U.S. patent application number 15/220574 was filed with the patent office on 2016-11-17 for fluorinated arylalkylaminocarboxamide derivatives.
The applicant listed for this patent is NEWRON PHARMACEUTICALS S.P.A.. Invention is credited to Paolo Pevarello.
Application Number | 20160332959 15/220574 |
Document ID | / |
Family ID | 46210226 |
Filed Date | 2016-11-17 |
United States Patent
Application |
20160332959 |
Kind Code |
A1 |
Pevarello; Paolo |
November 17, 2016 |
FLUORINATED ARYLALKYLAMINOCARBOXAMIDE DERIVATIVES
Abstract
Fluorinated arylalkylaminocarboxamide derivatives of formula (I)
are described wherein W, J, n, R.sup.1, R.sup.2, R.sup.2', R.sup.3,
R.sup.4, R.sup.5 and R.sup.6 have the meanings as defined in the
specification and pharmaceutically salts thereof, pharmaceutical
compositions containing them as active ingredients and their use as
sodium and/or calcium channel modulators useful in preventing,
alleviating and curing a wide range of pathologies, including
neurological, psychiatric, cardiovascular, inflammatory,
ophthalmic, urolological, and gastrointestinal diseases, where the
above mechanisms have been described as playing a pathological
role. ##STR00001##
Inventors: |
Pevarello; Paolo; (Pavia
(PV), IT) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
NEWRON PHARMACEUTICALS S.P.A. |
Bresso (MI) |
|
IT |
|
|
Family ID: |
46210226 |
Appl. No.: |
15/220574 |
Filed: |
July 27, 2016 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
14115723 |
Nov 5, 2013 |
9447029 |
|
|
PCT/EP2012/060006 |
May 29, 2012 |
|
|
|
15220574 |
|
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C07D 205/04 20130101;
C07D 207/06 20130101; A61K 31/495 20130101; A61P 25/28 20180101;
A61K 31/4453 20130101; A61P 1/04 20180101; A61P 1/16 20180101; C07C
237/14 20130101; A61K 31/165 20130101; C07D 217/06 20130101; A61K
31/5375 20130101; A61P 27/02 20180101; A61K 31/40 20130101; C07C
237/20 20130101; A61K 31/47 20130101; A61P 25/18 20180101; A61P
15/00 20180101; A61P 25/00 20180101; A61K 31/538 20130101; C07C
237/06 20130101; A61P 13/10 20180101; C07D 211/16 20130101; A61P
5/00 20180101; C07D 295/182 20130101; A61P 25/04 20180101; A61P
13/00 20180101; A61P 1/08 20180101; A61P 25/06 20180101; C07C
2601/14 20170501; A61K 31/445 20130101; A61K 31/167 20130101; C07D
265/36 20130101; A61P 9/00 20180101; C07D 295/195 20130101; C07D
277/24 20130101; C07D 295/185 20130101; A61P 27/16 20180101; A61K
31/426 20130101; A61K 31/397 20130101; A61P 1/14 20180101; C07D
209/08 20130101; A61K 31/4045 20130101; C07C 237/04 20130101; A61P
29/00 20180101; A61K 31/472 20130101; A61P 43/00 20180101; A61P
25/08 20180101; C07D 295/26 20130101 |
International
Class: |
C07C 237/06 20060101
C07C237/06; C07D 277/24 20060101 C07D277/24; A61K 31/426 20060101
A61K031/426; C07D 295/195 20060101 C07D295/195; A61K 31/40 20060101
A61K031/40; C07C 237/04 20060101 C07C237/04; A61K 31/167 20060101
A61K031/167; A61K 31/5375 20060101 A61K031/5375; A61K 31/538
20060101 A61K031/538; C07D 265/36 20060101 C07D265/36; A61K 31/495
20060101 A61K031/495; A61K 31/4453 20060101 A61K031/4453; A61K
31/445 20060101 A61K031/445; C07D 211/16 20060101 C07D211/16; C07D
217/06 20060101 C07D217/06; A61K 31/472 20060101 A61K031/472; C07D
295/26 20060101 C07D295/26; C07D 209/08 20060101 C07D209/08; A61K
31/4045 20060101 A61K031/4045; C07D 205/04 20060101 C07D205/04;
A61K 31/397 20060101 A61K031/397; A61K 31/165 20060101
A61K031/165 |
Foreign Application Data
Date |
Code |
Application Number |
Jun 27, 2011 |
EP |
11171522.3 |
Claims
1. A compound of general formula I ##STR00067## wherein: W is a
group A-[(CH.sub.2).sub.m--O]-- wherein: m is zero, 1, 2, or 3; A
is (C.sub.1-C.sub.4)alkyl optionally substituted with one to three
fluorine atoms; (C.sub.3-C.sub.6)cycloalkyl; phenyl optionally
substituted with a group selected from halo, methyl, methoxy,
trifluoromethyl, acetylamino, and dimethylaminomethyl; thienyl
optionally substituted with a chloro group; furanyl; isoxazolyl,
thiazolyl; piperidinyl; morpholinyl; pyridinyl or pyrimidinyl, the
pyridinyl and pyrimidinyl ring being optionally substituted with
one or two methoxy groups; J independently is hydrogen,
(C.sub.1-C.sub.4)alkyl; (C.sub.1-C.sub.4)alkoxy; or an halo group;
n is 1 or 2; R.sup.1 is hydrogen; (C.sub.1-C.sub.4)alkyl optionally
substituted with a hydroxy group or a (C.sub.1-C.sub.4)alkoxy
group; or (C.sub.3-C.sub.8)cycloalkyl; R.sup.2 and R.sup.2' are
independently hydrogen; (C.sub.1-C.sub.4)alkyl optionally
substituted with a (C.sub.1-C.sub.4)alkoxy group; phenyl optionally
substituted with a (C.sub.1-C.sub.4)alkyl, a
(C.sub.1-C.sub.4)alkoxy or an halo group; benzyl optionally
substituted with a (C.sub.1-C.sub.4)alkyl, a
(C.sub.1-C.sub.4)alkoxy or an halo group on the benzene ring; or
R.sup.2 and R.sup.2' taken together with the adjacent carbon atom
form a (C.sub.3-C.sub.6)cycloalkylidene group. R.sup.3 is hydrogen;
or (C.sub.1-C.sub.4)alkyl; R.sup.4 is hydrogen;
(C.sub.1-C.sub.4)alkyl; phenyl; cyclohexyl; or benzyl; or R.sup.3
and R.sup.4, taken together with the adjacent nitrogen atom, form
an azetidinyl, pyrrolidinyl, morpholinyl, piperidinyl or
piperazinyl ring, the piperidinyl ring being optionally substituted
with one or two (C.sub.1-C.sub.2)alkyl group(s), and the
piperazinyl ring being optionally substituted on the other N-atom
with a (C.sub.1-C.sub.4)alkyl, benzyl, or phenylsulfonyl group; or
a pyrrolidinyl, piperidinyl, morpholinyl or piperazinyl ring fused
with a benzene ring; R.sup.5 is hydrogen or fluoro; and R.sup.6 is
fluoro; if the case, either as single optical isomer in the
isolated form or as a mixture thereof in any proportion and its
pharmaceutically acceptable salt.
2. A compound of claim 1, wherein: W is a group
A-[(CH.sub.2).sub.m--O]-- wherein: m is zero, 1, 2 or 3; A is
(C.sub.1-C.sub.4)alkyl optionally substituted by one to three
fluorine atoms; (C.sub.3-C.sub.6)cycloalkyl; phenyl optionally
substituted with a halo group; or thiazolyl J independently is
hydrogen; C.sub.1-C.sub.4 alkyl; chloro; or fluoro; n is 1 or 2;
R.sup.1 is hydrogen; (C.sub.1-C.sub.4)alkyl optionally substituted
with a hydroxy group or a (C.sub.1-C.sub.4)alkoxy group; or
(C.sub.3-C.sub.6)cycloalkyl; R.sup.2 is hydrogen; or
(C.sub.1-C.sub.4)alkyl; R.sup.2' is hydrogen or
(C.sub.1-C.sub.4)alkyl optionally substituted with a
(C.sub.1-C.sub.4)alkoxy or a phenyl group, the phenyl group being
optionally substituted with a (C.sub.1-C.sub.4)alkoxy group;
R.sup.3 is hydrogen; or (C.sub.1-C.sub.4)alkyl; R.sup.4 is
hydrogen; (C.sub.1-C.sub.4)alkyl; phenyl; or cyclohexyl; or R.sup.3
and R.sup.4, taken together with the adjacent nitrogen atom, form
an azetidinyl, pyrrolidinyl, morpholinyl, piperidinyl or
piperazinyl, the piperydinyl ring being optionally substituted with
one or two (C.sub.1-C.sub.2)alkyl group(s) and the piperazinyl ring
being optionally substituted on the other N-atom with a
(C.sub.1-C.sub.4)alkyl, benzyl, or phenylsulfonyl group; or a
pirrolidinyl, piperidinyl, morpholinyl or piperazinyl ring fused
with a benzene ring; R.sup.5 is hydrogen or fluoro; and R.sup.6 is
fluoro; if the case, either as single optical isomer in the
isolated form or as a mixture thereof in any proportion and its
pharmaceutically acceptable salt.
3. A compound of claim 2 wherein: W is a group
A-[(CH.sub.2).sub.m--O]-- wherein: m is 1 or 2; A is
(C.sub.1-C.sub.4)alkyl optionally substituted by one to three
fluorine atoms; phenyl optionally substituted with a chloro or
fluoro group; or thiazolyl; J independently is hydrogen; methyl; or
fluoro; n is 1-2 R.sup.1 is hydrogen; (C.sub.1-C.sub.4)alkyl
optionally substituted with a hydroxy group or a
(C.sub.1-C.sub.4)alkoxy group; R.sup.2 is hydrogen; or methyl;
R.sup.2' is hydrogen; or (C.sub.1-C.sub.4)alkyl optionally
substituted with a methoxy or a phenyl group, the phenyl group
being optionally substituted with a methoxy group; R.sup.3 is
hydrogen; or (C.sub.1-C.sub.4)alkyl; R.sup.4 is hydrogen;
(C.sub.1-C.sub.4)alkyl; phenyl; or cyclohexyl; or R.sup.3 and
R.sup.4, taken together with the adjacent nitrogen atom, form an
azetidinyl, pyrrolidinyl, morpholinyl, piperidinyl, or piperazinyl
ring, the piperidinyl ring being optionally substituted with one or
two methyl group(s) and the piperazinyl ring being optionally
substituted on the other N-atom with a methyl, benzyl or
phenylsulfonyl group; or a pirrolidinyl, piperidinyl, morpholinyl,
or piperazinyl ring fused with a benzene ring; R.sup.5 is hydrogen
or fluoro; and R.sup.6 is fluoro; if the case, either as single
optical isomer in the isolated form or as a mixture thereof in any
proportion and its pharmaceutically acceptable salt.
4. A compound of claim 3, selected from:
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide;
2-[2,2-Difluoro-2-(3-pentyloxyphenyl)-ethylamino]-N,N-dimethyl-acetamide
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dipropyl-acetamide
2-[2,2-Difluoro-2-(3-butoxy-4-methylphenyl)-ethylamino]-N,N-dimethyl-acet-
amide;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dibutyl-acetamid-
e;
2-[2,2-Difluoro-2-(3-hexyloxyphenyl)-ethylamino]-N,N-dimethyl-acetamide-
;
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-N,N-dim-
ethyl-acetamide;
2-[2,2-Difluoro-2-(3-pentyloxyphenyl)-ethylamino]-N,N-dipropyl-acetamide;
2-{2,2-Difluoro-2-[3-(3-(3-fluorophenyl)-propoxy)-phenyl]-ethylamino}-N,N-
-dimethyl-acetamide;
2-{2,2-Difluoro-2-[3-(3-(3-chlorophenyl)-propoxy)-phenyl]-ethylamino}-N,N-
-dimethyl-acetamide;
2-[2,2-Difluoro-2-(3-butoxy-2-fluorophenyl)-ethylamino]-N,N-dimethyl-acet-
amide;
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-N,N-dime-
thyl-acetamide;
2-{2,2-Difluoro-2-[3-(3-thiazol-2-yl-propoxy)-phenyl]-ethylamino}-N,N-dim-
ethyl-acetamide;
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-N,N-dimethyl-acetamide;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(pyrrolidin-1-yl)-ethano-
ne;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N-methyl-N-phenyl-aceta-
mide;
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(pyrrol-
idin-1-yl)-ethanone;
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(pyrro-
lidin-1-yl)-ethanone;
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-1-(morpholin-4-yl)-etha-
none;
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(morpho-
lin-4-yl)-ethanone;
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(morph-
olin-4-yl)-ethanone;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(2H-benzo[b][1,4]oxazin--
4(3H)-yl)-ethanone;
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-1-(pyrrolidin-1-yl)-eth-
anone;
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-N-methyl-
-N-phenyl-acetamide;
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-N-methyl-
-N-phenyl-acetamide;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(4-methylpiperazin-1-yl)-
-ethanone;
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(4-
-methylpiperazin-1-yl)-ethanone;
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(4-met-
hylpiperazin-1-yl)-ethanone;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(piperidin-1-yl)-ethanon-
e;
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(piperidin-
-1-yl)-ethanone;
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(piper-
idin-1-yl)-ethanone;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-diethyl-acetamide;
2-{2,2-Difluoro-2-[3-(2-fluorobenzyloxy)-phenyl]-ethylamino}-N,N-dimethyl-
-acetamide;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(cis-3,5-dimethylpiperid-
in-1-yl)-ethanone;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(3,4-dihydroisoquinolin--
2(1H)-yl)-ethanone;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-diisopropyl-acetamide;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N-cyclohexyl-N-methyl-acet-
amide;
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-1-(piperidin-1-yl-
)-ethanone;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-[4-(phenylsulfonyl)-pipe-
razin-1-yl]-ethanone;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(indolin-1-yl)-ethanone;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(4-benzylpiperazin-1-yl)-
-ethanone;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(azetidin-1-yl-
)-ethanone;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-propanamide;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-3-methoxy-N,N-dimethyl-pro-
panamide;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-3-(4-methoxypheny-
l)-N,N-dimethyl-propanamide;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-2-N,N-trimethyl-propanamid-
e;
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-4-N,N-trimethyl-pentanam-
ide;
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-methylamino}-N,N-dimethyl--
acetamide;
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-(3-methoxypropyl)-am-
ino}-N,N-dimethyl-acetamide;
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-(2-methoxyethyl)-amino}-N,N-di-
methyl-acetamide;
2-[2-Fluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide;
2-{2-Fluoro-2-[3-(3-chlorobenzyloxy)-phenyl]-ethylamino}-N,N-dimethyl-ace-
tamide;
2-{2-Fluoro-2-[3-(3-fluorobenzyloxy)-phenyl]-ethylamino}-N,N-dimet-
hyl-acetamide; if the case, either as single optical isomer in the
isolated form or a mixture thereof in any proportion, and its
pharmaceutically acceptable salt.
5. A compound of claim 4 which is selected from
2-[2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide,
2-[2-fluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide,
its single optical isomer in the isolated form or a mixture thereof
in any proportion, and the pharmaceutically acceptable salts
thereof.
6. A compound of claim 5 which is
2-[2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide
or a pharmaceutically acceptable salt thereof.
7. A compound as in claim 1 wherein the pharmaceutically acceptable
salt is the hydrochloride.
8. A pharmaceutical composition containing a compound of claim 1 as
active ingredient together with a pharmaceutically acceptable
excipient.
9. A pharmaceutical composition of claim 8 containing a further
therapeutical agent.
10. A method for the treatment of a disorder caused by dysfunctions
of voltage gated sodium and/or calcium channels in a patient, said
method comprising the administration to a patient in need thereof
an effective amount of a sodium and/or calcium channel modulating
amount of a compound of claim 1.
11. A method as in claim 10 wherein the disorder caused by
dysfunctions of voltage gated sodium and/or calcium channels is
selected from neuropathic pain, chronic pain, acute pain,
headaches, neurological conditions, neurogenerative disorders,
cognitive disorders, psychiatric disorders, vertigo, tinnitus,
muscle spasm, cardiovascular diseases, endocrine disorders
involving excessive or hypersecretory or otherwise inappropriate
cellular secretion of an endogenous substance, liver diseases,
inflammatory processes affecting all body systems, disorders of the
gastrointestinal (GI) tract, disorders of the genito-urinary tract,
ophthalmic diseases and eating disorders.
12. A method as in claim 11 wherein the disorder is a neuropathic
pain, chronic pain and/or acute pain.
13. A method as claim 11 wherein the disorder is headache.
14. A method as in claim 11 wherein the disorder is a cognitive
and/or psychiatric disorder.
15. A method as in claim 11 wherein the disorder is a neurological
condition.
16. A method as claim 11 wherein the disorder is an inflammatory
process affecting all body systems, a disorder of the
gastrointestinal tract, a disorder of the genito-urinary tract, an
ophthalmic disease, a liver disease, a cardiovascular, and/or
neurodegenerative disorder caused by dysfunctions of voltage gated
sodium and/or calcium channels.
17. A method as in claim 11 wherein the patient is a poor
metabolizer, having very little or no CYP2D6 function, or is
assuming drug(s) that is(are) CYP2D6 inhibitor(s).
18. A method as in claim 15, wherein the neurological condition is
epilepsy.
19. A method as in claim 11, wherein the patient is administered a
further therapeutical agent.
20. A pharmaceutical composition of claim 8, wherein the compound
is in the form of a pharmaceutically acceptable salt.
21. A pharmaceutical composition of claim 20, wherein the
pharmaceutically acceptable salt is hydrochloride.
22. A method as in claim 10, wherein the compound is in the form of
a pharmaceutical acceptable salt.
23. A method as in claim 22, wherein the pharmaceutically
acceptable salt is hydrochloride.
24. A compound of claim 7 which is the hydrochloride salt of
2-[2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide.
25. A pharmaceutical composition of claim 21, wherein the compound
is the hydrochloride salt of
2-[2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide.
26. The method as in claim 23, wherein the pharmaceutically
acceptable salt is they hydrochloride salt of
2-[2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide.
Description
[0001] This is a non-provisional application of U.S. application
Ser. No. 14/115,723 filed Nov. 5, 2013, which is a U.S. National
stage application of PCT/EP2012/060006 filed on May 29, 2012, which
claims priority to and the benefit of European Application No.
11171522.3 filed on Jun. 27, 2011, the contents of which are all
incorporated herein by reference in their entireties.
[0002] The present invention relates to fluorinated
arylalkylaminocarboxamide derivatives, pharmaceutically acceptable
salts thereof, pharmaceutical compositions containing them and
their use as sodium and/or calcium channel modulators.
[0003] The fluorinated arylalkylaminocarboxamide derivatives,
object of the invention, are active as ion channel (in particular
as sodium and/or calcium channel) modulators, and therefore useful
in preventing, alleviating and curing a wide range of pathologies,
including but not limited to neurological, psychiatric,
cardiovascular, inflammatory, ophthalmic, urogenital and
gastrointestinal diseases where the above mechanisms have been
described as playing a pathological role.
BACKGROUND OF THE INVENTION
Chemical Background
[0004] WO 2007/071311 describes, substituted 2-phenylethylamino
derivatives as voltage-gated calcium and/or sodium channels
modulators of general formula I
##STR00002##
wherein: (a) [0005] J is a group A-[(CH.sub.2).sub.n--O].sub.r-- in
para position with respect to the ethylamino chain wherein: [0006]
n is zero or 1; [0007] r is 1; and [0008] A is trifluromethyl;
cyclopentyl; or phenyl optionally substituted with a halo group;
[0009] W is (C.sub.1-C.sub.4)alkoxy; [0010] R is hydrogen; [0011]
R.sup.0 is hydrogen; or (C.sub.1-C.sub.2)alkyl; [0012] R.sup.1 is
hydrogen; (C.sub.1-C.sub.4)alkyl optionally substituted with a
hydroxy group; cyclopropylmethyl; 2-propyn-1-yl; benzyl optionally
substituted with one or two (C.sub.1-C.sub.2)alkoxy groups on the
benzene ring; thiazolyl; a 5-6 membered saturated heterocyclyl
containing a nitrogen atom, optionally substituted with a
(C.sub.1-C.sub.2)alkyl group; or heterocyclylmethyl wherein the
heterocyclyl group is a 5-6 membered heterocylyl containing 1 to 3
hetero atoms selected from nitrogen, oxygen and sulfur, optionally
substituted with one or two groups selected from
(C.sub.1-C.sub.2)alkyl, hydroxymethyl and (C.sub.1-C.sub.2)alkoxy;
[0013] R.sup.2 is hydrogen; (C.sub.1-C.sub.4)alkyl; or phenyl;
[0014] R.sup.3 is hydrogen; or (C.sub.1-C.sub.4)alkyl; and [0015]
R.sup.4 is hydrogen; (C.sub.1-C.sub.4)alkyl optionally substituted
with a group selected from amino, (C.sub.1-C.sub.4)alkylamino,
di-(C.sub.1-C.sub.4)alkylamino, imidazolyl and pyrrolidinyl wherein
the imidazolyl and the pyrrolidinyl group is optionally substituted
with a (C.sub.1-C.sub.2)alkyl group; or benzyl; or [0016] R.sup.3
and R.sup.4, taken together with the adjacent nitrogen atom, form a
pyrrolidinyl, morpholinyl or piperazinyl ring optionally
substituted with a (C.sub.1-C.sub.2)alkyl group; or (b) [0017] J is
a group A-[(CH.sub.2).sub.n--O].sub.r-- in para position with
respect to the ethylamino chain wherein: [0018] n is 1; [0019] r is
1; and [0020] A is phenyl; or phenyl substituted with a halo group;
[0021] W is hydrogen; [0022] R is hydrogen; [0023] R.sup.0 is
(C.sub.1-C.sub.2)alkyl; [0024] R.sup.1 is hydrogen; [0025] R.sup.2
is (C.sub.1-C.sub.2)alkyl; [0026] R.sup.3 is hydrogen; or
(C.sub.1-C.sub.4) alkyl; and [0027] R.sup.4 is hydrogen; or
(C.sub.1-C.sub.4)alkyl; or (c) [0028] J is hydrogen; [0029] W is a
group A-[(CH.sub.2).sub.n--O].sub.r-- wherein: [0030] n is zero, 1
or 2; [0031] r is zero or 1; and [0032] A is
(C.sub.1-C.sub.4)alkyl, trifluoromethyl; cyclopropyl; cyclopentyl;
phenyl optionally substituted with a group selected from halo,
methyl, methoxy, trifluoromethyl, acetylamino, and
dimethylaminomethyl; thienyl optionally substituted with a chloro
group; furanyl; isoxazolyl optionally substituted with one or two
methyl groups; piperidinyl; morpholinyl; pyridinyl or pyrimidinyl,
the pyridinyl and pyrimidinyl ring being optionally substituted
with one or two methoxy groups; [0033] R is hydrogen; or fluoro;
[0034] R.sup.0 is hydrogen; or (C.sub.1-C.sub.2)alkyl; [0035]
R.sup.1 is isopropyl; cyclopropylmethyl; furanylmethyl;
tetrahydrofuranyl; or tetrahydrofuranylmethyl; [0036] R.sup.2 is
hydrogen; or (C.sub.1-C.sub.4)alkyl; [0037] R.sup.3 is hydrogen; or
(C.sub.1-C.sub.4)alkyl; and [0038] R.sup.4 is hydrogen;
(C.sub.1-C.sub.4)alkyl optionally substituted with a group selected
from (C.sub.1-C.sub.2)alkoxy, amino, (C.sub.1-C.sub.4)alkylamino
and di-(C.sub.1-C.sub.4)alkylamino; or heterocyclyl wherein the
heterocyclyl is selected from isoxazolyl, pyrazolyl, imidazolyl,
thiazolyl and 1,3,4 thiadiazolyl and may be optionally substituted
with a (C.sub.1-C.sub.2)alkyl group; or [0039] R.sup.3 and R.sup.4
taken together with the adjacent nitrogen atom form a pyrrolidine
ring; with the proviso that when A is (C.sub.1-C.sub.4)alkyl,
trifluoromethyl, cyclopropyl or cyclopentyl, then r is 1; and with
the further proviso that when R.sup.1 is isopropryl, then A is
trifluoromethyl and n is 1; used for the manufacture of a
medicament active as calcium and/or sodium channel modulators
against disorders caused by dysfunctions of voltage gated calcium
and/or sodium channels.
