U.S. patent application number 13/568754 was filed with the patent office on 2012-11-29 for derivatives of pyrimido [6,1-a] isoquinolin-4-one.
This patent application is currently assigned to Verona Pharma plc. Invention is credited to David Jack, Alexander William Oxford.
Application Number | 20120302533 13/568754 |
Document ID | / |
Family ID | 26315369 |
Filed Date | 2012-11-29 |
United States Patent
Application |
20120302533 |
Kind Code |
A1 |
Oxford; Alexander William ;
et al. |
November 29, 2012 |
DERIVATIVES OF PYRIMIDO [6,1-A] ISOQUINOLIN-4-ONE
Abstract
The present invention relates to derivatives of
pyrimido[6,1-a]isoquinolin-4-one and their application as
inhibitors of phosphodiesterase (PDE) isoenzymes. More particularly
the invention relates to derivatives of
pyrimido[6,1-a]isoquinolin-4-one and their use in medicine for
example as bronchodilators with anti-inflammatory properties.
Inventors: |
Oxford; Alexander William;
(Hertfordshire, GB) ; Jack; David; (Hertfordshire,
GB) |
Assignee: |
Verona Pharma plc
London
GB
|
Family ID: |
26315369 |
Appl. No.: |
13/568754 |
Filed: |
August 7, 2012 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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12150232 |
Apr 24, 2008 |
8242127 |
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13568754 |
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10786400 |
Feb 24, 2004 |
7378424 |
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12150232 |
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09964260 |
Sep 26, 2001 |
6794391 |
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10786400 |
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PCT/GB00/01193 |
Mar 29, 2000 |
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09964260 |
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Current U.S.
Class: |
514/171 ;
514/267 |
Current CPC
Class: |
A61P 43/00 20180101;
A61P 11/08 20180101; C07D 471/04 20130101; A61P 29/00 20180101;
A61P 11/00 20180101; A61P 11/06 20180101 |
Class at
Publication: |
514/171 ;
514/267 |
International
Class: |
A61K 31/519 20060101
A61K031/519; A61P 11/08 20060101 A61P011/08; A61P 29/00 20060101
A61P029/00; A61K 31/56 20060101 A61K031/56 |
Foreign Application Data
Date |
Code |
Application Number |
Mar 31, 1999 |
GB |
GB 9907454.4 |
Apr 28, 1999 |
GB |
GB 9909802.2 |
Claims
1-50. (canceled)
51. A composition comprising a compound of formula I: ##STR00033##
wherein each of R.sup.1 and R.sup.2 independently represents a
C.sub.1-6 alkyl or C.sub.2-7 acyl group; R.sup.5 represents a
hydrogen atom or a C.sub.1-3 alkyl, C.sub.2-3 alkenyl or C.sub.2-3
alkynyl group; R.sup.6 represents a hydrogen atom or a C.sub.1-6
alkyl, C.sub.2-6 alkenyl, C.sub.2-6 alkynyl, amino, C.sub.1-6
alkylamino, di(C.sub.1-6)alkylamino or C.sub.2-7 acylamino group;
each of R.sup.7 and R.sup.8 independently represents a hydrogen or
halogen atom or a hydroxy, trifluoromethyl, C.sub.1-6 alkyl,
C.sub.2-6 alkenyl, C.sub.2-6 alkynyl, C.sub.2-7 acyl, C.sub.1-6
alkylthio, C.sub.1-6 alkoxy, C.sub.3-6 cycloalkyl; R.sup.9
represents a hydrogen or halogen atom or a hydroxy,
trifluoromethyl, C.sub.1-6 alkyl, C.sub.2-4 alkenyl, C.sub.2-6
alkynyl, C.sub.2-7 acyl, C.sub.1-6 alkylthio, C.sub.1-6 alkoxy or
C.sub.3-6 cycloalkyl group; X represents a group CR.sup.3R.sup.4,
wherein each of R.sup.3 and R.sup.4 independently represents a
hydrogen atom or a C.sub.1-3 alkyl group; each of R.sup.10 and
R.sup.11 independently represents a hydrogen atom, a C.sub.1-3
alkyl, C.sub.3-6 cycloalkyl or phenyl group; Y represents an oxygen
atom or a group CHNO.sub.2, NCN, NH or NNO.sub.2; n is an integer
from 2 to 4; or a salt thereof; and further comprising an active
agent selected from the group consisting of a
.beta..sub.2-adrenoceptor agonist and a glucocorticoid steroid, and
a veterinarily or pharmaceutically acceptable carrier or
diluent.
52. The composition of claim 51, wherein each of R.sup.1 and
R.sup.2 represents a C.sub.1-6 alkyl; R.sup.1 and R.sup.2 are the
same as each other; each of R.sup.3 and R.sup.4 represents a
hydrogen atom; R.sup.5 represents a hydrogen atom; R.sup.6
represents a hydrogen atom; each of R.sup.7 and R.sup.8 represents
a O.sub.1-6 alkyl; R.sup.7 and R.sup.8 are the same as each other;
R.sup.9 represents a halogen atom or a methyl or acetyl group; Y
represents an oxygen atom or a group CHNO.sub.2; and n is 2.
53. The composition of claim 52, wherein each of R.sup.1 and
R.sup.2 represents a O.sub.1-4 alkyl, group; and each of R.sup.7
and R.sup.8 represents a methyl, ethyl or isopropyl group.
54. The composition of claim 51, wherein the composition is a
pharmaceutical composition for human medicine.
55. The composition of claim 54, adapted for administration by
aerosol.
Description
[0001] The present invention relates to derivatives of
pyrimido[6,1-a]isoquinolin-4-one and their application as
inhibitors of phosphodiesterase (PDE) isoenzymes. More particularly
the invention relates to derivatives of
pyrimido[6,1-a]isoquinolin-4-one and their use in medicine for
example as bronchodilators with anti-inflammatory properties.
[0002] In all cells where cyclic AMP (cAMP) is present as a
secondary messenger, intracellular concentrations of cAMP are
regulated by the two processes involved in its formation and
degradation. Stimulation of membrane bound receptors on the
external surface of the cells (e.g. by .beta.-adrenoceptor
agonists) results in activation of adenylyl cyclase to generate
cAMP from ATP. Phosphodiesterases present in the cell serve to
reduce the concentration of cAMP by hydrolysing it to adenosine
monophosphate (AMP).
[0003] In a disease such as asthma, the principal cells involved in
the associated bronchoconstriction and inflammatory processes are
subject to inhibitory control by cAMP. Inhibitors of type III
phosphodiesterase raise intracellular levels of cAMP, leading, to
relaxation of bronchial smooth muscle, whereas inhibitors of type
IV phosphodiesterase inhibit the release of damaging mediators from
pro-inflammatory cells. Thus, in principle, a combined PDE III/IV
inhibitor should have the desirable effects of a
.beta.-adrenoceptor agonist plus an inhaled anti-inflammatory
steroid which are currently the mainstay of treatment in severe
asthma. Moreover, a combined PDE III/IV inhibitor given by
inhalation should achieve beneficial effects similar to a
.beta.-agonist plus inhaled steroid and should be an unusually
effective treatment of asthma and other respiratory disorders
without the undesirable glucocorticoid effects of the steroid such
as osteoporosis and the stunting of growth.
[0004] The potential adverse effects of a PDE III/IV inhibitor
(e.g. nausea and vomiting, gastric acid secretion, cardiovascular
effects such as increased cardiac contractility, vasodilation and
potential arrhythmogenic activity) should be avoidable with a
compound that is delivered directly to the lungs by inhalation. It
is desirable that the substance is long acting, non irritant and
has a taste which is not so unpleasant as to have any adverse
effect on patient compliance.
[0005] An example of a pyrimido[6,1-a]isoquinolin-4-one derivative
with PDE III/IV inhibitory activity and known to possess
antihypertensive vasodilator activity is
trequinsin(9,10-dimethoxy-3-methyl-2-mesitylimino-2,3,6,7-tetrahydro-4H-p-
yrimido[6,1-a]isoquinolin-4-one), which is described by De Souza et
al., J. Med. Chem. 27 1470-1480 (1984) and in GB-A-1597717.
[0006] As described by De Souza et al. and in GB-A-1597717,
trequinsin has valuable pharmacological properties, and can be
administered to human subjects suffering from, for example,
respiratory disorders. However, it is unsuitable for administration
by inhalation because of its bitter taste and in vitro data
indicate its persistence of action is less than desirable.
[0007] It has now been found that it is possible to design certain
pyrimido[6,1-a]isoquinolin-4-one derivatives which are PDE
inhibitors, which have a longer duration of action relative to
trequinsin and other useful properties, such as improved taste.
[0008] According to a first aspect of the present invention there
is provided a compound of general formula I:
##STR00001##
wherein each of R.sup.1 and R.sup.2 independently represents a
C.sub.1-6 alkyl or C.sub.2-7 acyl group; R.sup.5 represents a
hydrogen atom or a C.sub.1-3 alkyl, C.sub.2-3 alkenyl or C.sub.2-3
alkynyl group; R.sup.6 represents a hydrogen atom or a C.sub.1-6
alkyl, C.sub.2-6 alkenyl, C.sub.2-6 alkynyl, amino, C.sub.1-6
alkylamino, di(C.sub.1-6)alkylamino or C.sub.2-7 acylamino group;
each of R.sup.7 and R.sup.8 independently represents a hydrogen or
halogen atom or a hydroxy, trifluoromethyl, C.sub.1-6 alkyl,
C.sub.2-6 alkenyl, C.sub.2-6 alkynyl, C.sub.2-7 acyl, C.sub.1-6
alkylthio, C.sub.1-6 alkoxy, C.sub.3-6 cycloalkyl; and R.sup.9
represents a hydrogen or halogen atom or a hydroxy,
trifluoromethyl, C.sub.1-6 alkyl, C.sub.2-6 alkenyl, C.sub.7-6
alkynyl, C.sub.2-7 acyl, C.sub.1-6 alkylthio, C.sub.1-6 alkoxy or
C.sub.3-6 cycloalkyl group; X represents OCH.sub.2 or a group
CR.sup.3R.sup.4, wherein each of R.sup.3 and R.sup.4 independently
represents a hydrogen atom or a C.sub.1-3 alkyl group; each of
R.sup.10 and R.sup.11 independently represents a hydrogen atom, a
C.sub.1-3 alkyl, C.sub.3-6 cycloalkyl or phenyl group; Y represents
an oxygen atom or a group CHNO.sub.2, NCN, NH or NNO.sub.2; n is an
integer from 2 to 4; or a salt thereof.
[0009] As used herein the term "halogen" or its abbreviation "halo"
means fluoro, chloro, bromo or iodo.
[0010] As used herein the term "C.sub.1-6 alkyl" refers to straight
chain or branched chain alkyl groups having from one to six carbon
atoms. Illustrative of such alkyl groups are methyl, ethyl, propyl,
isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl,
neopentyl and hexyl. C.sub.1-4 alkyl groups are preferred.
[0011] As used herein the term "C.sub.2-3 alkenyl" refers to
straight chain or branched chain hydrocarbon groups having from two
to three carbon atoms and having in addition one double bond, of
either E or Z stereochemistry where applicable. This term would
include for example, vinyl and 1-propenyl.
[0012] As used herein the term "C.sub.2-3 alkynyl" refers to
straight chain hydrocarbon groups having from two to three carbon
atoms and having in addition one triple bond. This term would
include for example, ethynyl and 1-propynyl.
