) of the

title compound as a white solid.

¹H NMR (δ, DMSO-d₆): 1.55 (3H, tr, CH₃); 2.1-3.2 (4H, m,

2CH₂); 3.50, 4.0 (1H, s, CH); 4.15, 4.4 (1H, m, CH); 4.8 (1H,

m, CH); 6.85-7.15 (3H, m, Ar); 9.6 (1H, s, OH); 9.5-10.0 (1H,

m, NH) ppm.

4. P r op a r gyla tion : 3-(N-Me,N-Et ca r ba m yloxy) N-P r o-

p a r gylp h en ylp r op yla m in e‚HCl (52b‚HCl). A solution of

propargyl bromide (1.1 g, 9.1 mmol) in acetonitrile (8.5 mL)

was added dropwise to a stirred mixture of 47b‚HCl (2.4 g,

8.8 mmol) and potassium carbonate (2.8 g) in acetonitrile (25

mL), and the mixture was stirred at room temperature for 7

h. The reaction mixture was filtered, and the filtrate was

removed under reduced pressure. The residue was purified by

column chromatography (hexane:EtOAc 2:1) to give 1.76 g of

the title compound as the free base (73%). The free base was

dissolved in dry ether (50 mL), and HCl/ether was added (to

pH 1). The mixture was stirred for 4 h at room temperature

and filtered, and the solid was washed with cold ether, to give,

after drying at 60° C in vacuo, 1.5 g (4.82 mmol, 55%).

P r ep a r a tion of Op tica l Isom er s of 3-(N-Meth yl,N-

cycloh exylca r ba m yloxy)-N-m eth yl-N-p r op a r gylp h en yl-

p r op yla m in e Mesyla te ((S)-53e Mesyla te). (1) (S)-3-(2-

Am in op r op yl)p h en ol ((S)-57): Racemic 56 (11.78 g, 77.92

mmol) and ^L-tartaric acid (11.70 g, 77.95 mmol) were heated

to reflux in methanol (520 mL). The yellow solution was then

concentrated to a volume of 200 mL, and the heat was

removed. The white suspension formed upon cooling to room

temperature was set aside for 24 h and filtered (18.9 g), the

solid thus obtained was heated to reflux in methanol (480 mL),

and the mixture was concentrated to a volume of 200 mL. On

cooling, a white solid precipitated, which was collected by

filtration after 3 h and dried (10.16 g), mp 174-176 °C, lit.²³

184-185 °C, [R]_D+18.08°(c ) 2, H₂O) lit.²³+29.5° (c ) 2, H₂O).

The tartrate salt was converted into its free base by dissolving

it in 22% ammonium hydroxide solution (1 L) and extracting

with CH₂Cl₂(50 × 70 mL). After drying and evaporation of

solvent, 4.35 g of (S)-57 was obtained as an off-white solid,

[R]_D+12° (c ) 1.3, methanol) lit.⁴¹+14.9° (c ) 1.3, methanol).

(R)-57 was similarly obtained from 56 and ^D-tartaric acid:

[R]_D-13.9° (c ) 1.3, methanol) lit.⁴¹-14.9° (c ) 1.3, methanol).

(2) (S)-3-H yd r oxy-N-m et h yl,N-p r op a r gylp h en ylp r o-

p yla m in e ((S)-55q). To a solution of (S)-58 (4.4 g, 21.8 mmol),

obtained from (S)-57 via N-formylation⁴⁰followed by reduc-

tion,⁴¹in dimethylacetamide (200 mL) stirred at 25 °C under

a nitrogen atmosphere, was added K₂CO₃(6.04 g, 43.64 mmol),

and the mixture was stirred for 10 min. Then a solution of

propargyl bromide (2.34 g, 19.64 mmol) in dimethylacetamide

(10 mL) was added over 2 min, and the mixture was stirred

at 25 °C for 24 h. Water (250 mL) was added, and the mixture

was stirred until all the solid material dissolved. The aqueous

layer was extracted with toluene (10 × 75 mL). The layers

were separated, and the combined toluene layer was washed

with saturated brine (2 × 150 mL) and dried (Na₂SO₄).

Removal of solvent at reduced pressure gave an orange oil,

which was purified by flash column chromatography using

ethyl acetate as the eluent. This gave 3.60 g (90.2%) of the

title compound as an orange oil.