[0040] WO 2008/151702 describes substituted
2-[2-(phenyl)-ethylamino]alkaneamide derivatives as voltage-gated
calcium and/or sodium channels modulators of general formula
(I)
##STR00003##
wherein: [0041] X is --O--, --S-- or --SO.sub.2--; [0042] Y is
hydrogen, OH or O(C.sub.1-C.sub.4)alkyl; [0043] Z is .dbd.O or
.dbd.S; [0044] R is (C.sub.3-C.sub.10)alkyl;
.omega.-trifluoro(C.sub.3-C.sub.10)alkyl; [0045] R.sub.1 and
R.sub.2 are, independently, hydrogen, hydroxy,
(C.sub.1-C.sub.8)alkoxy, (C.sub.1-C.sub.8) alkylthio, halo,
trifluoromethyl or 2,2,2-trifluoroethyl; or one of R.sub.1 and
R.sub.2 is in ortho position to R--X-- and, taken together with the
same R--X--, represents a
[0045] ##STR00004## group where R.sub.0 is (C.sub.2-C.sub.9)alkyl;
[0046] R.sub.3 and R'.sub.3 are, independently, hydrogen or
(C.sub.1-C.sub.4)alkyl; [0047] R.sub.4 and R.sub.5 are,
independently, hydrogen, (C.sub.1-C.sub.4)alkyl; or R.sub.4 is
hydrogen and R.sub.5 is a group selected from --CH.sub.2--OH,
--CH.sub.2--O--(C.sub.1-C.sub.6)alkyl, --CH(CH.sub.3)--OH, [0048]
--(CH.sub.2).sub.2--S--CH.sub.3, benzyl and 4-hydroxybenzyl; or
R.sub.4 and R.sub.5, taken together with the adjacent carbon atom,
form a (C.sub.3-C.sub.6)cycloalkyl residue; [0049] R.sub.6 and
R.sub.7 are independently hydrogen or (C.sub.1-C.sub.6)alkyl; or
taken together with the adjacent nitrogen atom form a 5-6 membered
monocyclic saturated heterocycle, optionally containing one
additional heteroatom chosen among --O--, --S-- and --NR.sub.8--
where R.sub.8 is hydrogen or (C.sub.1-C.sub.6) alkyl; with the
proviso that when X is --S-- or --SO.sub.2--, then Y is not OH or
O(C.sub.1-C.sub.4) alkyl; if the case, either as single optical
isomer in the isolated form or mixture thereof in any proportion
and its pharmaceutically acceptable salts.
[0050] The fluorinated compounds described in this application are
not comprised by either WO 2007/071311 or WO 2008/151702.
Biological Background
[0051] Sodium channels play an important role in the neuronal
network by transmitting electrical impulses rapidly throughout
cells and cell networks, thereby coordinating higher processes
ranging from locomotion to cognition. These channels are large
transmembrane proteins, which are able to switch between different
biophysical states to enable selective permeability for sodium
ions. For this process to occur an action potential is needed to
depolarize the membrane, and hence these channels are said
voltage-gated.
[0052] Voltage-gated sodium channels were originally classified
based on their sensitivity to tetrodotoxin, from low nanomolar
(Tetrodotoxin sensitive, TTXs) to high micromolar (Tetrodotoxin
resistant, TTXr). So far, 10 different sodium channel a subunits
have been identified and classified as Nav1.1 to Nav1.9.
[0053] Nav1.1 to Nav1.4, Nav1.6 and Nav1.7 are TTXs, whereas
Nav1.5, Nav1.8 and Nav.1.9 are TTXr, with different degrees of
sensitivity. Nav1.1 to Nav1.3 and Nav1.6, are primarily expressed
in the CNS, whereas Nav1.4 and Nav1.5 are mainly expressed in
muscle (skeletal and heart, respectively) and Nav1.8 and Nav1.9 are
predominantly expressed in small dorsal root ganglions (DRG).
[0054] Nav1.3, a TTX-s sodium channel normally absent in adult
neurons, is up-regulated following nerve injury as observed in the
sensory neurons and spinal cord neurons of rodents following
chronic nerve injuries (Waxman S. G., Kocsis J. D., Black J. A.:
"Type III sodium channel mRNA is expressed in embryonic but not in
adult spinal sensory neurons, and is reexpressed following
axotomy". J. Neurophysiol. 72, 466-470 (1994). Hains B. C., Klein
J. P., Saab C. Y. et al.: "Upregulation of sodium channel Nav1.3
and functional involvement in neuronal hyperexcitability associated
with central neuropathic pain after spinal cord injury". J.
Neurosci. 23, 8881-8892 (2003). Hains B. C., Saab C. Y., Klein J.
P. et al.: "Altered sodium channel expression in second-order
spinal sensory neurons contributes to pain after peripheral nerve
injury". J. Neurosci. 24, 4832-4839 (2004)) and confirmed in human
injured nerves after peripheral axotomy (Coward K., Aitken A.,
Powell A. et al.: "Plasticity of TTX-sensitive sodium channels PNI
and brain III in injured human nerves". Neuroreport 12, 495-500
(2001)) and in human painful neuromas (Black J. A., Nikolajsen L.,
Kroner K. et al.: "Multiple sodium channel isoforms and
mitogen-activated protein kinases are present in painful human
neuromas". Ann. Neurol. 64(6), 644-653 (2008)). Nav1.3 channels
exhibit several properties that can contribute to neuronal
hyperexcitability. The rapid recovery from inactivation, and the
ability to produce persistent current and ramp responses to
small/slow depolarizations can support high-frequency firing.
Interestingly, increased recovery rates have been described after
nerve injury that would contribute to increase neuronal
excitability in pain conditions (Cummins T. R., Waxman S. G.:
"Downregulation of Tetrodotoxin resistant sodium currents and
upregulation of a rapidly repriming tetrodotoxin-sensitive sodium
current in small spinal sensory neurons after nerve injury". J.
Neurosci. 17, 3503-3514 (1997). Cummins T. R., Aglieco F.,
Renganathan M. et al.: "Nav1.3 sodium channels: rapid repriming and
slow closed-state inactivation display quantitative differences
after expression in a mammalian cell line and in spinal sensory
neurons". J. Neurosci. 21, 5952-5961 (2001). Lampert A., Hains B.
C., Waxman S. G.: "Upregulation of persistent and ramp sodium
current in dorsal horn neurons after spinal cord injury". Exp.
Brain Res. 174, 660-666 (2006)).
[0055] Overall the specific expression and the biophysical
properties of the Nav1.3 would implicate this channel in the
generation of the TTX-sensitive ectopic discharges associated with
chronic pain.
[0056] Nav1.7 channel is a TTX-s channel preferentially expressed
in the primary DRG nociceptor neurons and in sympathetic ganglion
neurons. It displays slow kinetics of transition to and from the
inactivation state, that determine the possibility of generating
currents in response to small subthreshold depolarizations and
allow the channel to act as a threshold channel, thus amplifying
generator potentials (Catterall W. A., Goldin A. L., Waxman S. G.:
"International Union of Pharmacology. XLVII. Nomenclature and
Structure-Function Relationships of Voltage-Gated Sodium Channels".
Pharmacol. Rev. 57, 397-409 (2005)). Over the past few years,
Nav1.7 has gained a prominent role in pain research because human
genetic studies have directly linked single point mutations of the
SCN9A gene encoding for Nav1.7 to specific pain syndromes. Gain of
function mutations, that lower the threshold for channel
activation, are associated to a dominant-inherited neuropathy,
inherited erythromelalgia (IEM) whose hallmark symptom is severe
burning pain in the feet and hands in response to mild warmth and
exercise (Dib-Hajj S. D., Rush A. M., Cummins T. R. et al.:
"Gain-of-function mutation in Nav1.7 in familial erythromelalgia
induces bursting of sensory neurons". Brain 128(8), 1847-1854
(2005). Dib-Hajj S. D., Rush A. M., Cummins T. R., Waxman S. G.:
"Mutations in the sodium channel Nav1.7 underlie inherited
erythromelalgia". Drug Discovery Today: Disease Mechanisms 3(3),
343-350 (2006)).
[0057] One of the most compelling evidence that encouraged many
companies to pursue research programs towards Nav1.7 specific
inhibitors, has been the discovery that loss of function mutations
of Nav1.7 gene determines a congenital insensitivity to pain (CIP)
(Cox J. J., Reimann F., Nicholas A. K. et al.: "An SCN9A
channelopathy causes congenital inability to experience pain".
Nature 444, 894-8 (2006).
[0058] The TTX-r channel Nav1.8 is exclusively expressed in the
peripheral sensory neurons. Slow inactivation kinetic, rapid
repriming, depolarized threshold of activation and inactivation,
make it ideal for maintaining action potential firing in
depolarized fibers (Elliott A. A., Elliott J. R.: "Characterization
of TTX-sensitive and TTX-resistant sodium currents in small cells
from adult rat dorsal root ganglia". J. Physiol. 463, 39-56 (1993).
Akopian A. N., Souslova V., England S. et al.: "The
tetrodotoxin-resistant sodium channel SNS has a specialized
function in pain pathways". Nat. Neurosci. 2, 541-548 (1999).
Renganathan M., Cummins T. R., Waxman S. G.: "Contribution of
Nav1.8 sodium channels to action potential electrogenesis in DRG
neurons". J. Neurophysiol. 86, 629-640 (2001)). However, the
specific translocation and redistribution of Nav1.8 protein at the
peripheral site of injury observed in immunohistochemical studies
in animals and recently in humans (Novakovic S. D., Tzoumaka E.,
McGivern J. G. et al.: "Distribution of the tetrodotoxin-resistant
sodium channel PN3 in rat sensory neurons in normal and neuropathic
conditions". J. Neurosci. 15, 18(6) 2174-2187 (1998). Black J. A.,
Nikolajsen L., Kroner K. et al.: "Multiple sodium channel isoforms
and mitogen-activated protein kinases are present in painful human
neuromas". Ann. Neurol. 64(6), 644-653 (2008)), or the
redistribution and alterations of its activity in the remaining
uninjured neurons (Gold M., Weinreich D., Kim C. S. et al.:
"Redistribution of Nav 1.8 in uninjured axons enables neuropathic
pain". J. Neurosci. 23, 158-166 (2003)), suggests a dynamic
involvement of this channel in the generation and maintenance of
nociceptive impulses.
[0059] Another TTX-r channel, Nav1.9, is exclusively expressed in
small-diameter DRG neurons. It is still one of the least understood
members of the voltage gated sodium channels (VGSC) family, due to
the difficulty to express the recombinant form in heterologous
expression systems. Characterization of the biophysical properties
of this channel was done in sensory neurons from Nav 1.8-null mice
(Cummins T. R., Dib-Hajj S. D., Black J. A. et al.: "A novel
persistent tetrodotoxin-resistant sodium current in SNS-null and
wild-type small primary sensory neurons". J. Neurosci. 19 (24):RC43
(1999)). These neurons were shown to express a persistent
(non-inactivating) TTX-r current, with substantial overlap between
activation and steady-state inactivation centered close to resting
potential (Roza C., Laird J. M. A., Souslova V. et al.: "The
tetrodotoxin-resistant Na+ channel Nav1.8 is essential for the
expression of spontaneous activity in damaged sensory axons of
mice". J. Physiol. 550, 921-926 (2003)). As a result of these
properties, Nav1.9 channels behave as strong regulators of
excitability in cells in which they are present, playing a key role
in setting the resting membrane potential as well as contributing
to sub-threshold electrogenesis in small DRG neurons.
[0060] It has become clear that a number of drugs having a
previously unknown mechanism of action actually act by modulating
sodium channel conductance, including local anaesthetics (LA),
class I antiarrhythmics and anticonvulsants. Neuronal sodium
channel blockers have found application with their use in the
treatment of epilepsy (phenytoin and carbamazepine), bipolar
disorder (lamotrigine), preventing neurodegeneration, and in
reducing neuropathic pain. Various anti-epileptic drugs that even
through other mechanisms of action, stabilize neuronal excitability
are approved for different forms of neuropathic pain (gabapentin,
pregabalin and carbamazepine).
[0061] In addition, an increase in sodium channel expression and/or
activity has also been observed in several models of inflammatory
pain, suggesting a role of sodium channels in inflammatory
pain.
[0062] Calcium channels are membrane-spanning, multi-subunit
proteins that allow controlled entry of calcium ions into cells
from the extracellular fluid. Commonly, calcium channels are
voltage dependent and are referred to as voltage-gated calcium
channels (VGCC). VGCCs are found throughout the mammalian nervous
system, where they regulate the intracellular calcium ion levels
that are important for cell viability and function. Intracellular
calcium ion concentrations are implicated in a number of vital
processes in animals, such as neurotransmitter release, muscle
contraction, pacemaker activity and secretion of hormones. All
"excitable" cells in animals, such as neurons of the central
nervous system (CNS), peripheral nerve cells, and muscle cells,
including those of skeletal muscles, cardiac muscles and venous and
arterial smooth muscles, have voltage dependent calcium
channels.
[0063] Calcium channels are a large family with many genetically,
physiologically, and pharmacologically distinct subtypes. Based on
the biophysical properties of calcium currents recorded from
individual neurons, two super-families have been described: High
Voltage Activated (HVA) and Low Voltage Activated (LVA) calcium
channels. Calcium currents referred as L-type, P-type, Q-type,
N-type, R-type are HVA and as T-type are LVA. In particular, the
term "L-type" was originally applied to channels with a large
single channel conductance and long open time, and "T-type" was
applied to channels with a tiny single channel conductance and a
transient open time. Further exploration of functional calcium
channel diversity identified the "N-type" channel expressed in
neurons and the "P-type" channel, which is the dominant type
expressed in cerebellar Purkinje neurons and is pharmacologically
resistant to known blockers of L-type and N-type calcium channels.
From the molecular identity, ten distinct calcium channel subtypes
have been identified, cloned and expressed and grouped in three
families: Cav1 family (Cav 1.1, 1.2, 1.3, 1.4) is functionally
related to the L-type Ca current; Cav2 family (Cav 2.1, 2.2, 2.3)
is functionally related to the P/Q, N, R-type currents and Cav3
(Cav 3.1, 3.2, 3.3) family is functionally related to the T-type
current.
[0064] It is believed that calcium channels are relevant in certain
disease states. A number of compounds useful in treating various
cardiovascular diseases in mammals, including humans, are thought
to exert their beneficial effects by modulating functions of
voltage dependant calcium channels present in cardiac and/or
vascular smooth muscle. Compounds with activity against calcium
channels have also been implicated for the treatment of pain. In
particular N-type calcium channels (Cav2.2), responsible for the
regulation of neurotransmitter release, are thought to play a
significant role in nociceptive transmission, both due to their
tissue distribution as well as from the results of several
pharmacological studies. N-type calcium channels were found
up-regulated in the ipsilateral dorsal horn in neuropathic pain
models of injury (Cizkova D., Marsala J., Lukacova N., Marsala M.,
Jergova S., Orendacova J., Yaksh T. L. Exp. Brain Res. 147: 456-463
(2002)). Specific N-type calcium channel blockers were shown to be
effective in reducing pain responses in neuropathic pain models
(Mattews E. A., Dickenson A. H. Pain 92: 235-246 (2001)), in the
phase II of the formalin test (Diaz A., Dickenson A. H. Pain 69:
93-100 (1997)) and the hyperalgesia initiated by knee joint
inflammation (Nebe J., Vanegas H., Schaible H. G. Exp. Brain Res.
120: 61-69 (1998)). Mutant mice, lacking the N-type calcium
channels, were found to have a decreased response to persistent
pain as seen by a decrease in pain response during phase II of the
formalin test (Kim C., Jun K., Lee T., Kim S. S., Mcenery M. W.,
Chin H., Kim H. L, Park J. M., Kim D. K., Jung S. J., Kim J., Shin
H. S. Mol. Cell Neurosci. 18: 235-245 (2001); Hatakeyama S.,
Wakamori M., Ino M., Miyamoto N., Takahashi E., Yoshinaga T.,
Sawada K., Imoto K., Tanaka I., Yoshizawa T., Nishizawa Y., Mori
Y., Nidome T., Shoji S. Neuroreport 12: 2423-2427 (2001)) as well
as to neuropathic pain, assessed by a decrease in mechanical
allodynia and thermal hyperalgesia in the spinal nerve ligation
model (Yamamoto T., Takahara A.: "Recent updates of N-type calcium
channel blockers with therapeutic potential for neuropathic pain
and stroke". Curr. Top. Med. Chem. 9, 377-395 (2009)).
Interestingly, mice also showed lower levels of anxiety when
compared to wild type (Saegusa H., Kurihara T., Zong S., Kazuno A.,
Matsuda Y. Nonaka T., Han W., Toriyama H., Tanabe T., EMBO J. 20:
2349-2356 (2001)). The involvement of N-type calcium channels in
pain has been further validated in the clinic by ziconotide, a
peptide derived from the venom of the marine snail, Conus Magnus.
(Williams J. A., Day M., Heavner J. E.: "Ziconotide: an update and
review". Expert Opin. Pharmacother. 9(9), 1575-1583 (2008)). A
limitation in the therapeutic use of this peptide is that it has to
be administered intrathecally in humans (Bowersox S. S. and Luther
R. Toxicon, 36: 1651-1658 (1998); Vitale V., Battelli D., Gasperoni
E., Monachese N.: "Intrathecal therapy with ziconotide: clinical
experience and consideration on its use". Minerva Anestesiol. 74,
727-733 (2008)).
[0065] A comprehensive review on the role and usefulness of ion
channel modulators in neuropathic pain treatment has recently been
published. (E. Colombo et al.: "Ion channel modulators for the
treatment of neuropathic pain". Future Medicinal Chemistry, 2(5):
803-842 (2010)).
[0066] All together these findings indicate that compounds able to
block sodium and/or calcium channels have an important therapeutic
potential in preventing, alleviating and curing a wide range of
pathologies, including neurological, psychiatric, cardiovascular,
urogenital gastrointestinal and inflammatory diseases, where the
above mechanisms have been described as playing a pathological
role.
[0067] There are many papers and patents which describe sodium
channel and/or calcium channel modulators or antagonists for the
treatment or modulation of a plethora of disorders, such as their
use as local anaesthetics, antiarrhythmics, antiemetics, antimanic
anti-depressants, agents for the treatment of unipolar depression,
anxiety, cardiovascular diseases, urinary incontinence, diarrhoea,
inflammation, epilepsy, neurodegenerative conditions, nerve cell
death, neuropathic pain, migraine, acute hyperalgesia and
inflammation, renal disease, allergy, asthma, bronchospasm,
dysmenorrhea, esophageal spasm, glaucoma, urinary tract disorders,
gastrointestinal motility disorders, premature labour, obesity,
immune and endocrinological system disorders, including multiple
sclerosis.
[0068] A non-exhaustive list of patents/patent applications
describing sodium and/or calcium channel blockers and uses thereof
includes the references shown below.
[0069] U.S. Pat. No. 5,051,403 relates to a method of reducing
neuronal damage associated with an ischemic condition, such as
stroke, by administration of binding/inhibitory omega-conotoxin
peptide wherein the peptide is characterized by specific inhibition
of voltage-gated calcium channel currents selectively in neuronal
tissues.
[0070] U.S. Pat. No. 5,587,454 relates to compositions and methods
of producing analgesia particularly in the treatment of pain and
neuropathic pain.
[0071] U.S. Pat. No. 5,863,952 relates to calcium channel
antagonists for the treatment of ischaemic stroke.
[0072] U.S. Pat. No. 6,011,035 relates to calcium channel blockers,
useful in the treatment of conditions such as stroke and pain.
[0073] U.S. Pat. No. 6,117,841 relates to calcium channel blockers
and their use in the treatment of stroke, cerebral ischemia, pain,
head trauma or epilepsy.
[0074] U.S. Pat. No. 6,362,174 relates to N-type calcium channel
blockers in the treatment of stroke, cerebral ischemia, pain,
epilepsy, and head trauma.
[0075] U.S. Pat. No. 6,380,198 concerns the use of the calcium
channel blocker flunarizine for the topical treatment of
glaucoma.
[0076] U.S. Pat. No. 6,420,383 and U.S. Pat. No. 6,472,530 relate
to novel calcium channel blockers, useful for treating and
preventing a number of disorders such as hypersensitivity, allergy,
asthma, bronchospasm, dysmenorrhea, esophageal spasm, glaucoma,
premature labor, urinary tract disorders, gastrointestinal motility
disorders and cardiovascular disorders.
[0077] U.S. Pat. No. 6,458,781 relates to compounds that act to
block calcium channels and their use to treat stroke, cerebral
ischemia, pain, head trauma or epilepsy.
[0078] U.S. Pat. No. 6,521,647 relates to the use of calcium
channel blockers in the treatment of renal disease in animals,
especially chronic renal failure.
[0079] WO 97/10210 relates to tricyclic heterocyclic derivatives,
and their use in therapy, in particular as calcium channel
antagonists, e.g. for the treatment of ischaemia, in particular
ischaemic stroke.
[0080] WO 03/018561 relates to quinoline compounds as N-type
calcium channel antagonists and methods of using such compounds for
the treatment or prevention of pain or nociception.
[0081] WO 03/057219 relates to sodium channel blockers useful as
agents for treating or modulating a central nervous system
disorder, such as neuropathic pain, inflammatory pain,
inflammation-related pain or epilepsy.
[0082] WO 99/14199 discloses substituted
1,2,3,4,5,6-hexahydro-2,6-methano-3-benzazocines-10-oles as potent
sodium channel blockers useful for the treatment of several
diseases, such as stroke, neurodegenerative disorders, Alzheimer's
Disease, Parkinson's Disease and cardiovascular disorders.
[0083] WO 01/74779 discloses new aminopyridine sodium channel
blockers and their use as anticonvulsants, local anesthetics, as
antiarrythmics, for the treatment or prevention of
neurodegenerative conditions, such as amyotrophic lateral sclerosis
(ALS), for the treatment or prevention of both, acute or chronic
pain, and for the treatment or prevention of diabetic
neuropathy.