[0013] As used herein the term "C.sub.2-6 alkenyl" refers to
straight chain or branched chain hydrocarbon groups having from two
to six carbon atoms and having in addition one double bond, of
either E or Z stereochemistry where applicable. This term would
include for example, vinyl, 1-propenyl, 1- and 2-butenyl and
2-methyl-2-propenyl. C.sub.2-3 alkenyl groups are preferred.
[0014] As used herein the term "C.sub.2-6 alkynyl" refers to
straight chain or branched chain hydrocarbon groups having from two
to six carbon atoms and having in addition one triple bond. This
term would include for example, ethynyl, 1-propynyl, 1- and
2-butynyl, 2-methyl-2-propynyl, 2-pentanyl, 3-pentanyl, 4-pentanyl,
2-hexanyl, 3-hexanyl, 4-hexanyl and 5-hexanyl. C.sub.2-3 alkynyl
groups are preferred.
[0015] As used herein the term "C.sub.1-6 alkoxy" refers to
straight chain or branched chain alkoxy groups having from one to
six carbon atoms. Illustrative of such alkoxy groups are methoxy,
ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, sec-butoxy,
tert-butoxy, pentoxy, neopentoxy and hexoxy. C.sub.1-4 alkoxy
groups are preferred.
[0016] As used herein the term "C.sub.2-7 acyl" refers to straight
chain or branched chain acyl groups having from two to seven carbon
atoms. Illustrative of such acyl groups are acetyl, propionyl (or
propiono or propanoyl), isopropionyl (or isopropiono or
isopropanoyl), butyryl (or butanoyl), isobutyryl (or isobutanoyl),
pentanoyl (or valeryl), hexanoyl (or capronyl) and heptanoyl.
[0017] As used herein the term "C.sub.2-7 acyloxy" refers to
straight chain or branched chain acyloxy groups having from two to
seven carbon atoms. Illustrative of such acyloxy groups are
acetyloxy, propionyl (or propiono or propanoyl)oxy, isopropionyl
(or isopropiono or isopropanoyl)oxy, butyryl (or butanoyl)oxy,
isobutyryl (or isobutanoyl)oxy, pentanoyl (or valeryl)oxy, hexanoyl
(or capronyl)oxy and heptanoyloxy. C.sub.2-4 acyloxy groups are
preferred.
[0018] As used herein the term "C.sub.3-6 cycloalkyl" refers to an
alicyclic group having from three to six carbon atoms. Illustrative
of such cycloalkyl groups are cyclopropyl, cyclobutyl, cyclopentyl
and cyclohexyl. Cyclopentyl and cyclohexyl groups are
preferred.
[0019] As used herein the term "C.sub.1-6 alkylthio" refers to
straight chain or branched chain alkylthio groups having from one
to six carbon atoms. Illustrative of such alkylthio groups are
methylthio, ethylthio, propylthio, isopropylthio, butylthio,
isobutylthio, sec-butylthio, tert-butylthio, pentylthio,
neopentylthio and hexylthio. C.sub.1-4 alkylthio groups are
preferred.
[0020] As used herein the term "C.sub.1-6 alkylamino" refers to
straight chain or branched chain alkylamino groups having from one
to six carbon atoms. Illustrative of such alkylamino groups are
methylamino, ethylamino, propylamino, isopropylamino, butylamino,
isobutylamino, sec-butylamino, tert-butylamino, pentylamino,
neopentylamino and hexylamino. C.sub.1-4 alkylamino groups are
preferred.
[0021] As used herein, the term "di(C.sub.1-6)alkylamino" refers to
straight chain or branched chain di-alkylamino groups having from
one to six carbon atoms in each of the alkyl groups. Illustrative
of such dialkylamino groups are di-methylamino, di-ethylamino,
di-propylamino, di-isopropylamino, di-butylamino, di-isobutylamino,
di-sec-butylamino, di-tert-butylamino, di-pentylamino,
di-neopentylamino and di-hexylamino. Di(C.sub.1-4)alkylamino groups
are preferred.
[0022] As used herein, the term "C.sub.2-7 acylamino" refers to
straight chain or branched chain acylamino groups having from two
to seven carbon atoms. Illustrative of such acylamino groups are
acetylamino, propionyl (or propiono or propanoyl)amino,
isopropionyl (or isopropiono or isopropanoyl)amino, butyryl (or
butanoyl)amino, isobutyryl (or isobutanoyl)amino, pentanoyl (or
valeryl)amino, hexanoyl (or capronyl)amino and heptanoylamino.
C.sub.2-4 acylamino groups are preferred.
[0023] Where there is a substituent which renders a compound basic,
for example when R.sup.6 is an amino, alkylamino or dialkylamino
group, addition of an acid results in a salt. The acid may be any
suitable acid, and can be organic or inorganic.
[0024] Preferred compounds of general formula I include those in
which, independently or in any compatible combination:
each of R.sup.1 and R.sup.2 represents a C.sub.1-6 alkyl,
preferably a C.sub.1-4 alkyl, group; R.sup.1 and R.sup.2 are the
same as each other; each of R.sup.3 and R.sup.4 represents a
hydrogen atom; R.sup.5 represents a hydrogen atom; R.sup.6
represents a hydrogen atom; each of R.sup.7 and R.sup.8 represents
a C.sub.1-6 alkyl, preferably methyl, ethyl or isopropyl, group;
R.sup.7 and R.sup.8 are the same as each other; R.sup.9 represents
a hydrogen atom, a halogen atom or a methyl or acetyl group; Y
represents an oxygen atom or a group CHNO.sub.2; and n is 2.
[0025] Exemplary compounds include: [0026] 1.
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(N-carbamoyl-2-aminoethyl-
)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one; [0027] 2.
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N--(N'-isopropylcarbamoy-
l)-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one;
[0028] 3.
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N-[1-N'-methyl-
-2-nitroethenamine)]-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]-i-
soquinolin-4-one; [0029] 4.
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N-[1-(N'-isopropyl-2-nit-
roethenamine)]-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]-isoquin-
olin-4-one; [0030] 5.
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N-[1-(N',N'-dimethyl-2-n-
itroethenamine)]-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]-isoqu-
inolin-4-one; [0031] 6.
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N--(N'-phenylcarbamoyl)--
2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-2-one;
[0032] 7.
9,10-Dimethoxy-3-[2-guanidinoethyl]-2-(2,4,6-trimethylphenylimi-
no)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one; [0033]
8.
9,10-Dimethoxy-3-[N--(N'-nitro)-2-guanidinoethyl]-2-(2,4,6-trimethylpheny-
limino)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one;
[0034] 9.
3-[N--(N'-Cyclohexylcarbamoyl)-2-aminoethyl]-9,10-dimethoxy-2-(2,4,6-trim-
ethyl-phenylimino-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one;
[0035] 10.
3-(N-Carbamoyl-2-aminoethyl)-9,10-dimethoxy-2-(2-methylphenylimino)-3,4,6-
,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one; [0036] 11
3-(N-Carbamoyl-2-aminoethyl)-2-(2,6-diisopropylphenylimino)-9,10-dimethox-
y-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one; [0037]
12.
3-(N-Carbamoyl-4-aminobutyl)-9,10-dimethoxy-2-(2,4,6-trimethylphenylimino-
)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one; [0038]
13.
3-[N--(N'-Cyano-N''-methyl)-2-guanidinoethyl]-9,10-dimethoxy-2-(2,4,6-tri-
methyl-phenylimino)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one-
.
[0039] The compound:
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(N-carbamoyl
aminoethyl)-3,4,6,7 tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
is particularly preferred.
[0040] Compounds of general formula I may be prepared by any
suitable method known in the art and/or by the following process,
which itself forms part of the invention.
[0041] According to a second aspect of the invention, there is
provided a process for preparing a compound of general formula I as
defined above, the process comprising:
(a) derivatizing a compound of general formula II:
##STR00002##
wherein R.sup.1, R.sup.2, R.sup.5, R.sup.6, R.sup.7, R.sup.8,
R.sup.9, X and n are as defined for general formula I, with one or
more compounds capable of reacting at the primary amine group of
the aminoalkyl moiety (--(CH.sub.2).sub.n--NH.sub.2), to form a
compound of general formula I; or (b) when X in general formula I
represents a group CR.sup.3R.sup.4, wherein R.sup.3 represents a
hydrogen atom, R.sup.4 represents a hydrogen atom or a C.sub.1-3
alkyl group, and R.sup.5 represents a hydrogen atom or a C.sub.1-3
alkyl group, hydrogenating a compound of general formula III:
##STR00003##
wherein R.sup.1, R.sup.2, R.sup.6, R.sup.7, R.sup.8, R.sup.9,
R.sup.10, R.sup.11, Y and n are as defined for general formula I;
and (c) optionally converting a compound of general formula I so
formed into another compound of general formula I.
[0042] The reaction conditions of step (a) are generally such as to
favour the reaction, which may be a nucleophilic displacement or
addition and is carried out in a solvent which is suitable for the
particular reaction.
[0043] Compounds chosen for reacting with a compound of general
formula II are capable of reacting at the primary amine group of
the alkylamino moiety in the compound of general formula II, to
form a compound of general formula I. For example:
when Y represents an oxygen atom and each of R.sup.10 and R.sup.11
represents a hydrogen atom, a compound of general formula II may be
derivatised with sodium cyanate; when Y represents an oxygen atom,
R.sup.10 represents a hydrogen atom and R.sup.11 represents a
C.sub.1-3 alkyl, C.sub.3-6 cycloalkyl or phenyl group, a compound
of general formula II may be derivatised with an isocyanate of the
general formula R.sup.11NCO; when Y represents CHNO.sub.2, R.sup.10
represents a hydrogen atom and R.sup.11 represents a C.sub.1-3
alkyl or C.sub.3-6 cycloalkyl group, a compound of general formula
II may be derivatised with an N--C.sub.1-3 alkyl- or N--C.sub.3-6
cycloalkyl-1-(methylthio)-2-nitroethenamine of the general formula
CH.sub.3SC(.dbd.CHNO.sub.2)NR.sup.10R.sup.11; when Y represents
CHNO.sub.2, a compound of general formula II may be reacted first
with 1,1-bis(methylthio)-2-nitroethylene and the resulting compound
may then be reacted with an amine of the general formula
R.sup.10R.sup.11NH, wherein R.sup.10 and R.sup.11 are as defined
for general formula I; when Y represents NH, a compound of general
formula II may be derivatised with a compound of general formula
CH.sub.3SC(.dbd.NH)NR.sup.10R.sup.11 or a salt thereof, wherein
R.sup.10 and R.sup.11 are as defined for general formula I; and
when Y represents NCN, a compound of general formula II may be
derivatised with a compound of general formula
CH.sub.3SC(.dbd.NCN)NR.sup.10R.sup.11 or a salt thereof, wherein
R.sup.10 and R.sup.11 are as defined for general formula I.
