U.S. patent application number 12/755799 was filed with the patent office on 2010-08-05 for heteroaryl-substituted diazatricycloalkanes and methods of use thereof.
This patent application is currently assigned to Targacept, Inc.. Invention is credited to Jozef Klucik, Anatoly Mazurov, Lan Miao.
Application Number | 20100197720 12/755799 |
Document ID | / |
Family ID | 37508012 |
Filed Date | 2010-08-05 |
United States Patent
Application |
20100197720 |
Kind Code |
A1 |
Mazurov; Anatoly ; et
al. |
August 5, 2010 |
HETEROARYL-SUBSTITUTED DIAZATRICYCLOALKANES AND METHODS OF USE
THEREOF
Abstract
The present invention relates to amide and urea derivatives of
heteroaryl-substituted diazatricycloalkanes, pharmaceutical
compositions including the compounds, methods of preparing the
compounds, and methods of treatment using the compounds. More
specifically, the methods of treatment involve modulating the
activity of the .alpha.7 nAChR subtype by administering one or more
of the compounds to treat or prevent disorders mediated by the
.alpha.7 nAChR subtype. The diazatricycloalkanes typically consist
of a 1-azabicyclooctane fused to pyrrolidine ring. The substituent
heteroaryl groups are 5- or 6-membered ring heteroaromatics, such
as 3-pyridinyl and 5-pyrimidinyl moieties, which are attached
directly to the diazatricycloalkane. The secondary nitrogen of the
pyrrolidine moiety is substituted with an arylcarbonyl (amide type
derivative) or an arylaminocarbonyl (N-arylcarbamoyl) (urea type
derivative) group. The compounds are beneficial in therapeutic
applications requiring a selective interaction at certain nAChR
subtypes. That is, the compounds modulate the activity of certain
nAChR subtypes, particularly the .alpha.7 nAChR subtype, and do not
have appreciable activity toward muscarinic receptors. Radiolabeled
versions of the compounds can be used in diagnostic methods.
Inventors: |
Mazurov; Anatoly;
(Greensboro, NC) ; Miao; Lan; (Advance, NC)
; Klucik; Jozef; (Marietta, GA) |
Correspondence
Address: |
WOMBLE CARLYLE SANDRIDGE & RICE, PLLC
ATTN: PATENT DOCKETING, P.O. BOX 7037
ATLANTA
GA
30357-0037
US
|
Assignee: |
Targacept, Inc.
Winston-Salem
NC
|
Family ID: |
37508012 |
Appl. No.: |
12/755799 |
Filed: |
April 7, 2010 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
11465914 |
Aug 21, 2006 |
7732607 |
|
|
12755799 |
|
|
|
|
60710130 |
Aug 22, 2005 |
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Current U.S.
Class: |
514/292 ;
546/84 |
Current CPC
Class: |
A61P 25/28 20180101;
A61P 25/18 20180101; A61P 29/00 20180101; C07D 471/14 20130101;
A61P 25/04 20180101; A61P 25/22 20180101; A61P 9/00 20180101; A61P
35/00 20180101; A61P 31/04 20180101; C07D 471/18 20130101; A61P
25/14 20180101; A61P 25/16 20180101; A61P 25/24 20180101; A61P
25/00 20180101 |
Class at
Publication: |
514/292 ;
546/84 |
International
Class: |
A61K 31/439 20060101
A61K031/439; C07D 471/18 20060101 C07D471/18 |
Claims
1. A compound having a structure of the formula: ##STR00004##
wherein: Y is either oxygen or sulfur, Z is either NR' or a
covalent bond, A is either absent or a linker species selected from
the group consisting of --CR'R''--, --CR'R''--CR'R''--,
--CR'.dbd.CR'--, and --C.ident.C--, and wherein R' and R'' are as
hereinafter defined, Ar is a phenyl or a napthyl; and Cy is a
pyridine, wherein, individually, Ar, Cy, and the various open
valencies positions on the depicted
1,5-diazatricyclo[5.2.2.0<2,6>]undecane ring, can be
unsubstituted or can be substituted with one or more substituents
selected from the group consisting of alkyl, alkenyl, heterocyclyl,
cycloalkyl, aryl, substituted aryl, arylalkyl, substituted
arylalkyl, halo, --OR', --NR'R'', --CF.sub.3, --CN, --NO.sub.2,
--C.ident.CR'--, --SR', --N.sub.3, --C(.dbd.O)NR'R'',
--NR'C(.dbd.O)--R'', --C(.dbd.O)R', --C(.dbd.O)OR', --OC(.dbd.O)R',
--O(CR'R'').sub.rC(.dbd.O)R', --O(CR'R'').sub.rNR''C(.dbd.O)R',
--O(CR'R'').sub.rNR''SO.sub.2R', --OC(.dbd.O)NR'R'',
--NR'C(.dbd.O)O--R'', --SO.sub.2R', --SO.sub.2NR'R'', and
--NR'SO.sub.2R'', where each R' and R'' are individually hydrogen,
alkyl C.sub.1-C.sub.8 alkyl, cycloalkyl, heterocyclyl, aryl, or
arylalkyl, and r is an integer from 1 to 6, or R' and R'' combine
to form a cyclic functionality, or a radiolabeled version thereof,
or a pharmaceutically acceptable salt thereof.
2. The compound of claim 1 wherein Cy is 3-pyridinyl.
3. The compound of claim 1, wherein Y is O, and Z is NR'.
4. The compound of claim 1, wherein Y is O and Z is a covalent
bond.
5. A compound selected from the group consisting of:
5-benzoyl-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]undecane,
5-(2-fluorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane,
5-(3-fluorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane,
5-(4-fluorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane,
5-(2-chlorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane,
5-(3-chlorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane,
5-(4-chlorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane,
5-(2-bromobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane,
5-(3-bromobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6-
>]undecane,
5-(4-bromobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane,
5-(2-iodobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&-
gt;]undecane,
5-(3-iodobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]und-
ecane,
5-(4-iodobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane,
5-(2-methylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane,
5-(3-methylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane,
5-(4-methylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane,
5-(2-methoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
2,6>]undecane,
5-(3-methoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane,
5-(4-methoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
;2,6>]undecane,
5-(2-methylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane,
5-(3-methylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane,
5-(4-methylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane,
5-(2-phenylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane,
5-(3-phenylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane,
5-(4-phenylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane,
5-(2-phenoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
2,6>]undecane,
5-(3-phenoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane,
5-(4-phenoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
;2,6>]undecane,
5-(2-phenylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane,
5-(3-phenylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane,
5-(4-phenylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane,
5-(2-cyanobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane,
5-(3-cyanobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6-
>]undecane,
5-(4-cyanobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane,
5-(2-trifluoromethylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane,
5-(3-trifluoromethylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
2,6>]undecane,
5-(4-trifluoromethylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
2,6>]undecane,
5-(2-dimethylaminobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,-
6>]undecane,
5-(3-dimethylaminobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,-
6>]undecane,
5-(4-dimethylaminobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,-
6>]undecane,
5-(2-ethynylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane,
5-(3-ethynylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
;2,6>]undecane,
5-(4-ethynylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane,
5-(3,4-dichlorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(2,4-dimethoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&-
gt;]undecane,
5-(3,4,5-trimethoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane,
5-(naphth-1-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, and
5-(naphth-2-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, or a pharmaceutically acceptable salt thereof.
6. A compound selected from the group consisting of:
5-(phenylacetyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]unde-
cane,
5-(diphenylacetyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane,
5-(2-phenylpropanoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>-
]undecane, and
5-(3-phenylprop-2-enoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&-
gt;]undecane, or a pharmaceutically acceptable salt thereof.
7. A compound selected from the group consisting of
5-N-phenylcarbamoyl-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane,
5-(N-(2-fluorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[-
5.2.2.0<2,6>]undecane,
5-(N-(3-fluorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(4-fluorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(2-chlorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(3-chlorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(4-chlorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(2-bromophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane,
5-(N-(3-bromophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane,
5-(N-(4-bromophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane,
5-(N-(2-iodophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
;2,6>]undecane,
5-(N-(3-iodophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
;2,6>]undecane,
5-(N-(4-iodophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
;2,6>]undecane,
5-(N-(2-methylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(3-methylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(4-methylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(2-methoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(N-(3-methoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(N-(4-methoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(N-(2-methylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane,
5-(N-(3-methylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane,
5-(N-(4-methylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane,
5-(N-(2-phenylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(3-phenylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(4-phenylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane,
5-(N-(2-phenoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(N-(3-phenoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(N-(4-phenoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(N-(2-phenylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane,
5-(N-(3-phenylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane,
5-(N-(4-phenylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane,
5-(N-(2-cyanophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane,
5-(N-(3-cyanophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane,
5-(N-(4-cyanophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane,
5-(N-(2-trifluoromethylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo-
[5.2.2.0<2,6>]undecane,
5-(N-(3-trifluoromethylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo-
[5.2.2.0<2,6>]undecane,
5-(N-(4-trifluoromethylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo-
[5.2.2.0<2,6>]undecane,
5-(N-(2-dimethylaminophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5-
.2.2.0<2,6>]undecane,
5-(N-(3-dimethylaminophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5-
.2.2.0<2,6>]undecane,
5-(N-(4-dimethylaminophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5-
.2.2.0<2,6>]undecane,
5-(N-(2-ethynylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(N-(3-ethynylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(N-(4-ethynylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(N-(3,4-dichlorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane,
5-(N-(2,4-dimethoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2-
.2.0<2,6>]undecane,
5-(N-(3,4,5-trimethoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[-
5.2.2.0<2,6>]undecane,
5-(N-(1-naphthyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane, and
5-(N-(2-naphthyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane, or a pharmaceutically acceptable salt thereof.
8. A compound selected from the group consisting of
5-(N-benzylcarbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>-
]undecane,
5-(N-(4-bromobenzyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo-
[5.2.2.0<2,6>]undecane,
5-(N-(4-methoxybenzyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane,
5-(N-(1-phenylethyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane, and
5-(N-(diphenylmethyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, or a pharmaceutically acceptable salt
thereof.
9. The compound of claim 1, wherein A is absent.
10. A pharmaceutical composition comprising a pharmaceutical
carrier and a compound of claim 1.
11. The compound of claim 1, wherein the compound is
radiolabeled.
12. The compound of claim 11, wherein the compound comprises
.sup.11C, .sup.18F, .sup.76Br, .sup.123I, or .sup.125I.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a divisional application of U.S. patent
application Ser. No. 11/465,914 which claims benefit of U.S.
Provisional Patent Application No. 60/710,130, the contents of
which are each fully incorporated herein by reference.
FIELD OF THE INVENTION
[0002] The present invention relates to pharmaceutical compositions
incorporating compounds capable of affecting nicotinic
acetylcholinergic receptors (nAChRs), for example, as modulators of
specific nicotinic receptor subtypes (specifically, the .alpha.7
nAChR subtype). The present invention also relates to methods for
treating a wide variety of conditions and disorders, particularly
those associated with dysfunction of the central and autonomic
nervous systems.
BACKGROUND OF THE INVENTION
[0003] Nicotine has been proposed to have a number of
pharmacological effects. See, for example, Pullan et al., N. Engl.
J. Med. 330: 811 (1994). Certain of those effects may be related to
effects upon neurotransmitter release. See, for example, Sjak-shie
et al., Brain Res. 624: 295 (1993), where neuroprotective effects
of nicotine are proposed. Release of acetylcholine and dopamine by
neurons, upon administration of nicotine, has been reported by
Rowell et al., J. Neurochem. 43: 1593 (1984); Rapier et al., J.
Neurochem. 50: 1123 (1988); Sandor et al., Brain Res. 567: 313
(1991) and Vizi, Br. J. Pharmacol. 47: 765 (1973). Release of
norepinephrine by neurons, upon administration of nicotine, has
been reported by Hall et al., Biochem. Pharmacol. 21: 1829 (1972).
Release of serotonin by neurons, upon administration of nicotine,
has been reported by Hery et al., Arch. Int. Pharmacodyn. Ther.
296: 91 (1977). Release of glutamate by neurons, upon
administration of nicotine, has been reported by Toth et al.,
Neurochem Res. 17: 265 (1992). Confirmatory reports and additional
recent studies have included the modulation, in the central nervous
system (CNS), of glutamate, nitric oxide, GABA, tachykinins,
cytokines, and peptides (reviewed in Brioni et al., Adv. Pharmacol.
37: 153 (1997)). In addition, nicotine reportedly potentiates the
pharmacological behavior of certain pharmaceutical compositions
used for the treatment of certain disorders. See, for example,
Sanberg et al., Pharmacol. Biochem. & Behavior 46: 303 (1993);
Harsing et al., J. Neurochem. 59: 48 (1993) and Hughes, Proceedings
from Intl. Symp. Nic. S40 (1994). Furthermore, various other
beneficial pharmacological effects of nicotine have been proposed.
See, for example, Decina et al., Biol. Psychiatry 28: 502 (1990);
Wagner et al., Pharmacopsychiatry 21: 301 (1988); Pomerleau et al.,
Addictive Behaviors 9: 265 (1984); Onaivi et al., Life Sci. 54(3):
193 (1994); Tripathi et al., JPET 221: 91 (1982) and Hamon, Trends
in Pharmacol. Res. 15: 36 (1994).
[0004] Various compounds that target nAChRs have been reported as
being useful for treating a wide variety of conditions and
disorders. See, for example, Williams et al., DN&P 7(4): 205
(1994); Arneric et al., CNS Drug Rev. 1(1): 1 (1995); Arneric et
al., Exp. Opin. Invest. Drugs 5(1): 79 (1996); Bencherif et al.,
JPET 279: 1413 (1996); Lippiello et al., JPET 279: 1422 (1996);
Damaj et al., J. Pharmacol. Exp. Ther. 291: 390 (1999); Chiari et
al., Anesthesiology 91: 1447 (1999); Lavand'homme and Eisenbach,
Anesthesiology 91: 1455 (1999); Holladay et al., J. Med. Chem.