[0084] WO 04/087125 discloses amino acid derivatives as inhibitors
of mammalian sodium channels, useful in the treatment of chronic
and acute pain, tinnitus, bowel disorders, bladder dysfunction and
demyelinating diseases.
[0085] WO 06/028904 relates to quinazolines useful as modulators of
ion channels, and their preparation, pharmaceutical compositions,
and use as inhibitors of voltage-gated sodium channels, which is
useful in treatment of various diseases.
[0086] WO 06/024160 discloses the preparation of
piperazine-1-carboxamide derivatives as calcium channel
blockers.
[0087] WO 06/110917 describes spiro-oxindole compounds and their
preparation, pharmaceutical compositions and use as sodium channel
blockers.
[0088] WO 06/027052 describes the use of selected
(R)-2-[(halobenzyloxy)benzylamino]-propanamides and the
pharmaceutically acceptable salts thereof for the manufacture of
medicaments that are selectively active as sodium and/or calcium
channel modulators and therefore useful in preventing, alleviating
and curing a wide range of pathologies, including pain, migraine,
peripheral diseases, cardiovascular diseases, inflammatory
processes affecting all body systems, disorders affecting skin and
related tissues, disorders of the respiratory system, disorders of
the immune and endocrinological systems, gastrointestinal,
urogenital, metabolic and seizure disorders, where the above
mechanisms have been described as playing a pathological role.
[0089] WO 07/145922 discloses the preparation of benzazepinone
amino acids as sodium channel blockers.
[0090] WO 07/021941 relates to the preparation of N-thiazolyl
benzenesulfonamides as inhibitors of voltage-gated sodium
channels.
[0091] WO 08/141446 discloses amino acid derivatives as calcium
channel blockers.
[0092] WO 09/005460 describes the preparation and applications of
Nav1.7 sodium channel inhibitors for treatment of pain
disorders.
[0093] WO 09/039328 discloses pyridyl-sulfonamides as modulators of
sodium channels, their preparation, pharmaceutical compositions,
and use in treating various diseases.
[0094] WO 09/045381 relates to N-substituted oxindoline derivatives
as calcium channel blockers.
[0095] WO 10/007073 discloses the preparation of piperazine
derivatives as Cav2.2 calcium channel modulators.
[0096] WO 10/014257 describes the preparation of tetrahydropyridine
and dihydropyrrole compounds as calcium channel blockers for
treatment of pain and other disorders.
[0097] The cytochrome P450 superfamily (abbreviated as CYP) is a
large and diverse group of enzymes and the function of most CYP
enzymes is to catalyze the oxidation of organic substances. The
substrates of CYP enzymes include xenobiotic substances such as
drugs and other toxic chemicals. CYPs are the major enzymes
involved in drug metabolism and bioactivation, accounting for at
least 75% of the total metabolism. Human CYPs are primarily
membrane-associated proteins, located either in the inner membrane
of mitochondria or in the endoplasmic reticulum of cells (Smith G.,
Stubbins M. J. Xenobiotica 28 (12): 1129-65 (1998)). Many drugs may
increase or decrease the activity of various CYP isozymes either by
inducing the biosynthesis of an isozyme (enzyme induction) or by
directly inhibiting the activity of the CYP (enzyme inhibition).
This is a major source of adverse drug interactions, since changes
in CYP enzyme activity may affect the metabolism and clearance of
various drugs. For example, if one drug inhibits the CYP-mediated
metabolism of another drug, the second drug may accumulate within
the body to toxic levels. Hence, avoiding drug interactions may
necessitate dosage adjustments or the choice of drugs that do not
interact with the CYP system.
[0098] Cytochrome P450 2D6 (CYP2D6), a member of the cytochrome
P450 mixed-function oxidase system, is one of the most important
enzymes involved in the metabolism of xenobiotics in the body (Wolf
C. R., Smith G. IARC Sci. Publ.; 148: 209-29 (1999)). Whilst CYP2D6
is involved in the oxidation of a wide range of substrates of all
the CYPs, there is considerable variability in its expression in
the liver. The gene is located near two cytochrome P450 pseudogenes
on chromosome 22q13.1. Alternatively spliced transcript variants
encoding different isoforms have been found for this gene.
[0099] CYP2D6 shows the largest phenotypical variability among the
CYPs, largely due to genetic polymorphism. The genotype accounts
for normal, reduced, and non-existent CYP2D6 function in
subjects.
[0100] The CYP2D6 function in any particular subject may be
described as one of the following: [0101] poor metabolizers--these
subjects have little or no CYP2D6 function [0102] intermediate
metabolizers--these subjects metabolize drugs at a rate somewhere
between the poor and extensive metabolizers [0103] extensive
metabolizers--these subjects have normal CYP2D6 function [0104]
ultrarapid metabolizers--these subjects have multiple copies of the
CYP2D6 gene expressed, and therefore greater-than-normal CYP2D6
function.
[0105] Therefore, patients undergoing any therapeutical treatment
may be classified according to the above subject definitions.
[0106] Many antipsychotic drugs used for schizophrenia treatment
are CYP2D6 substrates: examples of these drugs include haloperidol,
risperidone, perphenazine, thioridazine, aripiprazole and
sertindole. If a drug is able to potently inhibit CYP2D6 the
subject taking said drug may become a poor metabolizer, i.e. may
experience an increase in plasma levels of a CYP2D6 metabolized
drug taken concomitantly. Quinidine, paroxetine, bupropion and
fluoxetine are powerful CYP2D6 inhibitors and the use of potent
inhibitors can render a patient that is a CYP2D6 extensive
metabolizer into a phenotypic poor metabolizer (De Leon J.,
Armstrong S. C., Cozza K. L. Psychosomatics; 47(1): 75-85 (2006)).
A CYP2D6 poor metabolizer phenotype may have a major role in
personalizing risperidone doses (De Leon J., Susce M. T., Pan R.
M., Wedlun P. J., Orrego M. L., Diaz F. J. Pharmacopsychiatry;
40(3), 93-102 (2007)). As a further example, sertindole undergoes
extensive hepatic metabolism by CYP2D6 and 3A4 to two principal
metabolites. CYP2D6 poor metabolizers may have sertindole clearance
reduced by 50-67%. The concomitant administration of sertindole and
CYP2D6 inhibitors should be used with extreme caution (Murdoch D.,
Keating G. M. CNS Drugs; 20(3): 233-255 (2006)).
[0107] It is therefore highly desirable, in view of avoiding undue
drug-drug interactions, to have compounds which are unable to
inhibit the major human CYPs, in particular CYP2D6, for example in
a psychosis and schizophrenia setting, but also in any pathology
treated with a drug that is also a CYP2D6 substrate (Foster A.
Mobley E., Wang Z. Pain Practice; 7(4): 352-356 (2007)).
DESCRIPTION OF THE INVENTION
[0108] The object of this invention is a new class of fluorinated
arylalkylamino carboxamide derivatives which are highly potent as
sodium and/or calcium channel modulators and therefore useful in
preventing, alleviating and curing a wide range of pathologies,
including, but not limited to psychiatric, neurological,
cardiovascular, inflammatory, ophthalmic, urogenital,
gastrointestinal diseases where the above mechanisms have been
described as playing a pathological role. Said compounds are also
characterized in that they are substantially free from any CYP2D6
inhibitory effect or exhibit a significantly reduced CYP2D6
inhibitory effect.
[0109] In this description and claims, the expression "sodium
and/or calcium channel modulator(s)" means compounds able to block
sodium and/or calcium currents in a voltage and/or use-dependent
manner.
[0110] In this description and claims the expression "substantially
free from any CYP2D6 inhibitory effect" means that the compound
exhibits a IC.sub.50 [.mu.M] value in the in vitro cytochrome
inhibition test according to Example 10 which is higher than 40
while the expression "reduced CYP2D6 inhibitory effect" means that
the compounds exhibits a IC.sub.50 [.mu.M] value which is higher
than 20.
[0111] In particular, the object of the present invention is a
compound of general formula I
##STR00005##
wherein: [0112] W is a group A-[(CH.sub.2).sub.m--O]-- wherein: m
is zero, 1, 2, or 3; A is (C.sub.1-C.sub.4)alkyl optionally
substituted with one to three fluorine atoms;
(C.sub.3-C.sub.6)cycloalkyl; phenyl optionally substituted with a
group selected from halo, methyl, methoxy, trifluoromethyl,
acetylamino, and dimethylaminomethyl; thienyl optionally
substituted with a chloro group; furanyl; isoxazolyl, thiazolyl;
piperidinyl; morpholinyl; pyridinyl or pyrimidinyl, the pyridinyl
and pyrimidinyl ring being optionally substituted with one or two
methoxy groups; [0113] J independently is hydrogen,
(C.sub.1-C.sub.4)alkyl; (C.sub.1-C.sub.4)alkoxy; or an halo group;
[0114] n is 1 or 2; [0115] R.sup.1 is hydrogen;
(C.sub.1-C.sub.4)alkyl optionally substituted with a hydroxy group
or a (C.sub.1-C.sub.4)alkoxy group; or (C.sub.3-C.sub.8)cycloalkyl;
[0116] R.sup.2 and R.sup.2' are independently hydrogen;
(C.sub.1-C.sub.4)alkyl optionally substituted with a
(C.sub.1-C.sub.4)alkoxy group; phenyl optionally substituted with a
(C.sub.1-C.sub.4)alkyl, a (C.sub.1-C.sub.4)alkoxy or an halo group;
benzyl optionally substituted with a (C.sub.1-C.sub.4)alkyl, a
(C.sub.1-C.sub.4)alkoxy or an halo group on the benzene ring; or
R.sup.2 and R.sup.2' taken together with the adjacent carbon atom
form a (C.sub.3-C.sub.6)cycloalkylidene group. [0117] R.sup.3 is
hydrogen; or (C.sub.1-C.sub.4)alkyl; [0118] R.sup.4 is hydrogen;
(C.sub.1-C.sub.4)alkyl; phenyl; cyclohexyl; or benzyl; or [0119]
R.sup.3 and R.sup.4, taken together with the adjacent nitrogen
atom, form an azetidinyl, pyrrolidinyl, morpholinyl, piperidinyl or
piperazinyl ring, the piperidinyl ring being optionally substituted
with one or two (C.sub.1-C.sub.2)alkyl group(s), and the
piperazinyl ring being optionally substituted on the other N-atom
with a (C.sub.1-C.sub.4)alkyl, benzyl, or phenylsulfonyl group; or
a pyrrolidinyl, piperidinyl, morpholinyl or piperazinyl ring fused
with a benzene ring; [0120] R.sup.5 is hydrogen or fluoro; and
[0121] R.sup.6 is fluoro; [0122] if the case, either as single
optical isomer in the isolated form or as a mixture thereof in any
proportion and its pharmaceutically acceptable salt.
[0123] The term "(C.sub.1-C.sub.4)alkyl" or the "(C.sub.1-C.sub.4)
alkyl" moiety in the other substitutents (e.g. in the terms alkoxy)
as used in this description and claims, when no otherwise
specified, identifies a straight or branched alkyl radical or
moiety; examples of said radicals or moieties include,
respectively: methyl, ethyl, propyl, isopropyl, butyl, isobutyl and
tert-butyl or methoxy, ethoxy, propoxy, isopropoxy, butoxy
isobutoxy and tert-butoxy.
[0124] The term "(C.sub.1-C.sub.4)alkyl" when substituted with "one
to three fluorine atoms" identifies a straight or branched alkyl
radical of 1 to 4 carbon atoms wherein one to three hydrogen atoms
attached to the same or different carbon atoms are independently
substituted by fluorine. Preferred representative examples of this
term are trifluoromethyl, 2,2,2-trifluoroethyl,
3,3,3-trifluoropropyl and 4,4,4-trifluorobutyl.
[0125] The terms "(C.sub.3-C.sub.6)cycloalkyl" and
"(C.sub.3-C.sub.6)cycloalkylidene" as used in this description and
claims, when not otherwise specified, identifies a cycle-forming
alkyl or alkylidene radical or moiety; examples of said radicals or
moieties include, respectively cyclopropyl, cyclobutyl,
cyclopentyl, and cyclohexyl, or cyclopropylidene, cyclobutylidene,
cyclopentylidene and cyclohexylidene.
[0126] The term "halo", when not otherwise specified herein, means
an halogen atom radical such as fluoro, chloro, bromo and iodo.
[0127] Where the compounds of this invention contain at least one
asymmetric carbon atom they can exist as single enantiomers or
diastereoisomers or a mixture thereof, the invention includes
within its scope all the possible single optical isomers in the
isolated form of said compounds and the mixtures thereof in any
proportion, e.g., the racemic mixtures.
[0128] Examples of pharmaceutically acceptable salts of the
compounds of formula I are salts with organic and inorganic acids
such as hydrochloric, hydrobromic, hydroiodic, nitric, sulfuric,
phosphoric, acetic, propionic, tartaric, fumaric, citric, benzoic,
succinic, cinnamic, mandelic, salicylic, glycolic, lactic, oxalic,
malic, maleic, malonic, fumaric, tartaric, p-toluenesulfonic,
methanesulfonic, glutaric acid and the like.
[0129] The compounds of formula I are active as calcium and/or
sodium channel modulators and therefore useful in preventing
alleviating and curing a wide range of pathologies, including but
not limited to psychiatric, neurological, cardiovascular,
inflammatory, ophthalmic, urogenital and gastrointestinal diseases
where the above mechanisms have been described as playing a
pathological role, said compounds being characterized in that they
are substantially free from any CYP2D6 inhibitory effect or exhibit
a significantly reduced CYP2D6 inhibitory effect.
[0130] A preferred group of compounds of formula I of this
invention comprises a compound wherein: [0131] W is a group
A-[(CH.sub.2).sub.m--O]-- wherein: m is zero, 1, 2 or 3; A is
(C.sub.1-C.sub.4)alkyl optionally substituted with one to three
fluorine atoms; (C.sub.3-C.sub.6)cycloalkyl; phenyl optionally
substituted with a halo group; or thiazolyl [0132] J independently
is hydrogen; C.sub.1-C.sub.4 alkyl; chloro; or fluoro; [0133] n is
1 or 2; [0134] R.sup.1 is hydrogen; (C.sub.1-C.sub.4)alkyl
optionally substituted with a hydroxy group or a
(C.sub.1-C.sub.4)alkoxy group; or (C.sub.3-C.sub.6)cycloalkyl;
[0135] R.sup.2 is hydrogen; or (C.sub.1-C.sub.4)alkyl; [0136]
R.sup.2' is hydrogen or (C.sub.1-C.sub.4)alkyl optionally
substituted with a (C.sub.1-C.sub.4)alkoxy or a phenyl group, the
phenyl group being optionally substituted with a
(C.sub.1-C.sub.4)alkoxy group; [0137] R.sup.3 is hydrogen; or
(C.sub.1-C.sub.4)alkyl; [0138] R.sup.4 is hydrogen;
(C.sub.1-C.sub.4)alkyl; phenyl; or cyclohexyl; or [0139] R.sup.3
and R.sup.4, taken together with the adjacent nitrogen atom, form
an azetidinyl, pyrrolidinyl, morpholinyl, piperidinyl or
piperazinyl, the piperydinyl ring being optionally substituted with
one or two (C.sub.1-C.sub.2)alkyl group(s) and the piperazinyl ring
being optionally substituted on the other N-atom with a
(C.sub.1-C.sub.4)alkyl, benzyl, or phenylsulfonyl group; or a
pirrolidinyl, piperidinyl, morpholinyl or piperazinyl ring fused
with a benzene ring; [0140] R.sup.5 is hydrogen or fluoro; and
[0141] R.sup.6 is fluoro; if the case, either as single optical
isomer in the isolated form or as a mixture thereof in any
proportion and its pharmaceutically acceptable salt.
[0142] A more preferred group of compounds of formula I comprises a
compound wherein; [0143] W is a group A-[(CH.sub.2).sub.m--O]--
wherein: m is 1 or 2; A is (C.sub.1-C.sub.4)alkyl optionally
substituted with one to three fluorine atoms; phenyl optionally
substituted with a chloro or fluoro group; or thiazolyl; [0144] J
independently is hydrogen; methyl; or fluoro; [0145] n is lor 2
[0146] R.sup.1 is hydrogen; (C.sub.1-C.sub.4)alkyl optionally
substituted with a hydroxy group or a (C.sub.1-C.sub.4)alkoxy
group; [0147] R.sup.2 is hydrogen; or methyl; [0148] R.sup.2' is
hydrogen; or (C.sub.1-C.sub.4)alkyl optionally substituted with a
methoxy or a phenyl group, the phenyl group being optionally
substituted with a methoxy group; [0149] R.sup.3 is hydrogen; or
(C.sub.1-C.sub.4)alkyl; [0150] R.sup.4 is hydrogen;
(C.sub.1-C.sub.4)alkyl; phenyl; or cyclohexyl; or [0151] R.sup.3
and R.sup.4, taken together with the adjacent nitrogen atom, form
an azetidinyl, pyrrolidinyl, morpholinyl, piperidinyl, or
piperazinyl ring, the piperidinyl ring being optionally substituted
with one or two methyl group(s) and the piperazinyl ring being
optionally substituted on the other N-atom with a methyl, benzyl or
phenylsulfonyl group; or a pirrolidinyl, piperidinyl, morpholinyl,
or piperazinyl ring fused with a benzene ring; [0152] R.sup.5 is
hydrogen or fluoro; and [0153] R.sup.6 is fluoro; if the case,
either as single optical isomer in the isolated form or as a
mixture thereof in any proportion and its pharmaceutically
acceptable salt.
[0154] Most preferably, a compound of formula I according to this
invention is selected from the group consisting of: [0155]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide;
(Example 1-1) [0156]
2-[2,2-Difluoro-2-(3-pentyloxyphenyl)-ethylamino]-N,N-dimethyl-acetamide
(Example 1-2) [0157]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dipropyl-acetamide
(Example 1-3) [0158]
2-[2,2-Difluoro-2-(3-butoxy-4-methylphenyl)-ethylamino]-N,N-dimethyl-acet-
amide; (Example 1-4) [0159]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dibutyl-acetamide;
(Example 1-5) [0160]
2-[2,2-Difluoro-2-(3-hexyloxyphenyl)-ethylamino]-N,N-dimethyl-acetamide;
(Example 1-6) [0161]
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-N,N-dime-
thyl-acetamide; (Example 1-7) [0162]
2-[2,2-Difluoro-2-(3-pentyloxyphenyl)-ethylamino]-N,N-dipropyl-acetamide;
(Example 1-8) [0163]
2-{2,2-Difluoro-2-[3-(3-(3-fluorophenyl)-propoxy)-phenyl]-ethylamino}-N,N-
-dimethyl-acetamide; (Example 1-9) [0164]
2-{2,2-Difluoro-2-[3-(3-(3-chlorophenyl)-propoxy)-phenyl]-ethylamino}-N,N-
-dimethyl-acetamide; (Example 1-10) [0165]
2-[2,2-Difluoro-2-(3-butoxy-2-fluorophenyl)-ethylamino]-N,N-dimethyl-acet-
amide; (Example 1-11) [0166]
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-N,N-dimethyl-a-
cetamide; (Example 1-12) [0167]
2-{2,2-Difluoro-2-[3-(3-thiazol-2-yl-propoxy)-phenyl]-ethylamino}-N,N-dim-
ethyl-acetamide; (Example 1-13) [0168]
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-N,N-dimethyl-acetamide;
(Example 1-14) [0169]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(pyrrolidin-1-yl)-ethano-
ne; (Example 1-15) [0170]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N-methyl-N-phenyl-acetamid-
e; (Example 1-16) [0171]
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(pyrrolidin--
1-yl)-ethanone; (Example 1-17) [0172]
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(pyrro-
lidin-1-yl)-ethanone; (Example 1-18) [0173]
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-1-(morpholin-4-yl)-etha-
none; (Example 1-19) [0174]
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(morpholin-4-
-yl)-ethanone; (Example 1-20) [0175]
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(morph-
olin-4-yl)-ethanone; (Example 1-21) [0176]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(2H-benzo[b][1,4]oxazin--
4(3H)-yl)-ethanone; (Example 1-22) [0177]
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-1-(pyrrolidin-1-yl)-eth-
anone; (Example 1-23) [0178]
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-N-methyl-N-phe-
nyl-acetamide; (Example 1-24) [0179]
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-N-methyl-
-N-phenyl-acetamide; (Example 1-25) [0180]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(4-methylpiperazin-1-yl)-
-ethanone; (Example 1-26) [0181]
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(4-methylpip-
erazin-1-yl)-ethanone; (Example 1-27) [0182]
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(4-met-
hylpiperazin-1-yl)-ethanone; (Example 1-28) [0183]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(piperidin-1-yl)-ethanon-
e; (Example 1-29) [0184]
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(piperidin-1-
-yl)-ethanone; (Example 1-30) [0185]
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(piper-
idin-1-yl)-ethanone; (Example 1-31) [0186]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-diethyl-acetamide;
(Example 1-32); [0187]
2-{2,2-Difluoro-2-[3-(2-fluorobenzyloxy)-phenyl]-ethylamino}-N,N-dimethyl-
-acetamide; (Example 1-33) [0188]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(cis-3,5-dimethylpiperid-
in-1-yl)-ethanone; (Example 1-34) [0189]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(3,4-dihydroisoquinolin--
2(1H)-yl)-ethanone; (Example 1-35) [0190]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-diisopropyl-acetamide;
(Example 1-36) [0191]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N-cyclohexyl-N-methyl-acet-
amide; (Example 1-37) [0192]
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-1-(piperidin-1-yl)-etha-
none; (Example 1-38) [0193]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-[4-(phenylsulfonyl)-pipe-
razin-1-yl]-ethanone; (Example 1-39) [0194]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(indolin-1-yl)-ethanone;
(Example 1-40) [0195]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(4-benzylpiperazin-1-yl)-
-ethanone; (Example 1-41) [0196]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(azetidin-1-yl)-ethanone-
; (Example 1-42) [0197]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-propanamide;
(Example 2-1) [0198]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-3-methoxy-N,N-dimethyl-pro-
panamide; (Example. 2-2) [0199]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-3-(4-methoxyphenyl)-N,N-di-
methyl-propanamide; (Example 2-3) [0200]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-2-N,N-trimethyl-propanamid-
e; (Example 2-4) [0201]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-4-N,N-trimethyl-pentanamid-
e; (Example 2-5) [0202]
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-methylamino}-N,N-dimethyl-acet-
amide; (Example. 3-1) [0203]
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-(3-methoxypropyl)-amino}-N,N-d-
imethyl-acetamide; (Example 3-2) [0204]
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-(2-methoxyethyl)-amino}-N,N-di-
methyl-acetamide; (Example 3-3) [0205]
2-[2-Fluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide;
(Example 4-1) [0206]
2-{2-Fluoro-2-[3-(3-chlorobenzyloxy)-phenyl]-ethylamino}-N,N-dimethyl-ace-
tamide; (Example 4-2) [0207]
2-{2-Fluoro-2-[3-(3-fluorobenzyloxy)-phenyl]-ethylamino}-N,N-dimethyl-ace-
tamide; Example 4-3 if the case, either as single optical isomer in
the isolated form or as a mixture thereof in any proportion and its
pharmaceutically acceptable salt.