P r ep a r a tion of Hyd r oxy P r op a r gyla m in oin d a n s 39. A

mixture of hydroxy 1-aminoindan 4,R1)H,R5)H (35 mmol),

propargyl bromide (35 mmol), and potassium carbonate (35

mmol) in DMA (100 mL) was stirred at room temperature for

24 h. The reaction mixture was filtered, diluted with water

(200 mL), and extracted with toluene (4 × 100 mL). The

organic extracts were combined, dried, and evaporated to

dryness under reduced pressure. The residue was then sub-

jected to flash column chromatography (hexane: EtOAc 1:1).

P r ep a r a tion of Hyd r oxy N-Me,N-P r op a r gyla m in oin -

d a n s 40. (1) A mixture of 39u (5.0 g, 26.7 mmol), paraform-

aldehyde (3.6 g, 30 mmol), and NaCNBH₃(1.96 g, 31.2 mmol)

in absol MeOH (90 mL) was refluxed under argon for 4 h. The

crude product (40u ) obtained after evaporation of the solvent

was purified by flash chromatography (hexane: EtOAc 70:30)

and converted to its HCl salt (Etheral HCl), 4.2 g (17.6 mmol,

66%). Analogously, 40w was prepared from 39w .

(2) N-Methylation of 7-hydroxy-1-aminoindan 4,R1)H,-

R5)H: Refluxing the latter (3.7 g, 24.8 mmol) in ethyl formate

(200 mL) for 18 h afforded, after removal of ethyl formate and

flash chromatography, 4.1 g (93%) of N-formyl-7-hydroxy-1-

aminoindan. A solution of the latter in dry THF (70 mL) was

added to an ice-cooled and stirred suspension of LAH in dry

THF (100 mL). The reaction mixture was stirred for 9 h at

room temperature, ice-cooled and treated with water (100 mL).

The pH was adjusted to ca. 8-9, water (200 mL) was added,

and the mixture was extracted with ether (6 × 250 mL). The

combined ethereal extracts were dried and evaporated to

dryness to give 3.2 g (94%) of 7-hydroxy-N-methyl-1-aminoin-

dan. Reaction with propargyl bromide (as described for 39)

afforded 40v in 53%.

Ser ies II. 1. Boc P r otection : 3-Hyd r oxy-N-Boc,N-m eth -

ylp h en eth yla m in e (43,R1)Me,R6)H). To a solution of

3-hydroxy-N-methylphenethylamine (42,R1)Me,R6)H) (8.33

g, 55.17 mmol) in dioxane (80 mL) and water (80 mL) were

added NaHCO₃(13.65 g) and di-tert-butyl dicarbonate (13.65

g, 62.54 mmol). The reaction mixture was stirred at room

temperature for 4 h and evaporated to dryness in vacuo. The

residue was taken up in a water:dioxane mixture (400 mL,

1:1), and the layers were separated. The aqueous layer was

re-extracted with ether (2 × 75 mL), and the combined ether

layer was dried (Na₂SO₄) and evaporated to dryness in vacuo.

The oily residue was purified by column chromatography

(hexane:EtOAc 2:1) to give 10.2 g (74%) of the title compound

as a viscous yellow oil.

2. Ca r ba m yla tion : 3-(N-Me,N-n P r ca r ba m yloxy) N-

Boc,N-m eth ylp h en eth yla m in e (44,R1)Me,R6)H). To an

ice-cooled solution of 43,R1)Me,R6)H) (5.0 g, 19.9 mmol) in

dry acetonitrile (65 mL) was added, under nitrogen, N-

methyl,N-n-propyl carbamoyl chloride (4.66 g, 34.43 mmol),

followed by the portionwise addition of NaH (60% disp. in oil,

1.03 g, 25.87 mmol). The reaction mixture was stirred at room

temperature under nitrogen for 6 h and evaporated to dryness

in vacuo. Water (200 mL) was added, the pH was adjusted to

∼9, and the aqueous layer was extracted with ether (4 × 100

mL). The combined ether layer was washed with NaOH

solution (pH 9.5) and water (150 mL), dried (Na₂SO₄), and

evaporated to dryness in vacuo to give an oil which was

Using this procedure, (R)-55q was obtained in 80%.

