U.S. patent application number 10/052320 was filed with the patent office on 2002-07-25 for combination therapy.
Invention is credited to Hill, Roger J., Tracey, Wayne R..
Application Number | 20020099075 10/052320 |
Document ID | / |
Family ID | 26730463 |
Filed Date | 2002-07-25 |
United States Patent
Application |
20020099075 |
Kind Code |
A1 |
Tracey, Wayne R. ; et
al. |
July 25, 2002 |
COMBINATION THERAPY
Abstract
The invention provdes methods of reducing tissue damage
resulting from ischemia which comprise administering to a mammal in
need of such reduction an effective amount of a combination, or a
pharmaceutical composition comprising such combination, of an NHE-1
inhibitor and a second compound selected from the group consisting
of: (a) a complement modulator, (b) a metabolic modulator, (c) an
anti-apoptotic agent, (d) a nitric oxide synthase-related agent,
and (e) an enzyme/protein modulator. The invention further provides
pharmaceutical compositions comprising an amount of an NHE-1
inhibitor; an amount of a second compound selected from the group
consisting of (a) a complement modulator, (b) a metabolic
modulator, (c) an anti-apoptotic agent, (d) a nitric oxide
synthase-related agent, and an enzyme/protein modulator selected
from the group consisting of a protein kinase C activator, an
endothelin converting enzyme inhibitor, a tissue-activated
fibrinolytic inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger
isoform-1 (NCX-1) inhibitor, and a poly (ADP ribose) synthetase
(PARS/PARP) inhibitor; and, preferably, a pharmaceutically
acceptable pharmaceutically acceptable carrier, vehicle, or
diluent. The invention further provides kits comprising an amount
of a sodium-hydrogen exchanger type-1 inhibitor, and a
pharmaceutically acceptable carrier, vehicle, or diluent in a first
unit dosage form; an amount of a second compound selected from the
group consisting of (a) a complement modulator, (b) a metabolic
modulator, (c) an anti-apoptotic agent, (d) a nitric oxide
synthase-related agent, and (e) an enzyme/protein modulator
selected from the group consisting of a protein kinase C activator,
an endothelin converting enzyme inhibitor, a tissue-activated
fibrinolytic inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger
isoform-1 (NCX-1) inhibitor, and a poly (ADP ribose) synthetase
(PARS/PARP) inhibitor, and a pharmaceutically acceptable carrier,
vehicle, or diluent in a second unit dosage form; and a
container.
Inventors: |
Tracey, Wayne R.; (Niantic,
CT) ; Hill, Roger J.; (Salem, CT) |
Correspondence
Address: |
Gregg C. Benson
Pfizer Inc.
Patent Department, MS 4159
Eastern Point Road
Groton
CT
06340
US
|
Family ID: |
26730463 |
Appl. No.: |
10/052320 |
Filed: |
January 17, 2002 |
Related U.S. Patent Documents
|
|
|
|
|
|
Application
Number |
Filing Date |
Patent Number |
|
|
60264173 |
Jan 25, 2001 |
|
|
|
Current U.S.
Class: |
514/310 ;
514/291; 514/314; 514/406; 514/407; 514/421 |
Current CPC
Class: |
A61K 31/4745 20130101;
A61K 31/4709 20130101; A61K 31/4745 20130101; A61K 31/4709
20130101; A61K 2300/00 20130101; A61K 2300/00 20130101; A61K
31/4725 20130101; A61K 45/06 20130101; A61K 31/4725 20130101; A61K
2300/00 20130101 |
Class at
Publication: |
514/310 ;
514/314; 514/406; 514/407; 514/291; 514/421 |
International
Class: |
A61K 031/4745; A61K
031/4709; A61K 031/4725 |
Claims
1. A method of reducing tissue damage resulting from ischemia which
method comprises administering to a mammal in need of such
reduction a therapeutically effective amount of a combination
comprising a sodium-hydrogen exchanger type 1 (NHE-1) inhibitor,
and an effective amount of a second compound selected from the
group consisting of (a) a compliment modulator, (b) a metabolic
modulator, (c) an anti-apoptotic agent, (d) a nitric oxide
synthase-related agent, and (e) an enzyme/protein modulator
selected from the group consisting of a protein kinase C activator,
an endothelin converting enzyme inhibitor, a tissue-activated
fibrinolytic inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger
isoform-1 (NCX-1) inhibitor, and a poly (ADP ribose) synthetase
(PARS/PARP) inhibitor.
2. A method according to claim 1 wherein said tissue is selected
from the group consisting of brain, cardiac, liver, kidney, lung,
gut, skeletal muscle, spleen, pancreas, nerve, spinal cord, retinal
tissue, vasculature, and intestinal tissue.
3. A method according to claim 2 wherein said tissue is cardiac
tissue.