[0208] The compounds of formula I, object of the present invention,
are prepared according to a synthetic process which comprises:
a) the reaction of a compound of formula II
##STR00006##
wherein J, W, and n have the same meanings defined in formula I
above, with a suitable fluorinating agent, such as
N-fluorobenzenesulfonimide, to give a compound of formula III
##STR00007##
wherein J, W, n, R.sup.5 and R.sup.6 have the same meanings as
defined in formula I above b) reaction of a compound of formula III
with a suitable reducing agent, such as lithium aluminium hydride,
to give a compound of formula IV
##STR00008##
wherein W, J, n, R.sup.5 and R.sup.6 have the same meanings as
defined in formula I above; c) protection of the amino group of a
compound of formula IV with a suitable protecting agent, such as
di-tert-butoxy-dicarbonate, to give a compound of formula V
##STR00009##
wherein W, J, n, R.sup.5 and R.sup.6 have the same meanings as
defined in formula I above and PROT is a suitable N-protecting
group, for example a tert-butoxycarbonyl group. d) reacting a
compound of formula V with a suitable haloalkylamide of formula
VI
##STR00010##
wherein X is a halogen atom and R.sup.2, R.sup.2', R.sup.3, and
R.sup.4 have the same meaning as defined in formula I above,
whereby a compound of formula I is obtained wherein R.sup.1 is
hydrogen. The compound of formula I wherein J, W, n, R.sup.1
R.sup.2, R.sup.2', R.sup.3, R.sup.4, R.sup.5, and R.sup.6 have the
same meanings as above and R.sup.1 has the same meanings as above,
apart from hydrogen, can be prepared through the reaction of a
compound of formula VII
##STR00011##
wherein J, W, n, R.sup.2, R.sup.2', R.sup.3, R.sup.4, R.sup.5, and
R.sup.6, have the same meanings as in formula I above, with a
compound R.sup.1--Z, wherein R.sup.1 has the meanings reported
above apart from hydrogen and Z is a halogen atom or a good leaving
group, e.g. methanesulfonyloxy, p-toluenesulfonyloxy or
trifluoromethanesulfonyloxy in the presence of a base or with a
carbonyl compound of the formula R.sup.7R.sup.8CO in the presence
of a reducing agent, wherein R.sup.7 and R.sup.8 both represent
hydrogen or, taken together with the adjacent carbonyl group,
represent a (C.sub.2-C.sub.4)aliphatic aldehyde or a
(C.sub.3-C.sub.4) aliphatic ketone, optionally substituted with a
hydroxyl group or a (C.sub.1-C.sub.4)alkoxy group, or R.sup.7 and
R.sup.8 taken together with the adjacent carbonyl group represent a
(C.sub.3-C.sub.8) alicyclic ketone.
[0209] A compound of the invention may be converted into another
compound of the invention. For instance, a compound of formula I
wherein W represents a benzyloxy radical may be transformed into
the corresponding hydroxy-derivative by catalytic hydrogenation and
then reacted with an appropriate reagent to replace the original
benzyl moiety with a different group, e.g., a
trifluoromethylbenzyl, phenylethyl, trifluoroethyl, cyclopentyl,
and cyclopropylmethyl. If desired, a compound of the invention may
be converted into a pharmaceutically acceptable salt and/or, if
desired, a salt may be converted into a free compound and/or, if
desired, a mixture of enantiomers or diastereoisomers of compounds
of the invention may be separated into the corresponding single
optical isomers.
[0210] The compounds of formula II and VI are commercially
available or are prepared from commercially available compounds
according to well-known methods.
[0211] When a compound of formula I is obtained wherein R.sup.1 is
hydrogen (i.e., a compound of formula VII) the introduction of a
radical R.sup.1 which is other than hydrogen defined above is
carried out according to conventional methods for the preparation
of secondary or tertiary amines such as alkylation or reductive
amination techniques as described above.
[0212] According to a preferred embodiment of the invention said
alkylation reaction is carried out in the presence of a base and,
more preferably, said base is selected from K.sub.2CO.sub.3,
triethylamine and diisopropylethylamine.
[0213] According to another preferred embodiment of the invention
said reductive amination with a compound R.sup.7R.sup.8CO, wherein
R.sup.7 and R.sup.8 have the same meanings as defined above is
carried out in the presence of a reducing agent selected from
NaBH.sub.4, NaBH.sub.3CN and (polystyrylmethyl)-trimethylammonium
cyanoborohydride.
[0214] In the preparation of the compounds of formula I and the
starting materials and/or intermediates described herein it may be
useful to protect certain groups which are sensitive to the
reaction conditions.
[0215] The evaluation of the usefullness of the optional
protection, as well as the selection of the suitable protecting
agent, according to the reaction carried out in the preparation of
the compounds of the invention and the functional group to be
protected, are within the common knowledge of the skilled
person.
[0216] The removal of the optional protective groups is carried out
according to conventional techniques.
[0217] For a general reference to the use of protective groups in
orgamic chemistry, see Theodora W. Greene and Peter G. M. Wuts
"Protective groups in organic synthesis", John Wiley & Sons,
Inc., II Ed., 1991.
[0218] The preparation of the salts of the compounds of formula I
is carried out according to known methods.
[0219] For the preparation of a single enantiomers or
diastereoisomers, if the case, of a compound of formula I, said
compound may be obtained through a sterically controlled synthesis
or by using reagents having the appropriate chirality or separating
the desired isomer from the enantiomeric or diastereoisomeric
mixture thereof according to conventional procedures. For instance,
single optically active enantiomers may be obtained from their
racemates by chiral chromatography or by converting them into a
mixture of diastereoisomeric derivatives, separating the
diastereoisomeric derivatives and restoring the respective
enantiomers.
[0220] Diastereoisomers can be separated from their mixtures by
means of conventional techniques based on their different
physico-chemical properties, such as chromatography, distillation,
or fractional crystallization.
Pharmacology
[0221] The compounds of the invention may be used for the
manufacture of a medicament active as sodium and/or calcium channel
modulators against disorders caused by dysfunctions of voltage
gated calcium and/or sodium channels being characterized in that
they are substantially free from any CYP2D6 inhibitory effect or
exhibit a significantly reduced CYP2D6 inhibitory effect.
[0222] The sodium channel modulating activity of the fluorinated
phenylalkylamino derivatives was measured through a
fluorescence-based sodium influx assay (Table 1), through patch
clamp techniques in constitutive and/or Nav 1.3 transfected cell
lines (Table 2) and in cortical neurons.
[0223] The CYP2D6 inhibition was assessed by performing in vitro
inhibition studies using Supersomes, microsomes derived from
baculovirus infected insect cells; the baculoviruses have been
engineered to express one or more drug metabolizing enzyme cDNAs
(Table 3).
[0224] The in vivo analgesic activity of the above compounds was
assessed in the "rat complete Freund's adjuvant model" and in the
"Bennett model of neuropathic pain in rats".
[0225] The in vivo sodium channel blocking and anticonvulsant
activity were measured using the "Maximal electroshock test" in
mice (Table 4).
[0226] The anti mania activity was measured using the "Amphetamine
and chlordiazepoxide-induced hyperlocomotion in mice" model.
[0227] The anti-schizophrenia and anti-addiction activities were
assessed using the "Test of cognitive impairment in schizophrenia"
and the "Cocaine-induced behavioural sensitization test" in
rats.
[0228] "Acute bladder irritation by acetic acid in rats" and
"Intermediate bladder irritation by cyclophosphamide in rats" tests
were used as models for urological diseases.
[0229] The anti migraine activity was measured using the "migraine
test" in rats.
[0230] Such substances exhibit also "use and frequency-dependency",
i.e. an enhancement of the block during a high frequency
stimulation when there is a large accumulation of channels in the
inactivated state, such as in neuronal pathological conditions.
Functionally, the use-dependent block results in depression of
neuronal activity at high frequency firing and with lower blocking
capacity at normal firing rate suggesting that the compounds of
this invention may selectively depress abnormal activity of the
calcium and/or sodium channels, leaving unaffected the
physiological activity, thus decreasing CNS depressant effects
(Catterall W. A., Trends Pharmacol. Sci. 8: 57-65 (1987)).
[0231] The compounds of the invention are active in vivo when
orally or intraperitoneally administered in the range of 0.1 to 100
mg/kg in different animal models hereafter described.
[0232] In view of the above described mechanisms of action, the
compounds of the present invention are useful in the prevention or
treatment of neuropathic pain. Neuropathic pain syndromes include,
and are not limited to: diabetic neuropathy; sciatica; non-specific
lower back pain; multiple sclerosis pain; fibromyalgia; HIV-related
neuropathy; neuralgia, such as post-herpetic neuralgia and
trigeminal neuralgia, Morton's neuralgia, causalgia; and pain
resulting from physical trauma, amputation, phantom limb, cancer,
toxins or chronic inflammatory conditions; central pain such as the
one observed in thalamic syndromes, mixed central and peripheral
forms of pain such as complex regional pain syndromes (CRPS) also
called reflex sympathetic dystrophies.
[0233] The compounds of the invention are also useful for the
treatment of chronic pain. Chronic pain includes, and is not
limited to, chronic pain caused by inflammation or an
inflammatory-related condition, ostheoarthritis, rheumatoid
arthritis, acute injury or trauma, upper back pain or lower back
pain (resulting from systematic, regional or primary spine disease
such as radiculopathy), bone pain (due to osteoarthritis,
osteoporosis, bone metastasis or unknown reasons), pelvic pain,
spinal cord injury-associated pain, cardiac chest pain, non-cardiac
chest pain, central post-stroke pain, myofascial pain, sickle cell
pain, cancer pain, Fabry's disease, AIDS pain, geriatric pain or
pain caused by headache, temporomandibular joint syndrome, gout,
fibrosis or thoracic outlet syndromes, in particular rheumatoid
arthritis and osteoarthritis.
[0234] The compounds of the invention are also useful in the
treatment of acute pain caused by acute injury, illness,
sport-medicine injuries, carpal tunnel syndrome, burns,
musculoskeletal sprains and strains, musculotendinous strain,
cervicobrachial pain syndromes, dyspepsis, gastric ulcer, duodenal
ulcer, dysmenorrhea, endometriosis or surgery (such as open heart
or bypass surgery), post operative pain, kidney stone pain,
gallbladder pain, gallstone pain, obstetric pain or dental
pain.
[0235] The compounds of the invention are also useful in the
treatment of headaches such as migraine, tension type headache,
transformed migraine or evolutive headache, cluster headache, as
well as secondary headache disorders, such as the ones derived from
infections, metabolic disorders or other systemic illnesses and
other acute headaches, paroxysmal hemicrania and the like,
resulting from a worsening of the above mentioned primary and
secondary headaches.
[0236] The compounds of the invention are also useful for the
treatment of neurological conditions such as epilepsy including
simple partial seizure, complex partial seizure, secondary
generalized seizure, further including absence seizure, myoclonic
seizure, clonic seizure, tonic seizure, tonic clonic seizure and
atonic seizure. The compounds of the invention are also useful for
the treatment of neurodegenerative disorders of various origins
such as Alzheimer's Disease and other dementia conditions such as
Lewys body, fronto-temporal dementia and taupathies; amyotrophic
lateral sclerosis, Parkinson's Disease and other parkinsonian
syndromes; essential tremors; other spino cerebellar degeneration
and Charcot-Marie-Toot neuropathy.
[0237] The compounds of the invention are also useful for the
treatment of cognitive disorders and of psychiatric disorders.
Psychiatric disorders include, and are not limited to major
depression, dysthymia, mania, bipolar disorder (such as bipolar
disorder type I, bipolar disorder type II), cyclothymic disorder,
rapid cycling, ultradian cycling, mania, hypomania, schizophrenia,
schizophreniform disorders, schizoaffective disorders, personality
disorders, attention disorders with or without hyperactive
behaviour, delusional disorders, brief psychotic disorders, shared
psychotic disorders, psychotic disorder due to a general medical
condition, substance-induced psychotic disorders or a psychotic
disorder not otherwise specified, anxiety disorders such as
generalised anxiety disorder, panic disorders, post-traumatic
stress disorder, impulse control disorders, phobic disorders,
dissociative states and moreover in smoke, drug addiction and
alcoholism. In particular bipolar disorders, psychosis, anxiety and
addiction.
[0238] Compounds of the invention are also useful in the treatment
of diseases such as vertigo, tinnitus, muscle spasm, and other
disorders including and not limited to cardiovascular diseases
(such as cardiac arrhythmia, cardiac infarction or angina pectoris,
hypertension, cardiac ischemia, cerebral ischemia) endocrine
disorders (such as acromegaly or diabetes insipidus) diseases in
which the pathophysiology of the disorder involves excessive or
hypersecretory or otherwise inappropriate cellular secretion of an
endogenous substance (such as catecholamine, a hormone or a growth
factor).
[0239] The compounds of the invention are also useful in the
selective treatment of liver disease, such as inflammatory liver
diseases, for example chronic viral hepatitis B, chronic viral
hepatitis C, alcoholic liver injury, primary biliary cirrhosis,
autoimmune hepatitis, non-alcoholic steatohepatitis and liver
transplant rejection.
[0240] The compounds of the invention inhibit inflammatory
processes affecting all body systems. Therefore are useful in the
treatment of inflammatory processes of the muscular-skeletal system
of which the following is a list of examples but it is not
comprehensive of all target disorders: arthritic conditions such as
alkylosing spondylitis, cervical arthritis, fibromyalgia, gout,
juvenile rheumatoid arthritis, lumbosacral arthritis,
osteoarthritis, osteoporosis, psoriatic arthritis, rheumatic
disease; disorders affecting skin and related tissues: eczema,
psoriasis, dermatitis and inflammatory conditions such as sunburn;
disorders of the respiratory system: asthma, allergic rhinitis and
respiratory distress syndrome, lung disorders in which inflammation
is involved such as asthma and bronchitis; chronic obstructive
pulmonary disease; disorders of the immune and endocrinological
systems: periarthritis nodosa, thyroiditis, aplastic anaemia,
scleroderma, myasthenia gravis, multiple sclerosis and other
demyelinizating disorders, encephalomyelitis, sarcoidosis,
nephritic syndrome, Bechet's syndrome, polymyositis,
gingivitis.
[0241] Compounds of the invention are also useful in the treatment
of gastrointestinal (GI) tract disorders such as inflammatory bowel
disorders including but not limited to ulcerative colitis, Crohn's
disease, ileitis, proctitis, celiac disease, enteropathies,
microscopic or collagenous colitis, eosinophilic gastroenteritis,
or pouchitis resulting after proctocolectomy and post ileonatal
anastomosis, and irritable bowel syndrome including any disorders
associated with abdominal pain and/or abdominal discomfort such as
pylorospasm, nervous indigestion, spastic colon, spastic colitis,
spastic bowel, intestinal neurosis, functional colitis, mucous
colitis, laxative colitis and functional dyspepsia; but also for
treatment of atrophic gastritis, gastritis varialoforme, ulcerative
colitis, peptic ulceration, pyrosis, and other damage to the GI
tract, for example, by Helicobacter pylori, gastroesophageal reflux
disease, gastroparesis, such as diabetic gastroparesis; and other
functional bowel disorders, such as non-ulcerative dyspepsia (NUD);
emesis, diarrhoea, and visceral inflammation.
[0242] Compounds of the invention are also useful in the treatment
of disorders of the genito-urinary tract such as overactive
bladder, prostatitis (chronic bacterial and chronic non-bacterial
prostatitis), prostadynia, interstitial cystitis, urinary
incontinence and benign prostatic hyperplasia, annexitis, pelvic
inflammation, bartholinitis and vaginitis. In particular overactive
bladder and urinary incontinence.
[0243] The compounds of the invention are also useful in the
treatment of ophthalmic diseases such as retinitis, retinopathies,
uveitis and acute injury to the eye tissue, macular degeneration or
glaucoma, conjunctivitis.
[0244] The compounds of the invention are also useful in the
treatment of eating disorders such as anorexia nervosa including
the subtypes restricting type and binge-eating/purging type;
bulimia nervosa including the subtypes purging type and nonpurging
type; obesity; compulsive eating disorders; binge eating disorder;
and eating disorder not otherwise specified.
[0245] In consideration of the fact that the compounds of this
invention are substantially free from any CYP2D6 inhibitory effect
or exhibit a significantly reduced CYP2D6 inhibitory effect, the
compounds of this invention are particularly useful for treating
the above described disorders caused by dysfunctions of voltage
gated sodium and/or calcium channel in patients that are defined as
poor metabolizers or are assuming drugs which are CYP2D6
inhibitors.
[0246] It will be appreciated that the compounds of the invention
may advantageously be used in conjunction with one or more other
therapeutic agents. Examples of suitable agents for adjunctive
therapy include a serotonin receptor modulator including a 5HT1B/1D
agonist, such as a triptan (e.g. sumatriptan or naratriptan); an
adenosine A1 agonist; an adenosine A2 antagonist; a purinergic P2X
antagonist, an EP ligand; an NMDA modulator, such as a glycine
antagonist; an AMPA modulator; a substance P antagonist (e.g. an
NK1 antagonist); a cannabinoid; a nicotinic receptor agonist; an
alpha-1 or 2 adrenergic agonist; acetaminophen or phenacetin; a
5-lipoxygenase inhibitor; a leukotriene receptor antagonist; a
DMARD (e.g. methotrexate); gabapentin, pregabalin and related
compounds; L-dopa and/or dopamine agonists; a
catechol-O-methyltransferase inhibitor; a tricyclic antidepressant
(e.g. amitryptiline); a neurone stabilising antiepileptic drug; a
monoaminergic uptake inhibitor (e.g. venlafaxine); a matrix
metalloproteinase inhibitor; a nitric oxide synthase (NOS)
inhibitor, such as an iNOS or an nNOS inhibitor; a free radical
scavenger; an alpha-synuclein aggregation inhibitor; a
cholinesterase inhibitor, a cholesterol lowering agent; an
alpha-secretase modulator; a beta-secretase modulator; a
beta-amyloid aggregation inhibitor; an inhibitor of the release, or
action, of tumor necrosis factor alpha; an antibody therapy, such
as monoclonal antibody therapy; an antiviral agent, such as a
nucleoside inhibitor (e.g. lamivudine) or an immune system
modulator (e.g. interferon); an opioid analgesic, such as morphine;
a vanilloid receptor antagonist; an analgesic, such as a
cyclooxygenase-1 and/or cyclooxygenase-2 inhibitor; a local
anaesthetic such as lidocaine and derivatives; a stimulant,
including caffeine; an H2-antagonist (e.g. ranitidine); a proton
pump inhibitor (e.g. omeprazole); an antacid (e.g. aluminium or
magnesium hydroxide; an antiflatulent (e.g. simethicone); a
decongestant (e.g. phenylephrine, phenylpropanolamine,
pseudoephedrine, oxymetazoline, epinephrine, naphazoline,
xylometazoline, propylhexedrine, or levo-desoxyephedrine,
naphazoline, xylometazoline, propylhexedrine, or
levo-desoxyephedrine); antitussive (e.g. codeine, hydrocodone,
carmiphen, carbetapentane, or dextramethorphan); a diuretic; or a
sedating or non-sedating antihistamine; an antipsychotic agent,
including typical and atypical antipsychotics (e.g. haloperidol,
risperidone, clozapine); an anti-depressant, such as a selective
serotonin re-uptake inhibitors, serotonin and noradrenaline
re-uptake inhibitors, MAO inhibitors and tryciclics antidepressant
drugs; a mood stabilizer (e.g. lithium, lamotrigine, valproate); an
anxyolitic agent (e.g. benzodiazepines, buspirone), beta-adrenergic
receptors antagonists, morphine or morphine derivatives, other
calcium or sodium channel blocker. It is to be understood that the
present invention covers also the use of a compound of formula (I)
or a pharmaceutically acceptable salt thereof in conjunction with
one or more other therapeutic agents. For said use, the compounds
of formula (I) and the other therapeutic agent(s) may be
administered either jointly or sequentially.
[0247] The compounds of the present invention are useful in human
and veterinary medicaments. It is to be understood that as used
herein the terms "treatment" or "treating" whenever not
specifically defined otherwise, include prevention, alleviation and
cure of pathological affection, in particular, they include both
treatment of established symptoms and prophylactic treatment. The
compounds of the present invention for their therapeutic or
preventive use in the above mentioned pathologies will be
preferably used as active ingredients in a pharmaceutical
composition.
[0248] Therefore, a further object of the present invention are
pharmaceutical compositions containing a therapeutically effective
amount of a compound of the invention or a salt thereof in
admixture with a pharmaceutically acceptable carrier.
[0249] Accordingly, the expression "therapeutically effective" when
referred to an "amount", a "dose" or "dosage" of the compounds of
this invention is intended as an "amount", a "dose" or "dosage" of
any said compounds sufficient for use in both treatment of the
established symptoms and the prophylactic treatment of the above
said pathological affections.
[0250] The pharmaceutical compositions object of the present
invention may be administered in a variety of immediate and
modified release dosage forms, e.g. orally, in the form of tablets,
troches, capsules, sugar or film coated tablets, liquid solutions,
emulsions or suspensions; rectally, in the form of suppositories;
parenterally, e.g. by intramuscular and/or depot formulations;
intravenous injection or infusion; locally and transdermally in
form of patch and gel and cream.
[0251] Suitable pharmaceutically acceptable, therapeutically inert
organic and/or inorganic carrier materials useful in the
preparation of such composition include, for example, water,
gelatin, arabic gum, lactose, starch, cellulose, magnesium
stearate, talc, vegetable oils, cyclodextrins, polyalkyleneglycols
and the like.
[0252] The composition comprising the fluorinated
arylalkylaminocarboxamide derivatives of formula I as above defined
can be sterilized and may contain further well known components,
such as, for example, preservatives, stabilizers, wetting or
emulsifying agents, e.g. paraffin oil, mannite monooleate, salts to
adjust osmotic pressure, buffers and the like. For example, the
solid oral forms may contain, together with the active ingredient,
diluents, e.g. lactose, dextrose, saccharose, cellulose, corn
starch or potato starch; lubricants, e.g. silica, talc, stearic
acid, magnesium or calcium stearate, and/or polyethylene glycols;
binding agents, e.g. starches, arabic gums, gelatin,
methylcellulose, carboxymethylcellulose or polyvinyl pyrrolidone;
disgregating agents, e.g. a starch, alginic acid, alginates or
sodium starch glycolate; effervescing mixtures; dyestuffs;
sweeteners; wetting agents such as lecithin, polysorbates,
laurylsulphates; and, in general, non-toxic and pharmacologically
inactive substances used in pharmaceutical formulations. Said
pharmaceutical preparations may be manufactured in known manner,
for example, by means of mixing, granulating, tabletting,
sugar-coating, or film-coating processes.
[0253] The preparation of the pharmaceutical compositions object of
the invention can be carried out according to common
techniques.
[0254] The oral formulations comprise sustained release
formulations that can be prepared in conventional manner, for
instance by applying an enteric coating to tablets and granules.
The liquid dispersion for oral administration may be e.g. syrups,
emulsions and suspensions.