(3) (S)-3-(N-Meth yl,N-cycloh exylca r ba m yloxy)-N-m e-

th yl,N-p r op a r gylp h en ylp r op yla m in e Mesyla te ((S)-53e

Mesyla te). To a solution of compound (S)-55q (1.80 g, 8.87

mmol) in dry acetonitrile (100 mL) cooled in an ice bath was

added under nitrogen N-methyl-N-cyclohexylcarbamyl chloride

(2.70 g, 15.39 mmol) followed by the portionwise addition of

NaH (60% oil dispersion, 0.467 g, 11.69 mmol). The mixture

was then stirred at room temperature under nitrogen for 18

Novel Dual Inhibitors of AChE and MAO

J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5277

h. Solvent was removed at reduced pressure, and water (100

mL) and ether (150 mL) were added. The mixture was stirred

until all the material dissolved. The layers were separated,

and the aqueous layer was reextracted with ether (4 × 70 mL).

The combined ether layers were washed with KOH solution

(pH 9.5), then water, and dried (Na₂SO₄). Removal of solvent

at reduced pressure gave 3.87 g of an orange oil which was

purified by flash column chromatography using ethyl acetate

as the eluent. This gave 2.57 g (84.5%) of the title compound

(free base) as a yellow oil. The free base was dissolved in dry

EtOAc (9 mL), and a solution of 95% ethanesulfonic acid (0.79

g, 6.81 mmol) in EtOAc (1.5 mL) was added. The solution was

cooled to 5 °C and stirred at this temperature. After 15 min,

a white solid precipitated, and the suspension was stirred at

5 °C for 3 h. The solid was collected by filtration using a

minimum amount of ice-cold ethyl acetate. This gave 2.3 g

(75%) of a white solid having a melting point of 118-120 °C.

P h a r m a cology. ACh E In h ibition . Human acetylcho-

linesterase (AChE; EC 3.1.1.7) purified from red blood cells

and equine butyrylcholinesterase (BuChE; EC 3.1.1.8) purified

from serum (Sigma Chemical Co., St Louis, MO) were used

for determination of cholinesterase inhibitory activity of the

compounds. Assays were performed as described by Ellman

et al.⁶⁷using 200 µM acetylcholine as substrate for AChE and

200 µM butyrylthiocholine as substrate for BuChE. Com-

pounds were preincubated with the enzyme for 60 min at 37

°C in 0.1 M phosphate buffer, pH 7.4, before addition of

substrate. Assays were performed in triplicate and four-point

dose-response curves were obtained for each compound. The

percent inhibition of enzyme activity in the absence of com-

pounds was used to calculate IC₅₀’s and their fiducial limits

by means of the statistical program Origin version 7 from a

plot of log(10) of concentration against % enzyme inhibition.

The bimolecular rate constant (k_i) for carbamylation of human

erythrocyte enzyme for four of the compounds and for rivastig-

mine were determined under the same conditions as those

alem, Israel. We also acknowledge Dr. H. Gottlieb from

Bar-Ilan University, Ramat Gan, Israel, for useful

discussions. We thank Mr. G. Attili and Dr. E. Bachar

for logistical assistance and helpful discussions.

Su p p or tin g In for m a tion Ava ila ble: Spectral (¹H NMR,

IR, MS) and analytical (C,H,N) data. This material is available

free of charge via the Internet at http://pubs.acs.org.

Refer en ces

(1) Coyle, J . T.; Price, D. L.; DeLong, M. R. Alzheimer’s Disease: a

Disorder of Cortical Cholinergic Innervation. Science 1983, 219,

1184-90.

(2) Newman, S. C. The Prevalence of Depression in Alzheimer’s

Disease and Vascular Dementia in a Population Sample. J .

Affect. Disord. 1999, 52, 169-176.

(3) Palmer, A. M.; Stratman, G. C.; Procter, A. W.; Bowen, D. M.

Possible Neurotransmitter Basis of Behavioral Changes in

Alzheimer’s Disease. Ann. Neurol. 1988, 23, 616-20.

(4) Good, P. F.; Werner, P.; Hsu, A.; Olanow, C. W.; Perl, D. P.

Evidence for Neuronal Oxidative Damage in Alzheimer’s Dis-

ease. Am. J . Pathol. 1996, 149, 21-28.

(5) Saura, J .; Luque, J . M.; Cesura, A. M.; Da Prada, M.; Chan-

Palay, V.; Hu Loffler, J .; Richards, J . G. Increased Monoamine

Oxidase B Activity in Plaque-Associated Astrocytes of Alzheimer

Brains Revealed by Quantitative Radioautography. Neuroscience

1994, 62, 15-30.