4. A method according to claim 1 wherein said sodium-hydrogen
exchanger type 1 (NHE-1) inhibitor is selected from the group
consisting of: (a) a compound of Formula (I) 7a prodrug thereof, or
a pharmaceutically acceptable salt of the compound or the prodrug
thereof; wherein: Z is carbon connected and is a five-membered,
diaza, diunsaturated ring having two contiguous nitrogens, said
ring optionally mono-, di-, or tri-substituted with up to three
substituents independently selected from R.sup.1, R.sup.2 and
R.sup.3; or Z is carbon connected and is a five-membered, triaza,
diunsaturated ring, said ring optionally mono- or di-substituted
with up to two substituents independently selected from R.sup.4 and
R.sup.5; wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4 and R.sup.5 are
each independently hydrogen, hydroxy(C.sub.1-C.sub.4)alkyl,
(C.sub.1-C.sub.4)alkyl, (C.sub.1-C.sub.4)alkylthio,
(C.sub.3-C.sub.4)cycloalkyl,
(C.sub.3-C.sub.7)cycloalkyl(C.sub.1-C.sub.4)- alkyl,
(C.sub.1-C.sub.4)alkoxy,
(C.sub.1-C.sub.4)alkoxy(C.sub.1-C.sub.4)al- kyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl, M or
M(C.sub.1-C.sub.4)alkyl, any of said previous
(C.sub.1-C.sub.4)alkyl moieties optionally having from one to nine
fluorines; said (C.sub.1-C.sub.4)alkyl or
(C.sub.3-C.sub.4)cycloalkyl optionally mono-or di-substituted
independently with hydroxy, (C.sub.1-C.sub.4)alkoxy,
(C.sub.1-C.sub.4)alklthio, (C.sub.1-C.sub.4)alkylsulfinyl,
(C.sub.1-C.sub.4)alkylsulfonyl, (C.sub.1-C.sub.4)alkyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl or mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylaminosulfonyl; and said
(C.sub.3-C.sub.4)cycloalkyl optionally having from one to seven
fluorines; wherein M is a partially saturated, fully saturated or
fully unsaturated five to eight membered ring optionally having one
to three heteroatoms selected independently from oxygen, sulfur and
nitrogen, or, a bicyclic ring consisting of two fused partially
saturated, fully saturated or fully unsaturated three to six
membered rings, taken independently, optionally having one to four
heteroatoms selected independently from nitrogen, sulfur and
oxygen; said M is optionally substituted, on one ring if the moiety
is monocyclic, or one or both rings if the moiety is bicyclic, on
carbon or nitrogen with up to three substituents independently
selected from R.sup.6, R.sup.7 and R.sup.8, wherein one of R.sup.6,
R.sup.7 and R.sup.8 is optionally a partially saturated, fully
saturated, or fully unsaturated three to seven membered ring
optionally having one to three heteroatoms selected independently
from oxygen, sulfur and nitrogen optionally substituted with
(C.sub.1-C.sub.4)alkyl and additionally R.sup.6, R.sup.7 and
R.sup.8 are optionally hydroxy, nitro, halo,
(C.sub.1-C.sub.4)alkoxy, (C.sub.1-C.sub.4)alkoxycarbonyl,
(C.sub.1-C.sub.4)alkyl, formyl, (C.sub.1-C.sub.4)alkanoyl,
(C.sub.1-C.sub.4)alkanoyloxy, (C.sub.1-C.sub.4)alkanoylamino,
(C.sub.1-C.sub.4)alkoxycarbonylamino, sulfonamido,
(C.sub.1-C.sub.4)alkylsulfonamido, amino, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylamino, carbamoyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl, cyano, thiol,
(C.sub.1-C.sub.4)alkylthio, (C.sub.1-C.sub.4)alkylsulfinyl,
(C.sub.1-C.sub.4)alkylsulfonyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkyla- minosulfonyl,
(C.sub.2-C.sub.4)alkenyl, (C.sub.2-C.sub.4)alkynyl or
(C.sub.5-C.sub.7)cycloalkenyl, wherein said
(C.sub.1-C.sub.4)alkoxy, (C.sub.1-C.sub.4)alkyl,
(C.sub.1-C.sub.7)alkanoyl, (C.sub.1-C.sub.4)alkylthio, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylamino or (C.sub.3-C.sub.7)cycloalkyl
R.sup.6, R.sup.7 and R.sup.8 substituents are optionally mono-
substituted independently with hydroxy,
(C.sub.1-C.sub.4)alkoxycarbonyl, (C.sub.3-C.sub.7)cycloalkyl,
(C.sub.1-C.sub.4)alkanoyl, (C.sub.1-C.sub.4)alkanoylamino,
(C.sub.1-C.sub.4)alkanoyloxy, (C.sub.1-C.sub.4)alkoxycarbonylamino,
sulfonamido, (C.sub.1-C.sub.4)alkylsulfonamido, amino, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylamino, carbamoyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl, cyano, thiol, nitro,
(C.sub.1-C.sub.4)alkylthio, (C.sub.1-C.sub.4)alkylsulfinyl,
(C.sub.1-C.sub.4)alkylsulfonyl or mono-N- or
di-N,N-(C.sub.1-C.sub.4)alky- laminosulfonyl or optionally
substituted with one to nine fluorines; (b) cariporide, or a
pharmaceutically acceptable salt thereof; (c) eniporide, or a
pharmaceutically acceptable salt thereof; (d) BIIB 513, or a
pharmaceutically acceptable salt thereof; (e) TY-12533, or a
pharmaceutically acceptable salt thereof; and (f) SM-15681, or a
pharmaceutically acceptable salt thereof.
5. A method according to claim 4 wherein said compound of Formula
(I) is a compound selected from the group consisting of:
[1-(2-chlorophenyl)-5-met- hyl-1H-pyrazole-4-carbonyl]guanidine;
[5-methyl-1-(2-trifluoromethylphenyl-
)-1H-pyrazole-4-carbonyl]guanidine;
[5-ethyl-1-phenyl-1H-pyrazole-4-carbon- yl]guanidine;
[5-cyclopropyl-1-(2-trifluoromethylphenyl)-1H-pyrazole-4-car-
bonyl]guanidine;
[5-cyclopropyl-1-phenyl-1H-pyrazole-4-carbonyl]guanidine;
[5-cyclopropyl-1-(2,6-dichlorophenyl)-1H-pyrazole-4-carbonyl]guanidine;
[5-methyl-1-(quinoli n-6-yl)-1H-pyrazole-4-carbonyl]guanidine;
[5-methyl-1-(naphthalen-1-yl)-1H-pyrazole-4-carbonyl]guanidine;
[5-cyclopropyl-1-(quinolin-5-yl)-1H-pyrazole-4-carbonyl]guanidine;
[5-cyclopropyl-1-(quinolin-8-yl)-1H-pyrazole-4-carbonyl]guanidine;
[3-methyl-1-phenyl-1H-pyrazole-4-carbonyl]guanidine;
[3-methyl-1-(naphthalen-1-yl)-1H-pyrazole-4-carbonyl]guanidine;
[3-methyl-1-(isoquinolin-5-yl)-1H-pyrazole-4-carbonyl]guanidine;
[2-methyl-5-phenyl-2H-pyrazole-3-carbonyl]guanidine;
[2-methyl-5-(naphthalen-1-yl)-2H-pyrazole-3-carbonyl]guanidine;
[5-methyl-2-phenyl-2H-1,2,3-triazole-4-carbonyl]guanidine;
[5-methyl-2-(3-methoxyphenyl)-2H-1,2,3-triazole-4-carbonyl]guanidine;
[2-(3-bromophenyl)-5-methyl-2H-1,2,3-triazole-4-carbonyl]guanidine;
[2-(naphthalen-1-yl)-5-methyl-2H-1,2,3-triazole-4-carbonyl]guanidine;
[2-(isoquinolin-5-yl)-5-methyl -2H-1,2,3-triazole-4-
carbonyl]guanidine;