[0255] The syrups may contain as carrier, for example, saccharose
or saccharose with glycerine and/or mannitol and/or sorbitol.
[0256] Suspensions and emulsions may contain as a carrier, for
example, a natural gum, agar, sodium alginate, pectin,
methylcellulose, carboxymethylcellulose, or polyvinyl alcohol. The
suspensions or solutions for intramuscular injections may contain,
together with the active compound, a pharmaceutically acceptable
carrier, e.g. sterile water, olive oil, ethyl oleate, glycols, e.g.
propylene glycol, and, if desired, a suitable amount of lidocaine
hydrochloride. The solutions for intravenous injections or infusion
may contain as carrier, for example, sterile water or preferably
they may be in the form of sterile, aqueous, isotonic saline
solutions.
[0257] The suppositories may contain, together with the active
ingredient, a pharmaceutically acceptable carrier, e.g. cocoa
butter, polyethylene glycol, a polyoxyethylene sorbitan fatty acid
ester surfactant or lecithin.
[0258] The pharmaceutical compositions comprising the fluorinated
arylalkylaminocarboxamide derivatives of formula I as above defined
will contain, per dosage unit, e.g., capsule, tablet, powder
injection, teaspoonful, suppository and the like from about 0.1 to
about 500 mg of one or more active ingredients most preferably from
1 to 10 mg.
[0259] Optimal therapeutically effective doses to be administered
may be readily determined by those skilled in the art and will
vary, basically, with the strength of the preparation, with the
mode of administration and with the advancement of the condition or
disorder treated. In addition, factors associated with the
particular subject being treated, including subject age, weight,
diet and time of administration, will result in the need to adjust
the dose to an appropriate therapeutically effective level.
Experimental Part
[0260] The .sup.1H-NMR spectra are stored in solution of CDCl.sub.3
or DMSO-d.sub.6 with a Varian Gemini 200 MHz spectrometer. The
chemical shifts are defined as d with CDCl.sub.3 or DMSO-d.sub.6
and D.sub.2O as internal standards.
[0261] The HPLC/MS analyses are performed with a Gilson instrument
by utilizing a X-Terra RP18 column (5 .mu.m, 4.6.times.50 mm)
coupled to a UV detector (220 nm) and a Finnigan Aqa mass
spectrometer (electron spray, positive ionization mode). General
conditions utilized for the analyses: flow: 1.2 ml/min; column
temperature: 50.degree. C.; AB elution gradient (eluent A: 0.1%
formic acid in water; eluent B: 0.1% formic acid in acetonitrile):
5-95% of B from 0 to 8.0 minutes, 95% of B from 8.0 to 9.5
minutes.
[0262] Abbreviations which are used in the description of the
Schemes and the Examples that follow are:
DCM: dichloromethane EtAc: ethyl acetate THF: tetrahydrofuran PE:
petroleum ether DMF: dimethylformamide DMSO: dimethylsulfoxide
DIPEA: diisopropylethylamine NaH: sodium hydride LiAlH.sub.4:
lithium aluminum hydride
LC/MS: Liquid Chromatography/Mass Spectrometry
TLC: Thin Layer Chromatography
[0263] RT: room temperature Boc.sub.2O:
di-tert-butyl-dicarbonate
EXAMPLES
[0264] For better illustrating the invention the following examples
are given.
Example 1-1
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide,
hydrochloride
##STR00012##
[0266] Formula: C.sub.16H.sub.24F.sub.2N.sub.2O.sub.2
[0267] MW: 314.36
[0268] Mass/charge ratio: 315.36 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.)
[0269] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.58 (bs, 2H),
7.47 (t, 1H), 6.98-7.29 (m, 3H), 4.08 (s, 2H), 4.04 (t, 2H), 3.86
(t, 2H), 2.93 (s, 3H), 2.91 (s, 3H), 1.62-1.82 (m, 2H), 1.34-1.54
(m, 2H), 0.95 (t, 3H).
[0270] The above compound is synthesized according to Scheme 1
##STR00013##
Step A
[0271] To a solution of 2-(3-methoxyphenyl)acetonitrile (2 g; 13.59
mmol) in 13 mL of dry DCM) cooled at 0.degree. C. under nitrogen
atmosphere, a 1M solution of BBr.sub.3 in DCM (28.54 mmol; 28.54
mL) is slowly added dropwise. The mixture is stirred at room
temperature for 20 hours. The reaction mixture is then poured into
ice, water is added and the organic phase is extracted three times
with DCM, washed with brine and dried over anhydrous
Na.sub.2SO.sub.4. After evaporation, the crude mixture is
chromatographed on silica gel using PE/EtAc (80/20) as an eluant,
yielding 1.28 g (71%) of 2-(3-hydroxyphenyl)acetonitrile.
Step B
[0272] To a solution of 2-(3-hydroxyphenyl)acetonitrile (2.29 g;
17.11 mmol) in dry DMF (25 mL), K.sub.2CO.sub.3 (7.08 g; 51.33
mmol), KI (0.61 g; 3.70 mmol) and 1-bromobutane (4.69 g; 3.69 mL;
34.22 mmol) are added and the mixture is stirred at 60.degree. C.
for 5 hours and then at room temperature overnight. The reaction
mixture is extracted with EtAc (150 mL) and washed with brine (150
mL): the aqueous phase is acidified with 0.1N HCl and extracted
again with ethyl acetate. The combined organic phases are dried
over anhydrous Na.sub.2SO.sub.4, filtered and evaporated. The crude
mixture is purified by flash-chromatography (eluant: PE/EtAc 99/1)
yielding, after evaporation, 3.07 g (95%) of
2-(3-butoxyphenyl)acetonitrile.
Step C
[0273] A solution of 2-(3-butoxyphenyl)acetonitrile (903 mg; 4.80
mmol) in dry THF (75 mL) is cooled at -78.degree. C. and
tert-butyllithium (1.6M in pentane; 6.6 mL; 10.56 mmol) is added
dropwise while maintaining the internal temperature between
-75.degree. C. and -78.degree. C. The solution is stirred for 10
minutes at -78.degree. C. then a solution of
N-fluorobenzensulfonimide (N-FSI; 3.78 g; 12.00 mmol) in dry THF
(12 mL) is added within 15 minutes. The reaction mixture is stirred
at -78.degree. C. for 2 hours then quenched with 0.01 N HCl at
-78.degree. C. and brought to room temperature. Ethyl acetate (50
mL) is then added and the mixture evaporated. 1.79 g of
benzenesulfonimide side-product precipitate (white solid) is
filtered off. The solution is washed with brine and dried over
anhydrous Na.sub.2SO.sub.4. After evaporation, the crude residue is
flash-chromatographed (eluant; petroleum ether/ethyl acetate,
99.5/0.5 then petroleum PE/EtAc, 99/1) to give 559 mg (52%) of pure
2,2-difluoro-[2-(-3-methoxyphenyl)]acetonitrile and further 355 mg
of the same product to be further purified.
Step D
[0274] A solution of AlCl.sub.3 (400 mg; 3.00 mmol) in dry ethyl
ether (6 mL) is stirred at 0.degree. C. for 30 minutes. A
pre-cooled (0.degree. C.) suspension of LiAlH.sub.4 (1M in THF;
3.00 mL; 3.00 mmol) is added to that mixture. After 5 minutes a
pre-cooled (0.degree. C.) solution of
2,2-difluoro-[2-(-3-methoxyphenyl)]acetonitrile in dry THF (9 mL)
is added. After 2 hours at 0.degree. C. the reaction is completed.
The solution is quenched with few drops of saturated NaHCO.sub.3,
extracted three times with EtAc, dried over anhydrous
Na.sub.2SO.sub.4, filtered and evaporated.
2,2-Difluoro-2-(3-butoxyphenyl)ethylamine (587 mg) is obtained and
used as a crude residue in Step E below.
Step E
[0275] 509 mg (2.22 mmol) of
2,2-difluoro-2-(3-butoxyphenyl)ethylamine in 41 mL of dry THF is
stirred at room temperature while di-tert-butyldicarbonate
(Boc.sub.2O) and Et.sub.3N are added. The mixture is stirred at
room temperature for 24 hours. The solvent is evaporated and the
crude residue is purified by flash-chromatography using PE/EtAc,
97/3, as an eluant.
N-(tert-butoxycarbonyl)-2,2-difluoro-2-(3-butoxyphenyl)ethylamine
was obtained as an off-white solid (481 mg, 66%).
Step F
[0276] 150 mg (0.46 mmol) of
N-(tert-butoxycarbonyl)-2,2-difluoro-2-(3-butoxyphenyl)ethylamine
are dissolved in dry DMF (2.5 mL) and the solution was cooled to
0.degree. C. NaH (60% in mineral oil; 22 mg; 0.55 mmol) is added
and the reaction mixture stirred for 10 minutes at 0.degree. C. and
for further 10 minutes at room temperature. The reaction mixture is
cooled again at 0.degree. C., then N,N-dimethylchloroacetamide (73
mg; 0.60 mmol) is added and stirring is continued for 24 hours at
room temperature. The reaction is quenched with water, extracted
three times with EtAc, washed with brine. The organic phases are
dried over anhydrous Na.sub.2SO.sub.4, filtered and evaporated. The
residue is flash-chromatographed on silica gel (eluant: DCM/EtAc,
from 98/2 to 95/5).
2-[N'-(tert-butoxycarbonyl)-2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N-
,N-dimethyl-acetamide (165 mg; 87%) is obtained as a white
solid.
Step G
[0277]
2-[N'-(tert-Butoxycarbonyl)-2,2-difluoro-2-(3-butoxyphenyl)-ethylam-
ino]-N,N-dimethyl-acetamide (160 mg; 0.39 mmol) is dissolved in DCM
(5 mL) then 0.6 mL (2.4 mmol) of 4M HCl in dioxane are added and
the reaction mixture allowed to stand overnight. Further 2 eq (0.2
mL) of 4M HCl in dioxane (total 0.8 mL) are added and the mixture
is stirred overnight. The solvent is evaporated, diethyl ether is
added and then evaporated to give 139 mg (100%) of white solid
2-[2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide,
hydrochloride (Example 1-1).
Examples 1-2 to 1-42
[0278] These compounds are prepared according to the same procedure
described in Scheme 1 using the suitable reagents.
Example 1-2
2-[2,2-Difluoro-2-(3-pentyloxyphenyl)-ethylamino]-N,N-dimethyl-acetamide,
hydrochloride
##STR00014##
[0280] Formula: C.sub.17H.sub.26F.sub.2N.sub.2O.sub.2
[0281] MW: 328.41
[0282] Mass/charge ratio: 329.25 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.) .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.37
(bs, 2H), 7.47 (t, 1H), 7.02-7.24 (m, 3H), 4.06 (s, 2H), 4.03 (t,
2H), 3.83 (t, 2H), 2.93 (s, 3H), 2.90 (s, 3H), 1.74 (q, 2H),
1.28-1.50 (m, 2H), 0.91 (t, 3H).
Example 1-3
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dipropyl-acetamide,
hydrochloride
##STR00015##
[0284] Formula: C.sub.20H.sub.32F.sub.2N.sub.2O.sub.2
[0285] MW: 370.49
[0286] Mass/charge ratio: 371.10 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.)
[0287] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 8.84 (bs, 2H),
7.48 (t, 1H), 7.02-7.26 (m, 3H), 4.03 (s, 2H), 3.94 (s, 2H), 3.78
(t, 2H), 3.21-3.29 (m, 2H), 3.06-3.19 (m, 2H), 1.64-1.79 (m, 2H),
1.37-1.61 (m, 2H), 0.95 (t, 3H), 0.86 (t, 3H), 0.64 (t, 3H).
Example 1-4
2-[2,2-Difluoro-2-(3-butoxy-4-methylphenyl)-ethylamino]-N,N-dimethyl-aceta-
mide, hydrochloride
##STR00016##
[0289] Formula: C.sub.17H.sub.26F.sub.2N.sub.2O.sub.2
[0290] MW: 328.41
[0291] Mass/charge ratio: 329.08 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.)
[0292] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.44 (bs, 2H),
7.15-7.17 (tdd, 3H), 4.03 (s, 2H), 4.03 (t, 2H), 3.81 (s, 2H), 3.09
(s, 2H), 2.80 (s, 3H), 2.78 (s, 3H), 2.13 (s, 3H), 1.77 (tt, 2H),
1.40 (tq, 2H), 0.88 (t, 3H).
Example 1-5
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dibutyl-acetamide,
hydrochloride
##STR00017##
[0294] Formula: C.sub.22H.sub.36F.sub.2N.sub.2O.sub.2
[0295] MW: 398.54
[0296] Mass/charge ratio: 399.33 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.)
Example 1-6
2-[2,2-Difluoro-2-(3-hexyloxyphenyl)-ethylamino]-N,N-dimethyl-acetamide,
hydrochloride
##STR00018##
[0298] Formula: C.sub.18H.sub.28F.sub.2N.sub.2O.sub.2
[0299] MW: 342.43
[0300] Mass/charge ratio: 343.31 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.)
[0301] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.47 (bs, 2H),
7.47 (t, 1H), 7.03-7.24 (m, 3H), 4.03 (s, 2H), 3.84 (t, 2H), 3.83
(s, 2H), 2.93 (s, 3H), 2.90 (s, 3H), 1.64-1.81 (m, 2H), 1.36-1.54
(m, 2H), 1.22-1.37 (m, 4H); 0.89 (t, 3H).
Example 1-7
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-N,N-dimet-
hyl-acetamide, hydrochloride
##STR00019##
[0303] Formula: C.sub.16H.sub.21F.sub.5N.sub.2O.sub.2
[0304] MW: 368.35
[0305] Mass/charge ratio: 369.20 (MH+, ESI pos, 3.2 KV, 25V,
400.degree. C.)
[0306] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.41 (bs, 2H),
7.43-7.58 (m, 1H), 6.99-7.29 (m, 3H), 4.11 (t, 2H), 4.06 (t, 2H),
3.84 (t, 2H), 2.93 (s, 3H), 2.90 (s, 3H), 2.31-2.48 (m, 2H),
1.86-2.09 (m, 2H), 1.22-1.37 (m, 4H); 0.89 (t, 3H).
Example 1-8
2-[2,2-Difluoro-2-(3-pentyloxyphenyl)-ethylamino]-N,N-dipropyl-acetamide,
hydrochloride
##STR00020##
[0308] Formula: C.sub.21H.sub.34F.sub.2N.sub.2O.sub.2
[0309] MW: 384.51
[0310] Mass/charge ratio: 385.22 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.)
[0311] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 8.82 (bs, 1H),
7.48 (t, 1H), 7.03-7.29 (m, 3H), 4.04 (s, 2H), 3.91 (s, 2H), 3.80
(t, 2H), 3.21-3.28 (m, 2H), 3.06-3.19 (m, 2H), 1.64-1.79 (m, 2H),
1.37-1.61 (m, 4H), 0.92 (t, 3H), 0.87 (t, 3H), 0.63 (t, 3H).
Example 1-9
2-{2,2-Difluoro-2-[3-(3-(3-fluorophenyl)-propoxy)-phenyl]-ethylamino}-N,N--
dimethyl-acetamide, hydrochloride
##STR00021##
[0313] Formula: C.sub.21H.sub.25F.sub.3N.sub.2O.sub.2
[0314] MW: 394.44
[0315] Mass/charge ratio: 395.19 (MH+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-10
2-{2,2-Difluoro-2-[3-(3-(3-chlorophenyl)-propoxy)-phenyl]-ethylamino}-N,N--
dimethyl-acetamide, hydrochloride
##STR00022##
[0317] Formula: C.sub.21H.sub.25ClF.sub.2N.sub.2O.sub.2
[0318] MW: 410.90
[0319] Mass/charge ratio: 411.75 (MH+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-11
2-[2,2-Difluoro-2-(3-butoxy-2-fluorophenyl)-ethylamino]-N,N-dimethyl-aceta-
mide, hydrochloride
##STR00023##
[0321] Formula: C.sub.16H.sub.23F.sub.3N.sub.2O.sub.2
[0322] MW: 332.37
[0323] Mass/charge ratio: 33.15 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.).
Example 1-12
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-N,N-dimethyl-ac-
etamide, hydrochloride
##STR00024##
[0325] Formula: C.sub.21H.sub.26F.sub.2N.sub.2O.sub.2
[0326] MW: 376.45
[0327] Mass/charge ratio: 377.28 (MH+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-13
2-{2,2-Difluoro-2-[3-(3-thiazol-2-yl-propoxy)-phenyl]-ethylamino}-N,N-dime-
thyl-acetamide, hydrochloride
##STR00025##
[0329] Formula: C.sub.18H.sub.23F.sub.2N.sub.3O.sub.2S
[0330] MW: 383.46
[0331] Mass/charge ratio: 384.22 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.).
Example 1-14
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-N,N-dimethyl-acetamide,
hydrochloride
##STR00026##
[0333] Formula: C.sub.19H.sub.22F.sub.2N.sub.2O.sub.2
[0334] MW: 348.40
[0335] Mass/charge ratio: 349.22 (MH+, ESI pos, 3.2 KV, 15V,
400.degree. C.).
[0336] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.40 (bs, 1H),
7.29-7.60 (m, 6H), 7.09-7.29 (m, 3H), 5.18 (s, 2H), 4.04 (s, 2H),
3.83 (t, 2H), 2.93 (s, 3H), 2.90 (s, 3H).
Example 1-15
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(pyrrolidin-1-yl)-ethanon-
e, hydrochloride
##STR00027##
[0338] Formula: C.sub.18H.sub.26F.sub.2N.sub.2O.sub.2
[0339] MW: 340.42
[0340] Mass/charge ratio: 341.02 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
[0341] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.34 (bs, 1H),
7.47 (t, 1H), 7.07-7.21 (m, 3H), 4.03 (t, 2H), 3.94 (s, 2H), 3.84
(t, 2H), 3.36 (t, 4H), 1.85-2.01 (m, 2H), 1.76-1.85 (m, 2H),
1.64-1.76 (m, 2H), 1.45 (m, 2H), 0.95 (t, 3H).
Example 1-16
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N-methyl-N-phenyl-acetamide-
, hydrochloride
##STR00028##
[0343] Formula: C.sub.21H.sub.26F.sub.2N.sub.2O.sub.2
[0344] MW: 376.45
[0345] Mass/charge ratio: 347.23 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-17
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(pyrrolidin-1-
-yl)-ethanone, hydrochloride
##STR00029##
[0347] Formula: C.sub.23H.sub.28F.sub.2N.sub.2O.sub.2
[0348] MW: 402.49
[0349] Mass/charge ratio: 403.26 (MH.sup.+, ESI pos, 3.2 KV, 15V,
400.degree. C.).
Example 1-18
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(pyrrol-
idin-1-yl)-ethanone
##STR00030##
[0351] Formula: C.sub.18H.sub.23F.sub.5N.sub.2O.sub.2
[0352] MW: 394.39
[0353] Mass/charge ratio: 395.23 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
[0354] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.48 (bs, 2H),
7.35-7.61 (m, 1H), 7.01-7.26 (m, 3H), 4.11 (t, 2H), 3.96 (s, 2H),
3.86 (t, 2H), 3.28-3.40 (m, 4H), 2.33-2.48 (m, 2H), 1.68-2.04 (m,
6H).
Example 1-19
2-[2,2-Difluoro-2-(3-benzyloxy-phenyl)-ethylamino]-1-(morpholin-4-yl)-etha-
none, hydrochloride
##STR00031##
[0356] Formula: C.sub.21H.sub.24F.sub.5N.sub.2O.sub.3
[0357] MW: 390.43
[0358] Mass/charge ratio: 391.22 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-20
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(morpholin-4--
yl)-ethanone, hydrochloride
##STR00032##
[0360] Formula: C.sub.23H.sub.28F.sub.2N.sub.2O.sub.3
[0361] MW: 418.49
[0362] Mass/charge ratio: 419.18 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-21
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(morpho-
lin-4-yl)-ethanone, hydrochloride
##STR00033##
[0364] Formula: C.sub.18H.sub.23F.sub.5N.sub.2O.sub.3
[0365] MW: 410.39
[0366] Mass/charge ratio: 411.22 (MH.sup.+, ESI pos, 3.2 KV, 15V,
400.degree. C.).
[0367] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.70 (bs, 2H),
7.50 (t, 1H), 6.88-7.37 (m, 3H), 4.15 (s, 2H), 4.12 (t, 2H), 3.87
(t, 2H), 3.54-3.67 (m, 4H), 3.44-3.54 (m, 2H), 3.32-3.44 (m, 2H),
2.32-2.47 (m, 2H), 1.86-2.06 (m, 2H).
Example 1-22
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino)1-(2H-benzo[b][1,4]oxazin-4(-
3H)-yl)-ethanone, hydrochloride
##STR00034##
[0369] Formula: C.sub.22H.sub.26F.sub.2N.sub.2O.sub.3
[0370] MW: 404.46
[0371] Mass/charge ratio: 405.29 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-23
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-1-(pyrrolidin-1-yl)-etha-
none, hydrochloride
##STR00035##
[0373] Formula: C.sub.21H.sub.24F.sub.2N.sub.2O.sub.2
[0374] MW: 374.43
[0375] Mass/charge ratio: 375.27 (MH.sup.+, ESI pos, 3.2 KV, 25V,
350.degree. C.).
Example 1-24
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-N-methyl-N-phen-
yl-acetamide, hydrochloride
##STR00036##
[0377] Formula: C.sub.26H.sub.28F.sub.2N.sub.2O.sub.2
[0378] MW: 438.52
[0379] Mass/charge ratio: 439.38 (MH.sup.+, ESI pos, 3.2 KV, 25V,
350.degree. C.).
Example 1-25
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-N-methyl--
N-phenyl-acetamide, hydrochloride
##STR00037##
[0381] Formula: C.sub.21H.sub.23F.sub.5N.sub.2O.sub.2
[0382] MW: 430.42
[0383] Mass/charge ratio: 431.29 (MH.sup.+, ESI pos, 3.2 KV, 25V,
350.degree. C.).
Example 1-26
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(4-methylpiperazin-1-yl)--
ethanone, hydrochloride
##STR00038##
[0385] Formula: C.sub.19H.sub.29F.sub.2N.sub.3O.sub.2
[0386] MW: 369.46
[0387] Mass/charge ratio: 370.07 (MH.sup.+, ESI pos, 3.2 KV, 15V,
400.degree. C.).
[0388] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 7.28-7.49 (m,
1H), 6.92-7.18 (m, 3H), 4.04 (t, 2H), 3.62-3.85 (m, 1H), 3.52 (s,
2H), 3.32 (t, 2H), 2.87-3.19 (m, 8H), 2.69 (s, 3H), 1.61-1.84 (m,
2H), 1.48 (dq, 2H), 0.96 (t, 3H).
Example 1-27
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl)-ethylamino}-1-(4-methylpipe-
razin-1-yl)-ethanone, hydrochloride
##STR00039##
[0390] Formula: C.sub.24H.sub.31F.sub.2N.sub.3O.sub.2
[0391] MW: 431.53
[0392] Mass/charge ratio: 431.37 (MH.sup.+, ESI pos, 3.2 KV, 15V,
400.degree. C.).