[0044] In specific cases:
for
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(N-carbamoyl-2-aminoe-
thyl)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one,
sodium cyanate may be chosen; for
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N--(N'-isopropylcarbamoy-
l)-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one,
isopropylisocyanate may be chosen; for
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N-[1-(N'-methyl-2-nitroe-
thenamine)]-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]-isoquinoli-
n-4-one, N-methyl-1-(methylthio)-2-nitroethenamine may be chosen;
for
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N-[1-(N'-isopropyl-2-nit-
roethenamine)]-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]-isoquin-
olin-4-one, 1,1-bis(methylthio)-2-nitroethylene and isopropylamine
may be chosen; for
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N-[1-(N',N'-dimethyl-2-n-
itroethenamine)]-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]-isoqu-
inolin-4-one, 1,1-bis(methylthio)-2-nitroethylene and dimethylamine
may be chosen; and for
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N--(N'-phenylcarbamoyl)--
2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-2-one,
phenylisocyanate may be chosen.
[0045] Compounds of general formula II may be prepared by reacting
a compound of general formula IV:
##STR00004##
wherein R.sup.1, R.sup.2, R.sup.5, R.sup.6, R.sup.7, R.sup.8,
R.sup.9 and X are as defined for general formula I, with a compound
of general formula V:
LG-(CH.sub.2).sub.n--N--PG V
wherein n is as defined for general formula I, LG represents a
leaving group, and PG represents a protecting group; and then
removing the protecting group.
[0046] The reaction between a compound of general formula IV and a
compound of general formula V is generally carried out in suitable
conditions for the reaction, which is a nucleophilic substitution.
A base such as K.sub.2CO.sub.3 may be used in the presence of NaI
and the reaction is performed in a suitable solvent such as
2-butanone.
[0047] The leaving group LG in general formula V may be any
suitable leaving group, but is preferably a halogen atom, such as
bromine. The protecting group PG in general formula V may be any
suitable protecting group, such as a phthaloyl group. If the
reaction between a compound of general formula IV and V is carried
out in a base such as K.sub.2CO.sub.3, the protecting group should
be base-stable. A suitable compound of general formula V is
N-(2-bromoethyl)phthalimide:
##STR00005##
[0048] The protecting group may then be removed by standard
deprotection procedures. For example, hydrazine hydrate may be
used. The reaction conditions are generally to favour the reaction,
for example in a suitable solvent such as ethanol and/or chloroform
at room temperature.
[0049] Compounds of general formula IV may be prepared by reacting
a compound of general formula VI:
##STR00006##
wherein R.sup.1, R.sup.2, R.sup.5, R.sup.6 and X are as defined for
general formula I and LG represents a leaving group; with a
compound of general formula VII:
##STR00007##
wherein R.sup.7, R.sup.8 and R.sup.9 are as defined for general
formula I.
[0050] Compounds of general formula VII are substituted anilines
which are either known in the art and available from commercial
sources or may readily be prepared by methods known per se.
[0051] The leaving group LG in compounds of general formula VI may
be chlorine, a thioalkyl group, preferably thiomethyl, or an
alkylsulphonyl group, preferably methylsulphonyl. Preferably it is
chlorine.
[0052] The reaction conditions are generally such as to favour the
reaction, which is a nucleophilic displacement which is preferably
carried out in a suitable solvent such as dimethylformamide or
isopropanol in the presence of a base such as potassium carbonate.
Suitable reaction conditions may be found in GB-A-1597717 and
EP-A-0124893, which disclose the preparation of related
compounds.
[0053] The reaction is generally applicable for producing compounds
of general formula I where R.sup.6 represents a hydrogen atom or a
C.sub.1-6 alkyl, C.sub.2-6 alkenyl, C.sub.2-6 alkynyl, amino,
C.sub.1-6 alkylamino or C.sub.2-7 acylamino group and R.sup.1 to
R.sup.5 and R.sup.7 to R.sup.9, X, Y and n have the meanings given
above.
[0054] Compounds of general formula VI where LG represents a
chlorine atom may be prepared by reacting a compound of general
formula VIII or a compound of general formula IX with phosphorous
oxychloride, or by heating a compound of general formula VIII with
phosphorous pentachloride:
##STR00008##
wherein R.sup.1, R.sup.2, R.sup.5 and R.sup.6 and X are as defined
for general formula I. Compounds of general formula VI where LG
represents a thioalkyl group may be prepared from compounds of
formula VIII by heating with phosphorous pentasulphide in a solvent
such as dioxan or pyridine to give initially the intermediate thio
derivative of VIII which, on treatment with an alkylating agent
such as an alkyl iodide eg. methyl iodide, in a suitable solvent
such as tetrahydrofuran or ethyl acetate, gives the thioalkyl
compound. Oxidation of the thioalkyl compound with, for example,
3-chloroperbenzoic acid in a solvent such as methylene chloride,
gives the alkylsulphone derivative.
[0055] Compounds of general formula VIII may be prepared by
reacting a compound of general formula IX, wherein R.sup.1,
R.sup.2, R.sup.5 and R.sup.6 are as defined for general formula I,
with a cyclodehydrating agent such as phosphorous oxychloride,
under less vigorous condition, ie lower temperatures, than those
required to give compounds of the general formula VI where LG
represents a chlorine atom. An alternative method has been
described in NL-A-6,401,827 (Hoffmann-La Roche) which involves
reacting the carbamoylmethylene-tetrahydroisoquinoline, general
formula XI (wherein R.sup.1, R.sup.2, R.sup.5 and X have the
meanings given above) with diethyl carbonate in ethanolic sodium
ethoxide:
##STR00009##
[0056] Compounds of general formula IX may be prepared by reacting
a compound of general formula XII
##STR00010##
wherein R.sup.1, R.sup.2, R.sup.3 and X are as defined for general
formula I, with R.sup.6CH(CO.sub.2Et).sub.2, wherein R.sup.6 is as
defined for general formula I, and a strong base such as sodium
ethoxide in a hot ethanolic solution. Alternatively, the
corresponding dimethyl ester can be employed in the presence of hot
methanolic sodium methoxide.
[0057] Compounds of general formula XII may be prepared by reacting
a compound of general formula XIII:
##STR00011##
wherein R.sup.1, R.sup.2, R.sup.5 and X are as defined for general
formula I, with urea by heating at 160.degree. C. Alternatively,
compounds of general formula XIII may be reacted with potassium
cyanate in the presence of acetic acid in a suitable solvent such
as ethanol.
[0058] Compounds of general formula XIII are either known in the
art or may readily be prepared by methods known per se. For
example, the preparation of 1-(3,4-dimethoxyphenethyl)barbituric
acid has been described by B. Lal et al. in J. Med. Chem. 27
1470-1480 (1984).
[0059] Turning to step (b), the reaction conditions of step (b) are
generally to favour the hydrogenation reaction, and the reaction is
generally carried out in a suitable solvent such as an alcohol, eg
ethanol, with a noble metal catalyst such as palladium, platinum,
rhodium or nickel, at room temperature. The catalyst may be
supported, for example on charcoal or alumina.
[0060] Compounds of general formula III may be prepared from a
compound of general formula XIV:
##STR00012##
wherein R.sup.1, R.sup.2 and R.sup.6 are as defined for general
formula I, and R.sup.4 and R.sup.5 independently represent a
hydrogen atom or a C.sub.1-3 alkyl group. The reactions are
conducted as described above for converting a compound of general
formula VIII to a compound of general formula II through compounds
of general formula VI and general formula IV, and the preferred
reaction conditions correspond accordingly.
[0061] Compounds of general formula XIV may be prepared from
compounds of general formula VIII (wherein X represents a CH.sub.2
group and R.sup.5 represents a hydrogen atom or a C.sub.1-3 alkyl
group) by heating with a noble metal catalyst such as palladium,
platinum, rhodium or nickel at a temperature of 300 to 350.degree.
C. The catalyst may be supported on charcoal or alumina and the
reaction carried out in an inert solvent such as an aromatic
hydrocarbon, eg p-cymene.
[0062] In optional step (c), a compound of general formula I may be
converted into another compound of general formula I. For example,
compounds of general formula I where R.sup.6 represents NH.sub.2
may be converted into compounds of general formula I where R.sup.6
represents a C.sub.1-6 alkylamino group by standard chemistry, such
as by alkylation of a protected derivative such as an acyl or a
p-toluenesulphonyl derivative followed by removal of the protecting
group, such as by acid hydrolysis. Compounds of general formula I
where R.sup.6 represents a di(C.sub.1-6)alkylamino group may be
prepared by direct alkylation of the alkylamino derivative.
Compounds of general formula I wherein R.sup.5, R.sup.6, R.sup.7,
R.sup.8 and/or R.sup.9 represent a C.sub.2-3 alkenyl, C.sub.2-6
alkenyl, C.sub.2-3 alkynyl or C.sub.2-6 alkynyl group may be
hydrogenated to give the corresponding compound with saturated
bonds. The reaction conditions for the hydrogenation are as
outlined above for step (b).
[0063] According to a third aspect, the present invention provides
a composition comprising a compound of general formula I and a
veterinarily or pharmaceutically acceptable carrier or diluent.
Preferably the composition is a pharmaceutical composition for
human medicine.
[0064] Compounds of the present invention are PDE inhibitors and
thus possess valuable pharmacological properties, such as
bronchodilator activity as demonstrated by the inhibition of
field-stimulated contraction of guinea-pig isolated trachea, and
anti-inflammatory activity as illustrated in studies on human
mononuclear cells stimulated by PHA (phytohaemagglutinin). In vitro
and in vivo data indicate the compounds have a long duration of
action, as demonstrated by their persistent protective effects
against histamine induced bronchospasm in the guinea-pig when
inhaled directly into the lungs as a dry powder. The invention
therefore also relates to acute, chronic or prophylactic treatment
of patients suffering from respiratory disorders including, in
particular, asthma, allergic asthma, hay fever, allergic rhinitis,
bronchitis, chronic obstructive pulmonary disease (COPD), adult
respiratory distress syndrome (ARDS), and cystic fibrosis. They may
also be used topically in skin disorders such as atopic dermatitis
and psoriasis, or in ocular inflammation or any other disease
including cerebral ischaemia or auto-immune diseases in which
increasing intracellular concentrations of cAMP is considered
beneficial.
[0065] One or more compounds as set out in the first aspect of the
invention may be present in association with one or more non-toxic
pharmaceutically and/or veterinarily acceptable carriers and/or
diluents and/or adjuvants and/or propellants and, if desired, other
active ingredients. Suitable carriers or diluents are known in the
art (eg Handbook of Pharmaceutical Excipients (1994) 2.sup.nd
Edition, Eds. A. Wade/P J Weller, The Pharmaceutical Press,
American Pharmaceutical Association).
[0066] Preferably, the compounds and the compositions of the
present invention are administered by inhalation, for example by
aerosols or sprays which can disperse the pharmacological active
ingredient in the form of a powder or in the form of a solution or
suspension. Pharmaceutical compositions with powder-dispersing
properties usually contain, in addition to the active ingredient, a
liquid propellant with a boiling point below room temperature and,
if desired, adjuncts, such as liquid or solid non-ionic or anionic
surfactants and/or wetting agent to form a stable dispersion.
Pharmaceutical compositions in which the pharmacological active
ingredient is in solution contain, in addition to this, a suitable
propellant, and furthermore, if necessary, an additional solvent
and/or a stabiliser. Instead of the propellant, compressed air can
also be use, it being possible for this to be produced as required
by means of a suitable compression and expansion device.
Pharmaceutical compositions may also be delivered by breath
activated inhalation devices. Dry powder compositions are preferred
for administration by inhalation.
[0067] According to a fourth aspect, the present invention provides
a compound of general formula I or a composition containing a
compound of general formula I for use in medicine.