40(28): 4169 (1997); Bannon et al., Science 279: 77 (1998); PCT WO
94/08992, PCT WO 96/31475, PCT WO 96/40682, and U.S. Pat. Nos.
5,583,140 to Bencherif et al., 5,597,919 to Dull et al., 5,604,231
to Smith et al., and 5,852,041 to Cosford et al. Nicotinic
compounds are reported as being particularly useful for treating a
wide variety of CNS disorders. Indeed, a wide variety of compounds
have been reported to have therapeutic properties. See, for
example, Bencherif and Schmitt, Current Drug Targets: CNS and
Neurological Disorders 1(4): 349 (2002); Levin and Rezvani, Current
Drug Targets: CNS and Neurological Disorders 1(4): 423 (2002);
O'Neill et al., Current Drug Targets: CNS and Neurological
Disorders 1(4): 399 (2002); U.S. Pat. Nos. 5,1871,166 to Kikuchi et
al., 5,672,601 to Cignarella, PCT WO 99/21834, and PCT WO 97/40049,
UK Patent Application GB 2295387, and European Patent Application
297,858.
[0005] CNS disorders are a type of neurological disorder. CNS
disorders can be drug induced; can be attributed to genetic
predisposition, infection or trauma; or can be of unknown etiology.
CNS disorders comprise neuropsychiatric disorders, neurological
diseases and mental illnesses, and include neurodegenerative
diseases, behavioral disorders, cognitive disorders and cognitive
affective disorders. There are several CNS disorders whose clinical
manifestations have been attributed to CNS dysfunction (i.e.,
disorders resulting from inappropriate levels of neurotransmitter
release, inappropriate properties of neurotransmitter receptors,
and/or inappropriate interaction between neurotransmitters and
neurotransmitter receptors). Several CNS disorders can be
attributed to a deficiency of choline, dopamine, norepinephrine
and/or serotonin. Relatively common CNS disorders include
pre-senile dementia (early-onset Alzheimer's disease), senile
dementia (dementia of the Alzheimer's type), micro-infarct
dementia, AIDS-related dementia, Creutzfeldt-Jakob disease, Pick's
disease, Parkinsonism including Parkinson's disease, Lewy body
dementia, progressive supranuclear palsy, Huntington's chorea,
tardive dyskinesia, hyperkinesia, mania, attention deficit
disorder, anxiety, dyslexia, schizophrenia, depression,
obsessive-compulsive disorders and Tourette's syndrome.
[0006] The nAChRs characteristic of the CNS have been shown to
occur in several subtypes, the most common of which are the
.alpha.4.beta.2 and .alpha.7 subtypes. See, for example, Schmitt,
Current Med. Chem. 7: 749 (2000). Ligands that interact with the
.alpha.7 nAChR subtype have been proposed to be useful in the
treatment of schizophrenia. There are a decreased number of
hippocampal nAChRs in postmortem brain tissue of schizophrenic
patients. Also, there is improved psychological effect in smoking
versus non-smoking schizophrenic patients. Nicotine improves
sensory gating deficits in animals and schizophrenics. Blockade of
the .alpha.7 nAChR subtype induces a gating deficit similar to that
seen in schizophrenia. See, for example, Leonard et al.,
Schizophrenia Bulletin 22(3): 431 (1996). Biochemical, molecular,
and genetic studies of sensory processing, in patients with the P50
auditory-evoked potential gating deficit, suggest that the .alpha.7
nAChR subtype may function in an inhibitory neuronal pathway. See,
for example, Freedman et al., Biological Psychiatry 38(1): 22
(1995).
[0007] More recently, .alpha.7 nAChRs have been proposed to be
mediators of angiogenesis, as described by Heeschen et al., J.
Clin. Invest. 100: 527 (2002). In these studies, inhibition of the
.alpha.7 subtype was shown to decrease inflammatory angiogenesis.
Also, .alpha.7 nAChRs have been proposed as targets for controlling
neurogenesis and tumor growth (Utsugisawa et al., Molecular Brain
Research 106(1-2): 88 (2002) and U.S. Patent Application
2002/0016371). Finally, the role of the .alpha.7 subtype in
cognition (Levin and Rezvani, Current Drug Targets: CNS and
Neurological Disorders 1(4): 423 (2002)), neuroprotection (O'Neill
et al., Current Drug Targets: CNS and Neurological Disorders 1(4):
399 (2002) and Jeyarasasingam et al., Neuroscience 109(2): 275
(2002)), and neuropathic pain (Xiao et al., Proc. Nat. Acad. Sci.
(US) 99(12): 8360 (2002)) has recently been recognized.
[0008] Various compounds have been reported to interact with
.alpha.7 nAChRs and have been proposed as therapies on that basis.
See, for instance, PCT WO 99/62505, PCT WO 99/03859, PCT WO
97/30998, PCT WO 01/36417, PCT WO 02/15662, PCT WO 02/16355, PCT WO
02/16356, PCT WO 02/16357, PCT WO 02/16358, PCT WO 02/17358,
Stevens et al., Psychopharm. 136: 320 (1998), Dolle et al., J.
Labeled Comp. Radiopharm. 44: 785 (2001) and Macor et al., Bioorg.
Med. Chem. Lett. 11: 319 (2001) and references therein. Among these
compounds, a common structural theme is that of the substituted
tertiary bicyclic amine (e.g., quinuclidine). Similar substituted
quinuclidine compounds have also been reported to bind at
muscarinic receptors. See, for instance, U.S. Pat. No. 5,712,270 to
Sabb and PCTs WO 02/00652 and WO 02/051841.
[0009] It would be desirable to provide a useful method for the
prevention and treatment of a condition or disorder by
administering a nicotinic compound to a patient susceptible to or
suffering from such a condition or disorder. It would be highly
beneficial to provide individuals suffering from certain disorders
(e.g., CNS diseases) with interruption of the symptoms of those
disorders by the administration of a pharmaceutical composition
containing an active ingredient having nicotinic pharmacology which
has a beneficial effect (e.g., upon the functioning of the CNS),
but does not provide any significant associated side effects. It
would be highly desirable to provide a pharmaceutical composition
incorporating a compound that interacts with nAChRs, such as those
that have the potential to affect the functioning of the CNS. It
would be highly desirable that such a compound, when employed in an
amount sufficient to affect the functioning of the CNS, would not
significantly affect those nAChR subtypes that have the potential
to induce undesirable side effects (e.g., appreciable activity at
cardiovascular and skeletal muscle receptor sites). In addition, it
would be highly desirable to provide a pharmaceutical composition
incorporating a compound which interacts with nicotinic receptors
but not muscarinic receptors, as the latter are associated with
side effects, such as hypersalivation, sweating, tremors,
cardiovascular and gastrointestinal disturbances, related to the
function of the parasympathetic nervous system (see Caulfield,
Pharmacol. Ther. 58: 319 (1993) and Broadley and Kelly, Molecules
6: 142 (2001)). Furthermore, it would be highly desirable to
provide pharmaceutical compositions, which are selective for the
.alpha.7 nAChR subtype, for the treatment of certain conditions or
disorders (e.g., schizophrenia, cognitive disorders, and
neuropathic pain) and for the prevention of tissue damage and the
hastening of healing (i.e., for neuroprotection and the control of
angiogenesis). The present invention provides such compounds,
compositions and methods.
SUMMARY OF THE INVENTION
[0010] The present invention relates to amide and urea derivatives
of heteroaryl-substituted diazatricycloalkanes, pharmaceutical
compositions including the compounds, methods of preparing the
compounds, and methods of treatment using the compounds. More
specifically, the methods of treatment involve modulating the
activity of the .alpha.7 nAChR subtype by administering one or more
of the compounds to treat or prevent disorders mediated by the
.alpha.7 nAChR subtype.
[0011] The diazatricycloalkanes typically consist of
1-azabicyclooctane fused to pyrrolidine ring. The substituent
heteroaryl groups are 5- or 6-membered ring heteroaromatics, such
as 3-pyridinyl and 5-pyrimidinyl moieties, which are attached
directly to the diazatricycloalkane. The secondary nitrogen of the
pyrrolidine moiety is substituted with an arylcarbonyl (amide type
derivative) or an arylaminocarbonyl (N-arylcarbamoyl) (urea type
derivative) group.
[0012] The compounds are beneficial in therapeutic applications
requiring a selective interaction at certain nAChR subtypes. That
is, the compounds modulate the activity of certain nAChR subtypes,
particularly the .alpha.7 nAChR subtype, and do not have
appreciable activity toward muscarinic receptors. The compounds can
be administered in amounts sufficient to affect the functioning of
the central nervous system (CNS) without significantly affecting
those receptor subtypes that have the potential to induce
undesirable side effects (e.g., without appreciable activity at
ganglionic and skeletal muscle nAChR sites and at muscarinic
receptors). The compounds are therefore useful towards modulating
release of ligands involved in neurotransmission, without
appreciable side effects.
[0013] The compounds can be used as therapeutic agents to treat
and/or prevent disorders characterized by an alteration in normal
neurotransmitter release. Examples of such disorders include
certain CNS conditions and disorders. The compounds can provide
neuroprotection, treat patients susceptible to convulsions, treat
depression, autism, and certain neuroendocrine disorders, and help
manage stroke patients. The compounds also are useful in treating
hypertension, type II diabetes and neoplasia and effecting weight
loss. As the compounds are selective for the .alpha.7 nAChR
subtype, they can be used to treat certain conditions or disorders
(e.g., schizophrenia, cognitive disorders, and neuropathic pain),
prevent tissue damage, and hasten healing (i.e., provide
neuroprotection and control of angiogenesis).
[0014] The pharmaceutical compositions provide therapeutic benefit
to individuals suffering from such conditions or disorders and
exhibiting clinical manifestations of such conditions or disorders.
The compounds, administered with the pharmaceutical compositions,
can be employed in effective amounts to (i) exhibit nicotinic
pharmacology and affect relevant nAChR sites (e.g., act as
pharmacological agonists at nicotinic receptors), and (ii) modulate
neurotransmitter secretion, and hence prevent and suppress the
symptoms associated with those diseases. In addition, the compounds
have the potential to (i) increase the number of nAChRs of the
brain of the patient, (ii) exhibit neuroprotective effects and
(iii) when employed in effective amounts, not cause appreciable
adverse side effects (e.g., significant increases in blood pressure
and heart rate, significant negative effects upon the
gastro-intestinal tract, and significant effects upon skeletal
muscle). The pharmaceutical compositions are believed to be safe
and effective with regards to prevention and treatment of various
conditions or disorders.
[0015] The foregoing and other aspects of the present invention are
explained in detail in the detailed description and examples set
forth below.
DETAILED DESCRIPTION OF THE INVENTION
[0016] The compounds described herein have structures that are
represented by Formula 1.
##STR00001##
[0017] In Formula 1, Y is either oxygen or sulfur, and Z is either
nitrogen (i.e., NR') or a covalent bond. A is either absent or a
linker species selected from the group --CR'R''--,
--CR'R''--CR'R''--, --CR'.dbd.CR'--, and --C.sub.2--, wherein R'
and R'' are as hereinafter defined. Ar is an aryl group, either
carbocyclic or heterocyclic, either monocyclic or fused polycyclic,
unsubstituted or substituted; and Cy is a 5- or 6-membered
heteroaromatic ring, unsubstituted or substituted. The junction
between the azacycle and the azabicycle can be characterized by any
of the various relative and absolute stereochemical configurations
at the junction sites (e.g., cis or trans, R or S). The invention
further includes pharmaceutically acceptable salts thereof. The
compounds have one or more asymmetric carbons and can therefore
exist in the form of racemic mixtures, enantiomers and
diastereomers. In addition, some of the compounds exist as E and Z
isomers about a carbon-carbon double bond. All these individual
isomeric compounds and their mixtures are also intended to be
within the scope of the present invention.
[0018] Thus, the invention includes compounds in which Ar is linked
to the diazatricycle, at the nitrogen of the pyrrolidine ring, by a
carbonyl group-containing functionality, forming an amide or a urea
functionality. Ar may be bonded directly to the carbonyl
group-containing functionality or may be linked to the carbonyl
group-containing functionality through linker A. Furthermore, the
invention includes compounds that contain a diazatricycle,
containing a 1-azabicyclo[2.2.2]octane.
[0019] As used herein, "alkoxy" includes alkyl groups from 1 to 8
carbon atoms in a straight or branched chain, also C.sub.3-8
cycloalkyl, bonded to an oxygen atom.
[0020] As used herein, "alkyl" includes straight chain and branched
C.sub.1-8 alkyl, preferably C.sub.1-6 alkyl. "Substituted alkyl"
defines alkyl substituents with 1-3 substituents as defined below
in connection with Ar and Cy.
[0021] As used herein, "arylalkyl" refers to moieties, such as
benzyl, wherein an aromatic is linked to an alkyl group that is
linked to the indicated position in the compound of Formulas 1 or
2. "Substituted arylalkyl" defines arylalkyl substituents with 1-3
substituents as defined below in connection with Ar and Cy.
[0022] As used herein, "aromatic" refers to 3- to 10-membered,
preferably 5- and 6-membered, aromatic and heteroaromatic rings and
polycyclic aromatics including 5- and/or 6-membered aromatic and/or
heteroaromatic rings.
[0023] As used herein, "aryl" includes both carbocyclic and
heterocyclic aromatic rings, both monocyclic and fused polycyclic,
where the aromatic rings can be 5- or 6-membered rings.
Representative monocyclic aryl groups include, but are not limited
to, phenyl, furanyl, pyrrolyl, thienyl, pyridinyl, pyrimidinyl,
oxazolyl, isoxazolyl, pyrazolyl, imidazolyl, thiazolyl,
isothiazolyl and the like. Fused polycyclic aryl groups are those
aromatic groups that include a 5- or 6-membered aromatic or
heteroaromatic ring as one or more rings in a fused ring system.
Representative fused polycyclic aryl groups include naphthalene,
anthracene, indolizine, indole, isoindole, benzofuran,
benzothiophene, indazole, benzimidazole, benzthiazole, purine,
quinoline, isoquinoline, cinnoline, phthalazine, quinazoline,
quinoxaline, 1,8-naphthyridine, pteridine, carbazole, acridine,
phenazine, phenothiazine, phenoxazine, and azulene.