[5-methyl-2-(quinolin-5-yl)-2H-1,2,3-triazole-4-carbonyl]guanidine;
[1-(naphthalen-1-yl)-5-cyclopropyl-2H-pyrazole-4-carbonyl]guanidine;
[1-(2-chloro-4-methylsulfonylphenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl-
]guanidine;
[1-(2-chlorophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guani-
dine;
[1-(2-trifluoromethyl-4-fluorophenyl)-5-cyclopropyl-1H-pyrazole-4-ca-
rbonyl]guanidine;
[1-(2-bromophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]-
guanidine;
[1-(2-fluorophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanid-
ine;
[1-(2-chloro-5-methoxyphenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]gu-
anidine;
[1-(2-chloro-4-methylaminosulfonylphenyl)-5-cyclopropyl-1H-pyrazo-
le-4-carbonyl]guanidine;
[1-(2,5-dichlorophenyl)-5-cyclopropyl-1H-pyrazole-
-4-carbonyl]guanidine;
[1-(2,3-dichlorophenyl)-5-cyclopropyl-1H-pyrazole-4-
-carbonyl]guanidine;
[1-(2-chloro-5-aminocarbonylphenyl)-5-cyclopropyl-1H--
pyrazole-4-carbonyl]guanidine;
[1-(2-chloro-5-aminosulfonylphenyl)-5-cyclo-
propyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(2-fluoro-6-trifluoromethylphe-
nyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(2-chloro-5-methylsulfonylphenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl-
]guanidine;
[1-(2-chloro-5-dimethylaminosulfonylphenyl)-5-cyclopropyl-1H-p-
yrazole-4-carbonyl]guanidine;
[1-(2-trifluoromethyl-4-chlorophenyl)-5-cycl-
opropyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(8-bromoquinolin-5-y1)-5-cycl-
opropyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(6-chloroquinolin-5-y1)-5-cyc-
lopropyl-1H-pyrazole4-carbonyl]guanidine;
[1-(indazol-7-yl)-5-cyclopropyl-- 1H-pyrazole-4-carbonyl]guanidine;
[1-(benzimidazol-5-yl)-5-cyclopropyl-1H--
pyrazole-4-carbonyl]guanidine;
[1-(1-isoquinolyl)-5-cyclopropyl-1H-pyrazol-
e-4-carbonyl]guanidine;
[5-cyclopropyl-1-(4-quinolinyl)-1H-pyrazole-4-carb- onyl]guanidine;
[1-(indazol-6-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine- ;
[1-(indazol-5-y1)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(benzimidazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(1-methylbenzimidazol-6-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine
[1-(5-quinolinyl)-5-n-propyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(5-quinolinyl)-5-isopropyl-1H-pyrazole-4-carbonyl]guanidine;
[5-ethyl-1-(6-quinolinyl)-1H-pyrazole4-carbonyl]guanidine;
[1-(2-methylbenzimidazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(1,4-benzodioxan-6-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(benzotriazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(3-chloroindazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(5-quinolinyl)-5-butyl-1H-pyrazole-4-carbonyl]guanidine;
[5-propyl-1-(6-quinolinyl)-1H-pyrazole-4-carbonyl]g uanidine;
[5-isopropyl-1-(6-quinolinyl)-1H-pyrazole-4-carbonyl]guanidine;
[1-(indazol-7-yl)-3-methyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(2,1,3-benzothiadiazol-4-yl)-3-methyl-1H-pyrazole-4-carbonyl]guanidine-
; and [3-methyl-1-(quinolin-5-yl)-1H-pyrazole-4-carbonyl]guanidine;
the prodrugs thereof, and the pharmaceutically acceptable salts of
the compounds, and the prodrugs.
6. A method according to claim 1 wherein said complement modulator
is selected from the group consisting of a C5a inhibitor, a soluble
complement receptor type 1 (sCR1) inhibitor, or an analog thereof,
and a C1 esterase inhibitor.
7. A method according to claim 6 wherein said C5a inhibitor is
L-747981, or a pharmaceutically acceptable salt thereof, and said
soluble complement receptor type 1 (sCR1) inhibitor is
amidinophenylpyruvic acid (APPA).
8. A method according to claim 1 wherein said metabolic modulator
is selected from the group consisting of a pyruvate dehydrogenase
complex up-regulator/activator, a pyruvate dehydrogenase kinase
inhibitor, a malonyl CoA decarboxylase inhibitor, an acetyl CoA
carboxylase activator, a partial fatty acid oxidation (pFOX)
inhibitor, a 5' AMP-activated protein kinase (AMPK) inhibitor, a
carnitine palmitoyl transferase inhibitor, and a fatty acid CoA
synthase inhibitor.
9. A method according to claim 8 wherein said pyruvate
dehydrogenase complex up-regulator/activator and said pyruvate
dehydrogenase kinase inhibitor are dichloroacetate (DCA), said
partial fatty acid oxidation (pFOX) inhibitor is ranolazine or
trimetazidine, said carnitine palmitoyl transferase inhibitor is
etomoxir, and said fatty acid CoA synthase inhibitor is triascin
C.
10. A method according to claim 1 wherein said anti-apoptotic agent
is a caspase inhibitor.
11. A method according to claim 10 wherein said caspase inhibitor
is a compound selected from the group consisting of a compound of
Formula (II) 8wherein R.sub.1 and R.sub.2, together with the
nitrogen to which they are attached, form a 4- to 7-membered ring;
R.sub.3 and R.sub.4, together with the nitrogen atom to which they
are attached, form a 4- to 7-membered ring; and R is benzoyl, or
(C.sub.1-6)alkyl; and a compound of Formula (III) 9wherein R.sub.1
is hydrogen, or (C.sub.1-4)alkyl; R.sub.2 is (C.sub.1-10)alkyl,
optionally substituted with aryl(C.sub.1-4)alkyl, optionally
substituted heteroaryl(C.sub.1-4)alkyl, optionally substituted
(C.sub.3-7) cycloalkyl, or R.sub.1 and R.sub.2, together with the
nitrogen to which they are attached, form a 3- to 10-membered ring
which optionally contains an additional heteroatom selected from
oxygen, nitrogen, or sulfur; R.sub.3 and R.sub.4 are
(C.sub.1-6)alkyl, hydrogen, nitro, or halogen; and R.sub.5 is
(C.sub.1-6)alkyl, hydrogen, arylalkyl, or heteroarylalkyl.
12. A method according to claim 1 wherein said nitric oxide
synthase-related agent is selected from the group consisting of
monophosporyl lipid A, or an analog thereof, a nitric oxide donor,
and a nitric oxide synthase activator.
13. A method according to claim 12 wherein said monophosporyl lipid
A, or said analog thereof is RC-552 (MPL-C) or ONO-4007, said
nitric oxide donor is nipride, and said nitric oxide synthase
inhibitor is aminoguanidine or N(G)-monomethyl-L-arginine
(L-NMMA).