[0068] Compounds of the present invention are useful as inhibitors
of phosphodiesterase isoenzymes. The compounds or compositions of
the present invention may be used to prevent or treat any disease
in which the compounds or compositions are useful, but particularly
a disease in which raising the intracellular concentration of cAMP
is desirable. Examples of diseases against which compounds are
useful include respiratory disorders including, in particular,
asthma, bronchitis, chronic obstructive pulmonary disease (COPD),
adult respiratory distress syndrome (ARDS), allergic asthma, hay
fever, allergic rhinitis, and cystic fibrosis. They may also be
used topically in skin disorders such as atopic dermatitis or
psoriasis, ocular inflammation, or any other disease including
cerebral ischaemia or auto-immune diseases in which increasing
intracellular concentrations of cAMP is considered beneficial.
[0069] This aspect of the invention is particularly relevant to the
treatment of humans, but is also applicable to general veterinary
industry, in particular domestic animals such as dogs and cats and
farm animals such as horses, pigs, cattle, sheep, etc.
[0070] Dosage levels of the order of about 0.02 mg to about 200 mg,
to be taken up to three times daily, are useful in the treatment of
the above-mentioned conditions. More particularly, a dosage range
of about 0.2 mg to about 20 mg, taken up to three times daily, is
effective. The particular dosage regime will however ultimately be
determined by the attending physician and will take into
consideration such factors as the medication being used, age,
weight, severity of symptoms and/or severity of treatment being or
to be applied, method of administration of the medication, adverse
reactions and/or other contraindications.
[0071] The medication according to this aspect of the invention may
be given to a patient together with other active agents, which may
for example be a different compound of the present invention, or
other compounds. Examples include .beta..sub.2-adrenoceptor
agonists, topical glucocorticoid steroids, xanthine derivatives,
antihistamine compounds, leukotriene antagonists, inhibitors of
leukotriene synthesis and/or combinations thereof.
[0072] According to a fifth aspect, the present invention provides
the use of a compound of general formula I in the manufacture of an
inhibitor of a type III/IV phosphodiesterase isoenzyme. The
invention encompasses the use of a compound of general formula I in
the manufacture of a bronchodilator and/or an anti-asthmatic
medication and/or a medicament for the prevention or treatment of
chronic obstructive pulmonary disease (COPD).
[0073] The invention also relates to a method for the treatment or
prevention of a disease in a mammal where a phosphodiesterase
isoenzyme inhibitor and/or a bronchodilator would be expected to be
of benefit, which method comprises administering to said mammal an
amount of an effective, non-toxic amount of a compound of general
formula I. The invention encompasses a method of treating or
preventing asthma and/or chronic obstructive pulmonary disease
(COPD) in a mammal.
[0074] Preferred features of each aspect of the invention apply to
each other aspect of the invention, mutatis mutandis.
[0075] FIG. 1, referred to in Preparations 1 to 4 below, shows the
route by which the compounds in Preparations 1 to 4 were
synthesised;
[0076] FIG. 2, referred to in Example A below, is a graph showing
the effect of DMSO on cholinergic contractile response in
superfused guinea pig trachea, wherein "n" is the number of
experiments;
[0077] FIG. 3, referred to in Example A below, is a graph showing
the effect of 10 .mu.M of the compound of Example 1 of the present
invention on contraction of guinea pig trachea to electrical field
stimulation over time (n=3), wherein the arrow denotes commencement
of washout period;
[0078] FIG. 4, referred to in Example A below, is a graph showing
the effect of 10 .mu.M of the compound of Example 9 on contraction
of guinea pig trachea to electrical field stimulation over time
(n=3);
[0079] FIG. 5, referred to in Example A below, is a graph showing
the effect of 10 .mu.M of the compound of Example 10 on contraction
of guinea pig trachea to electrical field stimulation over time
(n=3);
[0080] FIG. 6, referred to in Example A below, is a graph showing
the effect of 10 .mu.M of the compound of Example 11 on contraction
of guinea pig trachea to electrical field stimulation over time
(n=3);
[0081] FIG. 7, referred to in Example A below, is a graph showing
the effect of 10 .mu.M of the compound of Example 13 on contraction
of guinea pig trachea to electrical field stimulation over time
(n=3);
[0082] FIG. 8, referred to in Example A below, is a graph showing
the effect of 10 .mu.M of the compound of Example 8 on contraction
of guinea pig trachea to electrical field stimulation over time
(n=3);
[0083] FIG. 9 referred to in Example B below, is a graph showing
the effect of the compound of Example 1 of the present invention
against proliferation of human mononuclear cells stimulated by PHA,
wherein each point represents the mean of six experiments, and
vertical lines represent standard error of the mean.
PREPARATION 1
Synthesis of
2-Chloro-6,7-dihydro-9,10-Dimethoxy-4H-pyrimido-[6,1-a]isoquinolin-4-one
(shown as (1) in FIG. 1)
##STR00013##
[0085] A mixture of 1-(3,4-dimethoxyphenyl)barbituric acid (70 g,
0.24 mol), prepared according to the method described in B. Lal et
al. J. Med. Chem. 27 1470-1480 (1984), and phosphorus oxychloride
(300 ml, 3.22 mol) was refluxed for 2.5 h. The excess phosphorous
oxychloride was removed by distillation (20 mmHg) on warming. After
cooling the residue was slurried in dioxan (100 ml) and cautiously
added to a vigorously stirred ice/water solution (11). Chloroform
(11) was added and the resulting mixture was basified with 30%
sodium hydroxide solution. The organic layer was separated and the
aqueous phase further extracted with chloroform (2.times.750 ml).
The combined organic extracts were washed with water (1.51), dried
over magnesium sulphate and concentrated in vacuo to leave a gummy
material (90 g). This was stirred in methanol for a few minutes,
filtered and washed with methanol (200 ml), diethyl ether
(2.times.200 ml) and dried in vacuo at 40.degree. C. to yield the
title compound as a yellow/orange solid. 47 g, 62%
[0086] (300 MHz, CDCl.sub.3) 2.96 (2H, t, C.sub.(7)H.sub.2); 3.96
(6H, s, 2.times.OCH.sub.3; 4.20 (2H, t, C.sub.(6)H.sub.2); 6.61
(1H, s, C.sub.(1)H); 6.76 (1H, s, Ar--H); 7.10 (1H, s, Ar--H).
PREPARATION 2
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3,4,6,7-tetrahydro-2H-pyrimi-
do[6,1-a]isoquinolin-4-one (shown as (2) in FIG. 1)
[0087]
2-Chloro-9,10-dimethoxy-6,7-dihydro-4H-pyrimido[6,1-a]isoquinolin-4-
-one, prepared according to Preparation 1, (38.5 g, 0.13 mol) and
2,4,6-trimethylaniline (52.7 g, 0.39 mol) in propan-2-ol (3 l) was
stirred and heated at reflux, under nitrogen, for 24 h. After
cooling to room temperature, the solution was evaporated in vacuo
and the residue was purified by column chromatography on silica
gel, eluting with CH.sub.2Cl.sub.2/MeOH, initially 98:2, changing
to 96:4 once the product began to elute from the column. The title
compound was obtained with a slight impurity, (just above the
product on tlc). Yield 34.6 g, 67%.
PREPARATION 3
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(2-N-phthalimidoethyl)-3,4-
,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (shown as (3)
in FIG. 1)
[0088] A mixture of
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3,4,6,7-tetrahydro-2H-pyrim-
ido[6,1-a]isoquinolin-4-one (which was prepared according to
Preparation 2) (60.0 g, 0.153 mol), potassium carbonate (191 g,
1.38 mol), sodium iodide (137 g, 0.92 mol) and
N-(2-bromoethyl)phthalimide (234 g, 0.92 mol) in 2-butanone (1500
ml) was stirred and heated at reflux, under nitrogen, for 4 days.
After cooling to room temperature the mixture was filtered and the
filtrate was evaporated in vacuo. The residue was treated with
methanol (1000 ml) and the solid filtered off, washed with methanol
and recrystallised from ethyl acetate to obtain the title compound
as a pale yellow solid in yield 40.0 g, 46%. Evaporation of the
mother liquor and column chromatography of the residue on silica
gel (CH.sub.2Cl.sub.7/MeOH 95:5) provided further product 11.7 g,
13.5%.
PREPARATION 4
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(2-aminoethyl)-3,4,6,7-tet-
rahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (shown as (4) in FIG.
1)
[0089] A mixture of
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(2-N-phthalimidoethyl)-3,-
4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (22.0 g, 0.039
mol), prepared according to Preparation 3, and hydrazine hydrate
(11.3 g, 0.195 mol) in chloroform (300 ml) and ethanol (460 ml) was
stirred at room temperature, under nitrogen, for 18 h. Further
hydrazine hydrate (2.9 g, 0.05 mol) was added and the mixture was
stirred a further 4 h. After cooling in ice/water, the solid was
removed by filtration and the filtrate evaporated in vacuo. The
residue was dissolved in dichloromethane and the insoluble material
was removed by filtration. The filtrate was dried (MgSO.sub.4) and
evaporated in vacuo to afford the title compound as a yellow foam
in yield 16.2 g, 96%.
EXAMPLE 1
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(N-carbamoyl-2-aminoethyl)-
-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
##STR00014##
[0091] Sodium cyanate (6.0 g, 0.092 mol) in water (100 ml) was
added dropwise to a stirred solution of
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(2-aminoethyl)-3,4,6,7-te-
trahydro-2H-pyrimido[6,1-a]isoquinolin-4-one, prepared according to
Preparation 4 above (20.0 g, 0.046 mol) in water (600 ml) and 1N
HCl (92 ml) at 80.degree. C. After stirring for 2 h at 80.degree.
C. the mixture was cooled in an ice-bath and basified with 2N NaOH.
The mixture was extracted with dichloromethane (3.times.200 ml) and
the combined extract was dried (MgSO.sub.4) and evaporated in
vacuo. The resulting yellow foam was purified by column
chromatography on silica gel eluting with CH.sub.7Cl.sub.2/MeOH
(97:3) and triturated with ether to obtain the title compound as a
yellow solid, 11.9 g, 54%.
TABLE-US-00001 M.p.: 234-236.degree. C. m/z:
C.sub.26H.sub.31N.sub.5O.sub.4 requires M = 477 found (M + 1) = 478
HPLC: Area (%) 99.50 Column ODS (150 .times. 4.6 mm) MP pH3
KH.sub.2PO.sub.4/CH.sub.3CN (60/40) FR (ml/min) 1.0 RT (min) 9.25
Detection 250 nm
[0092] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta. 1.92 (1H, br s,
NH), 2.06 (6H, s, 2.times.CH.sub.3), 2.29 (3H, s, CH.sub.3), 2.92
(2H, t, CH.sub.2), 3.53 (2H, m, CH.sub.2), 3.77 (3H, s, OCH.sub.3),
3.91 (3H, s, OCH.sub.3), 4.05 (2H, t, CH.sub.2), 4.40 (2H, t,
CH.sub.2), 5.35 (2H, br s, NH.sub.2), 5.45 (1H, s, C.dbd.CH), 6.68
(1H, s, ArH), 6.70 (1H, s, ArH), 6.89 (2H, s, 2.times.ArH).