[0024] As used herein, a "carbonyl group-containing functionality"
is a moiety of the formula --C(.dbd.Y)--Z--, where Y are Z are as
defined herein.
[0025] As used herein, "Cy" groups are 5- and 6-membered ring
heteroaromatic groups. Representative Cy groups include pyridinyl,
pyrimidinyl, furanyl, pyrrolyl, thienyl, oxazolyl, isoxazolyl,
pyrazolyl, imidazolyl, thiazolyl, isothiazolyl and the like.
[0026] Individually, Ar and Cy, as well as the various positions on
the 1,5-diazatricyclo[5.2.2.0<2,6>]undecane ring, can be
unsubstituted or can be substituted with 1, 2 or 3 substituents,
such as alkyl, alkenyl, heterocyclyl, cycloalkyl, aryl, substituted
aryl, arylalkyl, substituted arylalkyl, halo (e.g., F, Cl, Br, or
I), --OR', --NR'R'', --CF.sub.3, --CN, --NO.sub.2, --C.sub.2R',
--SR', --N.sub.3, --C(.dbd.O)NR'R'', --NR'C(.dbd.O)--R'',
--C(.dbd.O)R', --C(.dbd.O)OR', --OC(.dbd.O)R',
--O(CR'R'').sub.rC(.dbd.O)R', --O(CR'R'').sub.rNR''C(.dbd.O)R',
--O(CR'R'').sub.rNR''SO.sub.2R', --OC(.dbd.O)NR'R'',
--NR'C(.dbd.O)O--R'', --SO.sub.2R', --SO.sub.2NR'R'', and
--NR'SO.sub.2R'', where R' and R'' are individually hydrogen, lower
alkyl (e.g., straight chain or branched alkyl including
C.sub.1-C.sub.8, preferably C.sub.1-C.sub.5, such as methyl, ethyl,
or isopropyl), cycloalkyl, heterocyclyl, aryl, or arylalkyl (such
as benzyl), and r is an integer from 1 to 6. R' and R'' can also
combine to form a cyclic functionality.
[0027] As used herein, cycloalkyl radicals contain from 3 to 8
carbon atoms. Examples of suitable cycloalkyl radicals include, but
are not limited to, cyclopropyl, cyclobutyl, cyclopentyl,
cyclohexyl, cycloheptyl and cyclooctyl. As used herein,
polycycloalkyl radicals are selected from adamantyl, bornanyl,
norbornanyl, bornenyl and norbornenyl.
[0028] As used herein, halogen is chlorine, iodine, fluorine or
bromine.
[0029] As used herein, heteroaryl radicals are rings that contain
from 3 to 10 members, preferably 5 or 6 members, including one or
more heteroatoms selected from oxygen, sulfur and nitrogen.
Examples of suitable 5-membered ring heteroaryl moieties include
furyl, pyrrolyl, imidazolyl, oxazolyl, thiazolyl, thienyl,
tetrazolyl, and pyrazolyl. Examples of suitable 6-membered ring
heteroaryl moieties include pyridinyl, pyrimidinyl, pyrazinyl, of
which pyridinyl and pyrimidinyl are preferred.
[0030] As used herein, "heterocyclic" or "heterocyclyl" radicals
include rings with 3 to 10 members, including one or more
heteroatoms selected from oxygen, sulfur and nitrogen. Examples of
suitable heterocyclic moieties include, but are not limited to,
piperidinyl, morpholinyl, pyrrolidinyl, imidazolidinyl,
pyrazolidinyl, isothiazolidinyl, thiazolidinyl, isoxazolidinyl,
oxazolidinyl, piperazinyl, tetrahydropyranyl and
tetrahydrofuranyl.
[0031] Examples of suitable pharmaceutically acceptable salts
include inorganic acid addition salts such as chloride, bromide,
sulfate, phosphate, and nitrate; organic acid addition salts such
as acetate, galactarate, propionate, succinate, lactate, glycolate,
malate, tartrate, citrate, maleate, fumarate, methanesulfonate,
p-toluenesulfonate, and ascorbate; salts with acidic amino acid
such as aspartate and glutamate; alkali metal salts such as sodium
salt and potassium salt; alkaline earth metal salts such as
magnesium salt and calcium salt; ammonium salt; organic basic salts
such as trimethylamine salt, triethylamine salt, pyridine salt,
picoline salt, dicyclohexylamine salt, and
N,N'-dibenzylethylenediamine salt; and salts with basic amino acid
such as lysine salt and arginine salt. The salts may be in some
cases hydrates or ethanol solvates. Representative salts are
provided as described in U.S. Pat. Nos. 5,597,919 to Dull et al.,
5,616,716 to Dull et al. and 5,663,356 to Ruecroft et al.
[0032] As used herein, neurotransmitters whose release is modulated
(i.e., increased or decreased, depending on whether the compounds
function as agonists, partial agonists or antagonists) by the
compounds described herein include, but are not limited to,
acetylcholine, dopamine, norepinephrine, serotonin and glutamate,
and the compounds described herein function as modulators of one or
more nicotinic receptors.
[0033] As used herein, an "agonist" is a substance that stimulates
its binding partner, typically a receptor. Stimulation is defined
in the context of the particular assay, or may be apparent in the
literature from a discussion herein that makes a comparison to a
factor or substance that is accepted as an "agonist" or an
"antagonist" of the particular binding partner under substantially
similar circumstances as appreciated by those of skill in the art.
Stimulation may be defined with respect to an increase in a
particular effect or function that is induced by interaction of the
agonist or partial agonist with a binding partner and can include
allosteric effects.
[0034] As used herein, an "antagonist" is a substance that inhibits
its binding partner, typically a receptor. Inhibition is defined in
the context of the particular assay, or may be apparent in the
literature from a discussion herein that makes a comparison to a
factor or substance that is accepted as an "agonist" or an
"antagonist" of the particular binding partner under substantially
similar circumstances as appreciated by those of skill in the art.
Inhibition may be defined with respect to a decrease in a
particular effect or function that is induced by interaction of the
antagonist with a binding partner, and can include allosteric
effects.
[0035] As used herein, a "partial agonist" is a substance that
provides a level of stimulation to its binding partner that is
intermediate between that of a full or complete antagonist and an
agonist defined by any accepted standard for agonist activity. It
will be recognized that stimulation, and hence, inhibition is
defined intrinsically for any substance or category of substances
to be defined as agonists, antagonists, or partial agonists. As
used herein, "intrinsic activity", or "efficacy," relates to some
measure of biological effectiveness of the binding partner complex.
With regard to receptor pharmacology, the context in which
intrinsic activity or efficacy should be defined will depend on the
context of the binding partner (e.g. receptor/ligand) complex and
the consideration of an activity relevant to a particular
biological outcome. For example, in some circumstances, intrinsic
activity may vary depending on the particular second messenger
system involved. See Hoyer and Boddeke, Trends Pharmacol Sci.,
14(7): 270 (1993). Where such contextually specific evaluations are
relevant, and how they might be relevant in the context of the
present invention, will be apparent to one of ordinary skill in the
art.
[0036] In one embodiment, Cy is 3-pyridinyl or 5-pyrimidinyl, Y is
oxygen, Z is a covalent bond and A is absent. In another
embodiment, Cy is 3-pyridinyl or 5-pyrimidinyl, Y is oxygen, Z is
nitrogen and A is absent. In a third embodiment, Cy is 3-pyridinyl
or 5-pyrimidinyl, Y is oxygen, Z is a covalent bond, and A is a
linker species. In a fourth embodiment, Cy is 3-pyridinyl or
5-pyrimidinyl, Y is oxygen, Z is nitrogen and A is a linker
species.
[0037] Representative compounds of the present invention include:
[0038]
5-benzoyl-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]undecane,
[0039]
5-(2-fluorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,-
6>]undecane, [0040]
5-(3-fluorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0041]
5-(4-fluorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0042]
5-(2-chlorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0043]
5-(3-chlorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0044]
5-(4-chlorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0045]
5-(2-bromobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane, [0046]
5-(3-bromobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane, [0047]
5-(4-bromobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane, [0048]
5-(2-iodobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]und-
ecane, [0049]
5-(3-iodobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]und-
ecane, [0050]
5-(4-iodobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]und-
ecane, [0051]
5-(2-methylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0052]
5-(3-methylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0053]
5-(4-methylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0054]
5-(2-methoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane, [0055]
5-(3-methoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane, [0056]
5-(4-methoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane, [0057]
5-(2-methylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, [0058]
5-(3-methylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, [0059]
5-(4-methylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, [0060]
5-(2-phenylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0061]
5-(3-phenylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0062]
5-(4-phenylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]u-
ndecane, [0063]
5-(2-phenoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane, [0064]
5-(3-phenoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane, [0065]
5-(4-phenoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane, [0066]
5-(2-phenylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, [0067]
5-(3-phenylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, [0068]
5-(4-phenylthiobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, [0069]
5-(2-cyanobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane, [0070]
5-(3-cyanobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane, [0071]
5-(4-cyanobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane, [0072]
5-(2-trifluoromethylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
2,6>]undecane, [0073]
5-(3-trifluoromethylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
2,6>]undecane, [0074]
5-(4-trifluoromethylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
2,6>]undecane, [0075]
5-(2-dimethylaminobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,-
6>]undecane, [0076]
5-(3-dimethylaminobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,-
6>]undecane, [0077]
5-(4-dimethylaminobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,-
6>]undecane, [0078]
5-(2-ethynylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane, [0079]
5-(3-ethynylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane, [0080]
5-(4-ethynylbenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]-
undecane, [0081]
5-(3,4-dichlorobenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, [0082]
5-(2,4-dimethoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&-
gt;]undecane, [0083]
5-(3,4,5-trimethoxybenzoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane, [0084]
5-(naphth-1-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, [0085]
5-(naphth-2-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, [0086]
5-(thien-2-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>-
;]undecane, [0087]
5-(thien-3-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>-
;]undecane, [0088]
5-(furan-2-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>-
;]undecane, [0089]
5-(benzothien-2-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane, [0090]
5-(benzofuran-2-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane, [0091]
5-(7-methoxybenzofuran-2-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2-
.2.0<2,6>]undecane, and [0092]
5-(1H-indol-3-ylcarbonyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6-
>]undecane.
[0093] Other compounds representative of the present invention
include: [0094]
5-(phenylacetyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&g-
t;]undecane, [0095]
5-(diphenylacetyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]un-
decane, [0096]
5-(2-phenylpropanoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>-
]undecane, and [0097]
5-(3-phenylprop-2-enoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6&-
gt;]undecane.
[0098] Other compounds representative of the present invention
include: [0099]
5-N-phenylcarbamoyl-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,-
6>]undecane, [0100]
5-(N-(2-fluorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0101]
5-(N-(3-fluorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0102]
5-(N-(4-fluorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0103]
5-(N-(2-chlorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0104]
5-(N-(3-chlorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0105]
5-(N-(4-chlorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0106]
5-(N-(2-bromophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane, [0107]
5-(N-(3-bromophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane, [0108]
5-(N-(4-bromophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane, [0109]
5-(N-(2-iodophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
;2,6>]undecane, [0110]
5-(N-(3-iodophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
;2,6>]undecane, [0111]
5-(N-(4-iodophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
;2,6>]undecane, [0112]
5-(N-(2-methylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0113]
5-(N-(3-methylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0114]
5-(N-(4-methylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0115]
5-(N-(2-methoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane, [0116]
5-(N-(3-methoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane, [0117]
5-(N-(4-methoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane, [0118]
5-(N-(2-methylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane, [0119]
5-(N-(3-methylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane, [0120]
5-(N-(4-methylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane, [0121]
5-(N-(2-phenylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0122]
5-(N-(3-phenylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0123]
5-(N-(4-phenylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane, [0124]
5-(N-(2-phenoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane, [0125]
5-(N-(3-phenoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane, [0126]
5-(N-(4-phenoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane, [0127]
5-(N-(2-phenylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane, [0128]
5-(N-(3-phenylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane, [0129]
5-(N-(4-phenylthiophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane, [0130]
5-(N-(2-cyanophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane, [0131]
5-(N-(3-cyanophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane, [0132]
5-(N-(4-cyanophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane, [0133] 5-(N-(2-tri
fluoromethylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
;2,6>]undecane, [0134]
5-(N-(3-trifluoromethylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo-
[5.2.2.0<2,6>]undecane, [0135]
5-(N-(4-trifluoromethylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo-
[5.2.2.0<2,6>]undecane, [0136]
5-(N-(2-dimethylaminophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5-
.2.2.0<2,6>]undecane, [0137]
5-(N-(3-dimethylaminophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5-
.2.2.0<2,6>]undecane, [0138]
5-(N-(4-dimethylaminophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5-
.2.2.0<2,6>]undecane, [0139]
5-(N-(2-ethynylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane, [0140]
5-(N-(3-ethynylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane, [0141]
5-(N-(4-ethynylphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane, [0142]
5-(N-(3,4-dichlorophenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.-
2.0<2,6>]undecane, [0143]
5-(N-(2,4-dimethoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2-
.2.0<2,6>]undecane, [0144]
5-(N-(3,4,5-trimethoxyphenyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[-
5.2.2.0<2,6>]undecane, [0145]
5-(N-(1-naphthyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane, and [0146]
5-(N-(2-naphthyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2-
,6>]undecane.
[0147] Other compounds representative of the present invention
include: [0148]
5-(N-benzylcarbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<-
2,6>]undecane, [0149]
5-(N-(4-bromobenzyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane, [0150]
5-(N-(4-methoxybenzyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0-
<2,6>]undecane, [0151]
5-(N-(1-phenylethyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&l-
t;2,6>]undecane, and [0152]
5-(N-(diphenylmethyl)carbamoyl)-3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0&-
lt;2,6>]undecane.