14. A method according to claim 1 wherein said endothelin
converting enzyme inhibitor is S-17162, said Na.sup.+/Ca.sup.+2
exchanger isoform-1 (NCX-1) inhibitor is KB-R7943, and said poly
(ADP ribose) synthetase (PARS/PARP) inhibitor is 3-aminobenzamide,
or a compound, or a pharmaceutically acceptable salt, prodrug,
active metabolite, or solvate thereof, of Formula (IV) 10wherein
R.sup.1 is H, halogen, cyano, an optionally substituted alkyl,
alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl
group; or --C(O)--R.sup.10, where R.sup.10 is hydrogen, an
optionally substituted alkyl, alkenyl, alkynyl, cycloalkyl,
heterocycloalkyl, aryl, or heteroaryl group; or --OR.sup.100 or
NR.sup.100R.sup.110, where R.sup.100 and R.sup.110 are each
independently H or an optionally substituted alkyl, alkenyl,
alkynyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl group;
R.sup.2 is H or alkyl; R.sup.3 is H or alkyl; R.sup.4 is H, halogen
or alkyl; X is O or S; and Y is (CR.sup.5R.sup.6)(CR.sup.7R.sup.8)n
or N.dbd.C(R.sup.5), where n is 0 or 1; R.sup.5 and R.sup.6 are
each independently H or an optionally substituted alkyl, alkenyl,
alkynyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl group;
and R.sup.7 and R.sup.8 are each independently H or an optionally
substituted alky, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,
aryl, or heteroaryl group.
15. A pharmaceutical composition comprising an amount of a
sodium-hydrogen exchanger type-1 inhibitor, and an amount of a
second compound selected from the group consisting of (a) a
complement modulator, (b) a metabolic modulator, (c) an
anti-apoptotic agent, (d) a nitric oxide synthase-related agent,
and (e) an enzyme/protein modulator selected from the group
consisting of a protein kinase C activator, an endothelin
converting enzyme inhibitor, a tissue-activated fibrinolytic
inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1)
inhibitor, and a poly (ADP ribose) synthetase (PARS/PARP)
inhibitor.
16. A pharmaceutical composition according to claim 15 further
comprising a pharmaceutically acceptable carrier, vehicle, or
diluent.
17. A pharmaceutical composition according to claim 15 wherein said
sodium-hydrogen exchanger type 1 (NHE-1) inhibitor is selected from
the group consisting of: (a) a compound of Formula (I) 11a prodrug
thereof, or a pharmaceutically acceptable salt of the compound or
the prodrug thereof; wherein: Z is carbon connected and is a
five-membered, diaza, diunsaturated ring having two contuguous
notrogens, said ring potionally mono-, di-, or tri-substituted with
up to three substituents independiently selected form R.sup.1 ,
R.sup.2 and R.sup.3; or Z is carbon connected and is a
five-membered, triaza, diunsaturated ring, said ring optionally
mono- or di-substituted with up to two substituents independently
selected from R.sup.4 and R.sup.5; wherein R.sup.1, R.sup.2,
R.sup.3, R.sup.4 and R.sup.5 are each independently hydrogen,
hydroxy(C.sub.1-C.sub.4)alkyl, (C.sub.1-C.sub.4)alkyl,
(C.sub.1-C.sub.4)alkylthio, (C.sub.3-C.sub.4)cycloalkyl,
(C.sub.3-C.sub.7)cycloalkyl(C.sub.1-C.sub.4)alkyl,
(C.sub.1-C.sub.4)alkoxy,
(C.sub.1-C.sub.4)alkoxy(C.sub.1-C.sub.4)alkyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl, M or
M(C.sub.1-C.sub.4)alkyl, any of said previous
(C.sub.1-C.sub.4)alkyl moieties optionally having from one to nine
fluorines; said (C.sub.1-C.sub.4)alkyl or
(C.sub.3-C.sub.4)cycloalkyl optionally mono-or di-substituted
independently with hydroxy, (C.sub.1-C.sub.4)alkoxy,
(C.sub.1-C.sub.4)alklthio, (C.sub.1-C.sub.4)alkylsulfinyl,
(C.sub.1-C.sub.4)alkylsulfonyl, (C.sub.1-C.sub.4)alkyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl or mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylaminosulfonyl; and said
(C.sub.3-C.sub.4)cycloalkyl optionally having from one to seven
fluorines; wherein M is a partially saturated, fully saturated or
fully unsaturated five to eight membered ring optionally having one
to three heteroatoms selected independently from oxygen, sulfur and
nitrogen, or, a bicyclic ring consisting of two fused partially
saturated, fully saturated or fully unsaturated three to six
membered rings, taken independently, optionally having one to four
heteroatoms selected independently from nitrogen, sulfur and
oxygen; said M is optionally substituted, on one ring if the moiety
is monocyclic, or one or both rings if the moiety is bicyclic, on
carbon or nitrogen with up to three substituents independently
selected from R.sup.6, R.sup.7 and R.sup.8, wherein one of R.sup.6,
R.sup.7 and R.sup.8 is optionally a partially saturated, fully
saturated, or fully unsaturated three to seven membered ring
optionally having one to three heteroatoms selected independently
from oxygen, sulfur and nitrogen optionally substituted with
(C.sub.1-C.sub.4)alkyl and additionally R.sup.6, R.sup.7 and
R.sup.8 are optionally hydroxy, nitro, halo,
(C.sub.1-C.sub.4)alkoxy, (C.sub.1-C.sub.4)alkoxycarbonyl,
(C.sub.1-C.sub.4)alkyl, formyl, (C.sub.1-C.sub.4)alkanoyl,
(C.sub.1-C.sub.4)alkanoyloxy, (C.sub.1-C.sub.4)alkanoylamino,
(C.sub.1-C.sub.4)alkoxycarbonylamino, sulfonamido,
(C.sub.1-C.sub.4)alkylsulfonamido, amino, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylamino, carbamoyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl, cyano, thiol,
(C.sub.1-C.sub.4)alkylthio, (C.sub.1-C.sub.4)alkylsulfinyl,
(C.sub.1-C.sub.4)alkylsulfonyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkyla- minosulfonyl,
(C.sub.2-C.sub.4)alkenyl, (C.sub.2-C.sub.4)alkynyl or
(C.sub.5-C.sub.7)cycloalkenyl, wherein said
(C.sub.1-C.sub.4)alkoxy, (C.sub.1-C.sub.4)alkyl,
(C.sub.1-C.sub.7)alkanoyl, (C.sub.1-C.sub.4)alkylthio, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylamino or (C.sub.3-C.sub.7)cycloalkyl
R.sup.6, R.sup.7 and R.sup.8 substituents are optionally mono-
substituted independently with hydroxy,
(C.sub.1-C.sub.4)alkoxycarbonyl, (C.sub.3-C.sub.7)cycloalkyl,
(C.sub.1-C.sub.4)alkanoyl, (C.sub.1-C.sub.4)alkanoylamino,
(C.sub.1-C.sub.4)alkanoyloxy, (C.sub.1-C.sub.4)alkoxycarbonylamino,
sulfonamido, (C.sub.1-C.sub.4)alkylsulfonamido, amino, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylamino, carbamoyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl, cyano, thiol, nitro,
(C.sub.1-C.sub.4)alkylthio, (C.sub.1-C.sub.4)alkylsulfinyl,
(C.sub.1-C.sub.4)alkylsulfonyl or mono-N- or
di-N,N-(C.sub.1-C.sub.4)alky- laminosulfonyl or optionally
substituted with one to nine fluorines; (b) cariporide, or a
pharmaceutically acceptable salt thereof; (c) eniporide, or a
pharmaceutically acceptable salt thereof; (d) BIIB 513, or a
pharmaceutically acceptable salt thereof; (e) TY-12533, or a
pharmaceutically acceptable salt thereof; and (f) SM-15681, or a
pharmaceutically acceptable salt thereof.