EXAMPLE 2
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N--(N'-isopropylcarbamoyl-
)-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
##STR00015##
[0094] Isopropylisocyanate (0.15 g, 1.77 mmol) was added dropwise
to a stirred solution of
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(2-aminoethyl)-3,4,6,7-te-
trahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (prepared according to
Preparation 4 above) (0.7 g, 1.61 mmol) in toluene (6 ml) at room
temperature, under nitrogen. After 2 h the solution was evaporated
in vacuo and the residue was purified by column chromatography on
silica gel (CH.sub.2Cl.sub.2/MeOH, 97:3) and triturated with ether
to obtain an off-white solid, 0.42 g, 50%.
TABLE-US-00002 M.p.: 181-182.degree. C. m/z:
C.sub.29H.sub.37N.sub.5O.sub.4 requires M = 519 found (M + 1) = 520
HPLC: Area (%) 94.99 Column ODS (150 .times. 4.6 mm) MP pH3
KH.sub.2PO.sub.4/CH.sub.3CN (40/60) FR (ml/min) 1.0 RT (min) 7.985
Detection 250 nm
[0095] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta. 0.89 (6H, d,
2.times.CH.sub.3), 2.05 (6H, s, 2.times.CH.sub.3), 2.29 (3H, s,
CH.sub.3), 1.94 (1H, br s, NH), 2.90 (2H, t, CH.sub.2), 3.49 (2H,
m, CH.sub.1), 3.77 (3H, s, OCH.sub.3), 3.91 (3H, s, OCH.sub.3),
4.05 (2H, t, CH.sub.2), 4.37 (2H, t, CH.sub.2), 5.02 (1H, br s,
NH), 5.46 (1H, s, C.dbd.CH), 6.67 (1H, s, ArH), 6.69 (1H, s, ArH),
6.87 (2H, s, 2.times.ArH).
EXAMPLE 3
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N-[1-(N'-methyl-2-nitroet-
henamine)]-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]-isoquinolin-
-4-one
##STR00016##
[0097] A mixture of
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(2-aminoethyl)-3,4,6,7-te-
trahydro-2H-pyrimido[6,1-a]isoquinolin-4-one, prepared according to
Preparation 4 above (0.8 g, 1.84 mmol) and
N-methyl-1-(methylthio)-2-nitroethenamine (0.30 g, 2.03 mmol) in
toluene (20 ml) was stirred and heated at reflux, under nitrogen,
for 2 h. After cooling to room temperature the solvent was
evaporated in vacuo and the residue was purified by column
chromatography on silica gel (CH.sub.2Cl.sub.2/MeOH, 97:3). The
title compound was obtained as a yellow foam in yield 0.61 g, 62%,
which on triturating with ether yielded a yellow solid 0.40 g,
41%.
TABLE-US-00003 M.p.: 126-130.degree. C. m/z:
C.sub.28H.sub.34N.sub.6O.sub.5 requires M = 534 found (M + 1) = 535
HPLC: Area (%) 98.98 Column ODS (150 .times. 4.6 mm) MP pH4
KH.sub.2PO.sub.4/CH.sub.3CN (45/55) FR (ml/min) 1.0 RT (min) 6.635
Detection 250 nm
[0098] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta. 2.07 (6H, s,
2.times.CH.sub.3), 2.29 (3H, s, CH.sub.3), 2.45 (3H, d,
NHCH.sub.3), 2.92 (2H, t, CH.sub.2), 3.65 (2H, m, CH.sub.2), 3.77
(3H, s, OCH.sub.3), 3.90 (3H, s, OCH.sub.3), 4.08 (2H, t,
CH.sub.2), 4.32 (2H, m, CH.sub.2), 5.46 (1H, s, .dbd.CH), 6.51 (1H,
s, CHNO.sub.2), 6.70 (2H, s, 2.times.ArH), 6.90 (2H, s,
2.times.ArH), 8.78 (1H, m, NH), 10.35 (1H, m, NH).
EXAMPLE 4
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N-[1-(N'-isopropyl-2-nitr-
oethenamine)]-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]-isoquino-
lin-4-one
[0099] 1,1-Bis(methylthio)-2-nitroethylene (5.3 g, 32.2 mmol) and
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(2-aminoethyl)-3,4,6,7-te-
trahydro-2H-pyrimido[6,1-a]isoquinolin-4-one, prepared according to
Preparation 4 above (1.4 g, 3.22 mmol) in toluene (20 ml) was
stirred and heated at reflux, under nitrogen, for 2 h. After
cooling to room temperature the solvent was evaporated in vacuo and
the residue was purified by column chromatography on silica gel
(CH.sub.2Cl.sub.2/MeOH, 99:1). This yielded intermediate compound
(shown as compound A below) as an oil which became a light beige
solid upon trituration with ether. Yield 0.95 g, 53%.
[0100] Isopropylamine (5 ml) was added to a stirred solution of (A)
(0.7 g, 1.27 mmol) in dichloromethane (10 ml) and heated at reflux,
under nitrogen, for 20 h. After cooling, the solution was
evaporated in vacuo and the residue was purified by column
chromatography on silica gel (CH.sub.2Cl.sub.2/MeOH, 98:2). The
title compound (shown as compound B below) was obtained as a foam
in yield 0.64 g, 89%. This became a pale yellow solid (0.38 g) upon
trituration with ether.
##STR00017##
TABLE-US-00004 M.p.: 144-146.degree. C. m/z:
C.sub.30H.sub.38N.sub.6O.sub.5 requires M = 562 found (M + 1) = 563
HPLC: Area (%) 97.57 Column ODS (150 .times. 4.6 mm) MP pH4
KH.sub.2PO.sub.4/CH.sub.3CN (40/60) FR (ml/min) 1.0 RT (min) 9.028
Detection 250 nm
[0101] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta. 0.87 (6H, d,
CH(CH.sub.3).sub.2), 2.05 (6H, s, 2.times.CH.sub.3), 2.29 (3H, s,
CH.sub.3), 2.93 (2H, m, CH.sub.2), 3.48 (1H, m CH(CH.sub.3).sub.2),
3.68 (2H, m, CH.sub.2), 3.78 (3H, s, OCH.sub.3), 3.91 (3H, s,
OCH.sub.3), 4.09 (2H, t, CH.sub.2), 4.34 (2H, m, CH.sub.2), 5.48
(1H, s, C.dbd.CH), 6.68 (1H, s, CHNO.sub.2), 6.69 (2H, s,
2.times.ArH), 6.90 (2H, s, 2.times.ArH), 7.04 (1H, d, NH), 10.75
(1H, m, NH).
EXAMPLE 5
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N-[1-(N',N'-dimethyl-2-ni-
troethenamine)]-2-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]-isoqui-
nolin-4-one
##STR00018##
[0103] A solution of compound A shown in Example 4 above (1.0 g,
1.81 mmol) and dimethylamine (33% in EtOH, 5.0 ml, 28 mmol) in
dichloromethane (10 ml) was stirred at 35.degree. C., under
nitrogen, for 18 h. The solution was then evaporated in vacuo and
the residue was purified by column chromatography on silica gel
(CH.sub.7Cl.sub.2/MeOH, 97:3) to obtain the title compound as a
yellow foam in yield 0.73 g, 73%. This became a pale yellow solid
(0.60 g) upon trituration with ether.
TABLE-US-00005 M.p.: 187-189.degree. C. m/z:
C.sub.29H.sub.36N.sub.6O.sub.5 requires M = 548 found (M + 1) = 549
HPLC: Area (%) 97.89 Column ODS (150 .times. 4.6 mm) MP pH4
KH.sub.2PO.sub.4/CH.sub.3CN (45/55) FR (ml/min) 1.0 RT (min) 6.768
Detection 250 nm
[0104] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta. 2.02 (6H, s,
2.times.CH.sub.3), 2.28 (3H, s, CH.sub.3), 2.95 (2H, m, CH.sub.2),
2.95 (6H, s, N(CH.sub.3).sub.2), 3.78 (3H, s, OCH.sub.3), 3.81 (2H,
t, CH.sub.2), 3.90 (3H, s, OCH.sub.3), 4.05 (2H, t, CH.sub.2), 4.55
(2H, t, CH.sub.2), 5.43 (1H, s, C.dbd.CH), 6.47 (1H, s, ArH), 6.67
(1H, s, CHNO.sub.2), 6.70 (1H, s, ArH), 6.89 (2H, s, 2.times.ArH),
9.35 (1H, m, NH).
EXAMPLE 6
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-[N--(N'-phenylcarbamoyl)-2-
-aminoethyl]-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-2-one
##STR00019##
[0106] Phenylisocyanate (0.16 g, 1.38 mmol) was added dropwise to a
stirred solution of
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(2-aminoethyl)-3,4,6,7-te-
trahydro-2H-pyrimido[6,1-a]isoquinolin-4-one, prepared according to
Preparation 4 above (0.6 g, 1.38 mmol) in toluene (5 ml) at room
temperature, under nitrogen. After 1 h the solvent was evaporated
in vacuo and the residue was purified by column chromatography on
silica gel (CH.sub.2Cl.sub.2/MeOH, 95:5). After trituration with
ether the title compound was obtained as a pale yellow solid 0.61
g, 80%.
TABLE-US-00006 M.p.: 116-118.degree. C. m/z:
C.sub.32H.sub.33N.sub.5O.sub.4 requires M = 553 found (M + 1) = 554
HPLC: Area (%) 98.80 Column ODS (150 .times. 4.6 mm) MP 0.02M
KH.sub.2PO.sub.4/CH.sub.3CN (42/58) FR (ml/min) 0.8 RT (min) 10.622
Detection 254 nm
[0107] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta. 2.05 (6H, s,
2.times.CH.sub.3), 2.30 (3H, s, CH.sub.3), 2.92 (2H, t, CH.sub.2),
3.67 (2H, m, CH.sub.2), 3.78 (3H, s, OCH.sub.3), 3.91 (3H, s,
OCH.sub.3), 4.06 (2H, t, CH.sub.2), 4.47 (2H, t, CH.sub.2), 5.51
(1H, s, C.dbd.CH), 5.60 (1H, s br, NH), 6.69 (1H, s, ArH), 6.72
(1H, s, ArH), 6.89 (2H, s, 2.times.ArH), 6.90-7.23 (5H, m,
5.times.ArH), 7.62 (1H, br s, NH).
EXAMPLE 7
9,10-Dimethoxy-3-[2-guanidinoethyl]-2-(2,4,6-trimethylphenylimino)-3,4,6,7-
-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
##STR00020##
[0108] 1,3-Di-(tert-butoxycarbonyl)thiourea (1)
[0109] Sodium hydride (60% in oil, 4.7 g, 0.117 mol) was washed
with petroleum ether to remove the oil, then added in portions to a
stirred solution of thiourea (2.0 g, 0.026 mol) in tetrahydrofuran
(400 ml) at 0.degree. C., under nitrogen. The mixture was stirred
for 5 minutes at 0.degree. C. then warmed to room temperature for
10 minutes. After re-cooling to 0.degree. C., di-tert-butyl
dicarbonate (16.1 g, 0.0585 mol) in tetrahydrofuran (100 ml) was
added dropwise and the mixture was stirred for 30 minutes at
0.degree. C. After a further 2 h at room temperature the reaction
was quenched by dropwise addition of saturated sodium bicarbonate
(40 ml) and poured into water (11). The solution was extracted with
ethyl acetate (3.times.200 ml) and the combined extract washed with
brine, dried (MgSO.sub.4) and evaporated in vacuo. The residual
solid was triturated with petroleum ether, removed by filtration
and dried in vacuo. The title compound (4.3 g, 60%) was obtained as
an off-white solid. M.p. 124-127.degree. C.