[0153] In each of these compounds, a
3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]undecane moiety
has the structure, with a partial numbering scheme provided, shown
below:
##STR00002##
[0154] The nitrogen at the position indicated above as the
5-position is the nitrogen involved in the formation of the amides,
thioamides, ureas and thioureas described herein.
[0155] In each of these compounds, individual isomers thereof,
mixtures thereof, including racemic mixtures, enantiomers,
diastereomers and tautomers thereof, and the pharmaceutically
acceptable salts thereof, are intended to be within the scope of
the present invention.
I. Methods of Preparing the Compounds
[0156] The manner in which compounds of the present invention can
be prepared can vary. While other synthetic strategies will be
apparent to those of skill in the art, the compounds of Formula 1
can be made by cyclization of aldol condensation products formed
from heteroaromatic aldehydes and 1-azabicyclo[2.2.2]octan-3-one.
Thus, when 3-quinuclidinone hydrochloride is reacted with
pyridine-3-carboxaldehyde (available from Aldrich Chemical
Company), in the presence of methanolic potassium hydroxide,
2-((3-pyridinyl)methylene)-1-azabicyclo[2.2.2]octan-3-one results.
A variety of heteroaromatic aldehydes can be used in the aldol
condensation, in place of pyridine-3-carboxaldehyde (variation at
Cy). Treatment of
2-((3-pyridinyl)methylene)-1-azabicyclo[2.2.2]octan-3-one with
nitromethane and sodium methoxide results in conjugate addition of
the nitromethane anion to the enone functionality. The nitro group,
of 2-(1-(3-pyridinyl)-2-nitroethyl)-1-azabicyclo[2.2.2]octan-3-one
thus produced, is then reduced with Raney nickel to give the
corresponding amine. Under the reaction conditions (Raney nickel in
ethanol), intramolecular reductive amination then takes place,
producing
3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]undecane. This
scaffold contains a secondary nitrogen, in the pyrrolidine ring,
will react with a variety of acylating agents (e.g., acid
chlorides, acid anhydrides, active esters, and carboxylic acids in
the presence of coupling reagents), to form amide derivatives, and
isocyanates, to produce urea derivatives (variation at Z-A-AR). The
amide and urea derivatives are thus easily prepared using methods
known to those skilled in the art of organic synthesis.
Commercially unavailable isocyanates can be prepared in situ from
corresponding amines and triphosgene in the presence of
triethylamine. This chemistry can be accomplished in 96-well plate
format to make libraries of such derivatives.
[0157] In some cases, reactive groups on Cy or AR may require
protection. Methods described by Greene and Wuts, Protective Groups
in Organic Synthesis 2.sup.nd ed., Wiley--Interscience Pub. (1991)
can be used to protect and deprotect these reactive groups.
[0158] The compounds can be isolated and purified using methods
well known to those of skill in the art, including, for example,
crystallization, chromatography and/or extraction.
[0159] The compounds of general Formula 1 can be obtained in
optically pure form by separating their racemates in accordance
with the customary methods.
[0160] The compounds of general Formula 1 can optionally be
converted into addition salts with a mineral or organic acid by the
action of such an acid in an appropriate solvent, for example, an
organic solvent such as an alcohol, a ketone, an ether or a
chlorinated solvent. These salts likewise form part of the
invention.
[0161] Representative pharmaceutically acceptable salts include,
but are not limited to, benzenesulphonate, hydrobromide,
hydrochloride, citrate, ethanesulphonate, fumarate, gluconate,
iodate, maleate, isethionate, methanesulphonate,
methylenebis(.beta.-oxynaphthoate), nitrate, oxalate, palmoate,
phosphate, salicylate, succinate, sulphate, tartrate,
theophyllinacetate, p-toluenesulphonate, hemigalactarate and
galactarate salts.
[0162] Imaging Agents
[0163] Certain compounds of the present invention can be
synthesized in such a manner as to incorporate a radionuclide
useful in diagnostic imaging. Of particular interest are those
compounds that include radioactive isotopic moieties such as
.sup.11C, .sup.18F, .sup.76Br, .sup.123I, .sup.125I, and the like.
The compounds can be radiolabeled at any of a variety of positions.
For example, a radionuclide of the halogen series may be used
within an alkyl halide or aryl halide moiety or functionality,
while a radionuclide such as .sup.11C may be used with an alkyl
(e.g., methyl) moiety or functionality.
[0164] For instance, commercially available
p-(dimethylamino)benzoic acid (Aldrich) is converted, by treatment
with iodomethane in methanol, into p-(trimethylammonium)benzoate,
as described by Willstaetter and Kahn, Chem. Ber. 37: 406 (1904).
The displacement of the trimethylammonium group by fluoride has
been reported, in similar compounds, by several researchers (see,
for instance, Mach et al., J. Med. Chem. 36: 3707 (1993) and
Jalalian et al., J. Labeled Compd. Radiopharm. 43: 545 (2000)).
These nucleophilic aromatic substitution reactions are typically
carried out in dimethylsulfoxide (with or without water cosolvent),
using KF or CsF as the source of fluoride ion (when KF is used,
often Kryptofix.RTM. 222 is added). When .sup.18F.sup.- is used in
such a displacement, p-.sup.18fluorobenzoic acid results. This
carboxylic acid can be rapidly coupled to the NH group at the
5-position of a compound of the formula:
##STR00003##
[0165] where Cy is as described above, and wherein the Cy and
1,5-diazatricyclo[5.2.2.0<2,6>]undecane ring can be
functionalized with the various substituents described above, to
form the desired p-.sup.18fluorobenzamide derivative, using any of
a variety of techniques known to those skilled in the art (some of
which are described previously). The resulting compound can be used
to specifically image .alpha.7 nAChRs. The related urea compound
can be prepared by replacing the p-.sup.18fluorobenzoic acid with a
compound that includes a .sup.18fluoroalkyl or
.sup.18fluoroarylalkyl N--C(O)--O-alkyl or other activated group
with the NH group at the 5-position of the starting material
described above. Similarly, the related thiourea or thioamide
compounds can be prepared by replacing the p-.sup.18fluorobenzoic
acid with ap-.sup.18fluorothiobenzoic acid, thiobenzoic acid or
with a compound that includes an N--C(S)--O-alkyl or other
activated group with the NH group at the 5-position of the starting
material described above.
[0166] This same starting material can be readily radiolabeled by
reacting the amine group at the 5-position with an activating agent
such as ethyl chloroformate to form an N--C(O)-ethoxy group (or
other activated carbonyl compound), which in turn is reacted with a
radiolabeled aryl or arylalkyl amine (i.e., to form aryl ureas or
arylalkyl ureas, where the radiolabel is on the aryl or arylalkyl
moiety). An example of a radiolabeled aryl amine is
aniline-UL-.sup.14C, which is commercially available from
SigmaAldrich. Alternatively, a radiolabeled aryl or arylalkyl
isocyanate can be reacted with the amine at the 5-position to form
a radiolabeled urea group. For example, bromophenyl-p-isocyanate
(carbonyl .sup.14C) is commercially available from American
Radiolabeled Chemicals, Inc.
[0167] The resulting radiolabeled compounds can be purified by
semi-preparative or preparative HPLC and briefly isolated for
reconstitution.
[0168] The required amine-containing precursor compounds are
described in detail above, and the resulting radiolabeled compounds
can be used to specifically image .alpha.7 nAChRs.
II. Pharmaceutical Compositions
[0169] The compounds described herein can be incorporated into
pharmaceutical compositions and used to prevent a condition or
disorder in a subject susceptible to such a condition or disorder,
and/or to treat a subject suffering from the condition or disorder.
The pharmaceutical compositions described herein include one or
more compounds of Formula 1 and/or pharmaceutically acceptable
salts thereof. Chiral compounds can be employed as racemic mixtures
or as pure enantiomers.
[0170] The manner in which the compounds are administered can vary.
The compositions are preferably administered orally (e.g., in
liquid form within a solvent such as an aqueous or non-aqueous
liquid, or within a solid carrier). Preferred compositions for oral
administration include pills, tablets, capsules, caplets, syrups,
and solutions, including hard gelatin capsules and time-release
capsules. Compositions can be formulated in unit dose form, or in
multiple or subunit doses. Preferred compositions are in liquid or
semisolid form. Compositions including a liquid pharmaceutically
inert carrier such as water or other pharmaceutically compatible
liquids or semisolids can be used. The use of such liquids and
semisolids is well known to those of skill in the art.
[0171] The compositions can also be administered via injection,
i.e., intravenously, intramuscularly, subcutaneously,
intraperitoneally, intraarterially, intrathecally; and
intracerebroventricularly. Intravenous administration is the
preferred method of injection. Suitable carriers for injection are
well known to those of skill in the art and include 5% dextrose
solutions, saline, and phosphate-buffered saline. The compounds can
also be administered as an infusion or injection (e.g., as a
suspension or as an emulsion in a pharmaceutically acceptable
liquid or mixture of liquids).
[0172] The formulations can also be administered using other means,
for example, rectal administration. Formulations useful for rectal
administration, such as suppositories, are well known to those of
skill in the art. The compounds can also be administered by
inhalation (e.g., in the form of an aerosol either nasally or using
delivery articles of the type set forth in U.S. Pat. No. 4,922,901
to Brooks et al., the disclosure of which is incorporated herein in
its entirety); topically (e.g., in lotion form); or transdermally
(e.g., using a transdermal patch, using technology that is
commercially available from Novartis and Alza Corporation).
Although it is possible to administer the compounds in the form of
a bulk active chemical, it is preferred to present each compound in
the form of a pharmaceutical composition or formulation for
efficient and effective administration.
[0173] Exemplary methods for administering such compounds will be
apparent to the skilled artisan. The usefulness of these
formulations can depend on the particular composition used and the
particular subject receiving the treatment. These formulations can
contain a liquid carrier that can be oily, aqueous, emulsified or
contain certain solvents suitable to the mode of
administration.
[0174] The compositions can be administered intermittently or at a
gradual, continuous, constant or controlled rate to a warm-blooded
animal (e.g., a mammal such as a mouse, rat, cat, rabbit, dog, pig,
cow, or monkey), but advantageously are administered to a human
being. In addition, the time of day and the number of times per day
that the pharmaceutical formulation is administered can vary.
[0175] Preferably, upon administration, the active ingredients
interact with receptor sites within the body of the subject that
affect the functioning of the CNS. More specifically, in treating a
CNS disorder, preferable administration is designed to optimize the
effect upon those relevant nicotinic acethylcholine receptor
(nAChR) subtypes that have an effect upon the functioning of the
CNS, while minimizing the effects upon muscle-type receptor
subtypes. Other suitable methods for administering the compounds of
the present invention are described in U.S. Pat. No. 5,604,231 to
Smith et al., the contents of which are hereby incorporated by
reference.
[0176] In certain circumstances, the compounds described herein can
be employed as part of a pharmaceutical composition with other
compounds intended to prevent or treat a particular disorder. In
addition to effective amounts of the compounds described herein,
the pharmaceutical compositions can also include various other
components as additives or adjuncts. Exemplary pharmaceutically
acceptable components or adjuncts which are employed in relevant
circumstances include antioxidants, free-radical scavenging agents,
peptides, growth factors, antibiotics, bacteriostatic agents,
immunosuppressives, anticoagulants, buffering agents,
anti-inflammatory agents, anti-pyretics, time-release binders,
anesthetics, steroids, vitamins, minerals and corticosteroids. Such
components can provide additional therapeutic benefit, act to
affect the therapeutic action of the pharmaceutical composition, or
act towards preventing any potential side effects that can be
imposed as a result of administration of the pharmaceutical
composition.
[0177] The appropriate dose of the compound is that amount
effective to prevent occurrence of the symptoms of the disorder or
to treat some symptoms of the disorder from which the patient
suffers. By "effective amount", "therapeutic amount" or "effective
dose" is meant that amount sufficient to elicit the desired
pharmacological or therapeutic effects, thus resulting in effective
prevention or treatment of the disorder.
[0178] When treating a CNS disorder, an effective amount of
compound is an amount sufficient to pass across the blood-brain
barrier of the subject, to bind to relevant receptor sites in the
brain of the subject and to modulate the activity of relevant nAChR
subtypes (e.g., provide neurotransmitter secretion, thus resulting
in effective prevention or treatment of the disorder). Prevention
of the disorder is manifested by delaying the onset of the symptoms
of the disorder. Treatment of the disorder is manifested by a
decrease in the symptoms associated with the disorder or an
amelioration of the recurrence of the symptoms of the disorder.
Preferably, the effective amount is sufficient to obtain the
desired result, but insufficient to cause appreciable side
effects.
[0179] The effective dose can vary, depending upon factors such as
the condition of the patient, the severity of the symptoms of the
disorder, and the manner in which the pharmaceutical composition is
administered. For human patients, the effective dose of typical
compounds generally requires administering the compound in an
amount sufficient to modulate the activity of relevant nAChRs to
effect neurotransmitter (e.g., dopamine) release, but the amount
should be insufficient to induce effects on skeletal muscles and
ganglia to any significant degree. The effective dose of compounds
will of course differ from patient to patient, but in general
includes amounts starting where CNS effects or other desired
therapeutic effects occur but below the amount where muscular
effects are observed.
[0180] The compounds, when employed in effective amounts in
accordance with the method described herein, are selective to
certain relevant nAChRs, but do not significantly activate
receptors associated with undesirable side effects at
concentrations at least greater than those required for eliciting
the release of dopamine or other neurotransmitters. By this is
meant that a particular dose of compound effective in preventing
and/or treating a CNS disorder is essentially ineffective in
eliciting activation of certain ganglionic-type nAChRs at
concentration higher than 5 times, preferably higher than 100
times, and more preferably higher than 1,000 times than those
required for modulation of neurotransmitter release. This
selectivity of certain compounds described herein against those
ganglionic-type receptors responsible for cardiovascular side
effects is demonstrated by a lack of the ability of those compounds
to activate nicotinic function of adrenal chromaffin tissue at
concentrations greater than those required for activation of
dopamine release.