Example 1-28
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl)-ethylamino}-1-(4-meth-
ylpiperazin-1-yl)-ethanone, hydrochloride
##STR00040##
[0394] Formula: C.sub.19H.sub.26F.sub.5N.sub.3O.sub.2
[0395] MW: 423.43
[0396] Mass/charge ratio: 424.28 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
[0397] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 11.56 (bs, 1H),
9.57 (bs, 1H), 7.37-7.60 (m, 1H), 6.99-7.28 (m, 3H), 4.29-4.59 (m,
1H), 4.16-4.30 (m, 1H), 4.11 (t, 2H), 3.80 (t, 2H), 2.87-3.94 (m,
8H), 2.77 (s, 3H), 2.33-2.47 (m, 2H), 1.85-2.06 (m, 2H).
Example 1-29
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(piperidin-1-yl)-ethanone-
, hydrochloride
##STR00041##
[0399] Formula: C.sub.19H.sub.28F.sub.2N.sub.2O.sub.2
[0400] MW: 354.44
[0401] Mass/charge ratio: 355.03 (MH.sup.+, ESI pos, 3.2 KV, 15V,
400.degree. C.).
[0402] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.37 (bs, 2H),
7.47 (t, 1H), 7.04-7.21 (m, 3H), 4.07 (s, 2H), 3.84 (t, 2H),
3.45-3.54 (m, 2H), 3.21-3.32 (m, 2H), 1.66-1.81 (m, 2H), 1.33-1.66
(m, 8H), 0.95 (t, 3H).
Example 1-30
2-{2,2-Difluoro-2-[3-(3-phenylpropoxy)-phenyl]-ethylamino}-1-(piperidin-1--
yl)-ethanone, hydrochloride
##STR00042##
[0404] Formula: C.sub.24H.sub.30F.sub.2N.sub.2O.sub.2
[0405] MW: 416.52
[0406] Mass/charge ratio: 417.34 (MH.sup.+, ESI pos, 3.2 KV, 15V,
350.degree. C.).
Example 1-31
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-1-(piperi-
din-1-yl)-ethanone, hydrochloride
##STR00043##
[0408] Formula: C.sub.19H.sub.25F.sub.5N.sub.2O.sub.2
[0409] MW: 408.41
[0410] Mass/charge ratio: 408.07 (MH.sup.+, ESI pos, 3.2 KV, 15V,
350.degree. C.).
Example 1-32
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-diethyl-acetamide,
hydrochloride
##STR00044##
[0412] Formula: C.sub.18H.sub.28F.sub.2N.sub.2O.sub.2
[0413] MW: 342.43
[0414] Mass/charge ratio: 343.05 (MH+, ESI pos, 3.2 KV, 15V,
350.degree. C.)
[0415] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 7.47 (t, 1H),
7.07-7.20 (m, 3H), 4.03 (t, 2H), 4.02 (s, 2H), 3.84 (t, 2H), 3.34
(q, 2H), 3.24 (q, 2H), 1.64-1.82 (m, 2H), 1.36-1.55 (m, 2H), 1.12
(t, 3H), 1.07 (t, 3H), 0.95 (t, 3H).
Example 1-33
2-{2,2-Difluoro-2-[3-(2-fluorobenzyloxy)-phenyl]-ethylamino}-N,N-dimethyl--
acetamide, hydrochloride
##STR00045##
[0417] Formula: C.sub.19H.sub.21F.sub.3N.sub.2O.sub.2
[0418] MW: 366.39
[0419] Mass/charge ratio: 367.18 (MH+, ESI pos, 3.2 KV, 15V,
400.degree. C.).
Example 1-34
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(cis-3,5-dimethylpiperidi-
n-1-yl)-ethanone, hydrochloride
##STR00046##
[0421] Formula: C.sub.21H.sub.32F.sub.2N.sub.2O.sub.2
[0422] MW: 382.50
[0423] Mass/charge ratio: 383.34 (MH.sup.+, ESI pos, 3.2 KV, 15V,
350.degree. C.).
Example 1-35
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(3,4-dihydroisoquinolin-2-
(1H)-yl)-ethanone, hydrochloride
##STR00047##
[0425] Formula: C.sub.23H.sub.28F.sub.2N.sub.2O.sub.2
[0426] MW: 402.49
[0427] Mass/charge ratio: 403.22 (MH.sup.+, ESI pos, 3.2 KV, 25V,
350.degree. C.).
Example 1-36
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-diisopropyl-acetamide,
hydrochloride
##STR00048##
[0429] Formula: C.sub.20H.sub.32F.sub.2N.sub.2O.sub.2
[0430] MW: 370.49
[0431] Mass/charge ratio: 371.19 (MH.sup.+, ESI pos, 3.2 KV, 25V,
350.degree. C.).
Example 1-37
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N-cyclohexyl-N-methyl-aceta-
mide, hydrochloride
##STR00049##
[0433] Formula: C.sub.21H.sub.32F.sub.2N.sub.2O.sub.2
[0434] MW: 382.50
[0435] Mass/charge ratio: 383.31 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-38
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-1-(piperidin-1-yl)-ethan-
one, hydrochloride
##STR00050##
[0437] Formula: C.sub.21H.sub.32F.sub.2N.sub.2O.sub.2
[0438] MW: 388.46
[0439] Mass/charge ratio: 389.21 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-39
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-[4-(phenylsulfonyl)-piper-
azin-1-yl]-ethanone, hydrochloride
##STR00051##
[0441] Formula: C.sub.24H.sub.31F.sub.2N.sub.3O.sub.4S
[0442] MW: 495.59
[0443] Mass/charge ratio: 496.24 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-40
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(indolin-1-yl)-ethanone,
hydrochloride
##STR00052##
[0445] Formula: C.sub.22H.sub.26F.sub.2N.sub.2O.sub.2
[0446] MW: 388.46
[0447] Mass/charge ratio: 389.25 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.).
Example 1-41
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(4-benzylpiperazin-1-yl)--
ethanone, dihydrochloride
##STR00053##
[0449] Formula: C.sub.25H.sub.33F.sub.2N.sub.3O.sub.2
[0450] MW: 445.56
[0451] Mass/charge ratio: 446.34 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 1-42
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino)-1-(azetidin-1-yl)-ethanone,
hydrochloride
##STR00054##
[0453] Formula: C.sub.17H.sub.24F.sub.2N.sub.2O.sub.2
[0454] MW: 326.39
[0455] Mass/charge ratio: 327.13 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
[0456] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 7.31-7.46 (m,
1H), 6.95-7.13 (m, 3H), 4.01 (t, 4H), 3.84 (t, 2H), 3.14-3.24 (m,
2H), 3.104 (dq, 2H), 2.18 (dt, 2H), 1.63-1.79 (m, 2H), 1.45 (dq,
2H), 0.94 (t, 3H).
Example 2-1
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-propanamide,
hydrochloride
##STR00055##
[0458] Formula: C.sub.17H.sub.26F.sub.2N.sub.2O.sub.2
[0459] MW: 328.41
[0460] Mass/charge ratio: 329.02 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.)
[0461] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.48 (bs, 1H),
7.47 (t, 1H), 7.04-7.24 (m, 3H), 4.24-4.51 (m, 1H), 4.03 (t, 2H),
3.71-3.95 (m, 1H), 3.51-3.71 (m, 1H), 2.98 (s, 3H), 2.89 (s, 3H),
1.62-1.82 (m, 2H), 1.42 (d, 3H), 1.34-1.54 (m, 2H), 0.95 (t,
3H).
[0462] The above compound was synthesized according to Scheme 2
##STR00056##
Step A
[0463] 75 mg (0.23 mmol) of
N-tert-butoxycarbonyl-2,2-difluoro-2-(3-butoxyphenyl)-ethylamine
are dissolved in dry DMF (5 mL). The solution is cooled to
0.degree. C. and NaH (6.7 mg; 1.2 eq) is added. The solution is
stirred at at RT for 10 min, then cooled again to 0.degree. C. and
2-chloro-N,N-dimethylpropanamide (31 mg; 0.23 mmol) are added.
After 4 hours a further 1.2 eq of NaH (6.7 mg) are added. The
reaction mixture is stirred overnight at RT, then quenched with
water, evaporated, the residue taken up with water and EtAc. The
organic layer is separated and the water layer extracted three
times with EtAc. The combined organic phases are dried over
anhydrous Na.sub.2SO.sub.4, filtered and evaporated. The resulting
crude residue is flash-chromatographed (eluant: petroleum
ether/EtAc, 9/1 to 8/2).
2-[N-tert-butoxycarbonyl-2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N--
dimethylpropanamide (84 mg; 81%) is obtained as pale yellow
oil.
Step B
[0464] 84 mg (0.196 mmol) of
2-[N-tert-butoxycarbonyl-2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N--
dimethylpropanamide are dissolved in 3 mL of DCM and 0.49 mL (1.96
mmmol; 10 eq) of a 4M HCl solution in dioxane are added. After 8
hours, further 10 eq (0.49 mL) of 4M HCll dioxane are added and the
mixture is stirred overnight. Further 5 eq (0.245 mL) of 4M HCl in
dioxane are the added and the mixture stirred for further 24 hours.
The solution is evaporated, the white residue suspended in diethyl
ether and then evaporated twice. The residue is
flash-chromatographed on silica gel (DCM/MeOH 1/1 as an eluant
followed by MeOH/conc. NH3 95/5). and the free base
2-[2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethylpropanamide
is isolated as a pale yellow fluid. The compound is dissolved in
DCM (3 mL) and 4M HCl in dioxane is added to bring the solution to
pH 2. The mixture is stirred for 10 minutes and then evaporated.
The white residue is taken up with ether and evaporated twice. 45
mg (49%) of
2-[2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethylpropanamide,
hydrochloride (Example 2-1) are obtained.
Examples 2-2 to 2-5
[0465] These compounds are prepared according to the procedure
described in Scheme 2.
Example 2-2
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-3-methoxy-N,N-dimethyl-prop-
anamide
##STR00057##
[0467] Formula: C.sub.18H.sub.28F.sub.2N.sub.2O.sub.3
[0468] MW: 358.43
[0469] Mass/charge ratio: 359.40 (MH.sup.+, ESI pos, 3.2 KV, 15V,
400.degree. C.).
[0470] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 7.39-7.59 (m,
1H), 7.06-7.21 (m, 3H), 4.64 (t, 1H), 4.03 (t, 2H), 3.58-3.87 (m,
4H), 3.30 (s, 3H), 3.02 (s, 3H), 2.91 (s, 3H), 1.59-1.81 (m, 2H),
1.36-1.57 (m, 2H), 0.94 (t, 3H).
Example 2-3
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-3-(4-methoxyphenyl)-N,N-dim-
ethyl-propanamide, hydrochloride
##STR00058##
[0472] Formula: C.sub.24H.sub.32F.sub.2N.sub.2O.sub.3
[0473] MW: 434.53
[0474] Mass/charge ratio: 435.36 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
Example 2-4
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-2-N,N-trimethyl-propanamide-
, hydrochloride
##STR00059##
[0476] Formula: C.sub.18H.sub.28F.sub.2N.sub.2O.sub.2
[0477] MW: 342.43
[0478] Mass/charge ratio: 342.31 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.).
Example 2-5
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-4-N,N-trimethyl-pentanamide-
, hydrochloride
##STR00060##
[0480] Formula: C.sub.20H.sub.32F.sub.2N.sub.2O.sub.2
[0481] MW: 370.49
[0482] Mass/charge ratio: 371.33 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.).
Example 3-1
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-methylamino}-N,N-dimethyl-aceta-
mide, hydrochloride
##STR00061##
[0484] Formula: C.sub.17H.sub.26F.sub.2N.sub.2O.sub.2
[0485] MW: 328.41
[0486] Mass/charge ratio: 329.17 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
[0487] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 7.45 (t, 1H),
7.04-7.18 (m, 3H), 4.03 (t, 2H), 3.73-3.91 (m, 2H), 3.70-4.13 (m,
2H), 2.86 (s, 3H), 2.84 (s, 3H), 2.80 (s, 3H), 1.62-1.82 (m, 2H),
1.37-1.57 (m, 2H), 0.94 (t, 3H).
[0488] The above compound is synthesized according to Scheme 3
##STR00062##
Examples 3-2 to 3-3
[0489] These compounds are prepared according to the procedure
described in Scheme 3.
Step A
[0490] 107.5 mg (0.34 mmol) of the free base of
2-[2,2-difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide
(see Example 1-1) are dissolved in dry THF (10 mL). To this
solution, formaldehyde (36.5% water solution, 52.1 .mu.L; 0.69
mmol), acetic acid (2.5 mL), and MP-CNBH.sub.3 (2.3 mmol/g; 325 mg;
0.75 mmol) are added sequentially. After stirring 1 hour the
reaction is completed. After further stirring for 1.5 hours, the
reaction mixture is evaporated. The crude residue is
flash-chromatographed on silica gel using DCM/MeOH (99.5/0.5) as an
eluant. 73.1 mg (65%) of pale yellow fluid
2-{[2,2-difluoro-2-(3-butoxyphenyl)-ethyl]-methylamino}-N,N-dimethyl-acet-
amide are obtained.
Step B
[0491] A solution of 73.1 mg of
2-{[2,2-difluoro-2-(3-butoxyphenyl)-ethyl]-methylamino}-N,N-dimethyl-acet-
amide in DCM (3 mL) is stirred and few drops of 4M HCl in dioxane
are added until reaching pH 2. The reaction mixture is stirred for
5 minutes and then evaporated. The white solid residue is suspended
in Et.sub.2O and evaporated twice to yield 77.4 mg (96%) of white
solid
2-{[2,2-difluoro-2-(3-butoxyphenyl)-ethyl]-methylamino}-N,N-dimethyl-acet-
amide, hydrochloride (Example 3-1).
Example 3-2
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-(3-methoxypropyl)-amino}-N,N-di-
methyl-acetamide, hydrochloride
##STR00063##
[0493] Formula: C.sub.20H.sub.32F.sub.2N.sub.2O.sub.3
[0494] MW: 386.49
[0495] Mass/charge ratio: 387.28 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
[0496] .sup.1H-NMR (300 MHz, DMSO-d6+TFA) .delta. ppm 7.38-7.50 (m,
1H), 7.08-7.17 (m, 3H), 3.98-4.11 (m, 4H), 3.85 (t, 2H), 3.33 (t,
2H), 3.21 (s, 3H), 3.12-3.20 (m, 2H), 2.88 (s, 3H), 2.86 (s, 3H),
1.78-1.91 (m, 2H), 1.65-1.78 (m, 2H), 1.36-1.54 (m, 2H), 0.94 (t,
3H).
Example 3-3
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-(2-methoxyethyl)-amino}-N,N-dim-
ethyl-acetamide, hydrochloride
##STR00064##
[0498] Formula: C.sub.19H.sub.30F.sub.2N.sub.2O.sub.3
[0499] MW: 372.46
[0500] Mass/charge ratio: 373.30 (MH.sup.+, ESI pos, 3.2 KV, 25V,
400.degree. C.).
[0501] .sup.1H-NMR (300 MHz, DMSO-d6+TFA) .delta. ppm 7.38 (t, 1H),
6.95-7.11 (m, 3H), 4.01 (t, 2H), 3.50 (s, 2H), 3.42 (t, 2H), 3.28
(t, 2H), 2.85 (s, 3H), 2.79 (s, 3H), 2.76 (s, 3H), 1.63-1.79 (m,
2H), 1.35-1.52 (m, 2H), 0.94 (t, 3H).
Example 4-1
2-[2-Fluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl-acetamide,
hydrochloride
##STR00065##
[0503] Formula: C.sub.16H.sub.25FN.sub.2O.sub.2
[0504] MW: 296.39
[0505] Mass/charge ratio: 297.04 (MH+, ESI pos, 3.2 KV, 25V,
350.degree. C.)
[0506] .sup.1H-NMR (300 MHz, DMSO-d6) .delta. ppm 9.35 (bs, 2H),
7.22-7.487 (m, 1H), 6.83-7.07 (m, 3H), 5.97 (ddd, 1H), 4.10 (s,
2H), 4.004 (t, 2H), 3.55 (td, 1H), 3.33-3.47 (m, 1H), 2.95 (s, 3H),
2.91 (s, 3H), 1.56-1.78 (m, 2H), 1.29-1.54 (m, 2H), 0.94 (t,
3H).
[0507] The above compound is synthesized according to Scheme 4
##STR00066##
Step A
[0508] To a solution of 2-(3-methoxyphenyl)acetonitrile (2 g; 13.59
mmol) in 13 mL of dry dichloromethane (DCM) cooled at 0.degree. C.
under an inert atmosphere, a 1M solution of BBr.sub.3 in DCM (28.54
mmol; 28.54 mL) is slowly added dropwise. The mixture is stirred at
room temperature for 20 hours. The reaction mixture is then poured
into ice, water is added and the organic phase is extracted three
times with dichloromethane, washed with brine and dried over
anhydrous Na.sub.2SO.sub.4. After evaporation, the crude mixture is
purified by flash-chromatography on silica gel using petroleum
ether/EtAc (80/20) as an eluant, affording 1.28 g (71%) of
2-(3-hydroxyphenyl)acetonitrile.
Step B
[0509] To a solution of 2-(3-hydroxyphenyl)acetonitrile (2.29 g;
17.11 mmol) in dry DMF (25 mL), K.sub.2CO.sub.3 (7.08 g; 51.33
mmol), KI (0.61 g; 3.70 mmol) and 1-bromobutane (4.69 g; 3.69 mL;
34.22 mmol) are added and the mixture is stirred at 60.degree. C.
for 5 hours and then at room temperature overnight. A TLC (DCM/EtAc
95/5) shows no presence of starting material. After evaporation,
the reaction mixture is extracted with ethyl acetate (150 mL) and
washed with brine (150 mL twice): the aqueous phase is acidified
with 0.1N HCl and extracted again with ethyl acetate. The combined
organic layers are dried over anhydrous Na.sub.2SO.sub.4, filtered
and evaporated. The crude mixture is purified by
flash-chromatography (eluant: petroleum ether/ethyl acetate 99/1)
yielding, after evaporation, 3.07 g (95%) of
2-(3-butoxyphenyl)acetonitrile as a pale yellow oil.
Step C
[0510] Tert-buthyllithium (1268 uL; 2.16 mmol) is added dropwise to
a solution of 2-(3-butoxyphenyl)acetonitrile (371 mg; 1.96 mmol) in
THF (16 mL) at -78.degree. C., under nitrogen atmosphere. The light
yellow solution turned into orange and stirring is continued for 1
hour. A solution of N-fluorobenzenesulfonimide (618 mg; 1.96 mmol)
in THF (2 mL) is added dropwise and the reaction is stirred at
-78.degree. C. for 2 hours. TLC (petroleum ether/EtAc 9:1) reveals
no presence of starting material and two more apolar spots. The
reaction is then quenched by adding 0.01N HCl, then more water is
added and extracted with DCM (three times). The combined organic
layers are dried over Na.sub.2SO.sub.4, filtered and evaporated.
The crude residue is purified by flash chromatography (petroleum
ether/EtAc, 99/1) affording 217 mg (53%) of
2-(3-butoxyphenyl)-2-fluoroacetonitrile as a colorless oil.
Step D
[0511] To a solution of 2-(3-butoxyphenyl)-2-fluoroacetonitrile
(109 mg: 0.53 mmol) in dry THF (5 mL), borane tetrahydrofurane
complex (2.10 mL; 2.10 mmol) is added and the reaction is stirred
at 0.degree. C. for 2 hours and then at RT for 6 hours. An LC/MS
shows almost complete conversion. The reaction is quenched adding
slowly few drops of EtOH and few drops of conc. HCl/EtOH (1:5) and
stirring is continued for 5 min. DCM was then added, followed by 5%
aqueous NaHCO.sub.3. The two phases are separated and the aqueous
phase is extracted twice with DCM. The combined organic layers are
dried over Na.sub.2SO.sub.4, filtered and evaporated. The crude
residue is purified using a SCX cartridge (eluant: DCM/MeOH 1/1 to
MeOH/conc. aq. NH.sub.3 95/5) affording
2-(3-butoxyphenyl)-2-fluoroethanamine (92 mg; 0.43 mmol; 83%) as a
pale yellow oil.
Step E
[0512] DIPEA (0.106 mL; 0.61 mmol) is added to a solution of
2-(3-butoxyphenyl)-2-fluoroethaneamine (92 mg; 0.43 mmol) and
Boc.sub.2O (0.121 mL; 0.52 mmol) in dry THF (6 mL) and the reaction
is stirred at RT for 2 hours. An LC/MS shows complete conversion.
DCM is added and the solution is washed with 5% aq. NaHCO.sub.3 and
1N HCl, dried over anhydrous Na.sub.2SO.sub.4, filtered and
evaporated to give tert-butyl
2-(3-butoxyphenyl)-2-fluoroethylcarbamate (136 mg; 0.437 mmol;
100%) as a pale yellow oil.
Step F
[0513] A solution of tert-butyl
2-(3-butoxyphenyl)-2-fluoroethylcarbamate (136 mg; 0.44 mmol) in
dry DMF (4 mL) under nitrogen atmosphere is cooled to 0.degree. C.
and sodium hydride (22.7 mg; 0.57 mmol) is added. The mixture is
stirred at RT for 10 min, then is cooled again to 0.degree. C. and
2-chloro-N,N-dimethyl-acetamide (0.054 mL; 0.524 mmol) is added.
The reaction mixture is stirred at RT for 4 hours. An LC/MS shows
very low conversion. Additional sodium hydride (38 mg; 0.96 mmol)
is added followed after 10 min by 2-chloro-N,N-dimethyl-acetamide
(0.09 mL; 0.87 mmol). Stirring is continued for 12 hours. An LC/MS
shows almost complete conversion. The solvent is evaporated, EtAc
is added and the solution is washed with brine, then dried over
anhydrous Na.sub.2SO.sub.4, filtered and evaporated. The crude
residue is purified by flash-chromatography (DCM/EtAc from 96/4 to
95/5) yielding tert-butyl
N-[2-(3-butoxyphenyl)-2-fluoroethyl]-N-[(2-dimethylamino)-2-oxoethyl)]-ca-
rbamate (100 mg; 0.25 mmol; 58%) as a colourless oil.
Step G
[0514] A mixture of tert-butyl
N-[2-(3-butoxyphenyl)-2-fluoroethyl]-N-[(2-dimethylamino)-2-oxoethyl)]-ca-
rbamate (96 mg; 0.24 mmol) and 4M HCl in dioxane (363 uL; 1.45
mmol) in dry DCM (6 mL) is stirred at RT for 4 hours. An LC/MS show
complete conversion. The solvent is evaporated affording
2-[2-(3-butoxyphenyl)-2-fluoroethylamino)]-N,N-dimethyl-acetamide
(50 mg; 0.17 mmol; 70%) as a pale yellow amorphous solid which is
triturated with EtAc, filtered and dried to give 22.2 mg (0.067
mmol; 44%) of white solid
2-[2-(3-butoxyphenyl)-2-fluoroethylamino]-N,N-dimethyl-acetamide,
hydrochloride. (Example 4-1).
Examples 4-2 to 4-3
[0515] These compounds were prepared according to the procedure
described in Scheme 4.