3-[N--(N',N''-Di-tert-butoxycarbonyl)-2-guanidinoethyl]-9,10-dimethoxy-2-(-
2,4,6-trimethylphenylimino)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinol-
in-4-one (3)
[0110] 1-Methyl-2-chloropyridinium iodide (0.77 g, 3.03 mmol) in
N,N-dimethylformamide (4 ml) was added dropwise to a stirred
mixture of N-di-(tert-butoxycarbonyl)thiourea (0.84 g, 3.03 mmol),
3-(2-aminoethyl)-9,10-dimethoxy-2-(2,4,6-trimethylphenylimino)-3,4,6,7-te-
trahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (2) (1.1 g, 2.53 mmol)
and triethylamine (0.56 g, 5.57 mmol) in N,N-dimethylformamide (8
ml). After 18 h the reaction was quenched by addition of water (40
ml) and extracted with ethyl acetate (3.times.25 ml). The combined
extract was washed with brine, dried (MgSO.sub.4) and evaporated in
vacuo. The residual oil was purified by column chromatography on
silica gel (petroleum ether/ethyl acetate, 2:1) to obtain the title
compound (1.05 g, 61%) as a yellow foam.
[0111] M/z [ES]+677.
9,10-Dimethoxy-3-[2-guanidinoethyl]-2-(2,4,6-trimethylphenylimino)-3,4,6,7-
-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
[0112] Trifluoroacetic acid (0.35 g, 3.1 mmol was added to a
stirred solution of 3 (0.95 g, 1.4 mmol) in dichloromethane (5 ml)
at room temperature. After 3 h further trifluoroacetic acid (0.35
g) was added and the mixture was stirred for an additional 16 h.
The solvent was then evaporated in vacuo and the residue was
treated with dichloromethane (20 ml) and basified to pH 10 with
saturated sodium bicarbonate. The organic phase was separated,
dried (MgSO.sub.4) and evaporated in vacuo. The residual oil was
purified by column chromatography on silica gel
(CH.sub.2Cl.sub.2/MeOH, 98:2.fwdarw.90:10) to obtain the title
compound, after trituration with diethyl ether, as an off-white
solid, 0.27 g, 40%.
[0113] M.p.: 226-228.degree. C.
[0114] M/z: C.sub.26H.sub.32N.sub.6O.sub.3 requires M=476. found
m/z [ES]+=477
[0115] HPLC:
[0116] Area (%) 98.73
[0117] Column ODS LUNA 3uC18(2) (100.times.4.6 mm)
[0118] MP 0.1% CF.sub.3CO.sub.2H/CH.sub.3CN (gradient 90%
aq.fwdarw.25% aq over 25 min)
[0119] RT (min) 11.413
[0120] FR (ml/min) 0.8
[0121] Detection 250 nm
[0122] .sup.1H NMR (250 MHz, CDCl.sub.3, 70.degree. C.): .delta.
2.03 (6H, s, 2.times.CH.sub.3), 2.26 (3H, s, CH.sub.3), 2.95 (2H,
t, CH.sub.3), 3.57 (2H, m, CH.sub.2), 3.67 (3H, s, OCH.sub.3), 3.85
(3H, s, OCH.sub.3), 3.98 (2H, t, CH.sub.2), 4.33 (2H, t, CH.sub.2),
5.40 (1H, s, C.dbd.CH), 6.73 (1H, s, ArH), 6.91 (2H, s,
2.times.ArH), 6.98 (1H, s, ArH), 7.25 (2H, br s, NH.sub.2), 7.73
(1H, m, NH).
[0123] The .sup.1H NMR was run at 70.degree. C. to obtain better
resolution because some of the signals were poorly resolved at room
temperature.
##STR00021##
EXAMPLE 8
9,10-Dimethoxy-3-[N--(N'-nitro)-2-guanidinoethyl]-2-(2,4,6-trimethylphenyl-
imino)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
##STR00022##
[0124] 2-Methyl-1-nitro-2-isothiourea (1)
[0125] S-Methylisothiouronium sulphate (3.0 g, 10.8 mmol) was added
in portions to a stirred mixture of fuming nitric acid (3 ml) and
concentrated sulphuric acid (9 ml) at -10 to +5.degree. C. After
stirring for a further 30 min at 5.degree. C. the solution was
poured onto ice (120 g) with stirring. The white solid was removed
by filtration, washed with water and dried in vacuo to obtain
2-methyl-1-nitro-2-isothiourea (2.0 g, 69%).
[0126]
9,10-Dimethoxy-3-[N--(N'-nitro)-2-guanidinoethyl]-2-(2,4,6-trimethy-
lphenylimino)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
[0127] 2-Methyl-1-nitro-2-isothiourea (0.405 g, 3.0 mmol) was added
to a stirred solution of
3-(2-aminoethyl)-9,10-dimethoxy-2-(2,4,6-trimethylphenylimino)-3,4,6,7-te-
trahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (1.29 g, 3.0 mmol) in
ethanol (12 ml) and heated to 70.degree. C. for 30 min. The solvent
was then evaporated in vacuo and the residue was purified by column
chromatography on silica gel [CH.sub.2Cl.sub.2/MeOH, 97:3)] to
obtain the title compound as a pale yellow solid (0.76 g, 49%).
TABLE-US-00007 M.p.: 253-256.degree. C. M/z:
C.sub.26H.sub.31N.sub.7O.sub.5 requires M = 521, found m/z [ES]+ =
522 HPLC: Area (%) 99.44 Column ODS LUNA 3uC18(2) (100 .times. 4.6
mm) MP 0.1% CF.sub.3CO.sub.2H/CH.sub.3CN (gradient 90% aq .fwdarw.
25% aq over 25 min) RT (min) 16.842 FR (ml/min) 1.0 Detection 250
nm
[0128] .sup.1H NMR (250 MHz, d.sub.6-DMSO, 70.degree. C.): .delta.
2.02 (6H, s, 2.times.CH.sub.3), 2.25 (3H, s, CH.sub.3), 2.94 (2H,
t, CH.sub.2), 3.63 (2H, m, CH.sub.2), 3.66 (3H, s, OCH.sub.3), 3.84
(3H, s, OCH.sub.3), 3.96 (2H, t, CH.sub.2), 4.37 (2H, t, CH.sub.2),
5.38 (1H, s, C.dbd.CH), 6.72 (1H, s, ArH), 6.88 (2H, s,
2.times.ArH), 6.96 (1H, s, ArH), 7.89 (1H, br s, NH).
[0129] The .sup.1H NMR was run at 70.degree. C. to obtain better
resolution because some of the signals were poorly resolved at room
temperature.
##STR00023##
EXAMPLE 9
3-[N--(N'-Cyclohexylcarbamoyl)-2-aminoethyl]-9,10-dimethoxy-2-(2,4,6-trime-
thyl-phenylimino-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
##STR00024##
[0131] Cyclohexylisocyanate (0.38 g, 3.04 mmol) in toluene (2 ml)
was added dropwise to a stirred solution of
3-(2-aminoethyl)-9,10-dimethoxy-2-(2,4,6-trimethylphenylimino)-3,4,6,7-te-
trahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (1.2 g, 2.76 mmol) in
toluene (8 ml) at room temperature, under nitrogen. After stirring
for 16 h the solution was evaporated in vacuo and the residue was
purified by column chromatography on silica gel
[dichloromethane/methanol (97:3)]. The product was triturated with
ether to obtain the title compound (0.61 g, 40%) as a pale yellow
solid.
TABLE-US-00008 M.p.: 120-122.degree. C. M/z:
C.sub.32H.sub.41N.sub.5O.sub.4 requires M = 559, found m/z [ES+] =
560 HPLC: Area (%) 98.59 Column ODS LUNA 3uC18(2) MP 0.1M
NH.sub.4OAc/CH.sub.3CN (40/60) RT (min) 9.145 FR (ml/min) 0.7
Detection 250 nm
[0132] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta. 0.7-1.8 (11H, m,
cyclohexyl), 2.05 (6H, s, 2.times.CH.sub.3), 2.27 (3H, s,
CH.sub.3), 2.93 (2H, t, CH.sub.2), 3.49 (2H, m, CH.sub.2), 3.78
(3H, s, OCH.sub.3), 3.91 (3H, s, OCH.sub.3), 4.05 (2H, t,
CH.sub.2), 4.37 (2H, t, CH.sub.2), 5.49 (1H, s, C.dbd.CH), 5.80
(1H, br s, NH), 6.69 (1H, s, ArH), 6.70 (1H, s, ArH), 6.90 (2H, s,
2.times.ArH).
EXAMPLE 10
3-(N-Carbamoyl-2-aminoethyl)-9,10-dimethoxy-2-(2-methylphenylimino)-3,4,6,-
7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
##STR00025##
[0133]
9,10-Dimethoxy-2-(2-methylphenylimino)-3,4,6,7-tetrahydro-2H-pyrimi-
do[6,1-a]isoquinolin-4-one (1)
[0134] 2-Methylaniline (5.44 ml, 51 mmol) and
2-chloro-9,10-dimethoxy-6,7-dihydro-4H-pyrimido[6,1-a]isoquinolin-4-one
(5 g, 17 mmol) were suspended in propan-2-ol (400 ml) and heated at
reflux, under nitrogen, for 24 h. After cooling to room
temperature, the solution was concentrated in vacuo and the residue
purified by flash column chromatography [dichloromethane/methanol
(98:2-96:4)] to afford the title compound (6.2 g, quantitative
yield) as a yellow/orange solid.
9,10-Dimethoxy-2-(2-methylphenylimino)-3-(N-phthalimidoethyl)-3,4,6,7-tetr-
ahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (21
[0135] A mixture of isoquinoline 1 (6.2 g, 17 mmol), potassium
carbonate (21.1 g, 153 mmol), sodium iodide (15.3 g, 102 mmol) and
N-(2-bromoethyl)phthalimide (25.9 g, 102 mmol) in 2-butanone (170
ml) was stirred at reflux, under nitrogen, for 7 days. After
cooling to room temperature, the mixture was filtered and the
residue washed with methanol (150 ml). The filtrate was
concentrated in vacuo and the resultant residue treated with
methanol (100 ml) and the solid filtered off and washed with
methanol. This solid was washed with ether to give the title
compound (2.67 g, 30%) as a pale yellow solid
3-(2-Aminoethyl)-9,10-dimethoxy-2-(2-methylphenylimino)-3,4,6,7-tetrahydro-
-2H-pyrimido[6,1-a]isoquinolin-4-one (3)
[0136] A mixture of phthalimide 2 (2.66 g, 4.96 mmol) and hydrazine
monohydrate (1.24 g, 24.8 mmol) in chloroform (35 ml) and ethanol
(60 ml) was stirred at room temperature, under nitrogen, for 18 h.
Additional hydrazine hydrate (0.25 g, 5 mmol) was added and the
mixture stirred for a further 5 h. After cooling to 0.degree. C. in
an ice/water bath, the solid was removed by filtration, the residue
washed with a little cold chloroform and the filtrate concentrated
in vacuo. This residue was taken up in dichloromethane and the
insoluble material removed by filtration. The filtrate was dried
(MgSO.sub.4), filtered and concentrated in vacuo to afford the
title compound (2.05 g, quantitative yield) as a yellow foam.