(6) Weinstock, M. Selectivity of Cholinesterase Inhibition: Clinical

Implications for the Treatment of Alzheimer’s Disease. CNS

Drugs 1999, 12, 307-323.

(7) Sano, M.; Ernesto, C.; Thomas, R. G.; Klauber, M. R.; Schafer,

K.; Grundman, M.; Woodbury, P.; Growdon, J .; Cotman, C. W.;

Pfeiffer, E.; Schneider, L. S.; Thal, L. J . A Controlled Trial of

Selegiline, Alpha-Tocopherol, or both as Treatment for Alzhei-

mer’s Disease. N. Engl. J . Med. 1997, 336, 1216-1222.

(8) Abu-Raya, S.; Blaugrund, E.; Trembovler, V.; Shilderman-Bloch,

E.; Shohami, E.; Lazarovici, P. Rasagiline, a Monoamine Oxi-

dase-B Inhibitor, Protects NGF-Differenciated PC12 Cells Against

Oxygen-Glucose Deprivation. J . Neurosci. Res. 1999, 58, 456-

463.

(9) Paterson, I. A.; Tatton, W. G. Antiapoptotic Actions of Monoam-

ine Oxidase B Inhibitors. Adv. Pharmacol. 1998, 42, 312-5.

(10) Youdim, M. B.; Wadia, A.; Tatton, W.; Weinstock, M. The Anti-

Parkinson Drug Rasagiline and its Cholinesterase Inhibitor

Derivatives Exert Neuroprotection Unrelated to MAO Inhibition

in Cell Culture and In Vivo. Ann. NY Acad. Sci. 2001, 939, 450-

8.

(11) O’Malley, G.; Bores, G.; Huger, F.; Kurys, B.; Merriman, M.;

Olsen, G.; Ong, H.; Petko, W.; Palermo, M. Synthesis and

Biological Evaluation of Combined AChE and MAO Inhibitors.

Abstracts of Papers, 205th American Chemical Society National

Meeting, 1993; American Chemical Society: Washington, DC,

1993; MEDI, abstract.

(12) Fink, D. M.; Palermo, M. G.; Bores, G. M.; Huger, F. P.; Kurys,

B. E.; Merriman, M. C.; Olsen, G. E.; Petko, W.; O’Malley, G. J .

Imino 1,2,3,4-Tetrahydrocyclopent[b]-indole Carbamates as Dual

Inhibitors of Acetylcholinesterase and Monoamine Oxidase.

Bioorg. Med. Chem. Lett. 1996, 6, 625-630.

(13) Palermo, M.; Bores, G.; Huger, F.; Kurys, B.; Merriman, M.;

Olsen, G.; Ong, H.; Petko, W.; O’Malley, G. Combined AChE and

Reversible MAO Inhibition as a Potential Therapeutic Approach

for Senile Dementia of the Alzheimer Type. Abstracts of Papers,

205th American Chemical Society National Meeting, 1993;

American Chemical Society: Washington, DC, 1993; MEDI,

abstract.

described above. The pseudo-first-order rate constants (k_obs

)

for progressive AChE inhibition at 4-5 concentrations of each

inhibitor were established as described by Bar-On et al.⁵¹by

means of the statistical program Origin version 7. From these

we calculated the apparent bimolecular rate constant k_ifor

each compound.

MAO In h ibition . Inhibition by the compounds of MAO-A

and MAO-B was determined by a method adapted from Tipton

and Youdim.⁶⁸Rat brain was homogenized in sucrose (0.3 M)

and served as the source of enzymes. The compound under

test was diluted from 10^-2M to 10^-9M in 0.05 M phosphate

buffer (pH 7.4) containing 0.15 µM selegiline for determination

of MAO-A, or in 0.05 M phosphate buffer containing 0.15 µM

clorgyline for determination of MAO-B. Fifty microliters of a

suitable dilution of the enzyme preparation was added to test

tubes containing 100 µL of phosphate buffer (control test

tubes); test tubes containing 100 µL of phosphate buffer and

10 µL of 0.01 M tranylcypromine (blank); test tubes containing

100 µL of various dilutions of the compounds. Incubation was

carried out for 60 min at 37 °C. Substrates (¹⁴C-5-hydroxy-

tryptamine creatinine disulfate (100 µM) for determination of

MAO-A) or (¹⁴C-phenylethylamine (10 µM) for determination

of MAO-B) were then added and the incubation continued for

a further 30 and 20 min, respectively. The reaction was

stopped with 250 µL of 2 M citric acid. Radioactive metabolites

were extracted into toluene/ethyl acetate (1:1 v/v), a solution

of 2,5-diphenyloxazole was added to a final concentration of

0.4%, and the metabolite content was estimated by liquid

scintillation counting. Activity in the presence of the drug was

expressed as a percentage of that of the control. Assays were

performed in duplicates, and variation between the duplicates

did not exceed 6%. Dose-response curves consisted of 6-7

points. Sigmoid curves and IC₅₀calculations were carried out

using “Prism” software.