Example 4-2
2-{2-Fluoro-2-[3-(3-chlorobenzyloxy)-phenyl]-ethylamino}-N,N-dimethyl-acet-
amide, hydrochloride
Example 4-3
2-{2-Fluoro-2-[3-(3-fluorobenzyloxy)-phenyl]-ethylamino}-N,N-dimethyl-acet-
amide, hydrochloride
Example 5
N-Type Calcium Channel Influx Assay
[0516] IMR32 human neuroblastoma cells constitutively express both
L- and N-type channels. Under differentiating conditions, IMR32
cells preferentially express on the membrane surface N-type calcium
channels. The remaining L-type calcium channels are blocked using
the selective L-type blocker nifedipine. In these experimental
conditions only N-type channels can be detected.
[0517] IMR32 cells are differentiated using 1 mM dibutyryl-cAMP and
2.5 .mu.M bromodeoxyuridine for 8 days (4 times) in 225 cm.sup.2
flask, then detached, seeded at 200,000 cells/well on 96
poly-L-lysine-coated plates and further incubated for 18-24 h in
the presence of differentiating buffer before use.
[0518] The Ca.sup.2+ Kit Assay (Molecular Devices, CA--USA), based
on a fluorescent calcium indicator and able to detect the calcium
influx determined by depolarizing conditions, is used for the
assay.
[0519] Differentiated cells are incubated with dye loading for 30
minutes at 37.degree. C. then, nifedipine alone (1 .mu.M) or in the
presence of co-conotoxin (as reference standard) or test compounds
are added for further 15 minutes.
[0520] The fluorescence (excitation: 485 nm, emission: 535 nm
wavelength) is measured before and after (30-40 s) the automated
injection of 100 mM KCl depolarizing solution using a Victor plate
reader (Perkin Elmer, MA--USA).
[0521] The inhibition curves are calculated from 5 concentrations,
each in triplicate, and the IC.sub.50 determined using a linear
regression analysis.
[0522] The compounds of the present invention inhibit N-type
calcium channels with pharmacologically significant IC.sub.50
values.
Example 6
TTXs-Sodium Channel Influx Assay
[0523] ND7/23 rat dorsal root ganglion-derived cell line
endogenously expresses a mixed population of TTXs sodium channels
(such as Nav1.3, Nav1.2, Nav1.1, Nav1.6). These cells lack of TTXr
sodium channels as shown by the absence of their respective
transcripts. ND7/23 cells are grown in Dulbecco's Modified Eagle
Medium (DMEM, Invitrogen, CA--USA) supplemented with 10% Foetal
Bovine Serum (FBS, Invitrogen, CA--USA) and 1 mM sodium piruvate.
The cells are seeded at 50,000 cells/well on 96
poly-L-lysine-coated plates and further incubated for 18-24 h
before use.
[0524] The Membrane Potential Kit Assay (Molecular Devices,
CA--USA), based on a negatively charged fluorescent dye able to
monitor changes in membrane potential caused by the sodium influx
due to the channel opening, is used for the assay.
[0525] Cells are incubated with the dye loading for 30 minutes at
25.degree. C. Then, 100 nM of the toxin Anemonia sulcata (used as
enhancer of the channel opener response) alone or in the presence
of TTX (as reference standard) or test compound are added for
further 15 minutes.
[0526] The fluorescence (excitation: 530 nm, emission: 565 nm
wavelength) is measured before and after (40-45 s) the automated
injection of the sodium channel opener veratridine (100 .mu.M)
using a Victor plate reader (Perkin Elmer, MA--USA).
[0527] The inhibition curves are calculated from 5 concentrations,
each in triplicate, and the IC.sub.50 determined using a linear
regression analysis.
[0528] The compounds of the present invention inhibit TTXs sodium
channels with pharmacologically significant IC.sub.50 values.
[0529] The results, obtained with some compounds which are
representative of the entire class of compounds of the invention
are reported in Table 1.
TABLE-US-00001 TABLE 1 IC.sub.50 COMPOUND [.mu.M]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl- 1.0
acetamide, hydrochloride (Example 1-1)
2-[2,2-Difluoro-2-(3-pentyloxyphenyl)-ethylamino]-N,N-dimethyl- 1.3
acetamide, hydrochloride (Example 1-2)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dipropyl- 0.58
acetamide, hydrochloride (Example 1-3)
2-[2,2-Difluoro-2-(3-hexyloxyphenyl)-ethylamino]-N,N-dimethyl- 0.14
acetamide, hydrochloride (Example 1-6)
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-
3.6 N,N-dimethyl-acetamide, hydrochloride (Example 1-7)
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-N,N-dimethyl- 1.2
acetamide, hydrochloride (Example 1-14)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(pyrrolidin-1- 1.1
yl)-ethanone, hydrochloride (Example 1-15)
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-
0.95 1-(pyrrolidin-1-yl)-ethanone (Example 1-18)
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-
7.2 1-(morpholin-4-yl)-ethanone, hydrochloride (Example 1-21)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(4- 23.2
methylpiperazin-1-yl)-ethanone, hydrochloride (Example 1-26)
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-
3.2 1-(4-methylpiperazin-1-yl)-ethanone, hydrochloride (Example
1-28)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(piperidin-1-yl)-
1.8 ethanone, hydrochloride (Example 1-29)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-diethyl- 2.8
acetamide, hydrochloride (Example 1-32)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(azetidin-1-yl)-
5.0 ethanone, hydrochloride (Example 1-42)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl- 20.0
propanamide, hydrochloride (Example 2-1)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-3-methoxy-N,N- 6.2
dimethyl-propanamide (Example 2-2)
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-methylamino}-N,N- 2.3
dimethyl-acetamide, hydrochloride (Example 3-1)
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-(3-methoxypropyl)- 2.4
amino}-N,N-dimethyl-acetamide, hydrochloride (Example 3-2)
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-(2-methoxyethyl)- 1.4
amino}-N,N-dimethyl-acetamide, hydrochloride (Example 3-3)
2-[2-Fluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl- 1.5
acetamide, hydrochloride (Example 4-1)
Example 7
Patch Clamp Studies of Calcium Currents Inhibition
Cells and Methods:
[0530] Functional inhibition of the N-type Ca currents is studied
using whole cell patch clamp methods (Hamill O. P., Marty A., Neher
E., Sakmann B., Sigworth F. J. Pflugers Arch. 391: 85-100 (1981))
on HEK293 cells expressing recombinant human N-type channels,
obtained after transient transfection of h .alpha.1B
(hCav2.2)+.beta.1b+.alpha.2.delta.-1 subunits.
[0531] Membrane currents are recorded and filtered at 5 kHz with an
Axon Axopatch 200B amplifier and digitized with an Axon Digidata
1322A (Axon Instruments, CA, USA). Voltage clamping of membrane
potentials and data acquisition are controlled online with Axon
pClamp8 software. Measuring and reference electrodes are AgCl--Ag
electrodes. Cells have initial seal resistances of >1 G.OMEGA.
and access resistances of 4.2.+-.0.2 M.OMEGA.. Cells are
continuously superfused with extracellular solutions using a
Biologic RSC-200 (Biologic SAS, France).
[0532] For calcium currents recording the control bath solution
contained (mM): choline chloride (70), MgCl.sub.2 (1), BaCl.sub.2
(20), TEA.Cl (50), Hepes (10), glucose (10). Internal pipette
solution consists of (mM): CsCl (140), EGTA (10), MgCl.sub.2 (2),
Hepes (10), MgATP (1), GTP Tris (0.3).
[0533] Compounds are dissolved as 20 mM stock solutions in DMSO and
then diluted to the final concentration in the external
solutions.
Voltage Protocols and Data Analyses:
[0534] A two-step protocol is used to determine the voltage
dependence of the block: N-type current is activated by a 600 ms
step pulse to +10 mV (test pulse) from a 5000 ms preconditioning
potential of -110 mV (resting condition) or -50/-55 mV (half
maximal steady-state inactivated condition), respectively.
[0535] The amplitude of calcium current peaks evoked by the
respective test pulses at a frequency of 0.06 Hz are measured
before and after exposure to the test substance. Tonic block of
currents is calculated as the difference between the peak calcium
current measured at the end of a stabilization period in the
control external bath solution and peak currents measured at the
end of test substance perfusion period (when steady state is
reached) divided by control peaks. Drug concentration-inhibition
curves are obtained by plotting tonic blocks versus drug
concentrations. Dose-response curves are fitted to the tonic block
data, according to the logistic equation:
y=A2+(A1-A2)/[1+(x/IC.sub.50).sup.p]. A1 and A2 are fixed values of
0 and 1 corresponding to 0 and 100% current inhibition, x is the
drug concentration, IC.sub.50 is the drug concentration resulting
in 50% current inhibition and p is the corresponding slope
factor.
[0536] The compounds of the present invention inhibit N-type
calcium channels with pharmacologically significant IC.sub.50
values.
Example 8
Patch Clamp Studies of Sodium Currents Inhibition
[0537] Cells and methods: Functional inhibition of the sodium
currents is studied using whole cell patch clamp methods (Hamill O.
P., Marty A., Neher E., Sakmann B., Sigworth F. J., Pflugers Arch.
391 (2): 85-100 (1981)) on the hybrid cell line ND7/23 (Wood J N,
Bevan S J, Coote P R, Dunn P M, Harmar A, Hogan P, Latchman D S,
Morrison C, Rougon G, Theveniau M.: "Novel cell lines display
properties of nociceptive sensory neurons". Proc. Biol. Sci.
September 22; 241(1302):187-94 (1990)), that express a mixed
population of voltage gated sodium channels.
[0538] Membrane currents are recorded as described in the example
above.
[0539] For sodium current recording control bath solution contained
(mM): NaCl (80), choline chloride (38), CaCl.sub.2 (1.3),
MgCl.sub.2 (2), KCl (2), CdCl.sub.2 (0.4), NiCl.sub.2 (0.3), TEA.Cl
(20), Hepes (10), glucose (10). Internal pipette solution consists
of (mM): CsF (65), CsCl (65), NaCl (10), CaCl.sub.2 (1.3),
MgCl.sub.2 (2), Hepes (10), EGTA (10), MgATP (1).
[0540] Compounds are dissolved as 20 mM stock solutions in DMSO and
then diluted to the final concentration in the external
solutions.
[0541] Voltage protocols and data analyses: A two-step protocol is
used to determine the voltage dependence of the block:
sodium current is activated by a 30 ms step pulse to 0 mV (test
pulse) from a 2000 ms preconditioning potential of -100 mV (resting
condition) or -70 mV (half maximal steady-state inactivated
condition), respectively.
[0542] Drug concentration-inhibition curves are obtained by
plotting tonic blocks in the resting and depolarized condition,
versus drug concentrations. Dose-response curves are fitted to the
tonic block data, according to the logistic equation:
y=A2+(A1-A2)/[1+(x/IC.sub.50)p]. A1 and A2 are fixed values of 0
and 1 corresponding to 0 and 100% current inhibition, x is the drug
concentration, IC.sub.50 is the drug concentration resulting in 50%
current inhibition and p is the corresponding slope factor.
[0543] Besides the voltage dependent block calculated as IC.sub.50s
from the resting and the depolarized membrane potential,
respectively, a better evaluation of the apparent affinity of drug
for the inactivated state is done by calculating the Ki according
to the equation 1/Kdep=h/Kr+(1-h)/Ki where Kr is the affinity of
drug for the resting/closed state; Kdep is the IC.sub.50 in the
depolarized condition, h and (1-h) are the fractions of channels
present at the rest and dep potentials, respectively (De Luca et
al. "Optimal requirements for high affinity and use-dependent block
of skeletal muscle sodium channel by N-benzyl analogs of
tocainidelike compounds". Mol Pharmacol 64:932-945.(2003)). In fact
although the IC.sub.50 value at resting (condition of maximal
availability current=Imax) can be considered as the affinity
constant for closed/resting (Kr) channels, the IC.sub.50 from
depolarized potential (the specific Vhalf was used as
preconditioning depolarized potential) is influenced by the
relative proportion of resting channels in equilibrium with
inactivated ones and by the ability of the drug to influence such
an equilibrium based on its affinity for inactivated state.
[0544] Ki represents a better estimation of the inactivated-state
block, cleaned from the closed/resting state block.
[0545] The results, obtained with compounds which are
representative of the entire class of compounds of the invention
are reported in Table 2.
TABLE-US-00002 TABLE 2 K.sub.i COMPOUND [.mu.M]
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl- 0.1
acetamide, hydrochloride (Example 1-1)
2-[2,2-Difluoro-2-(3-hexyloxyphenyl)-ethylamino]-N,N-dimethyl- 0.1
acetamide, hydrochloride (Example 1-6)
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-
0.1 N,N-dimethyl-acetamide, hydrochloride (Example 1-7)
2-[2,2-Difluoro-2-(3-benzyloxyphenyl)-ethylamino]-N,N- 0.5
dimethyl-acetamide, hydrochloride (Example 1-14)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(pyrrolidin-1- 0.2
yl)-ethanone, hydrochloride (Example 1-15)
2-{2,2-Difluoro-2-[3-(4,4,4-trifluorobutoxy)-phenyl]-ethylamino}-
0.2 1-(pyrrolidin-1-yl)-ethanone (Example 1-18)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-1-(piperidin-1- 0.7
yl)-ethanone, hydrochloride (Example 1-29)
2-[2,2-Difluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-diethyl- 0.3
acetamide, hydrochloride (Example 1-32)
2-{[2,2-Difluoro-2-(3-butoxyphenyl)-ethyl]-(2-methoxyethyl)- 1.7
amino}-N,N-dimethyl-acetamide, hydrochloride (Example 3-3)
2-[2-Fluoro-2-(3-butoxyphenyl)-ethylamino]-N,N-dimethyl- 1.1
acetamide, hydrochloride (Example 4-1)
[0546] Data expressed as K.sub.i values at .mu.M concentration
demonstrate that the compounds of the invention are potent as
inhibitors of sodium channels.
Example 9
Inhibition of Sodium Currents in Cortical Neurons
[0547] Cell Preparation and culturing: cortical neurons are
prepared from embryonic Wistar rats (E17-E19). Brains of E17/E19
rats are removed and placed in ice-cold Hank's solution (Hank's
solution (Invitrogen, CA--USA)+glucose 30%+Pen-Strep 100.times.
(Invitrogen, CA--USA) 100 U-100 .mu.g/ml and Hepes-NaOH 5 mM).
[0548] Cortex are isolated, cut in small parts and washed twice
with Hank's solution. The solution is removed except 1-2 ml and the
tissue is mechanically dissociated. After the mechanical
dissociation, 5 ml of complete DMEM (Dulbecco's modified Eagle
medium) (Invitrogen, CA--USA)+FBS (Invitrogen, CA--USA)
10%+Glutamine (Invitrogen, CA--USA) 2 mM+Pen-Strep 100 U-100
.mu.g/ml are added, and cell suspension is centrifuged for 5 min at
1000 rpm. Supernatant is removed and 5 ml of complete Neurobasal
(Invitrogen, CA--USA) medium is added+B27 supplement (code
17504044, Invitrogen, CA--USA) 2%+Glutamine 2 mM+Pen-Strep 100
U-100 .mu.g/ml).
[0549] Cells are counted and diluted in Neurobasal medium to a
concentration of 400000 cells per poly-D-lysine 5 .mu.g/ml treated
Petri dish.
[0550] Cortical neurons are used from day 6.sup.th till day
11.sup.th after plating, and once a week Neurobasal medium is
changed.
[0551] Whole Cell Patch Clamp Recordings: Experiments on cortical
neurons are carried out using standard whole cell patch clamp
methods (Hamill O. P., Marty A., Neher E., Sakmann B., Sigworth F.
J., Pflugers Arch. 391 (2): 85-100 (1981)). Membrane currents are
recorded and filtered at 5 kHz with an Axon Axopatch 200B amplifier
and data digitized with an Axon Digidata 1322A (Axon Instruments,
CA, USA). Protocol playing and data acquisition are controlled
online with Axon pClamp8 software. Measuring and reference
electrodes are AgCl--Ag electrodes. A Sutter Instrument (Sutter
Instrument, CA, USA) P-87 Puller is used for pulling patch clamp
pipettes with a resistance of 2-3 M.OMEGA. from Harward
borosilicate glass tubes. Cells are continuously superfused with
extracellular solutions, using a solution changer Biologic RSC-200
(Bio-Logic Sas, France).
[0552] Solutions: Sodium current recording control bath solution
contains (mM): NaCl (60), choline Cl (60), CaCl.sub.2 (1.3),
MgCl.sub.2 (2), KCl (2), CdCl.sub.2 (0.4), NiCl.sub.2 (0.3), TEACl
(20), Hepes (10), glucose (10). Internal pipette solution consists
of (mM): CsF (65), CsCl (65), NaCl (10), CaCl.sub.2 (1.3),
MgCl.sub.2 (2), Hepes (10), EGTA (10), MgATP (1).
[0553] Voltage protocols and data analyses: cells are clamped at
-90 mV, then a two step protocol is used to determine the voltage
dependence of the block. Sodium currents are activated by a 30 ms
step pulse to -10 mV (test pulse) from a 2000 ms preconditioning
potential of -110 mV (resting condition) and a potential of
.about.-50 mV (half maximal steady-state condition).
[0554] Drug concentration-inhibition curves are obtained by
plotting tonic blocks in the resting and depolarized condition,
versus drug concentrations. Dose-response curves are fitted to the
tonic block data, according to the logistic equation:
y=A2+(A1-A2)/[1+(x/IC.sub.50)p]. A1 and A2 are fixed values of 0
and 1 corresponding to 0 and 100% current inhibition, x is the drug
concentration, IC.sub.50 is the drug concentration resulting in 50%
current inhibition and p is the corresponding slope factor.
[0555] The compounds of the present invention inhibit sodium
currents of cortical neurons with pharmacologically significant
IC.sub.50 values.
Example 10
Inhibition of Cytochrome P4502D6 (CYP2D6)
[0556] The inhibition of Cytochrome P4502D6 (CYP2D6) is assessed by
performing in vitro inhibition studies using Supersomes, microsomes
derived from baculovirus infected insect cells; the baculoviruses
are engineered to express one or more drug metabolizing enzyme
cDNAs. Supersomes catalyze the same enzymatic reactions as human
liver microsome enzymes, but they contain much higher enzyme
activity than other microsome sources (Crespi C. L. and Penman B.
W., Advances Pharmacology, 43, 171-188 (1997); Crespi C. L. and
Miller V. P., Analytical Biochemistry, 248, 188-190 (1997)).
[0557] Kits with Supersomes are supplied by GENTEST (MA, USA).
Serial Dilution of Test Compound and Positive Control in a 96-Well
Plate
[0558] Test compound is dissolved in DMSO 500.times. the highest
final concentration desired in the IC.sub.50 assay. 30 ml of
deionized water is pre-warmed to 37.degree. C. and all kit
components are placed on ice. For each well of column 1, 149.4
.mu.L of NADPH-Cofactor Mix, (187.5 .mu.l of Cofactors, 150 .mu.l
of G6PDH, 100 .mu.l of Control Protein and 14.56 ml of 37.degree.
C. water) are added.
[0559] In each well from Column 2 to 12, 100 .mu.l of Cofactor/DMSO
mix (40 .mu.L DMSO in 9.96 ml of NADPH-Cofactor Mix) are added. To
each well of column 1, 0.6 .mu.l of test compound or positive
control are added. 50 .mu.l from each well of column 1 are serially
diluted to column 8. The extra 50 .mu.l from column 8 are
discarded. The plate is covered and pre-incubated at 37.degree. C.
for 10 minutes.
[0560] Preparation of enzyme/substrate mix: 7.92 ml of pre-warmed
deionized water, 75 .mu.l of enzyme, 3 .mu.l of 10 mM AMMC and 2 ml
of pre-warmed buffer are mixed.
Reaction Initiation and Termination
[0561] After the pre-incubation time (10'), 100 .mu.l of
enzyme/substrate mix to each well from column 1 to 10 are added.
The plate is incubated at 37.degree. C. for 30 minutes. After this
time, 75 .mu.l of Stop Reagent to each well are added. For blank
controls, 100 .mu.l of enzyme/substrate mix are added to columns 11
and 12.
Reading the Results
[0562] Plates are read at the Victor plate reader (Perkin Elmer,
MA--USA) at 390 nm excitation and 460 nm emission wavelengths.
[0563] The results, obtained with some compounds which are
representative of the entire class of compounds of the invention
are reported in Table 3, compared with the corresponding
de-fluorinated reference standards of the closest prior art.
TABLE-US-00003 TABLE 3 IC.sub.50 Corresponding IC.sub.50
Fluorinated derivative [.mu.M] de-fluorinated derivative [.mu.M]
2-[2,2-Difluoro-2- >40 2-[2-(3-Butoxyphenyl)- 5.8
(3-butoxyphenyl)-ethyl- ethylamino]-N,N-dimethyl-
amino]-N,N-dimethyl- acetamide, hydrochloride acetamide,
hydrochloride (Example 1-1) 2-[2,2-Difluoro-2- >40
2-[2-(3-Butoxyphenyl)- 3.0 (3-butoxyphenyl)-ethyl- ethylamino]-1-
amino]-1- (pyrrolidin-1-yl)- (pyrrolidin-1-yl)- ethanone,
hydrochloride ethanone, hydrochloride (Example 1-15) 2-[2-Fluoro-2-
>40 2-[2-(3-Butoxyphenyl)- 5.8 (3-butoxyphenyl)-ethyl-
ethylamino]-N,N-dimethyl- amino]-N,N-dimethyl- acetamide,
hydrochloride acetamide, hydrochloride (Example 4-1)
2-[2,2-Difluoro-2- 34.0 2-[2-(3-Butoxyphenyl)- 2.5
(3-butoxyphenyl)-ethyl- ethylamino]-N,N-diethyl-
amino]-N,N-diethyl- acetamide, hydrochloride acetamide,
hydrochloride (Example 1-32) 2-[2,2-Difluoro-2- 23.9
2-[2-(3-Butoxyphenyl)- 0.7 (3-butoxyphenyl)-ethyl-
ethylamino]-N,N-dipropyl- amino]-N,N-dipropyl- acetamide,
hydrochloride acetamide, hydrochloride (Example 1-3)
2-[2,2-Difluoro-2- 38.3 2-[2-(3-Butoxyphenyl)- 1.1
(3-butoxyphenyl)-ethyl- ethylamino]-1- amino]-1-(piperidin-1-yl)-
(piperidin-1-yl)- ethanone, hydrochloride ethanone, hydrochloride
(Example 1-29) 2-{[2,2-Difluoro-2- >40 2-{[2-(3-Butoxyphenyl)-
7.3 (3-butoxyphenyl)-ethyl]- ethyl]-(3-methoxypropyl)-
(3-methoxypropyl)- amino}-N,N-dimethyl- amino}-N,N-dimethyl-
acetamide, hydrochloride acetamide, hydrochloride (Example 3-2)
2-[2,2-Difluoro-2- >40 2-[2-(3-Butoxyphenyl)- 5.8
(3-butoxyphenyl)-ethyl- ethylamino]-N,N-dimethyl-
amino]-N,N-dimethyl- propanamide, hydrochloride propanamide,
hydrochloride (Example 2-1) 2-[2,2-Difluoro-2- >40
2-[2-(3-Pentyloxyphenyl)- 3.5 (3-pentyloxyphenyl)-ethyl-
ethylamino]-N,N-dimethyl- amino]-N,N-dimethyl- acetamide,
hydrochloride acetamide, hydrochloride (Example 1-2)
2-[2,2-Difluoro-2- 37.8 2-[2-(3-Hexyloxyphenyl)- 0.9
(3-hexyloxyphenyl)-ethyl- ethylamino]-N,N-dimethyl-
amino]-N,N-dimethyl- acetamide, hydrochloride acetamide,
hydrochloride (Example 1-6) 2-{2,2-Difluoro-2- >40
2-{2-[3-(4,4,4- 5.0 [3-(4,4,4-trifluoro- trifluorobutoxy)-phenyl]-
butoxy)-phenyl]-ethyl- ethylamino}-1-(pyrrolidin-
amino}-1-(pyrrolidin- 1-yl)-ethanone, hydrochloride 1-yl)-ethanone
(Example 1-18) 2-{2,2-Difluoro-2- >40 2-{2-[3-(4,4,4- 1.8
[3-(4,4,4-trifluoro- trifluorobutoxy)phenyl]-
butoxy)-phenyl]-ethyl- ethylamino}-1- amino}-1-(morpholin-4-yl)-
(morpholin-4-yl)- ethanone, hydrochloride ethanone, hydrochloride
(Example 1-21) 2-[2,2-Difluoro-2- >40 2-[2-(3-Benzyloxyphenyl)-
10.6 (3-benzyloxyphenyl)-ethyl- ethylamino]-N,N-dimethyl-
amino]-N,N-dimethyl- acetamide, hydrochloride acetamide,
hydrochloride (Example 1-14) 2-{[2,2-Difluoro-2- >40
2-{[2-(3-Butoxyphenyl)- 3.4 (3-butoxyphenyl)-ethyl]-
ethyl]-(2-methoxyethyl)- (2-methoxyethyl)]- amino}-N,N-dimethyl-
amino}-N,N-dimethyl- acetamide, hydrochloride acetamide,
hydrochloride (Example 3-3)
[0564] From the data presented in Table 3 it is apparent the
difluoro-substituted derivatives always display inhibitory activity
on CYP2D6 with IC.sub.50 values above 20 .mu.m and, in most cases,
near or above 40 .mu.M, whilst the corresponding unsubstituted
analogs from the prior art are endowed with inhibitory activities,
most often in the single digit micromolar range.