3-(N-Carbamoyl-2-aminoethyl)-9,10-dimethoxy-2-(2-methylphenylimino)-3,4,6,-
7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
[0137] Sodium cyanate (0.64 g, 9.8 mmol) in water (13 ml) was added
dropwise to a stirred solution of amine 3 (2 g, 4.9 mmol) in water
(63 ml) and 1M HCl (9.8 ml) at 80.degree. C. After stirring for 3 h
at 80.degree. C. the mixture was cooled and basified with 2M NaOH.
The mixture was extracted with CH.sub.2Cl.sub.2 until no more
product remained in the organic phase. The organic phases were
combined, dried (MgSO.sub.4), filtered and concentrated in vacuo.
The residue was purified by column chromatography
[dichloromethane/methanol (97:3)] and the product triturated with
ether to afford the title compound (1.0 g, 45%) as a pale yellow
solid.
TABLE-US-00009 M.p.: 236-238.degree. C. m/z:
C.sub.24H.sub.27N.sub.5O.sub.4 requires M = 449, found (M + 1) =
450 HPLC: Area 100% Column ODS LUNA (150 4.6 mm) MP pH3
KH.sub.2PO.sub.4/CH.sub.3CN gradient 90% aq going to 50% over 25
mins FR 1.0 mlmin.sup.-1 RT 14.284 min Detection 250 nm
[0138] .sup.1H NMR (300 MHz; d.sub.6-DMSO): .delta. 2.08 (3H, s,
CH.sub.3), 2.89 (2H, t, CH.sub.2), 3.60 (3H, s, OMe), 3.79 (3H, s,
OMe), 3.91 (2H, t, CH.sub.2), 4.14 (2H, t, CH.sub.2), 5.44 (2H, br
s, NH.sub.2), 5.64 (1H, s, vinylic H), 6.07 (1H, t, NH), 6.71 (1H,
s, ArH), 6.73 (1H, d, ArH), 6.92 (1H, t, ArH), 6.95 (1H, s, ArH),
7.13 (1H, t, ArH), 7.19 (1H, d, ArH).
[0139] NB. The proton NMR spectrum above does not show the chemical
shift for one CH.sub.2 group as it is believed that this signal is
obscured by the water signal present in the d.sub.6-DMSO at
3.31-3.35.
##STR00026##
EXAMPLE 11
3-(N-Carbamoyl-2-aminoethyl)-2-(2,6-diisopropylphenylimino)-9,10-dimethoxy-
-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
##STR00027##
[0140]
2-(2,6-Diisopropylphenylimino)-9,10-dimethoxy-3,4,6,7-tetrahydro-2H-
-pyrimido-isoquinolin-4-one (1)
[0141] 2,6-Diisopropylaniline (9.62 ml, 51 mmol) and
2-chloro-9,10-dimethoxy-6,7-dihydro-4H-pyrimido[6,1-a]isoquinolin-4-one
(5 g, 17 mmol) were suspended in propan-2-ol (400 ml) and heated at
reflux, under nitrogen, for 4 days. After cooling to room
temperature, the solution was concentrated in vacuo and the residue
purified by flash column chromatography [dichloromethane/methanol
(98:2-96:4)] to afford the title compound (5.8 g, 79%) as a
yellow/orange solid.
2-(2,6-Diisopropylphenylimino)-9,10-dimethoxy-3-(N-phthalimidoethyl)-3,4,6-
,7-tetrahydro-2H-pyrimido[6,1-a] isoquinolin-4-one (2)
[0142] A mixture of isoquinoline 1 (5.8 g, 13.4 mmol), potassium
carbonate (16.7 g, 121 mmol), sodium iodide (12.1 g, 80 mmol) and
N-(2-bromoethyl)phthalimide (25.9 g, 80 mmol) in 2-butanone (150
ml) was stirred at reflux, under nitrogen, for 51/2 days. After
cooling to room temperature, the mixture was filtered and the
residue washed with methanol (150 ml). The filtrate was
concentrated in vacuo and the resultant residue treated with
methanol (100 ml) and the solid filtered off and washed thoroughly
with methanol. The resultant solid was dried to give the title
compound (4.9 g, 60%) as a pale yellow solid.
3-(2-Aminoethyl)-2-(2,6-diisopropylphenylimino)-9,10-dimethoxy-3,4,6,7-tet-
rahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (3)
[0143] A mixture of phthalimide 2 (4.9 g, 8.08 mmol) and hydrazine
monohydrate (2.01 g, 40.4 mmol) in chloroform (70 ml) and ethanol
(105 ml) was stirred at room temperature, under nitrogen, for 18 h.
Additional hydrazine hydrate (0.5 g, 10 mmol) was added and the
mixture stirred for a further 3 h. After cooling to 0.degree. C. in
an ice/water bath, the solid was removed by filtration, the residue
washed with a little cold chloroform and the filtrate concentrated
in vacuo. This residue was taken up in dichloromethane and the
insoluble material removed by filtration. The filtrate was dried
(MgSO.sub.4), filtered and concentrated in vacuo to afford the
title compound (3.24 g, 84%) as a yellow foam.
3-(N-Carbamoyl-2-aminoethyl)-2-(2,6-diisopropylphenylimino)-9,10-dimethoxy-
-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
[0144] Sodium cyanate (0.87 g, 13.4 mmol) in water (20 ml) was
added dropwise to a stirred solution of amine 3 (3.2 g, 6.7 mmol)
in water (100 ml) and 1M HCl (13.4 ml) at 80.degree. C. After
stirring for 4 h at 80.degree. C. the mixture was cooled and
basified with 2M NaOH. The mixture was extracted with
CH.sub.2Cl.sub.2 until no more product remained in the organic
phase. The organic phases were combined, dried (MgSO.sub.4),
filtered and concentrated in vacuo. The residue was purified by
column chromatography [dichloromethane/methanol (97:3)] and the
isolated product taken up in a mixture of dichloromethane and ether
which on concentration afforded the title compound (0.6 g, 17%) as
a pale yellow foam.
TABLE-US-00010 M.p.: 213-215.degree. C. m/z:
C.sub.29H.sub.37N.sub.5O.sub.4 requires M = 519, found (M + 1) =
520 HPLC: Area 97.59% Column ODS LUNA (150 4.6 mm) MP pH3
KH.sub.2PO.sub.4/CH.sub.3CN gradient 90% aq going to 50% over 25
mins FR 1.0 mlmin.sup.-1 RT 10.723 min Detection 250 nm
[0145] .sup.1H NMR (300 MHz; DMSO): 1.07 (12H, dd, 4 CH.sub.3),
2.82-2.94 (4H, m, CH.sub.2 and 2 CH(CH.sub.3).sub.2, 3.55 (3H, s,
OMe), 3.78 (3H, s, OMe), 3.91 (2H, t, CH.sub.2), 4.17 (2H, t,
CH.sub.2), 5.32 (1H, s, vinylic H), 5.45 (2H, br s, NH.sub.2), 6.13
(1H, t, NH), 6.56 (1H, s, ArH), 6.95 (1H, s, ArH), 7.00 (1H, m,
ArH), 7.10 (1H, s, ArH), 7.12 (1H, br s, ArH).
[0146] NB. The proton NMR spectrum above does not show the chemical
shift for one CH.sub.2 group as it is believed that this signal is
obscured by the water signal present in the d.sub.6-DMSO at
3.30-3.33.
##STR00028##
EXAMPLE 12
3-(N-Carbamoyl-4-aminobutyl)-9,10-dimethoxy-2-(2,4,6-trimethylphenylimino)-
-3,4,62-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
##STR00029##
[0147]
9,10-Dimethoxy-3-(4-N-phthalimidobutyl)-2-(2,4,6-trimethylphenylimi-
no)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (1)
[0148] A mixture of
9,10-dimethoxy-2-(2,4,6-trimethylphenylimino)-3,4,6,7-tetrahydro-2H-pyrim-
ido[6,1-a]isoquinolin-4-one (4.0 g, 10.2 mmol),
N-(4-bromobutyl)phthalimide (8.6 g, 30.6 mmol), potassium carbonate
(12.7 g, 91.8 mmol) and sodium iodide (4.6 g, 30.6 mmol) in
2-butanone (100 ml) was stirred at reflux, under nitrogen for 4
days. After cooling to room temperature, the solid was removed by
filtration and the filtrate was evaporated in vacuo. The residual
solid was purified by column chromatography on silica gel
[petroleum ether/ethyl acetate (2:1)-(1:1)]. The title compound
(1.45 g, 24%) was obtained as a yellow solid.
3-(4-Aminobutyl)-9,10-dimethoxy-2-(2,4,6-trimethylphenylimino)-3,4,6,7-tet-
rahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (2)
[0149] A solution of 1 (1.4 g, 2.36 mmol) in ethanol (30 ml) and
chloroform (20 ml) was treated with hydrazine hydrate (0.42 g, 7.09
mmol) and stirred at room temperature, under nitrogen. After 18 h
further hydrazine hydrate (0.42 g) was added and stirred for an
additional 5 h. The reaction mixture was then cooled to 0.degree.
C. and the solid was removed by filtration. The filtrate was dried
(MgSO.sub.4) and evaporated in vacuo to obtain the title compound
(1.0 g, 92%) as a yellow solid.
3-(N-Carbamoyl-4-aminobutyl)-9,10-dimethoxy-2-(2,4,6-trimethylphenylimino)-
-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
[0150] Sodium cyanate (0.28 g, 4.32 mmol) in water (6 ml) was added
dropwise to a stirred solution of 2 (1.0 g, 2.16 mmol) in water (30
ml) and 1N HCl (4.3 ml) at 80.degree. C. After 2 h at 80.degree. C.
the mixture was cooled and extracted with dichloromethane
(3.times.20 ml) and the extract dried (MgSO.sub.4) and evaporated
in vacuo. The residue was purified by column chromatography on
silica gel [dichloromethane/methanol (97:3)] to obtain the title
compound (0.70 g, 64%) as a yellow solid.
TABLE-US-00011 M.p: 234-235.degree. C. M/z
C.sub.28H.sub.35N.sub.5O.sub.4 requires M = 505, found m/z [ES+] =
506 HPLC: Area (%) 98.94 Column ODS LUNA 3uC18(2) (100 .times. 4.6
mm) MP 0.02M KH.sub.2PO.sub.4/CH.sub.3CN (gradient 90% aq .fwdarw.
50% aq over 25 min) RT (min) 17.201 FR (ml/min) 1.0 Detection 250
nm
[0151] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta. 1.64 (2H, m,
CH.sub.2), 1.90 (2H, m, CH.sub.2), 2.07 (6H, s, 2.times.CH.sub.3),
2.26 (3H, s, CH.sub.3), 2.92 (2H, t, CH.sub.2), 3.30 (2H, m,
CH.sub.2), 3.73 (3H, s, OCH.sub.3), 3.88 (3H, s, OCH.sub.3), 4.06
(2H, t, CH.sub.2), 4.27 (4H, m, CH.sub.2+NH.sub.2), 5.44 (1H, s,
C.dbd.CH), 6.65 (1H, s, ArH), 6.67 (1H, s, ArH), 6.89 (2H, s,
2.times.ArH).