[0181] The compounds described herein, when employed in effective
amounts in accordance with the methods described herein, can
provide some degree of prevention of the progression of CNS
disorders, ameliorate symptoms of CNS disorders, and ameliorate to
some degree of the recurrence of CNS disorders. The effective
amounts of those compounds are typically below the threshold
concentration required to elicit any appreciable side effects, for
example those effects relating to skeletal muscle. The compounds
can be administered in a therapeutic window in which certain CNS
disorders are treated and certain side effects are avoided.
Ideally, the effective dose of the compounds described herein is
sufficient to provide the desired effects upon the CNS but is
insufficient (i.e., is not at a high enough level) to provide
undesirable side effects. Preferably, the compounds are
administered at a dosage effective for treating the CNS disorders
but less than 1/5, and often less than 1/10, the amount required to
elicit certain side effects to any significant degree.
[0182] Most preferably, effective doses are at very low
concentrations, where maximal effects are observed to occur, with a
minimum of side effects. Typically, the effective dose of such
compounds generally requires administering the compound in an
amount of less than 5 mg/kg of patient weight. Often, the compounds
of the present invention are administered in an amount from less
than about 1 mg/kg patent weight and usually less than about 100
.mu.g/kg of patient weight, but frequently between about 10 .mu.g
to less than 100 .mu.g/kg of patient weight. For compounds that do
not induce effects on muscle-type nicotinic receptors at low
concentrations, the effective dose is less than 5 mg/kg of patient
weight; and often such compounds are administered in an amount from
50 .mu.g to less than 5 mg/kg of patient weight. The foregoing
effective doses typically represent that amount administered as a
single dose, or as one or more doses administered over a 24-hour
period.
[0183] For human patients, the effective dose of typical compounds
generally requires administering the compound in an amount of at
least about 1, often at least about 10, and frequently at least
about 100 mg/24 hr/patient. For human patients, the effective dose
of typical compounds requires administering the compound which
generally does not exceed about 500, often does not exceed about
400, and frequently does not exceed about 300 mg/24 hr/patient. In
addition, the compositions are advantageously administered at an
effective dose such that the concentration of the compound within
the plasma of the patient normally does not exceed 50 ng/mL, often
does not exceed 30 ng/mL, and frequently does not exceed 10
ng/mL.
III. Methods of Using the Compounds and/or Pharmaceutical
Compositions
[0184] The compounds can be used to treat those types of conditions
and disorders for which other types of nicotinic compounds have
been proposed as therapeutics. See, for example, Williams et al.,
Drug News Perspec. 7(4): 205 (1994); Arneric et al., CNS Drug Rev.
1(1): 1 (1995); Arneric et al., Exp. Opin. Invest. Drugs 5(1): 79
(1996); Bencherif et al., J. Pharmacol. Exp. Ther. 279: 1413
(1996); Lippiello et al., J. Pharmacol. Exp. Ther. 279: 1422
(1996); Damaj et al., J. Pharmacol. Exp. Ther. 291: 390 (1999);
Chiari et al., Anesthesiology 91: 1447 (1999); Lavand'homme and
Eisenbach, Anesthesiology 91: 1455 (1999); Holladay et al., J. Med.
Chem. 40(28): 4169 (1997); Bannon et al., Science 279: 77 (1998),
PCT WO 94/08992, PCT WO 96/31475, and U.S. Pat. Nos. 5,583,140 to
Bencherif et al., 5,597,919 to Dull et al., and 5,604,231 to Smith
et al., the disclosures of each of which are incorporated herein by
reference in their entirety.
[0185] More particularly, the compounds can be used to treat those
types of conditions and disorders for which nicotinic compounds
with selectivity for the .alpha.7 nAChR subtype have been proposed
as therapeutics. See, for example, Leonard et al., Schizophrenia
Bulletin 22(3): 431 (1996); Freedman et al., Biological Psychiatry
38(1): 22 (1995); Heeschen et al., J. Clin. Invest. 100: 527
(2002); Utsugisawa et al., Molecular Brain Research 106(1-2): 88
(2002); U.S. Patent Application 2002/0016371, Levin and Rezvani,
Current Drug Targets: CNS and Neurological Disorders 1(4): 423
(2002); O'Neill et al., Current Drug Targets: CNS and Neurological
Disorders 1(4): 399 (2002); Jeyarasasingam et al., Neuroscience
109(2): 275 (2002); Xiao et al., Proc. Nat. Acad. Sci. (US) 99(12):
8360 (2002); PCT WO 99/62505, PCT WO 99/03859, PCT WO 97/30998, PCT
WO 01/36417, PCT WO 02/15662, PCT WO 02/16355, PCT WO 02/16356, PCT
WO 02/16357, PCT WO 02/16358, PCT WO 02/17358, Stevens et al.,
Psychopharm. 136: 320 (1998); Dolle et al., J. Labeled Comp.
Radiopharm. 44: 785 (2001) and Macor et al., Bioorg. Med. Chem.
Lett. 11: 319 (2001) and references therein, the contents of each
of which are hereby incorporated by reference in their
entirety.
[0186] The compounds can also be used as adjunct therapy in
combination with existing therapies in the management of the
aforementioned types of diseases and disorders. In such situations,
it is preferably to administer the active ingredients in a manner
that minimizes effects upon nAChR subtypes such as those that are
associated with muscle and ganglia. This can be accomplished by
targeted drug delivery and/or by adjusting the dosage such that a
desired effect is obtained without meeting the threshold dosage
required to achieve significant side effects. The pharmaceutical
compositions can be used to ameliorate any of the symptoms
associated with those conditions, diseases and disorders.
Representative classes of disorders that can be treated are
discussed in detail below.
[0187] Treatment of CNS Disorders
[0188] Examples of conditions and disorders that can be treated
include neurological disorders and neurodegenerative disorders,
and, in particular, CNS disorders. CNS disorders can be drug
induced; can be attributed to genetic predisposition, infection or
trauma; or can be of unknown etiology. CNS disorders comprise
neuropsychiatric disorders, neurological diseases and mental
illnesses, and include neurodegenerative diseases, behavioral
disorders, cognitive disorders and cognitive affective disorders.
There are several CNS disorders whose clinical manifestations have
been attributed to CNS dysfunction (i.e., disorders resulting from
inappropriate levels of neurotransmitter release, inappropriate
properties of neurotransmitter receptors, and/or inappropriate
interaction between neurotransmitters and neurotransmitter
receptors). Several CNS disorders can be attributed to a deficiency
of choline, dopamine, norepinephrine and/or serotonin.
[0189] Examples of CNS disorders that can be treated in accordance
with the present invention include pre-senile dementia (early onset
Alzheimer's disease), senile dementia (dementia of the Alzheimer's
type), Lewy Body dementia, micro-infarct dementia, AIDS-related
dementia, HIV-dementia, multiple cerebral infarcts, Parkinsonism
including Parkinson's disease, Pick's disease, progressive
supranuclear palsy, Huntington's chorea, tardive dyskinesia,
hyperkinesia, mania, attention deficit disorder, anxiety,
depression, dyslexia, schizophrenia, obsessive-compulsive
disorders, Tourette's syndrome, mild cognitive impairment (MCI),
age-associated memory impairment (AAMI), premature amnesic and
cognitive disorders which are age-related or a consequence of
alcoholism, or immunodeficiency syndrome, or are associated with
vascular disorders, with genetic alterations (such as, for example,
trisomy 21) or with attention deficiencies or learning
deficiencies, acute or chronic neurodegenerative conditions such as
amyotrophic lateral sclerosis, multiple sclerosis, peripheral
neurotrophies, and cerebral or spinal traumas. In addition, the
compounds can be used to treat nicotine addiction and/or other
behavioral disorders related to substances that lead to dependency
(e.g., alcohol, cocaine, heroin and opiates, psychostimulants,
benzodiazepines and barbiturates).
[0190] Schizophrenia is an example of a CNS disorder that is
particularly amenable to treatment by modulating the .alpha.7 nAChR
subtype. The compounds can also be administered to improve
cognition and/or provide neuroprotection, and these uses are also
particularly amenable to treatment with compounds, such as the
compounds of the present invention, that are specific for the
.alpha.7 nAChR subtype.
[0191] The disorders can be treated and/or prevented by
administering to a patient in need of treatment or prevention
thereof an effective treatment or preventative amount of a compound
that provides some degree of prevention of the progression of a CNS
disorder (i.e., provides protective effects), ameliorating the
symptoms of the disorder, and ameliorating the recurrence of the
disorder.
[0192] Anti-Inflammatory Uses
[0193] Excessive inflammation and tumor necrosis factor (TNF)
synthesis cause morbidity and even mortality in a variety of
diseases. These diseases include, but are not limited to,
endotoxemia, sepsis, rheumatoid arthritis, and irritable bowel
disease. The nervous system, primarily through the vagus nerve, is
known to regulate the magnitude of the innate immune response by
inhibiting the release of macrophage tumor necrosis factor. This
physiological mechanism is known as the "cholinergic
anti-inflammatory pathway" (see, for example, Tracey, Nature.
420(6917): 853 (2002)).
[0194] The nAChR .alpha.7 subunit is required for acetylcholine
inhibition of macrophage TNF release, and also inhibits release of
other cytokines. Agonists (or, at elevated dosages, partial
agonists) at the .alpha.7-specific nAChR subtype can inhibit the
TNF-modulated inflammatory response. Accordingly, those compounds
described herein that are .alpha.7 agonists can be used to treat
inflammatory disorders characterized by excessive synthesis of TNF
(see also Wang et al., Nature, 421(6921): 384 (2003)).
[0195] Inflammatory conditions that can be treated or prevented by
administering the compounds described herein include, but are not
limited to chronic and acute inflammation, psoriasis, gout, acute
pseudogout, acute gouty arthritis, arthritis, rheumatoid arthritis,
osteoarthritis, allograft rejection, chronic transplant rejection,
asthma, atherosclerosis, mononuclear-phagocyte dependent lung
injury, idiopathic pulmonary fibrosis, atopic dermatitis, chronic
obstructive pulmonary disease, adult respiratory distress syndrome,
acute chest syndrome in sickle cell disease, inflammatory bowel
disease, Crohn's disease, ulcerative colitis, acute cholangitis,
aphteous stomatitis, glomerulonephritis, lupus nephritis,
thrombosis, and graft vs. host reaction.
[0196] Minimizing the Inflammatory Response Associated with
Bacterial and/or Viral Infection
[0197] Many bacterial and/or viral infections are associated with
side effects brought on by the formation of toxins, and the body's
natural response to the bacteria or virus and/or the toxins.
Examples of such bacterial infections include anthrax, botulism,
and sepsis. As discussed above, the body's response to infection
often involves generating a significant amount of tumor necrosis
factor and/or other cytokines. The over-expression of these
cytokines can result in significant injury, such as septic shock
(when the bacteria is sepsis), endotoxic shock, urosepsis and toxic
shock syndrome.
[0198] Cytokine expression is mediated by the .alpha.7 nAChR and
can be inhibited by administering agonists or partial agonists of
these receptors. Those compounds described herein that are agonists
or partial agonists of these receptors can therefore be used to
minimize the inflammatory response associated with bacterial
infection, as well as viral and fungal infections. Certain of the
compounds themselves may also have antimicrobial properties.
[0199] These compounds can also be used as adjunct therapy in
combination with existing therapies to manage bacterial, viral and
fungal infections, such as antibiotics, antivirals and antifungals.
Antitoxins can also be used to bind to toxins produced by the
infectious agents and allow the bound toxins to pass through the
body without generating an inflammatory response. Examples of
antitoxins are disclosed, for example, in U.S. Pat. No. 6,310,043
to Bundle et al., incorporated herein by reference. Other agents
effective against bacterial and other toxins can be effective and
their therapeutic effect can be complimented by co-administration
with the compounds described herein.
[0200] Analgesic Uses
[0201] The compounds can be administered to treat and/or prevent
pain, including neurologic, neuropathic and chronic pain. The
analgesic activity of compounds described herein can be
demonstrated in models of persistent inflammatory pain and of
neuropathic pain, performed as described in U.S. Published Patent
Application No. 20010056084 A1 to Allgeier et al. (e.g., mechanical
hyperalgesia in the complete Freund's adjuvant rat model of
inflammatory pain and mechanical hyperalgesia in the mouse partial
sciatic nerve ligation model of neuropathic pain).
[0202] The analgesic effect is suitable for treating pain of
various genesis or etiologies, in particular in treating
inflammatory pain and associated hyperalgesia, neuropathic pain and
associated hyperalgesia, chronic pain (e.g., severe chronic pain,
post-operative pain and pain associated with various conditions
including cancer, angina, renal or biliary colic, menstruation,
migraine and gout) and fibromyalgia syndrome. Inflammatory pain may
be of diverse genesis, including arthritis and rheumatoid disease,
teno-synovitis and vasculitis. Neuropathic pain includes trigeminal
or herpetic neuralgia, diabetic neuropathy pain, causalgia, low
back pain and deafferentation syndromes such as brachial plexus
avulsion.
[0203] Inhibition of Neovascularization
[0204] The .alpha.7 nAChR is also associated with
neovascularization. Inhibition of neovascularization, for example,
by administering antagonists (or at certain dosages, partial
agonists) of the .alpha.7 nAChR can inhibit neovascularization and,
accordingly, treat or prevent conditions characterized by
undesirable neovascularization or angiogenesis. Such conditions can
include those characterized by inflammatory angiogenesis and/or
ischemia-induced angiogenesis. Neovascularization associated with
tumor growth can also be inhibited by administering those compounds
described herein that function as antagonists or partial agonists
of .alpha.7 nAChR.
[0205] Specific antagonism of .alpha.7 nAChR-specific activity
reduces the angiogenic response to inflammation, ischemia, and
neoplasia. Guidance regarding appropriate animal model systems for
evaluating the compounds described herein can be found, for
example, in Heeschen, et al., J. Clin Invest, 110(4): 527 (2002),
incorporated herein by reference regarding disclosure of
.alpha.7-specific inhibition of angiogenesis and cellular (in
vitro) and animal modeling of angiogenic activity relevant to human
disease, especially the Lewis lung tumor model (in vivo, in
mice--see, in particular, pages 529, and 532-533).