18. A pharmaceutical composition according to claim 17 wherein said
compound of Formula (I) is a compound selected from the group
consisting of:
[1-(2-chlorophenyl)-5-methyl-1H-pyrazole-4-carbonyl]guanidine;
[5-methyl-1-(2-trifluoromethylphenyl)-1H-pyrazole-4-carbonyl]guanidine;
[5-ethyl-1-phenyl-1H-pyrazole-4-carbonyl]guanidine;
[5-cyclopropyl-1-(2-trifluoromethylphenyl)-1H-pyrazole-4-carbonyl]guanidi-
ne; [5-cyclopropyl-1-phenyl-1H-pyrazole-4-carbonyl]guanidine;
[5-cyclopropyl-1-(2,6-dichlorophenyl)-1H-pyrazole-4-carbonyl]guanidine;
[5-methyl-1-(quinolin-6-yl)-1H-pyrazole-4-carbonyl]guanidine;
[5-methyl-1-(naphthalen-1-yl)-1H-pyrazole-4-carbonyl]guanidine;
[5-cyclopropyl-1-(quinolin-5-yl)-1H-pyrazole-4-carbonyl]guanidine;
[5-cyclopropyl-1-(quinolin-8-yl)-1H-pyrazole-4-carbonyl]guanidine;
[3-methyl-1-phenyl-1H-pyrazole-4-carbonyl]guanidine;
[3-methyl-1-(naphthalen-1-yl)-1H-pyrazole-4-carbonyl]guanidine;
[3-methyl-1-(isoquinolin-5-yl)-1H-pyrazole-4-carbonyl]guanidine;
[2-methyl-5-phenyl-2H-pyrazole-3-carbonyl]guanidine;
[2-methyl-5-(naphthalen-1-yl)-2H-pyrazole-3-carbonyl]guanidine;
[5-methyl-2-phenyl-2H-1,2,3-triazole-4-carbonyl]guanidine;
[5-methyl-2-(3-methoxyphenyl)-2H-1,2,3-triazole-4-carbonyl]guanidine;
[2-(3-bromophenyl)-5-methyl-2H-1,2,3-triazole-4-carbonyl]guanidine;
[2-(naphthalen-1-yl)-5-methyl-2H-1,2,3-triazole-4-carbonyl]guanidine;
[2-(isoquinolin-5-yl)-5-methyl -2H-1,2,3-triazole-4-
carbonyl]guanidine; [5-methyl-2-(quinolin-5-yl)-2H-
1,2,3-triazole-4-carbonyl]guanidine;
[1-(naphthalen-1-yl)-5-cyclopropyl-2H-pyrazole-4-carbonyl]guanidine;
[1-(2-chloro-4-methylsulfonylphenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl-
]guanidine;
[1-(2-chlorophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guani-
dine;
[1-(2-trifluoromethyl-4-fluorophenyl)-5-cyclopropyl-1H-pyrazole-4-ca-
rbonyl]guanidine;
[1-(2-bromophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]-
guanidine;
[1-(2-fluorophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanid-
ine;
[1-(2-chloro-5-methoxyphenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]gu-
anidine;
[1-(2-chloro-4-methylaminosulfonylphenyl)-5-cyclopropyl-1H-pyrazo-
le-4-carbonyl]guanidine;
[1-(2,5-dichlorophenyl)-5-cyclopropyl-1H-pyrazole-
-4-carbonyl]guanidine;
[1-(2,3-dichlorophenyl)-5-cyclopropyl-1H-pyrazole-4-
-carbonyl]guanidine;
[1-(2-chloro-5-aminocarbonylphenyl)-5-cyclopropyl-1H--
pyrazole-4-carbonyl]guanidine;
[1-(2-chloro-5-aminosulfonylphenyl)-5-cyclo-
propyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(2-fluoro-6-trifluoromethylphe-
nyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(2-chloro-5-methylsulfonylphenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl-
]guanidine;
[1-(2-chloro-5-dimethylaminosulfonylphenyl)-5-cyclopropyl-1H-p-
yrazole-4-carbonyl]guanidine;
[1-(2-trifluoromethyl-4-chlorophenyl)-5-cycl-
opropyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(8-bromoquinolin-5-y1)-5-cycl-
opropyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(6-chloroquinolin-5-y1)-5-cyc-
lopropyl-1H-pyrazole4-carbonyl]guanidine;
[1-(indazol-7-yl)-5-cyclopropyl-- 1H-pyrazole-4-carbonyl]guanidine;
[1-(benzimidazol-5-yl)-5-cyclopropyl-1H--
pyrazole-4-carbonyl]guanidine;
[1-(1-isoquinolyl)-5-cyclopropyl-1H-pyrazol-
e-4-carbonyl]guanidine;
[5-cyclopropyl-1-(4-quinolinyl)-1H-pyrazole-4-carb- onyl]guanidine;
[1-(indazol-6-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine- ;
[1-(indazol-5-yI)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(benzimidazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(1-methylbenzimidazol-6-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine
[1-(5-quinolinyl)-5-n-propyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(5-quinolinyl)-5-isopropyl-1H-pyrazole-4-carbonyl]guanidine;
[5-ethyl-1-(6-quinolinyl)-1H-pyrazole4-carbonyl]guanidine;
[1-(2-methylbenzimidazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(1,4-benzodioxan-6-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(benzotriazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(3-chloroindazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(5-quinolinyl)-5-butyl-1H-pyrazole-4-carbonyl]guanidine;
[5-propyl-1-(6-quinolinyl)-1H-pyrazole-4-carbonyl]g uanidine;
[5-isopropyl-1-(6-quinolinyl)-1H-pyrazole-4-carbonyl]guanidine;
[1-(indazol-7-yl)-3-methyl-1H-pyrazole-4-carbonyl]guanidine;
[1-(2,1,3-benzothiadiazol-4-yl)-3-methyl-1H-pyrazole-4-carbonyl]guanidine-
; and [3-methyl-1-(quinolin-5-yl)-1H-pyrazole-4-carbonyl]guanidine;
the prodrugs thereof, and the pharmaceutically acceptable salts of
the compounds, and the prodrugs.