(14) Kimura, T.; Kuroki, Y. Pyrimidine Compound. EP 664291 (UBE

Industries, 1994).

(15) Bruhlmann, C.; Ooms, F.; Carrupt, P.-A.; Testa, B.; Catto, M.;

Leonetti, F.; Altomare, C.; Carotti, A. Coumarins Derivatives

as Dual Inhibitors of Acetylcholinesterase and Monoamine

Oxidase. J . Med. Chem. 2001, 44, 3195-3198.

(16) Knollema, S.; Aukema, W.; Hom, H.; Korf, J .; Horst, G. J .T.

L-Deprenyl Reduces Brain Damage in Rats Exposed to Transient

Hypoxia-Ischemia. Stroke 1995, 26, 1883-1887.

(17) Lahtinen, H.; Koistinaho, J .; Kauppinen, R.; Haapalinna, A.;

Keinanen, R.; Sivenius, J . Selegiline Treatment After Transient

Global Ischemia in Gerbils Enhances the Survival of CA1

Pyramidal Cells in the Hippocampus. Brain Res. 1997, 757,

260-267.

(18) inberg, J . P. M.; Takeshima, T.; J ohnston, J . M.; Commissiong,

J . W. Increased Survival of Dopaminergic Neurons by Rasa-

giline, a Monoamine Oxidase-B Inhibitor. NeuroReport 1998, 9,

703-707.

(19) Huang, W.; Chen, Y.; Shohami, E.; Weinstock, M. Neuroprotec-

tive Effect of Rasagiline, a Selective Monoamine Oxidase-B

Inhibitor, Against Closed Head Injury in the Mouse. Eur. J .

Pharmacol. 1999, 366, 127-135.

Ack n ow led gm en t. We gratefully acknowledge the

dynamic NMR studies performed by Prof. I. Ringel from

the School of Pharmacy, the Hebrew University, J erus-

5278 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24

Sterling et al.

(20) Gulinnaz, A.; Girgin, F. K.; Ozgonul, M.; Mentes, G.; Ersoz, B.

MAO Inhibitors and Oxidant Stress in Aging Brain Tissue. Eur.

Neuropsychopharmacology 1999, 9, 247-252.

(21) Dostert, P.; O’Brien, E. M.; Tipton, K. F.; Meroni, M.; Melloni,

P.; Strolin, M.; Benedetti, M. Inhibition of Monoamine Oxidase

by the R and S Enantiomers of N[3-(2,4-Dichlorophenoxy)-

propyl]-N-Methyl-3-Butyn-2-Amine. Eur. J . Med. Chem. 1992,

27, 45-52.

(46) Harel, M.; Sussman, J . L.; Krejci, E.; Bon, S.; Chanal, P.;

Massoulie, J .; Silman, I. Conversion of Acetylcholinesterase to

Butyrylcholinesterase: Modeling and Mutagenesis. Proc. Natl.

Acad. Sci. U.S.A. 1992, 89, 10827-10831.

(47) Lieske, C. N.; Gepp, R. T.; Clark, J . H.; Meyer, H. G.; Blumberg,

P.; Tseng, C. C. Anticholinesterase Activity of Potential Thera-

peutic 5-(1,3,3-Trimethylindolinyl) Carbamates. J . Enzyme In-

hib. 1991, 5, 215-223.

(22) Kawasaki, I.; Matsuda, K.; Kaneko, T. Preparation of 1,7-Bis-

(p-hydroxyphenyl) heptane. Bull. Chem. Soc. J pn. 1971, 44,

1986-7.

(23) Oshiro, Y.; Sakurai, Y.; Tanaka, T.; Ueda, H.; Kikuchi, T.;

Tottori, K. Novel Cerebroprotective Agents with CNS Stimulat-

ing Activity. 1. Synthesis and Pharmacology of the 1-Amino-7-

Hydroxyindan Derivatives. J . Med. Chem. 1991, 34, 2004-13.

