8/3/2019 Anne Blow et al- Synthesis of Cocaine Analogues by Multicomponent Grignard Reactions

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Synthesis of Cocaine Analogues by MulticomponentGrignard Reactions

Anne Blow and Mikael Bols, Department of Chemistry, University of Aarhus, Langelandsgade 140, DK-8000 Aarhus, DenmarkSteffen Sinning and Ove Wiborg, Lab. of Molecular Neurobiology, Psychiatric Hospital in Aarhus, Skovagervej 2, DK-8240 Risskov, Denmark

IntroductionCocaine abuse is a growing problem in the US and in Western Europe, but at present nomedical therapy is available for the treatment of cocaine addiction. It is suggested that it is

possible to design cocaine analogues that could be useful in abuse treatment.1

This hasresulted in synthesis of numerous cocaine analogues though yet without the desired biologicaleffect. 2 Combinatorial chemistry is an efficient way to generate many compounds in fewreactions. Even though very useful in drug discovery combinatorial chemistry has not yetbeen used for synthesis of cocaine analogues. In the present study small libraries of cocaineanalogues were synthesized by multicomponent Grignard reactions followed by 2 dimensionalscreening for biological activity .

Cocaine is a stimulant of the central nervous system. When a nerve terminal in the restingstate (A) is stimulated (B), dopamine ( ) is released and diffuses across the synaptic cleftwhere a dopamine receptor ( ) is stimulated. The stimulating action of dopamine ends with itsreuptake by dopamine transporters ( ), into the presynaptic neuron (C). Cocaine ( ) binds to and blocks the transporter function, flooding thesynapse with excess dopamine (D), which prolongs the signalling at key brain synapses. Though, a potential drug candidate is desired to inhibitcocaine binding without affecting the normal dopamine reuptake.

ResultsThe multicomponent Grignard reactions were carried out on R -anhydroecgonine methyl ester ( 1),which was prepared from R -cocaine in three steps. First the ester functionalities werehydrolysed by aqueous HCl followed by dehydration with POCl 3. The last step was esterificationusing MeOH and acid. 3 Grignard reagents were freshly prepared from 5 alkyl- or aryl bromidesbefore addition of 2 equivalents to the ,-unsaturated ester 1. By quenching the reaction at-78 C using anhydrous TFA we mainly obtained the desired products 2 and only small amounts ofthe epimer with a 2-equatorial carbomethoxy group.

In this way 25 different cocaine analogues were synthesized in libraries (1-10) containing 5compounds each according to the matrix to the right (Each library contain the compounds of arow or column). 24 of the compounds are new but the last compound (WIN35065-2) is known tobind to the dopamine transporter and included in the library to have a positive control in thescreening procedure. The libraries were synthesized in two dimensions both horizontally(Library 1-5 ) and vertically ( Library 6-10 ) in order to facilitate identification of a possible leadcompound, since such a compound must be active in both dimensions.

The 10 libraries were screened for binding affinity to the humandopamine, serotonin, and norepinephrine transporters and formonoamine reuptake. Here only results concerning the dopamine

transporter will be presented. In the diagrams to the left a high columnindicates high affinity (low K i) and a low column low affinity (high K i). Asseen in the diagrams, the compounds that bind to the dopaminetransporter do also inhibit dopamine reuptake. The positive controlcame up and two new high affinity compounds were identified. Theseand three other compounds were resynthesized individually andevaluated biologically (see below).

The results show that an aromatic group in the 3 position isnecessary to obtain high binding affinity. If an alkyl group is presentin that position the affinity decreases dramatically. But thearomatic group can be too bulky, since the presence of a t -butylphenyl substituent totally destroys binding and uptake affinity.

Conclusion25 cocaine analogues were synthesized using solution-phasecombinatorial chemistry. This was done by a 1,4-conjugate additionof a multicomponent Grignard reagent to an ,-unsaturated ester.By screening for biological activity in 2 dimensions two new highaffinity compounds were identified and resynthesized.

References1. Carroll, F.I.; Lewin, A.H.; Boja, J.W.; Kuhar, M.J. J. Med. Chem . 1992 , 35 , 969-9812. Singh, S. Chem. Rev. 2000 , 100 , 925-10243. Zheng, Q-H.; Mulholland, G.K. Nucl. Med. Biol. 1996 , 23 , 981-986

Library 6 Library 7 Library 8 Library 9 Library 10

L i b r a r y

5 N O OCH3

N

OOCH 3

NO

OCH 3 N

OOCH 3

N

OOCH 3

L i b r a r y

4 N O OCH 3

N

OOCH 3

N

OOCH3

NO

OCH3

NO

OCH3

L i b r a r y

3

NO

OCH 3 N O OCH 3

N

OOCH3

N O OCH3

N O OCH 3

L i b r a r y

2N

OOCH 3

NO

OCH 3 N

OOCH 3

N

OOCH 3

N

OOCH 3

L i b r a r y

1N

OOCH3

Positivecontrol

NO

OCH 3

N

OOCH 3

N

OOCH3

High affinity N

OOCH 3

High affinity

N

O

OOCH 3

R -cocaine

NO

OCH 3

WIN 35065-2

N

R

OOCH 3

O

Analogues

Postsynaptic Neuron

B. StimulatedC. Normal dopamine reuptake

Presynaptic Neuron

A. Rest state

D. Cocaine inhibition of reuptake

N

O

OOCH 3

O

N

OH

OOH N

OOH N

OOCH 3

1M aq HCl

reflux 15h

POCl 3 MeOH

reflux 1h rt 2h

N

R

OOCH 31. RMgBr, Et 2O,

4h -40 oC

2. TFA in Et 2O, -78oC

R -cocaine 1 2

L i b r

a r y 6

L i b r

a r y 7

L i b r

a r y 8

L i b r

a r y 9

L i b r

a r y 1 0

Library1

Library2Library3

Library4Library5

0

100

200

300

400

500

600

700

1 0 0 0 0 0 / K i ( n M )

Dopamine reuptake

L i b r

a r y 6

L i b r

a r y 7

L i b r

a r y 8

L i b r

a r y 9

L i b r

a r y 1

0

Library1

Library2Library3

Library4Library5

0

100

200

300

400

500

600

700

1 0 0 0 0 0 / K i ( n M )

Dopamine transporter binding

NO

OCH 3N

OOCH 3

Ki (uptake)Ki (binding)

20 nM25 nM

Ki (uptake)Ki (binding)

85 nM140 nM

NO

OCH 3

Ki (uptake)Ki (binding)

13500 nM39000 nM

NO

OCH 3

Ki (uptake)Ki (binding)

2300 nM6700 nM

NO

OCH 3

Ki (uptake)Ki (binding)

5300 nM18000 nM