19. A pharmaceutical composition according to claim 15 wherein said
complement modulator is selected from the group consisting of a C5a
inhibitor, a soluble complement receptor type-1 (sCR1) inhibitor,
or an analog thereof, and a C1 esterase inhibitor.
20. A pharmaceutical composition according to claim 19 wherein said
C5a inhibitor is L-747981, or a pharmaceutically acceptable salt
thereof, and said soluble complement receptor type 1 (sCR1)
inhibitor is amidinophenylpyruvic acid (APPA).
21. A pharmaceutical composition according to claim 15 wherein said
metabolic modulator is selected from the group consisting of a
pyruvate dehydrogenase complex up-regulator/activator, a pyruvate
dehydrogenase kinase inhibitor, a malonyl CoA decarboxylase
inhibitor, an acetyl CoA carboxylase activator, a partial fatty
acid oxidation (pFOX) inhibitor, a 5' AMP-activated protein kinase
(AMPK) inhibitor, a carnitine palmitoyl transferase inhibitor, and
a fatty acid CoA synthase inhibitor.
22. A method according to claim 21 wherein said pyruvate
dehydrogenase complex up-regulator/activator and said pyruvate
dehydrogenase kinase inhibitor are dichloroacetate (DCA), said
partial fatty acid oxidation (PFOX) inhibitor is ranolazine or
trimetazidine, said carnitine palmitoyl transferase inhibitor is
etomoxir, and said fatty acid CoA synthase inhibitor is triascin
C.
23. A pharmaceutical composition according to claim 15 wherein said
anti-apoptotic agent is a caspase inhibitor.
24. A pharmaceutical composition according to claim 23 wherein said
caspase inhibitor is a compound selected from the group consisting
of a compound of Formula (II) 12wherein R.sub.1 and R.sub.2,
together with the nitrogen to which they are attached, form a 4- to
7-membered ring; R.sub.3 and R.sub.4, together with the nitrogen
atom to which they are attached, form a 4- to 7-membered ring; and
R is benzoyl, or (C.sub.1-6)alkyl; and a compound of Formula (III)
13wherein R.sub.1 is hydrogen, or (C.sub.1-4)alkyl; R.sub.2 is
(C.sub.1-10)alkyl, optionally substituted with
aryl(C.sub.1-4)alkyl, optionally substituted
heteroaryl(C.sub.1-4)alkyl, optionally substituted
(C.sub.3-7)cycloalkyl, or R.sub.1 and R.sub.2, together with the
nitrogen to which they are attached, form a 3- to 10-membered ring
which optionally contains an additional heteroatom selected from
oxygen, nitrogen, or sulfur; R.sub.3 and R.sub.4 are
(C.sub.1-6)alkyl, hydrogen, nitro, or halogen; and R.sub.5 is
(C.sub.1-6,)alkyl, hydrogen, arylalkyl, or heteroarylalkyl.
25. A pharmaceutical composition according to claim 15 wherein said
nitric oxide synthase-related agent is selected from the group
consisting of monophosporyl lipid A, or an analog thereof, a nitric
oxide donor, and a nitric oxide synthase activator.
26. A pharmaceutical composition according to claim 25 wherein said
monophosporyl lipid A, or said analog thereof is RC-552 (MPL-C) or
ONO-4007, said nitric oxide donor is nipride, and said nitric oxide
synthase inhibitor is aminoguanidine or N(G)-monomethyl-L-arginine
(L-NMMA).
27. A pharmaceutical composition according to claim 15 wherein said
endothelin converting enzyme inhibitor is S-17162, said
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor is
KB-R7943, and said poly (ADP ribose) synthetase (PARS/PARP)
inhibitor is 3-aminobenzamide, or a compound, or a pharmaceutically
acceptable salt, prodrug, active metabolite, or solvate thereof, of
Formula (IV) 14wherein R.sup.1 is H, halogen, cyano, an optionally
substituted alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,
aryl, or heteroaryl group; or --C(O)--R.sup.10, where R.sup.10 is
hydrogen, an optionally substituted alkyl, alkenyl, alkynyl,
cycloalkyl, heterocycloalkyl, aryl, or heteroaryl group; or
--OR.sup.100or NR.sup.100 R.sup.110, where R.sup.100 and R.sup.110
are each independently H or an optionally substituted alkyl,
alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl
group; R.sup.2 is H or alkyl; R.sup.3 is H or alkyl; R.sup.4 is H,
halogen or alkyl; X is O or S; and Y is
(CR.sup.5R.sup.6)(CR.sup.7R.sup.8)n or N.dbd.C(R.sup.5), where n is
0 or 1; R.sup.5 and R.sup.6 are each independently H or an
optionally substituted alkyl, alkenyl, alkynyl, cycloalkyl,
heterocycloalkyl, aryl, or heteroaryl group; and R.sup.7 and
R.sup.8 are each independently H or an optionally substituted alky,
alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl
group.
28. A method of reducing tissue damage resulting from ischemia
which method comprises administering to a mammal in need of such
treatment an effective amount of a pharmaceutical composition of
claim 15.
29. A method according to claim 28 wherein said tissue is selected
from the group consisting of brain, cardiac, liver, kidney, lung,
gut, skeletal muscle, spleen, pancreas, nerve, spinal cord, retinal
tissue, vasculature, and intestinal tissue.
30. A method according to claim 29 wherein said tissue is cardiac
tissue.
31. A kit comprising an amount of a sodium-hydrogen exchanger
type-1 inhibitor, and a pharmaceutically acceptable carrier,
vehicle, or diluent in a first unit dosage form; an amount of a
second compound selected from the group consisting of (a) a
complement modulator, (b) a metabolic modulator, (c) an
anti-apoptotic agent, (d) a nitric oxide synthase-related agent,
and (e) an enzyme/protein modulator selected from the group
consisting of a protein kinase C activator, an endothelin
converting enzyme inhibitor, a tissue-activated fibrinolytic
inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1)
inhibitor, and a poly (ADP ribose) synthetase (PARS/PARP)
inhibitor, and a pharmaceutically acceptable carrier, vehicle, or
diluent in a second unit dosage form; and a container.