(24) Borch, R. F.; Bernstein, M. D.; Durst, H. D. The Cyanohydri-

doborate Anion as a Selective Reducing Agent. J . Am. Chem.

Soc. 1971, 93, 2897-2904.

(25) Cannon, J . G.; Dushin, R. G.; Long, J . P.; Ilhan, M.; J ones, N.

D.; Swartzendruber, J . K. Synthesis and Dopaminergic Activity

of (R) and (S)-4-Hydroxy-2-(Di-n-propylamino)indan. J . Med.

Chem. 1985, 28, 515-18.

(26) Singh, T.; Stein, R. G.; Hoops, J . F.; Biel, J . H.; Hoya, W. K.;

Cruz, D. R. Antimalarials. 7-Chloro-4-(Substituted amino)-

quinolines. J . Med. Chem. 1971, 14, 283-286.

(48) Iyo, M.; Namba, H.; Fukushi, K.;. Measurement of Acetylcho-

linesterase by Positron Emission Tomography in the Brains of

Healthy Controls and Patients with Alzheimer’s Disease. Lancet

1997, 349, 1805-1809.

(49) Arendt, T.; Bruckner, M.; Lange, M.; Bigl, V. Changes in

Acetylcholinesterase and Butyrylcholinesterase in Alzheimer’s

Disease Resemble Embryonic Development - a Study of Molec-

ular Forms. J . Neurochem. 1992, 21, 231-244.

(50) Bolognesi, M. L.; Andrisano, V.; Bartolini, M.; Minarini, A.;

Rosini, M.; Tumiatti, V.; Melchiorre, C. Hexahydrochromeno-

[4,3-b]pyrrole Derivatives as AChE Inhibitors. J . Med. Chem.

2001, 44, 105-9.

(51) Bar-On, P.; Millard, C. B.; Harel, M.; Dvir, H.; Enz, A.; Sussman,

J . L.; Silman, I. Kinetic and Structural Studies on the Interaction

of Cholinesterases with the Anti-Alzheimer Drug Rivastigmine.

Biochemistry, 2002, 41, 3555-64, was published while this

manuscript was in the final stages of submission. In their crystal

structure, the rivastigmine carbamoyl bond has already been

cleaved and the two fragments (NAP and carbamoyl) are bound

by the enzyme some distance from one another. Thus, it is

impossible to reconstruct the conformation of rivastigmine at

the time of initial binding. Although the 4-hydroxy dimeth-

ylaminoindan resulting from 32a superimposes more success-

fully on their NAP, there is no apparent reason that 6-hydroxy

aminoindan from 31a will not also fit in the site. We are

continuing to investigate this issue.

(52) Hazelhoff, B.; De Vries, J . B.; Dijkstra, D.; de J ong, W.; Horn,

A. S. The Neuropharmacological Profile of N-Methyl-N-Propar-

gyl-2-Aminotetralin: a Potent Monoamine Oxidase Inhibitor.

Naunyn-Schmiedeberg’s Arch. Pharmacol. 1985, 330, 50-58.

(53) Sterling, J .; Veinberg, A.; Lerner, D.; Goldenberg, W.; Levy, R.;

Youdim, M.; Finberg, J . (R) (+)-N-Propargyl-1-Aminoindan

(Rasagiline) and Derivatives: Highly Selective and Potent

Inhibitors of Monoamine Oxidase B. J . Neural Transm. [Suppl]

1998, 52, 301-305.

(27) Huebner, C. F.; Donoghue, E. M.; Plummer, A. J .; Furness, P.

A. N-Methyl-N-2-Propynyl-1-indanamine, a Potent MAO Inhibi-

tor. J . Med. Chem. 1966, 9, 830-832.

(28) Toth, G., unpublished results.

(29) Lidor, R.; Bahar, E. Method for Preparing Optically Active

1-Aminoindan Derivatives. US 5639913, 1997.

(30) Teranishi, K.; Nakatsuka, S.; Goto, T. Facile synthesis of

6-Hydroxyindole and 6-Methoxyindole via Regioselective Friedel-

Crafts Acylation and Baeyer-Villiger Oxidation. Synthesis 1994,

1018-20.

(31) (a) Riggs, R. M.; Nichols, D. E.; Foreman, M. M.; Truex, L. L.

Evaluation of Isomeric 4-(Chlorohydroxyphenyl)-1,2,3,4-tet-

rahydroisoquinolines as Dopamine D-1 Antagonists. J . Med.