[0206] Representative tumor types that can be treated using the
compounds described herein include NSCLC, ovarian cancer,
pancreatic cancer, breast carcinoma, colon carcinoma, rectum
carcinoma, lung carcinoma, oropharynx carcinoma, hypopharynx
carcinoma, esophagus carcinoma, stomach carcinoma, pancreas
carcinoma, liver carcinoma, gallbladder carcinoma, bile duct
carcinoma, small intestine carcinoma, urinary tract carcinoma,
kidney carcinoma, bladder carcinoma, urothelium carcinoma, female
genital tract carcinoma, cervix carcinoma, uterus carcinoma,
ovarian carcinoma, choriocarcinoma, gestational trophoblastic
disease, male genital tract carcinoma, prostate carcinoma, seminal
vesicles carcinoma, testes carcinoma, germ cell tumors, endocrine
gland carcinoma, thyroid carcinoma, adrenal carcinoma, pituitary
gland carcinoma, skin carcinoma, hemangiomas, melanomas, sarcomas,
bone and soft tissue sarcoma, Kaposi's sarcoma, tumors of the
brain, tumors of the nerves, tumors of the eyes, tumors of the
meninges, astrocytomas, gliomas, glioblastomas, retinoblastomas,
neuromas, neuroblastomas, Schwannomas, meningiomas, solid tumors
arising from hematopoietic malignancies (such as leukemias,
chloromas, plasmacytomas and the plaques and tumors of mycosis
fungoides and cutaneous T-cell lymphoma/leukemia), and solid tumors
arising from lymphomas.
[0207] The compounds can also be administered in conjunction with
other forms of anti-cancer treatment, including co-administration
with antineoplastic antitumor agents such as cis-platin,
adriamycin, daunomycin, and the like, and/or anti-VEGF (vascular
endothelial growth factor) agents, as such are known in the
art.
[0208] The compounds can be administered in such a manner that they
are targeted to the tumor site. For example, the compounds can be
administered in microspheres, microparticles or liposomes
conjugated to various antibodies that direct the microparticles to
the tumor. Additionally, the compounds can be present in
microspheres, microparticles or liposomes that are appropriately
sized to pass through the arteries and veins, but lodge in
capillary beds surrounding tumors and administer the compounds
locally to the tumor. Such drug delivery devices are known in the
art.
[0209] Other Disorders
[0210] In addition to treating CNS disorders, inflammatory
disorders, and neovascular disorders, and inhibiting the pain
response, the compounds can be also used to prevent or treat
certain other conditions, diseases, and disorders. Examples include
autoimmune disorders such as Lupus, disorders associated with
cytokine release, cachexia secondary to infection (e.g., as occurs
in AIDS, AIDS related complex and neoplasia), as well as those
indications set forth in PCT WO 98/25619. The compounds can also be
administered to treat convulsions such as those that are
symptomatic of epilepsy, and to treat conditions such as syphilis
and Creutzfeldt-Jakob disease.
[0211] Diagnostic Uses
[0212] The compounds can be used in diagnostic compositions, such
as probes, particularly when they are modified to include
appropriate labels. The probes can be used, for example, to
determine the relative number and/or function of specific
receptors, particularly the .alpha.7 receptor subtype. The
compounds of the present invention most preferably are labeled with
a radioactive isotopic moiety such as .sup.11C, .sup.18F,
.sup.76Br, .sup.123I or .sup.125I, as discussed above.
[0213] The administered compounds can be detected using known
detection methods appropriate for the label used. Examples of
detection methods include position emission topography (PET) and
single-photon emission computed tomography (SPECT). The radiolabels
described above are useful in PET (e.g., .sup.11C, .sup.18F or
.sup.76Br) and SPECT (e.g., .sup.123I) imaging, with half-lives of
about 20.4 minutes for .sup.11C, about 109 minutes for .sup.18F,
about 13 hours for .sup.123I, and about 16 hours for .sup.76Br. A
high specific activity is desired to visualize the selected
receptor subtypes at non-saturating concentrations. The
administered doses typically are below the toxic range and provide
high contrast images. The compounds are expected to be capable of
administration in non-toxic levels. Determination of dose is
carried out in a manner known to one skilled in the art of
radiolabel imaging. See, for example, U.S. Pat. No. 5,969,144 to
London et al.
[0214] The compounds can be administered using known techniques.
See, for example, U.S. Pat. No. 5,969,144 to London et al. The
compounds can be administered in formulation compositions that
incorporate other ingredients, such as those types of ingredients
that are useful in formulating a diagnostic composition. Compounds
useful in accordance with carrying out the present invention most
preferably are employed in forms of high purity. See, for example,
U.S. Pat. No. 5,853,696 to Elmalch et al.
[0215] After the compounds are administered to a subject (e.g., a
human subject), the presence of that compound within the subject
can be imaged and quantified by appropriate techniques in order to
indicate the presence, quantity, and functionality of selected
nicotinic cholinergic receptor subtypes. In addition to humans, the
compounds can also be administered to animals, such as mice, rats,
dogs, and monkeys. SPECT and PET imaging can be carried out using
any appropriate technique and apparatus. See Villemagne et al., In:
Arneric et al., (Eds.) Neuronal Nicotinic Receptors Pharmacology
and Therapeutic Opportunities, 235-250 (1998) and U.S. Pat. No.
5,853,696 to Elmalch et al. for a disclosure of representative
imaging techniques.
[0216] The radiolabeled compounds bind with high affinity to
selective nAChR subtypes (e.g., .alpha.7) and preferably exhibit
negligible non-specific binding to other nicotinic cholinergic
receptor subtypes (e.g., those receptor subtypes associated with
muscle and ganglia). As such, the compounds can be used as agents
for noninvasive imaging of nAChR subtypes within the body of a
subject, particularly within the brain for diagnosis associated
with a variety of CNS diseases and disorders.
[0217] In one aspect, the diagnostic compositions can be used in a
method to diagnose disease in a subject, such as a human patient.
The method involves administering to that patient a detectably
labeled compound as described herein, and detecting the binding of
that compound to selected nicotinic receptor subtypes (e.g.,
.alpha.7 receptor subtype). Those skilled in the art of using
diagnostic tools, such as PET and SPECT, can use the radiolabeled
compounds described herein to diagnose a wide variety of conditions
and disorders, including conditions and disorders associated with
dysfunction of the central and autonomic nervous systems. Such
disorders include a wide variety of CNS diseases and disorders,
including Alzheimer's disease, Parkinson's disease, and
schizophrenia. These and other representative diseases and
disorders that can be evaluated include those that are set forth in
U.S. Pat. No. 5,952,339 to Bencherif et al., the contents of which
are hereby incorporated by reference.
[0218] In another aspect, the diagnostic compositions can be used
in a method to monitor selective nAChR subtypes of a subject, such
as a human patient. The method involves administering a detectably
labeled compound as described herein to that patient and detecting
the binding of that compound to selected nAChR subtypes (e.g., the
.alpha.7 receptor subtype).
[0219] The following examples are provided to further illustrate
the present invention, and should not be construed as limiting
thereof.
IV. Synthetic Examples
[0220] The following synthetic examples are provided to illustrate
the present invention and should not be construed as limiting the
scope thereof. In these examples, all parts and percentages are by
weight, unless otherwise noted. Reaction yields are reported in
mole percentage.
[0221] Compounds of the present invention are derivatives of
3-pyrid-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]undecane, the
synthesis of which is described below:
2-((3-Pyridinyl)methylene)-1-azabicyclo[2.2.2]octan-3-one
[0222] Potassium hydroxide (56 g, 0.54 mole) was dissolved in
methanol (420 mL). 3-Quinuclidinone hydrochloride (75 g, 0.49 mole)
was added and the mixture was stirred for 30 min at ambient
temperature. 3-Pyridinecarboxaldehyde (58 g, 0.54 mole) was added
and the mixture stirred for 16 h at ambient temperature. The
reaction mixture became yellow during this period, with solids
caking on the walls of the flask. The solids were scraped from the
walls and the chunks broken up. With rapid stirring, water (390 mL)
was added. When the solids dissolved, the mixture was cooled at
4.degree. C. overnight. The crystals were collected by filtration,
washed with water, and air dried to obtain 80 g of yellow solid. A
second crop (8 g) was obtained by concentration of the filtrate to
.about.10% of its former volume and cooling at 4.degree. C.
overnight. Both crops were sufficiently pure for further
transformation (88 g, 82%).
2-(1-(3-Pyridinyl)-2-nitroethyl)-1-azabicyclo[2.2.2]octan-3-one
[0223] 2-((3-Pyridinyl)methylene)-1-azabicyclo[2.2.2]octan-3-one
(6.4 g, 0.024 mol) in dry methanol (45 mL) was added drop-wise to
sodium methoxide (produced in situ, 0.036 mol). Nitromethane (3.7
mL, 0.068 mol) was then added, and the mixture was heated at reflux
for 3 h. After cooling to room temperature, 1 N HCl was slowly
added to adjust pH to 8. The mixture was concentrated by rotary
evaporation to yield a solid brown residue. The residue was
purified by column chromatography, using ethyl acetate/hexane (1:1,
v/v), followed by chloroform/methanol/ammonia (90:10:1, v/v), as
eluent, to obtain a yellow oil (4.2 g, 64%).
3-Pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]undecane
[0224]
6-(1-(3-Pyridinyl)-2-nitroethyl)-1-azabicyclo[2.2.2]octan-3-one
(14.0 g, 0.046 mol) was dissolved in ethanol (200 mL), and then
Raney nickel was added under nitrogen. The mixture was subjected to
hydrogenolysis (40 psi H.sub.2) for 48 h and then filtered through
Celite and concentrated by rotary evaporation to a crude brown
residue. The residue was purified by column chromatography, using
chloroform/methanol/ammonia (80:20:1, v/v) as eluent, to yield
3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]undecane as a
yellow oil (8.0 g, 67%).
[0225] The following example describes the synthesis of various
amide derivatives of
3-(3-pyridinyl)-1,5-diazatricyclo[5.2.2.0<2,6>]undecane.
Example 1
Amide derivatives of
3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]undecane
[0226] Benzotriazol-1-yloxytris(dimethylamino)phosphonium
hexafluorophosphate (BOP, 0.097 g, 0.22 mmol) was added to a
solution of the carboxylic acid (0.22 mmol) and triethylamine (0.66
mmol) in dichloromethane (1 mL), and then
3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]undecane (0.046
g, 0.20 mmol) was added. The mixture was stirred for 48 h at room
temperature, then treated with 10% NaOH (0.2 mL). The biphasic
mixture was separated by phase filtration, and the organic phase
was concentrated on the Genevac centrifugal evaporator. The crude
residue was dissolved in methanol (1 mL) and purified by HPLC on a
C18 silica gel column, using acetonitrile/water gradient containing
0.05% trifluoroacetic acid.
[0227] Compounds made by this procedure were isolated as
trifluoroacetate salts and characterized by LC/MS. Compounds
exhibiting appropriate molecular ions and fragmentation patterns,
and purities of 90% or greater were submitted for biological
assessment. Selected compounds were analyzed by NMR spectroscopy,
which confirmed their structural assignments. Table 1 lists
molecular weights, calculated and measured by LC/MS, for some
representative compounds, all of which bind at the .alpha.7 nAChR
subtype with Ki values of <100 nM.
TABLE-US-00001 TABLE 1 LCMS Calc. Mass Compound # Compound Name FB
Mass (MH.sup.+) 1 5-(Benzofuran-2-ylcarbonyl)-3- 373.459 374.32
pyridin-3-yl-1,5- diazatricyclo[5.2.2.0<2,6>]undecane 2
5-(7-Methoxybenzofuran-2- 403.485 404.35
ylcarbonyl)-3-pyridin-3-yl-1,5- diazatricyclo[5.2.2.0<2,
6>]undecane 3 5-(Naphth-2-ylcarbonyl)-3-pyridin- 383.498 384.35
3-yl-1,5- diazatricyclo[5.2.2.0<2,6>]undecane 4
5-(1H-Indol-3-ylcarbonyl)-3-pyridin- 372.474 373.38 3-yl-1,5-
diazatricyclo[5.2.2.0<2,6>]undecane
[0228] The following example describes the synthesis of various
urea derivatives of
3-(3-pyridinyl)-1,5-diazatricyclo[5.2.2.0<2,6>]undecane.
Example 2
Urea derivatives of
3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2,6>]undecane
[0229] A mixture of
3-pyridin-3-yl-1,5-diazatricyclo[5.2.20<2,6>]undecane (0.20
mmol) and the appropriate isocyanate (0.22 mmol) were stirred in
dry dichloromethane (1 mL) for 48 h at ambient temperature. Then
the mixture was concentrated under reduced pressure and the residue
was dissolved in methanol (0.75 mL) and purified by HPLC on a C18
silica gel column, using acetonitrile/water gradients containing
0.05% trifluoroacetic acid.
[0230] Compounds made by this procedure were isolated as
trifluoroacetate salts and characterized by LC/MS. Compounds
exhibiting appropriate molecular ions and fragmentation patterns,
and purities of 90% or greater were submitted for biological
assessment. Selected compounds were analyzed by NMR spectroscopy,
which confirmed their structural assignments. Table 2 lists
molecular weights, calculated and measured by LC/MS, for some
representative compounds, all of which bind at the .alpha.7 nAChR
subtype with Ki values of <100 nM.
TABLE-US-00002 TABLE 2 LCMS Com- Calc. Mass pound # Compound Name
FB Mass (MH.sup.+) 5 5-(N-(3,4- 417.342 417.22
Dichlorophenyl)carbamoyl)-3-
pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2, 6>]undecane 6
5-(N-(4-Bromophenyl)carbamoyl)- 427.348 (.sup.81Br)
3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2, 6>]undecane 7
5-(N-(4-Chlorophenyl)carbamoyl)- 382.897 383.27
3-pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2, 6>]undecane 8
5-(N-(4- 394.543 395.29 Methylthiophenyl)carbamoyl)-3-
pyridin-3-yl-1,5-diazatricyclo[5.2.2.0<2, 6>]undecane
V. Biological Assays
Example 3
Radioligand Binding at CNS nAChRs
[0231] .alpha.4.beta.2 nAChR Subtype
[0232] Rats (female, Sprague-Dawley), weighing 150-250 g, were
maintained on a 12 h light/dark cycle and were allowed free access
to water and food supplied by PMI Nutrition International, Inc.