Description
CROSS REFERENCE TO RELATED APPLICATION
[0001] This application claims the benefit of U.S. Provisional
Application No. 60/264,173, Jan.25, 2001.
FIELD OF THE INVENTION
[0002] The invention relates to methods of reducing tissue damage
resulting from ischemia using a combination, or a pharmaceutical
composition of such combination, of a sodium/hydrogen exchange
type-1 (NHE-1) inhibitor and a second compound selected from the
group consisting of: (a) a complement modulator, (b) a metabolic
modulator, (c) an antiapoptotic agent, (d) a nitric oxide
synthase-related agent, and (e) an enzyme/protein modulator. The
invention further provides kits directed to such combinations.
BACKGROUND OF THE INVENTION
[0003] Ischemic injury, particularly to that of the myocardium, can
occur in out patients as well as in perioperative settings and can
lead to the development of sudden death, myocardial infarction, or
congestive heart failure. There is currently an unmet medical need
to prevent or minimize myocardial ischemic injury, particularly
perioperative myocardial infarction. Such therapy is anticipated to
be lifesaving and reduce the need for hospitalization, enhance
quality of life and reduce overall health care costs of high risk
patients.
[0004] Pharmacological cardioprotection would reduce the incidence
and progression of myocardial infarction and dysfunction occurring
in these perioperative surgical settings. In addition to reducing
myocardial damage and improving postischemic myocardial function in
patients with ischemic heart disease, cardioprotection would also
decrease the incidence of cardiac morbidity and mortality due to
myocardial infarction and dysfunction in "at risk" patients (i.e.,
those patients greater than 65 years of age, exercise intolerant,
those suffering from coronary artery disease, diabetes mellitus, or
hypertension, and the like) that require non-cardiac surgery.
[0005] In response thereto, numerous therapeutic regimens have been
developed, for example, the use of compounds that inhibit the
sodium/hydrogen exchange type-1 (NHE-1) transport system. The
mechanism by which NHE-1 inhibitors elicit protective effects
against ischemia, particularly that affecting the myocardium,
consists of a reduction in the increased sodium ion influx which is
caused in reperfused/hypoperfused tissues due to intracellular
acidification and subsequent activation of the sodium/hydrogen
exchange transport system. This results in a delay of sodium
overload of the tissue. Since sodium and calcium ion transport are
coupled in cardiac tissue, this also prevents the life-threatening
calcium overload of myocardial cells.
[0006] The use of NHE-1 inhibitors in combination with certain
other therapeutic agents is generally known. For example, EPO 0 918
515 discloses the use of NHE-1 inhibitors with blood pressure
reducing agents, ACE-inhibitors, angiotensin receptors antagonists,
fat level reducing agents, and HMG-CoA reductase inhibitors; CA
2,227,112 discloses the use of NHE-1 inhibitors with
sodium-dependent bicarbonate/chloride exchanger (NCBE) inhibitors;
CA 2,245,776 discloses the use of NHE-1 inhibitors with, inter
alia, .beta.-receptor blockers, calcium antagonists, loop
diuretics, thiazide diuretics, potassium-sparing diuretics,
aldosterone antagonists, cardiac glycosides, antiarrythmics,
K.sub.ATP channel openers, K.sub.ATP channel blockers, and
veratride-activatable sodium channel inhibitors; and commonly
assigned PCT International Application Publication No. WO 99/43663
discloses the use of NHE-1 inhibitors with, inter alia, adenosine,
adenosine agonists, nitrates, platelet inhibitors, aspirin,
dipyridamol, potassium chloride, clonidine, prazosin, and adenosine
A.sub.3-receptor agonists.
[0007] In accordance with the practice of the kits, methods and
pharmaceutical compositions of the instant invention, it is
believed that the administration of a combination of an NHE-1
inhibitor and a second compound selected from the group consisting
of: (a) a complement modulator, (b) a metabolic modulator, (c) an
anti-apoptotic agent, (d) a nitric oxide synthase-related agent,
and (e) an enzyme/protein modulator selected from the group
consisting of a protein kinase C activator, an endothelin
converting enzyme inhibitor, a tissue-activated fibrinolytic
inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1)
inhibitor, and a poly (ADP ribose) synthetase (PARS/PARP)
inhibitor, will afford greater therapeutic advantages than either
of the combination components administered alone.
SUMMARY OF THE INVENTION
[0008] The invention provdes methods of reducing tissue damage
resulting from ischemia which comprise administering to a mammal in
need of such reduction an effective amount of a combination, or a
pharmaceutical composition comprising such combination, of an NHE-1
inhibitor and a second compound selected from the group consisting
of: (a) a complement modulator, (b) a metabolic modulator, (c) an
anti-apoptotic agent, (d) a nitric oxide synthase-related agent,
and (e) an enzyme/protein modulator.
[0009] The invention further provides pharmaceutical compositions
comprising an amount of an NHE-1 inhibitor; an amount of a second
compound selected from the group consisting of (a) a complement
modulator, (b) a metabolic modulator, (c) an anti-apoptotic agent,
(d) a nitric oxide synthase-related agent, and (e) an
enzyme/protein modulator selected from the group consisting of a
protein kinase C activator, an endothelin converting enzyme
inhibitor, a tissue-activated fibrinolytic inhibitor (TAFI), a
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor, and a
poly (ADP ribose) synthetase (PARS/PARP) inhibitor; and,
preferably, a pharmaceutically acceptable pharmaceutically
acceptable carrier, vehicle, or diluent.
[0010] The invention further provides a kit comprising an amount of
a sodium-hydrogen exchanger type-1 inhibitor, and a
pharmaceutically acceptable carrier, vehicle, or diluent in a first
unit dosage form; an amount of a second compound selected from the
group consisting of (a) a complement modulator, (b) a metabolic
modulator, (c) an anti-apoptotic agent, (d) a nitric oxide
synthase-related agent, and (e) an enzyme/protein modulator
selected from the group consisting of a protein kinase C activator,
an endothelin converting enzyme inhibitor, a tissue-activated
fibrinolytic inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger
isoform-1 (NCX-1) inhibitor, and a poly (ADP ribose) synthetase
(PARS/PARP) inhibitor, and a pharmaceutically acceptable carrier,
vehicle, or diluent in a second unit dosage form; and a
container.