Chem. 1987, 30, 1887-1889. (b) Ginsburg D. The Action of tert-

Butyl Hypochlorite on Organic Compounds. II. Aromatic Alde-

hydes. J . Am. Chem. Soc. 1951, 73, 702-704.

(32) Toth, G.; Kover, K. E. Simple, Safe, Large Scale synthesis of

5-Arylmethyl-2,2-dimethyl-1,3-dioxane-4,6-diones and 3-Aryl-

Propanoic Acids. Synth. Commun. 1995, 25, 3067-3074.

(33) Meldrum, A. N. A â-Lactonic Acid from Acetone and Malonic

Acid. J . Chem. Soc. 1908, 93, 598-601. Davidson, D; Bernhard,

S. A. The Structure of Meldrum’s supposed â-Lactonic Acid. J .

Am. Chem. Soc. 1948, 70, 3426-8.

(54) Binda, C.; Newton-Ginson, P.; Hubalek, F.; Edmondson, D. E.;

Mattevi, A. Structure of Human Monoamine Oxidase B, a Drug

Target for the Treatment of Neurological Disorders. Nature

Struct. Biol. 2002, 9, 22, was published while this manuscript

was in the final stages of submission.

(34) Demir, A. S.; Tanyeli, C.; Sesenoglu, O.; Demic, S.; Evin, O. O.

A Simple Synthesis of 1-Aminophosphonic Acids from 1-Hy-

droxy-iminophosphonates with NaBH₄in the Presence of Tran-

sition Metal Compounds. Tetrahedron Lett. 1996, 37, 407-410.

(35) Copinga, S.; Tepper, P.; Grol, C. J .; Horn, A. S.; Dubocovich, M.

L. 2-Amino-8-Methoxytetralins: A Series of Nonindolic Mela-

tonin-like Agents. J . Med. Chem. 1993, 36, 2891-98.

(36) Ellenstein, A.; Ringel, I. (School of Pharmacy, the Hebrew

University, J erusalem, Israel), unpublished results, personal

communication.

(37) Weinstock, M.; Razin, M.; Ringel, I.; Tashma, Z.; Chorev, M.

Acetylcholinesterase Inhibition by Novel Carbamates: A Kinetic

and NMR Study. In Multidiscipinary Approaches to Choline-

sterase Functions, 36th; Shafferman, A., Velan, B., Eds.; Ple-

num: New York, NY, 1992; pp 251-9.

(38) Buck, J . S.; Baltzly, R.; Ide, W. S. â-Phenylethylamine Deriva-

tives. Tertiary and Quaternary salts. J . Am. Chem. Soc. 1938,

60, 1789-92.

(39) Organic Syntheses; Wiley: New York, 1963; Collect. Vol. IV, pp

573-6.

(40) Carlsson, A.; Lindquist, M.; Wysokowski, J . Substituted Meta-

tyramines as Brain Monoamine Depletors. Acta Pharm. Suecica

1970, 7, 293-302.

(41) Acid addition Salts of D-(+)-1-(3-Hydroxyphenyl)-2-Aminopro-

pane and their Manufacture and Use. GB 1527479, 1977.

(42) Saari, W. S.; Raab, A. W.; Engelhardt, E. L. The Stereoisomers

of Alpha-(1-Aminoethyl)-m-Hydroxybenzyl Alcohol. J . Med. Chem.

1968, 11, 1115-7.

(55) Kalir, A.; Sabbagh, A.; Youdim, M. B. H. Selective Acetylenic

‘Suicide’ and Reversible Inhibitors of MAO Types A and B. Br.

J . Pharmacol. 1981, 73, 55-64.

(56) Riederer, P.; Reynolds, G. P.; Youdim, M. B. H.; J ellinger, K. In

Vitro Tests of MAO Inhibitors in Human Brain Tissue: Chemi-

cal Structure and Pharmacological Action. Int. Congr. Ser.-

Excerpta Med. 1982, 564 (MAO), 345-350.

(57) Polymeropoulos, P. l-Deprenyl: A Unique MAO-B Inhibitor. In

Inhibitors of MAO B, Pharmacology and Clinical Use in Neu-

rodegenerative Disorders; Szelenyi, I. Ed.; Birkhauser Verlag:

Basel, 1993.

(58) Moron, J . A.; Campillo, M; Perez, V.; Unzeta, M.; Pardo, L.