Animals were anesthetized with 70% CO.sub.2, then decapitated.
Brains were removed and placed on an ice-cold platform. The
cerebral cortex was removed and placed in 20 volumes
(weight:volume) of ice-cold preparative buffer (137 mM NaCl, 10.7
mM KCl, 5.8 mM KH.sub.2PO.sub.4, 8 mM Na.sub.2HPO.sub.4, 20 mM
HEPES (free acid), 5 mM iodoacetamide, 1.6 mM EDTA, pH 7.4); PMSF,
dissolved in methanol to a final concentration of 100 .mu.M, was
added and the suspension was homogenized by Polytron. The
homogenate was centrifuged at 18,000.times.g for 20 min at
4.degree. C. and the resulting pellet was re-suspended in 20
volumes of ice-cold water. After 60 min incubation on ice, a new
pellet was collected by centrifugation at 18,000.times.g for 20 min
at 4.degree. C. The final pellet was re-suspended in 10 volumes of
buffer and stored at -20.degree. C. On the day of the assay, tissue
was thawed, centrifuged at 18,000.times.g for 20 min, and then
re-suspended in ice-cold PBS (Dulbecco's Phosphate Buffered Saline,
138 mM NaCl, 2.67 mM KCl, 1.47 mM KH.sub.2PO.sub.4, 8.1 mM
Na.sub.2HPO.sub.4, 0.9 mM CaCl.sub.2, 0.5 mM MgCl.sub.2,
Invitrogen/Gibco, pH 7.4) to a final concentration of approximately
4 mg protein/mL. Protein was determined by the method of Lowry et
al., J. Biol. Chem. 193: 265 (1951), using bovine serum albumin as
the standard.
[0233] The binding of [.sup.3H]nicotine was measured using a
modification of the methods of Romano et al., Science 210: 647
(1980) and Marks et al., Mol. Pharmacol. 30: 427 (1986). The
[.sup.3H]nicotine (Specific Activity=81.5 Ci/mmol) was obtained
from NEN Research Products. The binding of [.sup.3H]nicotine was
measured using a 3 h incubation at 4.degree. C. Incubations were
conducted in 48-well micro-titre plates and contained about 400
.mu.g of protein per well in a final incubation volume of 300
.mu.L. The incubation buffer was PBS and the final concentration of
[.sup.3H]nicotine was 5 nM. The binding reaction was terminated by
filtration of the protein containing bound ligand onto glass fiber
filters (GF/B, Brandel) using a Brandel Tissue Harvester at
4.degree. C. Filters were soaked in de-ionized water containing
0.33% polyethyleneimine to reduce non-specific binding. Each filter
was washed with ice-cold buffer (3.times.1 mL). Non-specific
binding was determined by inclusion of 10 .mu.M non-radioactive
L-nicotine (Acros Organics) in selected wells.
[0234] The inhibition of [.sup.3H]nicotine binding by test
compounds was determined by including seven different
concentrations of the test compound in selected wells. Each
concentration was replicated in triplicate. IC.sub.50 values were
estimated as the concentration of compound that inhibited 50
percent of specific [.sup.3H]nicotine binding. Inhibition constants
(Ki values), reported in nM, were calculated from the IC.sub.50
values using the method of Cheng et al., Biochem. Pharmacol. 22:
3099 (1973).
[0235] For initial screening, a single concentration of test
compounds was tested in the above assay format with the following
modifications. The binding of [.sup.3H]epibatidine was measured.
The [.sup.3H]epibatidine (Specific Activity=48 Ci/mmol) was
obtained from NEN Research Products. The binding of
[.sup.3H]epibatidine was measured using a 2 h incubation at
21.degree. C. (room temperature). Incubations were conducted in
96-well Millipore Multiscreen (MAFB) plates containing about 200
.mu.g of protein per well in a final incubation volume of 150
.mu.L. The incubation buffer was PBS and the final concentration of
[.sup.3H]epibatidine was 0.3 nM. The binding reaction was
terminated by filtration of the protein containing bound ligand
onto the glass fiber filter base of the Multiscreen plates. Filters
were soaked in de-ionized water containing 0.33% polyethyleneimine
to reduce non-specific binding. Each filter was washed with
ice-cold buffer (3.times.0.25 mL). Non-specific binding was
determined by inclusion of 10 .mu.M non-radioactive L-nicotine
(Acros Organics) in selected wells. The single concentration of
test compound was 5 .mu.M and testing was performed in triplicate.
`Active` compounds were defined as compounds that inhibited the
binding of [.sup.3H]epibatidine to the receptor by at least 50%
compared with the binding of [.sup.3H]epibatidine in the absence of
competitor. For those compounds found to be active in the single
point screen, the inhibition constants (Ki values) were determined
as described in the previous paragraphs of this section.
.alpha.7 nAChR Subtype
[0236] Rats (female, Sprague-Dawley), weighing 150-250 g, were
maintained on a 12 h light/dark cycle and were allowed free access
to water and food supplied by PMI Nutrition International, Inc.
Animals were anesthetized with 70% CO.sub.2, then decapitated.
Brains were removed and placed on an ice-cold platform. The
hippocampus was removed and placed in 10 volumes (weight:volume) of
ice-cold preparative buffer (137 mM NaCl, 10.7 mM KCl, 5.8 mM
KH.sub.2PO.sub.4, 8 mM Na.sub.2HPO.sub.4, 20 mM HEPES (free acid),
5 mM iodoacetamide, 1.6 mM EDTA, pH 7.4); PMSF, dissolved in
methanol to a final concentration of 100 .mu.M, was added and the
tissue suspension was homogenized by Polytron. The homogenate was
centrifuged at 18,000.times.g for 20 min at 4.degree. C. and the
resulting pellet was re-suspended in 10 volumes of ice-cold water.
After 60 min incubation on ice, a new pellet was collected by
centrifugation at 18,000.times.g for 20 min at 4.degree. C. The
final pellet was re-suspended in 10 volumes of buffer and stored at
-20.degree. C. On the day of the assay, tissue was thawed,
centrifuged at 18,000.times.g for 20 min, and then re-suspended in
ice-cold PBS (Dulbecco's Phosphate Buffered Saline, 138 mM NaCl,
2.67 mM KCl, 1.47 mM KH.sub.2PO.sub.4, 8.1 mM Na.sub.2HPO.sub.4,
0.9 mM CaCl.sub.2, 0.5 mM MgCl.sub.2, Invitrogen/Gibco, pH 7.4) to
a final concentration of approximately 2 mg protein/mL. Protein was
determined by the method of Lowry et al., J. Biol. Chem. 193: 265
(1951), using bovine serum albumin as the standard.
[0237] The binding of [.sup.3H]MLA was measured using a
modification of the methods of Davies et al., Neuropharmacol. 38:
679 (1999). [.sup.3H]MLA (Specific Activity=25-35 Ci/mmol) was
obtained from Tocris. The binding of [.sup.3H]MLA was determined
using a 2 h incubation at 21.degree. C. Incubations were conducted
in 48-well micro-titre plates and contained about 200 .mu.g of
protein per well in a final incubation volume of 300 .mu.L. The
incubation buffer was PBS and the final concentration of
[.sup.3H]MLA was 5 nM. The binding reaction was terminated by
filtration of the protein containing bound ligand onto glass fiber
filters (GF/B, Brandel) using a Brandel Tissue Harvester at room
temperature. Filters were soaked in de-ionized water containing
0.33% polyethyleneimine to reduce non-specific binding. Each filter
was washed with PBS (3.times.1 mL) at room temperature.
Non-specific binding was determined by inclusion of 50 .mu.M
non-radioactive MLA in selected wells.
[0238] The inhibition of [.sup.3H]MLA binding by test compounds was
determined by including seven different concentrations of the test
compound in selected wells. Each concentration was replicated in
triplicate. IC.sub.50 values were estimated as the concentration of
compound that inhibited 50 percent of specific [.sup.3H]MLA
binding. Inhibition constants (Ki values), reported in nM, were
calculated from the IC.sub.50 values using the method of Cheng et
al., Biochem. Pharmacol. 22: 3099 (1973).
[0239] For initial screening, a single concentration of test
compounds was tested in the above assay format with the following
modifications. Incubations were conducted in 96-well plates in a
final incubation volume of 150 .mu.L. Once the binding reaction was
terminated by filtration onto glass fiber filters, the filters were
washed four times with approximately 250 .mu.L of PBS at room
temperature. Non-specific binding was determined by inclusion of 10
.mu.M non-radioactive MLA in selected wells. The single
concentration of test compound was 5 .mu.M and testing was
performed in triplicate. `Active` compounds were defined as
compounds that inhibited the binding of [.sup.3H]MLA to the
receptor by at least 50% compared with the binding of [.sup.3H]MLA
in the absence of competitor. For those compounds found to be
active in the single point screen, the inhibition constants (Ki
values) were determined as described in the previous paragraphs of
this section.
Determination of Dopamine Release
[0240] Dopamine release was measured using striatal synaptosomes
obtained from rat brain, according to the procedures set forth by
Rapier et al., J. Neurochem. 54: 937 (1990). Rats (female,
Sprague-Dawley), weighing 150-250 g, were maintained on a 12 h
light/dark cycle and were allowed free access to water and food
supplied by PMI Nutrition International, Inc. Animals were
anesthetized with 70% CO.sub.2, then decapitated. The brains were
quickly removed and the striata dissected. Striatal tissue from
each of 2 rats was pooled and homogenized in ice-cold 0.32 M
sucrose (5 mL) containing 5 mM HEPES, pH 7.4, using a glass/glass
homogenizer. The tissue was then centrifuged at 1,000.times.g for
10 min. The pellet was discarded and the supernatant was
centrifuged at 12,000.times.g for 20 min. The resulting pellet was
re-suspended in perfusion buffer containing monoamine oxidase
inhibitors (128 mM NaCl, 1.2 mM KH.sub.2PO.sub.4, 2.4 mM KCl, 3.2
mM CaCl.sub.2, 1.2 mM MgSO.sub.4, 25 mM HEPES, 1 mM ascorbic acid,
0.02 mM pargyline HCl and 10 mM glucose, pH 7.4) and centrifuged
for 15 min at 25,000.times.g. The final pellet was resuspended in
perfusion buffer (1.4 mL) for immediate use.
[0241] The synaptosomal suspension was incubated for 10 min at
37.degree. C. to restore metabolic activity. [.sup.3H]Dopamine
([.sup.3H]DA, specific activity=28.0 Ci/mmol, NEN Research
Products) was added at a final concentration of 0.1 .mu.M and the
suspension was incubated at 37.degree. C. for another 10 min.
Aliquots of tissue (50 .mu.L) and perfusion buffer (100 .mu.L) were
loaded into the suprafusion chambers of a Brandel Suprafusion
System (series 2500, Gaithersburg, Md.). Perfusion buffer (room
temperature) was pumped into the chambers at a rate of 3 mL/min for
a wash period of 8 min. Test compound (10 .mu.M) or nicotine (10
.mu.M) was then applied in the perfusion stream for 40 sec.
Fractions (12 sec each) were continuously collected from each
chamber throughout the experiment to capture basal release and
agonist-induced peak release and to re-establish the baseline after
the agonist application. The perfusate was collected directly into
scintillation vials, to which scintillation fluid was added.
[.sup.3H]DA released was quantified by scintillation counting. For
each chamber, the integrated area of the peak was normalized to its
baseline.
[0242] Release was expressed as a percentage of release obtained
with an equal concentration of L-nicotine. Within each assay, each
test compound was replicated using 2-3 chambers; replicates were
averaged. When appropriate, dose-response curves of test compound
were determined. The maximal activation for individual compounds
(Emax) was determined as a percentage of the maximal activation
induced by L-nicotine. The compound concentration resulting in half
maximal activation (EC.sub.50) of specific ion flux was also
defined.
Example 4
Selectivity vs. Peripheral nAChRs
[0243] Interaction at the Human Muscle nAChR Subtype
[0244] Activation of muscle-type nAChRs was established on the
human clonal line TE671/RD, which is derived from an embryonal
rhabdomyosarcoma (Stratton et al., Carcinogen 10: 899 (1989)).
These cells express receptors that have pharmacological (Lukas, J.
Pharmacol. Exp. Ther. 251: 175 (1989)), electrophysiological
(Oswald et al., Neurosci. Lett. 96: 207 (1989)), and molecular
biological profiles (Luther et al., J. Neurosci. 9: 1082 (1989))
similar to the muscle-type nAChR.
[0245] TE671/RD cells were maintained in proliferative growth phase
according to routine protocols (Bencherif et al., Mol. Cell.
Neurosci. 2: 52 (1991) and Bencherif et al., J. Pharmacol. Exp.
Ther. 257: 946 (1991)). Cells were cultured in Dulbecco's modified
Eagle's medium (Gibco/BRL) with 10% horse serum (Gibco/BRL), 5%
fetal bovine serum (HyClone, Logan Utah), 1 mM sodium pyruvate, 4
mM L-Glutamine, and 50,000 units penicillin-streptomycin (Irvine
Scientific). When cells were 80% confluent, they were plated to 6
well polystyrene plates (Costar). Experiments were conducted when
the cells reached 100% confluency.
[0246] Nicotinic acetylcholine receptor (nAChR) function was
assayed using .sup.86Rb.sup.+ efflux according to the method
described by Lukas et al., Anal. Biochem. 175: 212 (1988). On the
day of the experiment, growth media was gently removed from the
well and growth media containing .sup.86Rubidium chloride (10.sup.6
.mu.Ci/mL) was added to each well. Cells were incubated at
37.degree. C. for a minimum of 3 h. After the loading period,
excess .sup.86Rb.sup.+ was removed and the cells were washed twice
with label-free Dulbecco's phosphate buffered saline (138 mM NaCl,
2.67 mM KCl, 1.47 mM KH.sub.2PO.sub.4, 8.1 mM Na.sub.2HPO.sub.4,
0.9 mM CaCl.sub.2, 0.5 mM MgCl.sub.2, Invitrogen/Gibco, pH. 7.4),
taking care not to disturb the cells. Next, cells were exposed to
either 100 .mu.M of test compound, 100 .mu.M of L-nicotine (Acros
Organics) or buffer alone for 4 min. Following the exposure period,
the supernatant containing the released .sup.86Rb.sup.+ was removed
and transferred to scintillation vials. Scintillation fluid was
added and released radioactivity was measured by liquid
scintillation counting.