Example 11
Complete Freund's Adjuvant Model of Chronic Inflammatory Pain
[0565] Monoarthritis is induced in rats (200 g weight) by an
intra-plantar injection into the left hind paw of 100 .mu.l of
complete Freund's adjuvant (CFA) containing heat-killed and dried
Mycobacterium tubercolosis in a mixture of paraffin oil and an
emulsifying agent, mannite monooleate. The CFA injection produces
an area of localized edema and inflammation starting from few hours
after injection, with a progressive reduction in the mechanical
withdrawal threshold.
[0566] Each animal is allowed to develop the arthritis over a
period of 8-9 days before testing.
Mechanical Allodynia
[0567] Mechanical allodynia thresholds is determined according to
the method of Chaplan et al. (Chaplan S. R., Bach F. W., Pogrel J.
W., Chung J. M., Yaksh T. L. J. Neurosci. Methods 53: 55-63
(1994)). Rats are placed in individual plastic boxes of
24.times.10.times.15 cm on a mesh metal floor and allowed to
acclimate for about 30 minutes before testing. A series of
calibrated von Frey hairs (Stoelting, Wood Dale, Ill., USA) with
logarithmically incremental stiffness ranging from 2.83 to 5.88
expressed Log.sub.10 of [10.times. force in (mg)] are applied to
the paw with a modified up-down method (Dixon W. J. Am. Stat.
Assoc. 60: 967-978 (1965)). In the absence of a paw withdrawal
response to the initially selected hair, a thicker hair
corresponding to a stronger stimulus is presented until a sharp
withdrawal is recorded. The procedure is repeated twice. Each hair
is presented perpendicularly against the paw, with sufficient force
to cause slight bending, and held 2-3 s. The stimulation of the
same intensity is applied five/six times to the hind paw at
intervals of few seconds. The mechanical threshold is expressed as
Log.sub.10 of [10.times. force in (mg)] indicating the force of the
von Frey hair to which the animal react (paw withdrawn, licking or
shaking).
[0568] The mechanical allodynia thresholds are measured before
(pre-drug) and at 30, 60, 90, 120, 240 and 360 minutes after the
treatment. A 24 h threshold is also measured.
[0569] The compounds of the invention are administered in a range
of doses of 0.1-100 mg/kg.
Example 12
Bennett Model of Neuropathic Pain in Rats
[0570] Effects on neuropathic pain are tested in the chronic
constriction injury model in the rat (Bennett, G. J. and Xie, Y.
K., "A peripheral mononeuropathy in rat that produces disorders of
pain sensation like those seen in man". Pain, 33, 87-107 (1988)).
Under pentobarbital anesthesia (Nembutal, 50 mg/kg, i.p.),
unilateral multiple ligations are performed on male Sprague-Dawley
rats (140-160 g) at the right common sciatic nerve. The sciatic
nerve is exposed by blunt dissection at the level of mid-thigh and
four loose ligatures (5-0 chromic catgut) are placed around the
nerve taking care not to interrupt the epineural circulation. After
operation, animals are allowed to recover for one week. Animals
develop a cold allodynia which is stable for at least five weeks.
Cold allodynia is tested on a metal plate cooled by a water bath to
a constant temperature of 4.degree. C. The animals, randomly
assigned to groups of 10 for each test dose and vehicle, are
observed for periods of 2 minutes before and after application of
test compound and the number of brisk withdrawal reactions is
counted. Several time points after application are tested. Percent
maximal possible effect (% MPE) and standard error of the mean
(SEM) of each time point is determined with the pre-test value used
as 100% MPE. The area under the data (AUD) is calculated for the
observation period and expressed as percent inhibition of vehicle
control. Significance is calculated by paired t-test on the percent
AUD values.
Example 13
Maximal Electroshock Test (MES) in Mice
[0571] The maximal electroshock test (MES) is used commonly in the
screening of anti-epileptic drugs in rodent models.
[0572] Animals and Apparatus:
[0573] Male CD1 mice weighing 25 g are used. The procedure
described by White et al. (White H. S., Woodhead J. H., Franklin M.
R., Swinyard E. A., and Wolf H. H. Antiepileptic Drugs 4th ed:
99-110 (1995), Raven Press, Ltd., New York) is followed. An Ugo
Basile electroconvulsive generator Model ECT UNIT 7801 (Ugo Basile,
Italy) is used to deliver an electrical stimulus sufficient to
produce a hindlimb tonic extensor response in at least 97% of
control animals. The stimulus is delivered intra-aurally through
clip electrodes in mice (0.7 s of a 40 mA shock, with a pulse train
of 80 Hz having a pulse duration of 0.4 ms). The acute effect of
compounds administered intraperitoneally or orally 15-60 minutes
before MES induction are examined and compared with a vehicle
control group. Ten mice are studied per group. Complete suppression
of the hindlimb tonic extensor component of seizures is taken as
evidence of anticonvulsant activity.
[0574] The compounds of the invention are administered
intravenously (iv), orally (os) or intraperitoneally (ip) at the
doses of 0.1-100 mg/kg.
[0575] The results, obtained with a compound representative of the
entire chemical class of the invention, administered iv and/or po,
15 minutes before testing, and reported in Table 4, demonstrate
that these compounds are active as anticonvulsant drugs.
TABLE-US-00004 TABLE 4 50% COMPOUND Protection
2-[2,2-Difluoro-2-(3-butoxyphenyl)- (9.8 mg/kg, po)
ethylamino]-N,N-dimethyl- acetamide, hydrochloride (Example 1-1)
2-[2-Fluoro-2-(3-butoxyphenyl)- (4.0 mg/kg, iv)
ethylamino]-N,N-dimethyl- acetamide, hydrochloride (Example
4-1)
Example 14
Amphetamine and Chlordiazepoxide-Induced Hyperlocomotion in
Mice
[0576] In this model, mice are treated with a mixture of
d-amphetamine plus an anxiolytic dose of the benzodiazepine,
chlordiazepoxide (Rushton R., Steinberg H. "Combined effects of
chlordiazepoxide and d-amphetamine on activity of rats in an
unfamiliar environment". Nature 211:1312-3 (1966); Arban R., Maraia
G., Brackenborough K., Winyard L., Wilson A., Gerrard P., Large C.
"Evaluation of the effects of lamotrigine, valproate and
carbamazepine in a rodent model of mania". Behavioural Brain
Research, 158: 123-132 (2005)). The model has been claimed to mimic
some aspects of mania in bipolar disorder. Importantly, the
hyperactivity induced by the mixture of d-amphetamine and
chlordiazepoxide could be prevented by prior administration of the
established mood stabilizer, lithium, as well as other mood
stabilizers drugs (e.g. magnesium valproate and carbamazepine).
Therefore, this model has face and predictive validity as a model
of bipolar disorder and represents a valuable tool to determine, if
a test compound could be a potential mood stabilizer drug
candidate.
[0577] Amphetamine (AMP) (2.5 mg/kg) plus chlordiazepoxide
hydrochloride (CDZ) (3 mg/kg/ip) are administered to male Albino
Swiss mice (25-32 g) in a volume of 10 ml/kg. The locomotor
activity is recorded using Opto-M3 System (Columbus Instruments,
OH--USA) which is multi-channel activity monitor. Opto-M3 system
has 10 infrared emitters and respective amount of receivers (0.5''
beam spacing), attached to the PC computer and calculating both
ambulatory activity and total counts. Thus the system
differentiates forward locomotion (ambulation) from stereotyped
like movement (total counts). Mice are pretreated with the test
compound (5 mg/kg) and 10 min later, with AMP (2.5 mg/kg) or AMP
jointly with CDZ (3 mg/kg). After successive 30 min. the mice are
treated again with the same dose of the test compound and are
placed individually in the motor activity cages. The locomotor
activity (ambulation and total activity count) is evaluated for 30
min. Each group consists of 8-10 mice.
[0578] Statistical analysis: the data are evaluated by an analysis
of variance (ANOVA), followed, when appropriate, by individual
comparison with the control using Dunnett's test.
Amphetamine-chlordiazepoxide administration induces a significant
increase in locomotor activity.
Example 15
Model of Cognitive Impairment in Schizophrenia
[0579] Cognitive impairment is often associated with schizophrenia
and it has come to be recognized as a core element of the disorder,
bearing on patient's recovery and re-integration into society.
[0580] Particular interest has recently attracted a pharmacological
model of cognitive dysfunctions in schizophrenia, which is based on
the effects of glutamate NMDA receptor antagonists such as
phencyclidine (PCP) and ketamine (Javitt D C, Zukin S R. Am. J.
Psychiatry. 148:1301-1308. (1991)) which impair attention and
increase "impulsivity" and "compulsive" perseveration in mice
performing a complex task (Greco B, Invernizzi R W, Carli M.
Psychopharmacology (Berl) 179(1):68-76 (2005)).
Materials and Methods
[0581] Animals:
[0582] Male DBA/2N mice (Charles River, Italy) are used. The mice
weigh 25-30 g at the start of the experiments, and are housed under
temperature-controlled conditions (21.degree. C.) with a 12 hours
light 12 hours dark cycle (light on 7:00 am-7:00 pm). Food (Rieper,
Italy) is available ad libitum. The animals have two hours of
access to water at the end of each day's testing.
[0583] The Five-Choice Serial Reaction Time Task Apparatus:
[0584] The test apparatus consists of four
21.6.times.17.8.times.12.7 cm chambers (Med Associates Inc.
GA--USA), as previously described (Greco B, Invernizzi R W, Carli
M. Psychopharmacology (Berl) 179(1):68-76 (2005)). Stimuli and
recording of responses, are managed by a SmartCtrl.TM. Package 8
In/16 Out (Med Associates Inc. GA--USA) with additional interfacing
by MED-PC for Windows (Med Associates Inc. GA--USA). The running
program for the 5-Choice Serial Reaction Time (5-CSRT) task is
custom-written.
[0585] Behavioural Procedures: Habituation to Liquid Reinforcer and
Nose-Poking in the Holes.
[0586] Mice are handled for one week and their body weight
recorded. They are then water-deprived by allowing them 2-hours
access to water in the early evening until their body weight has
stabilised (8 days). Then, over the next two days the mice are
habituated in their home cages to the reinforcer (10% sucrose
solution) used afterwards in the operant procedures. On the
following two days mice are habituated to the operant boxes. During
this stage, 10% sucrose solution is available in a small bowl
placed below the receptacle hole of the box. First, mice have to
learn that every 5 seconds the liquid reward is available in a
small cup in the receptacle hole. During this period head entries
are recorded. During the next period, mice are trained to poke
their noses into the illuminated holes. Immediately after a poke in
the water receptacle a LED at the rear of one of the holes is
turned on. A nose-poke in the lighted hole extinguishes the light
stimulus and the liquid dipper provides a 0.01 mL liquid reward in
the receptacle hole. Any response in one of the other four holes
have no consequence and is not recorded. The light stimulus is
presented in all five holes in random order. A mouse is switched to
the 5-CSRT task after it has completed at least 50 rewarded
nose-poke trials in one 30-min session.
[0587] The Five-Choice Serial Reaction Time Task.
[0588] The start of the session is signalled by illumination of the
house-light and the delivery of a 0.01 mL liquid reward. Nose
poking in the receptacle hole begins the first trial. After a fixed
delay (the inter-trial interval, ITI), the LED at the rear of one
of the holes comes on for a short period. The LED stimulus is
presented the same number of times in each hole during a complete
session, with the order of presentation randomised by the computer.
While the light is on, and for a short period afterwards (the
limited hold), responses in the hole that is illuminated (correct
response) result in the liquid reward. Responses in the holes that
have not been illuminated (incorrect responses) or failure to
respond within the limited hold (omissions) cause the house-lights
to be turned off for a short period (time out). Responses in the
holes while the house-light is off restart the time out. After the
delivery of the liquid reward, or at the end of time out, the mouse
starts the next trial by poking its nose into the receptacle hole.
Responses made in the holes after a correct response (perseverative
responses), or after the end of time out before nose-poking into
the receptacle hole, result in a period of time out. Responses in
the holes during the ITI (anticipatory responses) also result in a
period of time out. After anticipatory responses a nose-poke into
the receptacle hole restart the current trial. Each daily session
consists of 100 trials or 30 min of testing, whichever is completed
sooner, after which all lights are turned off and further responses
have no effect. In the first session of the test schedule, the
stimulus and limited hold each last 1 minute and, depending on
individual performance, they are progressively reduced to 1 second.
The stimulus duration is reduced in the following sequence: 60, 30,
10, 5, 2.5, 2, 1.5 and 1 second (baseline). The ITI and time out
both lasts 2 seconds during the first session and the ITI is raised
to 5 seconds in subsequent sessions; time out is not changed.
Throughout the whole period of training and experiments each mouse
has one session per day on a 5-CSRT task.
[0589] Drugs and Treatment Schedules.
[0590] The test compound is dissolved in water and is administered
intraperitoneally (IP) at the dose of 10 mg/kg. Five minutes after
the treatment mice are injected with vehicle (saline) or PCP (1.5
mg/kg) and 10 minutes later they start the test session. In each
experiment the various combination of the test compound with
vehicle or PCP are administered according to a Latin-square design.
At least 48 hours are left between the drug testing days. During
these intervening days the mice are tested on the 5-CSRT task to
re-establish baseline performance and to check for any residual
effects of drugs.
[0591] Statistical Analysis:
[0592] The main dependent variables selected for analysis are: (a)
the percentage of correct responses (total correct responses/total
correct+total incorrect responses.times.100); (b) percentage of
omissions (total omissions/total correct responses+total incorrect
responses+total omissions.times.100); (c) the number of
anticipatory responses in the holes during the ITI; (d) the number
of perseverative responses in the holes after a correct response.
Correct responses and omissions, as percentages, are transformed
according to the formula 2 arcsin (SQRT (% X/100)), to normalize
the distributions in accordance with the ANOVA model.
[0593] The effects of the test compound (n=12) on PCP induced
deficits in the 5-CSRT task are analysed independently by a within
subjects 2.times.2 ANOVA with factors Drug (test compound) and PCP.
Subsequently the treatment group means are compared using a
post-hoc Tukey-Kramer test. Statistical software (SAS Institute
Inc., NC--USA) is run on Micro VAX 3500 computer (Digital,
MA--USA).
Example 16
Cocaine-Induced Behavioural Sensitization Test
[0594] Drug addiction is a pathological behaviour characterized by
compulsive drug seeking and intake. One animal model of these
behavioral changes is the long-lasting increase in locomotor
activity induced by repeated administration of psychostimulant
drugs in rodents (Robinson T. E. and Berridge K. C. Brain Res.
Brain Res. Rev. 18, 247-91 (1993)) known as drug-induced
behavioural sensitization. The effect of test compounds are
evaluated in a model of cocaine-induced behavioral sensitization in
rat.
[0595] Locomotor activity apparatus: Male Wistar rats weighing
200-250 g upon arrival are used. Locomotor activity is measured in
sixteen identical metal wire hanging cages each measuring 36 cm
(L).times.25 cm (W).times.20 cm (H). Each cage contains two sets of
infrared emitter-detector photocells positioned along the long axis
1 cm above the grid floor and 8 cm from the front and back of the
cage. Background noise is provided by a white noise generator.
Movement within the cages produces photocell interruptions, which
are automatically recorded by an IBM-compatible computer.
[0596] Sensitization procedure and treatment: Animals are
habituated to the locomotor activity chambers for 2-3 consecutive
days before the experiment. Rats receive 5 daily i.p. injections of
cocaine (15 mg/kg) or saline and either the test compound (0.1-100
mg/kg) or its vehicle and locomotor activity is recorded for 3 h.
Ten days after the last injection of cocaine or saline (day 15),
the animals are challenged with 15 mg/kg of cocaine in absence of
the test compound and locomotor activity is again monitored for 3
hours.
[0597] By the fifth day of treatment with cocaine, animals
pretreated i.p. with vehicle show an increased locomotor response
(20% higher then the first day, p<0.05). Ten days after the last
injection of cocaine or saline, the animals are challenged with 15
mg/kg of cocaine in absence of the test compound and locomotor
activity is again monitored for 3 h. The rats previously treated
with cocaine and that have not received the test compound are
expected to show an increased locomotor activity response to
cocaine (30% higher then first day, p<0.05). If the rats that
have been pretreated with the test compound during the 5
day-cocaine treatment do not show an increase in locomotor activity
the test compound is considered to have an effect in preventing
psychostimulant drugs addiction. (Koob G. F., Sanna P. P., Bloom F.
E. Neuron 21: 467-476 (1998); Robinson T. E., Berridge K. C. Brain
Res. Brain Res. Rev. 18: 247-291 (1993))
[0598] Statistical analysis: Data (total number of beam breaks in 3
hours) are analyzed using a two way ANOVA with repeated measures on
one factor including the four experimental groups (i.e.,
saline/vehicle, saline/test compound, cocaine/vehicle and
cocaine/test compound) and two time points (day 1 and day 5)
followed by a simple effects analysis. A second two way ANOVA with
repeated measures on one factor is used to compare day 1 and the
challenge day followed by a Newman-Keuls post hoc test.
Example 17
Acute Bladder Irritation by Acetic Acid in Rats
[0599] Experiments are performed using adult anesthetized female
Sprague Dawley rats (170-200 g). A catheter (PE-50) is inserted via
a midline abdominal incision into the bladder through the bladder
dome, and then intravescical pressure is measured to monitor
bladder activity during continuous infusion of 0.15% of acetic
acid. Continuous intravesical infusion of acetic acid irritates the
bladder and reduces the intercontraction intervals (ICI) in
anesthetized rats. ICIs, maximal contraction pressure, and pressure
thresholds inducing reflex bladder contraction are measured before
and after intravesical infusion of acetic acid in rats treated with
compounds of the invention.
Example 18
Intermediate Bladder Irritation by Cyclophosphamide (CYP) in
Rats
[0600] Experiments are performed using both adult awake and
anesthetized female Sprague Dawley rats (170-200 g). Chemical
cystitis is induced by CYP, which is metabolized to acrolein, an
irritant eliminated in the urine. CYP (150 mg/kg/i.p.) is
administered one day before the experiment. Pre-treatment with CYP
causes bladder irritation and very frequent voidings with an ICI of
about 150-200 seconds between voids.
[0601] Active compounds increase the ICI in both awake and
anesthetized rats used in this experimental model.
Example 19
Migraine Test in Rats
[0602] Animals and Surgery:
[0603] Male Wistar rats (250-350 g) are anesthetized with sodium
pentobarbital (50 mg/kg i.p.) dissolved in saline.
[0604] The trachea and left femoral artery are cannulated
for artificial ventilation (55 strokes/minute) and for measurement
of mean blood pressure (MBP) respectively. The femoral vein is
cannulated for the intravenous administration of test agents.
[0605] Body temperature is maintained at 37-38.degree. C. by
automatic control of a heating pad. Animals are placed in a
stereotaxic frame and a longitudinal incision is made in the scalp.
A burr hole is drilled in the skull and a stainless steel bipolar
electrode Plastics One MS 306 (Plastics One Inc. VA--USA) is
lowered into left ophthalmic branch of the trigeminal ganglion (3.8
mm dorsal to bregma, 2.5 mm lateral from the midline and 9.5 mm
below the dural surface) and secured with dental cement. Correct
placement of the electrode is confirmed by a brief electrical
stimulation, which cause movement of the jaw due to activation of
the trigeminal fiber. Following removal of the brain, the correct
position of the electrode into the fiber, is visually checked at
the end of each experiment.
[0606] A second hole is drilled ipsilateral of the electrode (1.5
mm rostral to bregma, and 1.5 mm lateral from the sagittal suture)
and a needle probe (tip diameter 0.8 mm) of a laser doppler
flowmeter is fixed pointing with its tip onto a branch of the
middle cerebral artery (MCA) and Cerebral Blood Flow (CBF) change
recorded on-line by the PeriFlux 4001 Laser Doppler system
(Perimed, Italy).
[0607] Artefacts of the laser Doppler reading during electrical
stimulation of the trigeminal ganglion due to muscular movements
are prevented by a bolus of iv injection of the neuromuscular
blocker pancuronium bromide (0.6 mg/kg iv).
[0608] Anesthesia and neuromuscular blockade are maintained all
over the experiment with an infusion of sodium pentobarbital and
pancuronium (12.5 mg/kg/h+2.4 mg/kg/h, respectively).
[0609] Experimental Protocol:
[0610] At the end of the surgery, a pause of thirty minutes is
taken in order to stabilize the measured parameters.
[0611] Rest CBF is increased by electrical stimulation with
rectangular pulse of 0.5 ms length, 1-10 Hz, 0.5-1 mA for periods
of 30 s. After two averaged pre-drug stimulations, vehicle or drugs
are administered.
[0612] Active compounds reduce the increase in blood flow induced
by trigeminal stimulation.
* * * * *