Molecular Determinants of MAO Selectivity in

a Series of

Indolylmethylamine Derivatives: Biological Activities, 3D-QSAR/

CoMFA Analysis, and Computational Simulation of Ligand

Recognition. J . Med. Chem. 2000, 43, 1684-1691.

(59) For the parent compounds (R ) H), the N-methyl derivative

showed about 3 orders of magnitude more MAOI activity than

the secondary amine for both A and B, whereas in the hydroxy

derivative (R ) OH) solely A was enhanced by this factor and B

by only 1 order of magnitude. On the other hand, N-methylation

of the methoxy analogue (R ) OCH₃) decreased A activity by a

factor of 2 and B by a factor of 10. All of these compounds are A

selective by factors ranging from 2 to 200.

(60) Swett, L. R.; Martin, W. B.; Taylor, J . D.; Everett, G. M.; Wykes,

A. A.; Gladish, Y. C. Structure-Activity Relations in the

Pargyline Series. Ann. N.Y. Acad. Sci. 1963, 891-898.

(61) Borbe, H.; Niebch, G.; Nickel, B. Kinetic Evaluation of MAO-B

Activity Following Oral Administration of Selegiline and Des-

methyl Selegiline in Rat. J . Neural Transm. 1990, 32 (suppl)

131-7.

(43) Buckley, T. F., III; Rapoport, H. R-Amino Acids as Chiral Educts

for Asymmetric Products. Amino Acylation with N-Acylamino

Acids. J . Am. Chem. Soc. 1981, 103, 6157-63.

(44) Marco, J . L.; Royer, J .; Husson, H.-P. Asymmetric Synthesis

IX: Preparation of Chiral R-Substituted Phenethylamines.

Synth. Commun. 1987, 17, 669-76.

(45) Weinstock, M.; Razin, M.; Chorev, M.; Tashma, Z. Pharmacologi-

cal Activity of Novel Anticholinesterase Agents of Potential Use

in Treatment of Alzheimer’s Disease. J . Neural Transm. 1994,

s43, 219-225.

(62) Maycock, A. L.; Abeles, R. H.; Salch, J . I.; Singer, T. P. The Action

of Acetylenic Inhibitors on Mitochondrial Monoamine Oxidase:

Structure of the Flavin Site in the Inhibited Enzyme. In

Monoamine Oxidase and its Inhibition; Wolstenholme, G. E. W.,

Knight, J ., Eds.; Elsevier: Amsterdam, 1976; pp 33-47.

(63) Rudenko, V. A.; Yakubovitsch, A. Y.; Nikiforova, T. Y. J . Gen.

Chem. USSR (Engl. Transl.) 1947, 17, 2256.

Novel Dual Inhibitors of AChE and MAO

J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5279

(64) McGhee, W. D.; Pan, Y.; Talley, J . J . Conversion of Amines to

Carbamoyl Chlorides Using Carbon Dioxide as a Phosgene

Replacement. Tetrahedron Lett. 1994, 35, 839-842.

(65) Lieber, E.; Rao, C. N. R.; Layer, C. B.; Trivedi, J . P. 5-(Disub-

stituted) Amino-1,2,3,4-Thiatriazoles. Can. J . Chem. 1963, 41,

1643-1644.

(66) Lidor, R.; Bahar, E.; Zairi, O.; Atili, G.; Amster, D. A Facile

Synthesis for Racemic and Optically Active 1-Aminoindans. Org.

Prep. Proced. Int. 1997, 29, 701-706.

(67) Ellman, G. L.; Courtney, K. D.; Anders, F.; Featherstone, R. M.

A New and Rapid Colorimetric Determination of Acetylcho-

linesterase Activity. Biochem. Pharmacol. 1961, 7, 88-95.

(68) Tipton, K. F.; Youdim, M. B. H. The Assay of Monoamine

Oxidase Activity. In Methodes in Biogenic Amine Research;

Parvez, S., Nagatsu, T., Nagatsu, I.; Parvez, H., Eds.; Elsevier:

Amsterdam, 1983; pp 441-467.

J M020120C

Article Doi

Novel dual inhibitors of AChE and MAO derived from hydroxy aminoindan and phenethylamine as potential treatment for Alzheimer's disease

DOI: 10.1021/jm020120c

Source and publish data:

Authors:

Article abstract of DOI:10.1021/jm020120c

Full text of DOI:10.1021/jm020120c

Products guided by the article

R&D Labs maybe for 479206-14-9

Relevant to this article

Hot Product