[0247] Within each assay, each point had 2 replicates, which were
averaged. The amount of .sup.86Rb.sup.+ release was compared to
both a positive control (100 .mu.M L-nicotine) and a negative
control (buffer alone) to determine the percent release relative to
that of L-nicotine.
[0248] When appropriate, dose-response curves of test compound were
determined. The maximal activation for individual compounds (Emax)
was determined as a percentage of the maximal activation induced by
L-nicotine. The compound concentration resulting in half maximal
activation (EC.sub.50) of specific ion flux was also
determined.
Interaction at the Rat Ganglionic nAChR Subtype
[0249] Activation of rat ganglion nAChRs was established on the
pheochromocytoma clonal line PC12, which is a continuous clonal
cell line of neural crest origin, derived from a tumor of the rat
adrenal medulla. These cells express ganglion-like nAChRs (see
Whiting et al., Nature 327: 515 (1987); Lukas, J. Pharmacol. Exp.
Ther. 251: 175 (1989); Whiting et al., Mol. Brain. Res. 10: 61
(1990)).
[0250] Rat PC12 cells were maintained in proliferative growth phase
according to routine protocols (Bencherif et al., Mol. Cell.
Neurosci. 2: 52 (1991) and Bencherif et al., J. Pharmacol. Exp.
Ther. 257: 946 (1991)). Cells were cultured in Dulbecco's modified
Eagle's medium (Gibco/BRL) with 10% horse serum (Gibco/BRL), 5%
fetal bovine serum (HyClone, Logan Utah), 1 mM sodium pyruvate, 4
mM L-Glutamine, and 50,000 units penicillin-streptomycin (Irvine
Scientific). When cells were 80% confluent, they were plated to 6
well Nunc plates (Nunclon) and coated with 0.03% poly-L-lysine
(Sigma, dissolved in 100 mM boric acid). Experiments were conducted
when the cells reached 80% confluency.
[0251] Nicotinic acetylcholine receptor (nAChR) function was
assayed using .sup.86Rb.sup.+ efflux according to a method
described by Lukas et al., Anal. Biochem. 175: 212 (1988). On the
day of the experiment, growth media was gently removed from the
well and growth media containing .sup.86Rubidium chloride (10.sup.6
.mu.Ci/mL) was added to each well. Cells were incubated at
37.degree. C. for a minimum of 3 h. After the loading period,
excess .sup.86Rb.sup.+ was removed and the cells were washed twice
with label-free Dulbecco's phosphate buffered saline (138 mM NaCl,
2.67 mM KCl, 1.47 mM KH.sub.2PO.sub.4, 8.1 mM Na.sub.2HPO.sub.4,
0.9 mM CaCl.sub.2, 0.5 mM MgCl.sub.2, Invitrogen/Gibco, pH. 7.4),
taking care not to disturb the cells. Next, cells were exposed to
either 100 .mu.M of test compound, 100 .mu.M of nicotine or buffer
alone for 4 min. Following the exposure period, the supernatant
containing the released .sup.86Rb.sup.+ was removed and transferred
to scintillation vials. Scintillation fluid was added and released
radioactivity was measured by liquid scintillation counting.
[0252] Within each assay, each point had 2 replicates, which were
averaged. The amount of .sup.86Rb.sup.+ release was compared to
both a positive control (100 .mu.M nicotine) and a negative control
(buffer alone) to determine the percent release relative to that of
L-nicotine.
[0253] When appropriate, dose-response curves of test compound were
determined. The maximal activation for individual compounds (Emax)
was determined as a percentage of the maximal activation induced by
L-nicotine. The compound concentration resulting in half maximal
activation (EC.sub.50) of specific ion flux was also
determined.
Interaction at the Human Ganglionic nAChR Subtype
[0254] The cell line SH-SY5Y is a continuous line derived by
sequential subcloning of the parental cell line, SK-N-H, which was
originally obtained from a human peripheral neuroblastoma. SH-SY5Y
cells express a ganglion-like nAChR (Lukas et al., Mol. Cell.
Neurosci. 4: 1 (1993)).
[0255] Human SH-SY5Y cells were maintained in proliferative growth
phase according to routine protocols (Bencherif et al., Mol. Cell.
Neurosci. 2: 52 (1991) and Bencherif et al., J. Pharmacol. Exp.
Ther. 257: 946 (1991)). Cells were cultured in Dulbecco's modified
Eagle's medium (Gibco/BRL) with 10% horse serum (Gibco/BRL), 5%
fetal bovine serum (HyClone, Logan Utah), 1 mM sodium pyruvate, 4
mM L-Glutamine, and 50,000 units penicillin-streptomycin (Irvine
Scientific). When cells were 80% confluent, they were plated to 6
well polystyrene plates (Costar). Experiments were conducted when
the cells reached 100% confluency.
[0256] Nicotinic acetylcholine receptor (nAChR) function was
assayed using .sup.86Rb.sup.+ efflux according to a method
described by Lukas et al., Anal. Biochem. 175: 212 (1988). On the
day of the experiment, growth media was gently removed from the
well and growth media containing .sup.86Rubidium chloride (10.sup.6
.mu.Ci/mL) was added to each well. Cells were incubated at
37.degree. C. for a minimum of 3 h. After the loading period,
excess .sup.86Rb.sup.+ was removed and the cells were washed twice
with label-free Dulbecco's phosphate buffered saline (138 mM NaCl,
2.67 mM KCl, 1.47 mM KH.sub.2PO.sub.4, 8.1 mM Na.sub.2HPO.sub.4,
0.9 mM CaCl.sub.2, 0.5 mM MgCl.sub.2, Invitrogen/Gibco, pH 7.4),
taking care not to disturb the cells. Next, cells were exposed to
either 100 .mu.M of test compound, 100 .mu.M of nicotine, or buffer
alone for 4 min. Following the exposure period, the supernatant
containing the released .sup.86Rb.sup.+ was removed and transferred
to scintillation vials. Scintillation fluid was added and released
radioactivity was measured by liquid scintillation counting.
[0257] Within each assay, each point had 2 replicates, which were
averaged. The amount of .sup.86Rb.sup.+ release was compared to
both a positive control (100 .mu.M nicotine) and a negative control
(buffer alone) to determine the percent release relative to that of
L-nicotine.
[0258] When appropriate, dose-response curves of test compound were
determined. The maximal activation for individual compounds (Emax)
was determined as a percentage of the maximal activation induced by
L-nicotine. The compound concentration resulting in half maximal
activation (EC.sub.50) of specific ion flux was also defined.
Example 5
Determination of Binding at Non-Nicotinic Receptors
[0259] Muscarinic M3 Subtype
[0260] The human clonal line TE671/RD, derived from an embryonal
rhabdomyosarcoma (Stratton et al., Carcinogen 10: 899 (1989)), was
used to define binding to the muscarinic M3 receptor subtype. As
evidenced through pharmacological (Bencherif et al., J. Pharmacol.
Exp. Ther. 257: 946 (1991) and Lukas, J. Pharmacol. Exp. Ther. 251:
175 (1989)), electrophysiological (Oswald et al., Neurosci. Lett.
96: 207 (1989)), and molecular biological studies (Luther et al.,
J. Neurosci. 9: 1082 (1989)) these cells express muscle-like
nicotinic receptors.
[0261] TE671/RD cells were maintained in proliferative growth phase
according to routine protocols (Bencherif et al., Mol. Cell.
Neurosci. 2: 52 (1991) and Bencherif et al., J. Pharmacol. Exp.
Ther. 257: 946 (1991)). They were grown to confluency on 20-150 mm
tissue culture treated plates. The media was then removed and cells
scraped using 80 mL of PBS (Dulbecco's Phosphate Buffered Saline,
138 mM NaCl, 2.67 mM KCl, 1.47 mM KH.sub.2PO.sub.4, 8.1 mM
Na.sub.2HPO.sub.4, 0.9 mM CaCl.sub.2, 0.5 mM MgCl.sub.2,
Invitrogen/Gibco, pH 7.4) and then centrifuged at 1000 rpm for 10
mM. The supernatant was then suctioned off and the pellet(s) stored
at -20.degree. C. until use.
[0262] On the day of the assay, the pellets were thawed,
re-suspended with PBS and centrifuged at 18,000.times.g for 20 min,
then re-suspended in PBS to a final concentration of approximately
4 mg protein/mL and homogenized by Polytron. Protein was determined
by the method of Lowry et al., J. Biol. Chem. 193: 265 (1951),
using bovine serum albumin as the standard.
[0263] The binding of [.sup.3H]QNB was measured using a
modification of the methods of Bencherif et al., J. Pharmacol. Exp.
Ther. 257: 946 (1991). [.sup.3H]QNB (Specific Activity=30-60
Ci/mmol) was obtained from NEN Research Products. The binding of
[.sup.3H]QNB was measured using a 3 h incubation at 4.degree. C.
Incubations were conducted in 48-well micro-titre plates and
contained about 400 .mu.g of protein per well in a final incubation
volume of 300 .mu.L. The incubation buffer was PBS and the final
concentration of [.sup.3H]QNB was 1 nM. The binding reaction was
terminated by filtration of the protein containing bound ligand
onto glass fiber filters (GF/B, Brandel) using a Brandel Tissue
Harvester at 4.degree. C. Filters were pre-soaked in de-ionized
water containing 0.33% polyethyleneimine to reduce non-specific
binding. Each filter was washed with ice-cold buffer (3.times.1
mL). Non-specific binding was determined by inclusion of 10 .mu.M
non-radioactive atropine in selected wells.
[0264] The inhibition of [.sup.3H]QNB binding by test compounds was
determined by including seven different concentrations of the test
compound in selected wells. Each concentration was replicated in
triplicate. IC.sub.50 values were estimated as the concentration of
compound that inhibited 50 percent of specific [.sup.3H]QNB
binding. Inhibition constants (Ki values), reported in nM, were
calculated from the IC.sub.50 values using the method of Cheng et
al., Biochem. Pharmacol. 22: 3099 (1973).
Example 6
Determination of Activity at the .alpha.7 nAChR subtype
[0265] Selective .alpha.7 agonists can be found using a functional
assay on FLIPR (see, for example, PCT WO 00/73431 A2, the contents
of which are hereby incorporated by reference), which is a
commercially available high throughput assay (Molecular Devices
Corporation, Sunnyvale, Calif.). FLIPR is designed to read the
fluorescent signal from each well of a 96 or 384 well plate as fast
as twice a second for up to 30 minutes. This assay can be used to
accurately measure the functional pharmacology of .alpha.7 nAChR
and 5HT.sub.3R subtypes. Cell lines that express functional forms
of the .alpha.7 nAChR subtype, using the .alpha.7/5-HT.sub.3
channel as the drug target and/or cell lines that express
functional 5-HT.sub.3, are used to conduct the assay. In both
cases, the ligand-gated ion channels are expressed in SH-EP1 cells.
Both ion channels can produce a robust signal in the FLIPR assay.
Using the FLIPR assay, the compounds described herein can be
evaluated for their ability to function as agonists, partial
agonists or antagonists at the .alpha.7 nAChR subtype.
Example 7
Summary of Biological Activity
[0266] Compounds of the present invention exhibit Ki values at the
.alpha.7 subtype in the nM-M range, indicating that they have very
high affinity for the .alpha.7 nAChR subtype. High-throughput
screening indicated that none of the compounds bound to
.alpha.4.beta.2 nAChR subtypes with any significant affinity (Ki
values>10 .mu.M).
[0267] Compounds of the present invention exhibited little or no
agonist activity in functional models bearing muscle-type receptors
(.alpha.1.beta.1.gamma..delta. subtype in human TE671/RD clonal
cells), or ganglion-type receptors (.alpha.3.beta.4 subtype in the
Shooter subclone of rat pheochromocytoma PC12 cells and in human
SHSY-5Y clonal cells), generating only 1-12% (human muscle), 1-19%
(rat ganglion) and 1-15% (human ganglion) of nicotine's response at
these subtypes. These data indicate selectivity for CNS over PNS
nAChRs. Because similar compounds had been described by others as
exhibiting muscarinic activity (see, for instance, U.S. Pat. No.
5,712,270 to Sabb and PCTs WO 02/00652 and WO 02/051841),
representative compounds (#s 1, 2, 4, 9 and 11) were evaluated for
their ability to inhibit [.sup.3H]QNB binding at muscarinic sites
in the human clonal line TE671/RD. None of the compounds was able
to inhibit [.sup.3H]QNB binding, indicating that these compounds do
not bind to human M3 receptors. Thus, compounds of the present
invention are distinguished in their in vitro pharmacology from
reference compounds (see, for instance, U.S. Pat. No. 5,712,270 to
Sabb and PCTs WO 02/00652 and WO 02/051841) by virtue of the
inclusion, in their structure, of the 3-pyridinylmethyl substituent
in the 2 position of the 1-azabicycle.
[0268] The data show that the compounds of the present invention
are potent .alpha.7 nicotinic ligands that selectively bind at
.alpha.7 nAChR subtypes. In contrast, the compounds of the present
invention do not bind well at those subtypes of the nAChR that are
characteristic of the peripheral nervous system or at M3 muscarinic
receptors. Thus, the compounds of the present invention possess
therapeutic potential in treating central nervous system disorders
without producing side effects associated with interaction with the
peripheral nervous system. The affinity of these ligands for
.alpha.7 nAChR subtypes is tolerant of a wide variety of aryl (Ar
in Formula 1) groups and substituents thereon. Furthermore, the
synthesis is straightforward, efficient and amenable to massively
parallel protocols.
[0269] Having disclosed the subject matter of the present
invention, it should be apparent that many modifications,
substitutions and variations of the present invention are possible
in light thereof. It is to be understood that the present invention
can be practiced other than as specifically described. Such
modifications, substitutions and variations are intended to be
within the scope of the present application.
* * * * *