##STR00030##
EXAMPLE 13
3-[N--(N'-Cyano-N''-methyl)-2-guanidinoethyl]-9,10-dimethoxy-2-(2,4,6-trim-
ethyl-phenylimino)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
##STR00031##
[0152]
3-[N--(N'-Cyano-S-methyl)-2-isothioureidoethyl]-9,10-dimethoxy-2-(2-
,4,6-trimethylphenylimino)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinoli-
n-4-one (1)
[0153] Dimethyl N-cyanodithioiminocarbonate (7.45 g, 46.1 mmol) was
added to a solution of
3-(2-aminoethyl)-9,10-dimethoxy-2-(2,4,6-trimethylphenylimino)-3,4,6,7-te-
trahydro-2H-pyrimido[6,1-a]isoquinolin-4-one (2.0 g, 4.61 mmol) in
toluene (50 ml) and stirred at 90.degree. C. under nitrogen. After
2 h the solvent was evaporated in vacuo and the residue was
purified by column chromatography on silica gel
[dichloromethane/methanol (100:0)-(95:5)]. The title compound (2.30
g, 94%) was obtained as a yellow solid.
3-[N--(N'-Cyano-N''-methyl)-2-guanidinoethyl]-9,10-dimethoxy-2-(2,4,6-trim-
ethylphenylimino)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one
[0154] A solution of 1 (2.0 g, 3.76 mmol) in dichloromethane (30
ml) was treated with 2M methylamine/THF (9.4 ml, 18.8 mmol) and
stirred at reflux, under nitrogen. After 16 h additional 2M
methylamine/THF (9.4 ml) was added, followed by a further two
portions of 2M methylamine/THF (9.4 ml) at 2 h intervals. After 24
h at reflux the reaction was cooled and evaporated in vacuo. The
residue was purified by column chromatography on silica gel
[dichloromethane/methanol (98:2)] to obtain, after trituration with
ether, the title compound (1.20 g, 62%) as a pale yellow solid.
TABLE-US-00012 M.p.: 223-224.degree. C. M/z :
C.sub.28H.sub.33N.sub.7O.sub.3 requires M = 515, found m/z [ES+] =
516 HPLC: Area (%) 100 Column ODS LUNA 3uC18(2) (100 .times. 4.6
mm) MP 0.02M KH.sub.2PO.sub.4/CH.sub.3CN (gradient 90% aq .fwdarw.
50% aq over 25 min) RT (min) 17.838 FR (ml/min) 1.0 Detection 250
nm
[0155] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta. 1.99 (6H, s,
2.times.CH.sub.3), 2.22 (3H, s, CH.sub.3), 2.43 (3H, d,
NHCH.sub.3), 2.86 (2H, t, CH.sub.2), 3.52 (2H, m, CH.sub.2), 3.69
(3H, s, OCH.sub.3), 3.83 (3H, s, OCH.sub.3), 4.02 (2H, t,
CH.sub.2), 4.28 (2H, m, CH.sub.2), 5.39 (1H, s, C.dbd.CH), 6.61
(1H, s, ArH), 6.63 (1H, s, ArH), 6.82 (2H, s, 2.times.ArH).
##STR00032##
[0156] The pharmacological utility of the compounds of the present
invention has been demonstrated in studies using compounds
previously synthesised from the above Examples. The results below
serve to illustrate the generic application of the compounds of the
present invention.
EXAMPLE A
Efficacy of a Compound of the Invention Against Electrical-Induced
Contraction of Guinea-Pig Isolated Trachea
[0157] The efficacy of the compounds of Example 1, 8, 9, 10, 11 and
13 were tested against electrical-induced contraction of guinea-pig
isolated trachea. The results demonstrate that the compounds of the
present invention inhibit the contractile responses with a long
duration of action.
Method
[0158] Superfusion of guinea-pig tracheal rings was performed
according to a previously described method (Coleman et al. 1996;
Pulmonary Pharmacology, 9, 107-117). Briefly, guinea-pig tracheal
preparations were cut into rings then opened by sectioning the ring
opposite the smooth muscle and suspended between two platinum
electrodes under 1 g tension. The tissues were superfused at a rate
of 3.25 mL/min with Krebs-Henseleit solution at 37.degree. C.
containing the cyclooxygenase inhibitor, indomethacin (5 .mu.M) and
bubbled with 95% O.sub.2 and 5% CO.sub.2. Tracheal preparations
were allowed to equilibrate for 40 min before commencement of
electrical stimulation delivered as a 10 s train of square wave
pulses at 3 Hz, 0.1 ms duration and 20V (approx. 400 mAmps)
generated every 100 sec by means of physiological square
wave-stimulator.
[0159] The compound of Example 1 was dissolved in DMSO containing
Tween 80 (10%) and distilled water (0.01M), which were then added
to the organ bath to give a final concentration of 10 .mu.M. The
other compounds were prepared in DMSO and diluted in
Krebs-Henseleit solution which yielded a final superfusion
concentration of 0.05% DMSO, and superfused at a rate of 0.3
mL/min; contractile responses to electrical field stimulation was
recorded on a Macintosh computer using MacLab software.
Results
[0160] The vehicle, DMSO, failed to significantly inhibit the
contractile response to electrical field stimulation (FIG. 2). The
results for the compounds are shown in FIGS. 3 for compound of
Example 1; 4 for compound of Example 9; 5 for compound of Example
10; 6 for compound of Example 11; 7 for compound of Example 13; and
8 for compound of Example 8.
[0161] The compounds caused complete inhibition of the contractile
response to electrical field stimulation and the effect was
maintained for more than 2-4 hours.
EXAMPLE B
Efficacy of a Compound of the Invention Against Proliferation of
Human Mononuclear Cells Stimulated by PHA
[0162] The effect of the compound of Example 1,
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(N-carbamoyl-2-aminoethyl-
)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one, against
proliferation of human mononuclear cells stimulated by PHA was
investigated. Proliferation was significantly inhibited by the
compound, indicating that it possesses anti-inflammatory activity.
The result below serves to illustrate the generic application of
the novel compounds of the present invention.
Method
[0163] Normal healthy volunteers underwent phlebotomy and 25 ml of
blood was collected. Mononuclear cells were separated and purified
according to the method of Banner et al. (Banner et al. Br. J.
Pharmacol. 116 3169-3174 (1995)). Human peripheral mononuclear
cells (100,000 per well) were stimulated for 24 h with
phytohaemagglutinin (PHA, 2 .mu.g/ml) in the absence or presence of
the compound of Example 1 (0.001-100 .mu.M) at 37.degree. C. in a
95% air, 5% CO.sub.2 atmosphere. Twenty four hours later,
[.sup.3H]-thymidine (0.1 .mu.Ci) was added to each well and the
cells were incubated for a further 24 h period. Cells were then
harvested onto glass fibre filters using a cell harvester (ICN
Flow, Buckinghamshire) and counted in a scintillation counter.
Results
[0164] The compound under test caused a concentration dependent
inhibition of the proliferation of human mononuclear cells
stimulated with PHA (number of experiments is 6; FIG. 8). The
IC.sub.50 values and 95% confidence limits for these compounds are
indicated in Table 1. The result is also shown in the graph of FIG.
8.
TABLE-US-00013 TABLE 1 IC.sub.50 value for the compound of Example
1 against proliferation of human mononuclear cells stimulated with
PHA Compound from IC.sub.50 n Example 1 0.46 .mu.M (0.239-0.897)
6
EXAMPLE C
Inhibition of Phosphodiesterase (PDE) Type 3 and 4 Isozymes
[0165] The compound of Example 1,
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(N-carbamoyl-2-aminoethyl-
)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one, has been
shown to be a potent inhibitor of phosphodiesterase (PDE) type 3
and 4 isozymes. The IC.sub.50 values are shown below.
TABLE-US-00014 PDE3 (nM) PDE4 (nM) (human platelet) (human
neutrophil) Compound of Example 1 0.43 1479 Rolipram ND 6412
Cilostamide 89 ND Rolipram is a known PDE 4 inhibitor and
cilostamide is a known PDE3 inhibitor ND--Not determined
EXAMPLE D
Effect of the Compound of EXAMPLE 1,
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(N-carbamoyl-2-aminoethyl-
)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one, on LPS
Induced TNF-.alpha. Release from Human Monocytes
TABLE-US-00015 [0166] IC50 (nM) Compound of Example 1 7.5 n = 6 CDP
840 (PDE4 inhibitor) 92 n = 6 Siguazodan (PDE3 inhibitor) >100
.mu.M
EXAMPLE E
In Vivo Tests
[0167] 1. The compound of Example 1,
9,10-Dimethoxy-2-(2,4,6-trimethylphenylimino)-3-(N-carbamoyl-2-aminoethyl-
)-3,4,6,7-tetrahydro-2H-pyrimido[6,1-a]isoquinolin-4-one, was
tested in a model of histamine induced bronchospasm. Conscious
guinea-pigs were exposed to dry powder (micronised) compound. The
drug was blended with lactose so that the final concentration was
0.25, 2.5 and 25%. At various times after exposure to drug the
animals were anaesthetized and challenged with varying doses of
histamine. Total airway resistance and mean arterial blood pressure
were recorded.
[0168] Exposure to dry powder (2.5 and 25%) provided significant
protection against histamine induced bronchospasm over a 5.5 hour
period and reduced mean blood pressure over this period.
[0169] 2. Intravenous administration of the compound (1 to 100
.mu.g/kg) to urethane anaesthetized guinea-pigs produced a dose
dependant reduction in mean arterial blood pressure. The compound
was dissolved in DMSO then diluted with saline (there was evidence
that the compound had come out of solution).
[0170] 3. In a model of antigen induced eosinophilia in the
ovalbumin sensitized guinea-pig, the compound (10 mg/kg)
administered orally 1 hour prior to antigen challenge,
significantly inhibited the recruitment of eosinophils to the lungs
following antigen challenge (aerosol) in sensitized guinea-pigs.
Exposure to dry powder (25%), 1.5 hours prior to antigen challenge,
significantly inhibited recruitment of eosinophils to the lungs
(measurements were made 24 hours after challenge).
[0171] Further experiments were carried out to characterise the
duration of action in this model. Compound (25%) administration 5.5
hours before antigen challenge failed to significantly inhibit
eosinophil recruitment to the lungs.
EXAMPLE F
Taste of Compounds
[0172] For pharmaceutical compounds which are administered orally,
taste is a very important factor in ensuring patient compliance.
Unexpectedly, the compounds of the present invention are
substantially tasteless. They are therefore particularly suitable
for oral administration, for example as dry powder to be
inhaled.
Method
[0173] Small amounts of the compound of Example 1, trequinsin
(9,10-dimethoxy-3-methyl-2-mesitylimino-2,3,6,7-tetrahydro-4H-pyrimido[6,-
1-a]isoquinolin-4-one) and desmethyl trequinsin
(9,10-dimethoxy-2-mesitylimino-2,3,6-tetrahydro-4H-pyrimido[6,1-a]isoquin-
olin-4-one) were placed on the tip of the tongue of an informed,
healthy male volunteer and the taste of each compound was
assessed.
Results
[0174] The results, displayed in Table 3 below, show that the
compound of Example 1 has significantly improved taste compared
with trequinsin or desmethyl trequinsin.
TABLE-US-00016 Compound (from) Taste.sup.1 Example 1 *** Trequinsin
* Desmethyl trequinsin * .sup.1Scale: * Very bitter ** Bitter ***
Tasteless
* * * * *