DETAILED DESCRIPTION OF THE INVENTION
[0011] The invention provides methods of reducing tissue damage
(e.g., substantially preventing tissue damage and/or inducing
tissue protection) resulting from ischemia which methods comprise
administering to a mammal (e.g., human male or female) in need of
such reduction a therapeutically effective amount of a combination,
or a pharmaceutical composition comprising such combination, of a
sodium-hydrogen exchanger type 1 (NHE-1) inhibitor, and a second
compound selected from the group consisting of (a) a complement
modulator, (b) a metabolic modulator, (c) an anti-apoptotic agent,
(d) a nitric oxide synthase-related agent, and (e) an
enzyme/protein modulator selected from the group consisting of a
protein kinase C activator, an endothelin converting enzyme
inhibitor, a tissue-activated fibrinolytic inhibitor (TAFI), a
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor, and a
poly (ADP ribose) synthetase (PARS/PARP) inhibitor.
[0012] Preferred ischemic tissues that may be treated in accordance
with the methods of the present invention comprise those tissues
selected from the group consisting of brain, cardiac, liver,
kidney, lung, gut, skeletal muscle, spleen, pancreas, nerve, spinal
cord, retinal tissue, vasculature, and intestinal tissue. An
especially preferred tissue comprises cardiac tissue.
[0013] Although any NHE-1 inhibitor may be employed in the methods
and pharmaceutical compositions of the present invention, it is
generally preferred that such inhibitor be selected from the group
consisting of:
[0014] (a) a compound of Formula (I) 1
[0015] a prodrug thereof, or a pharmaceutically acceptable salt of
the compound or the prodrug thereof; wherein:
[0016] Z is carbon connected and is a five-membered, diaza,
diunsaturated ring having two contiguous nitrogens, said ring
optionally mono-, di-, or tri-substituted with up to three
substituents independently selected from R.sup.1, R.sup.2 and
R.sup.3;
[0017] or
[0018] Z is carbon connected and is a five-membered, triaza,
diunsaturated ring, said ring optionally mono- or di-substituted
with up to two substituents independently selected from R.sup.4 and
R.sup.5;
[0019] wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4 and R.sup.5 are
each independently hydrogen, hydroxy(C.sub.1-C.sub.4)alkyl,
(C.sub.1-C.sub.4)alkyl, (C.sub.1-C.sub.4)alkylthio,
(C.sub.3-C.sub.4)cycloalkyl,
(C.sub.3-C.sub.7)cycloalkyl(C.sub.1-C.sub.4)- alkyl,
(C.sub.1-C.sub.4)alkoxy,
(C.sub.1-C.sub.4)alkoxy(C.sub.1-C.sub.4)al- kyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl, M or
M(C.sub.1-C.sub.4)alkyl, any of said previous
(C.sub.1-C.sub.4)alkyl moieties optionally having from one to nine
fluorines; said (C.sub.1-C.sub.4)alkyl or
(C.sub.3-C.sub.4)cycloalkyl optionally mono-or di-substituted
independently with hydroxy, (C.sub.1-C.sub.4)alkoxy,
(C.sub.1-C.sub.4)alklthio, (C.sub.1-C.sub.4)alkylsulfinyl,
(C.sub.1-C.sub.4)alkylsulfonyl, (C.sub.1-C.sub.4)alkyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl or mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylaminosulfonyl; and said
(C.sub.3-C.sub.4)cycloalkyl optionally having from one to seven
fluorines;
[0020] wherein M is a partially saturated, fully saturated or fully
unsaturated five to eight membered ring optionally having one to
three heteroatoms selected independently from oxygen, sulfur and
nitrogen, or, a bicyclic ring consisting of two fused partially
saturated, fully saturated or fully unsaturated three to six
membered rings, taken independently, optionally having one to four
heteroatoms selected independently from nitrogen, sulfur and
oxygen;
[0021] said M is optionally substituted, on one ring if the moiety
is monocyclic, or one or both rings if the moiety is bicyclic, on
carbon or nitrogen with up to three substituents independently
selected from R.sup.6, R.sup.7 and R.sup.8, wherein one of R.sup.6,
R.sup.7 and R.sup.8 is optionally a partially saturated, fully
saturated, or fully unsaturated three to seven membered ring
optionally having one to three heteroatoms selected independently
from oxygen, sulfur and nitrogen optionally substituted with
(C.sub.1-C.sub.4)alkyl and additionally R.sup.6, R.sup.7 and
R.sup.8 are optionally hydroxy, nitro, halo,
(C.sub.1-C.sub.4)alkoxy, (C.sub.1-C.sub.4)alkoxycarbonyl,
(C.sub.1-C.sub.4)alkyl, formyl, (C.sub.1-C.sub.4)alkanoyl,
(C.sub.1-C.sub.4)alkanoyloxy, (C.sub.1-C.sub.4)alkanoylamino,
(C.sub.1-C.sub.4)alkoxycarbonylamino, sulfonamido,
(C.sub.1-C.sub.4)alkylsulfonamido, amino, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylamino, carbamoyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl, cyano, thiol,
(C.sub.1-C.sub.4)alkylthio, (C.sub.1-C.sub.4)alkylsulfinyl,
(C.sub.1-C.sub.4)alkylsulfonyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkyla- minosulfonyl,
(C.sub.2-C.sub.4)alkenyl, (C.sub.2-C.sub.4)alkynyl or
(C.sub.5-C.sub.7)cycloalkenyl,
[0022] wherein said (C.sub.1-C.sub.4)alkoxy,
(C.sub.1-C.sub.4)alkyl, (C.sub.1-C.sub.7)alkanoyl,
(C.sub.1-C.sub.4)alkylthio, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylamino or (C.sub.3-C.sub.7)cycloalkyl
R.sup.6, R.sup.7 and R.sup.8 substituents are optionally mono-
substituted independently with hydroxy,
(C.sub.1-C.sub.4)alkoxycarbonyl, (C.sub.3-C.sub.7)cycloalkyl,
(C.sub.1-C.sub.4)alkanoyl, (C.sub.1-C.sub.4)alkanoylamino,
(C.sub.1-C.sub.4)alkanoyloxy, (C.sub.1-C.sub.4)alkoxycarbonylamino,
sulfonamido, (C.sub.1-C.sub.4)alkylsulfonamido, amino, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylamino, carbamoyl, mono-N- or
di-N,N-(C.sub.1-C.sub.4)alkylcarbamoyl, cyano, thiol, nitro,
(C.sub.1-C.sub.4)alkylthio, (C.sub.1-C.sub.4)alkylsulfinyl,
(C.sub.1-C.sub.4)alkylsulfonyl or mono-N- or
di-N,N-(C.sub.1-C.sub.4)alky- laminosulfonyl or optionally
substituted with one to nine fluorines;
[0023] (b) cariporide, or a pharmaceutically acceptable salt
thereof;
[0024] (c) eniporide, or a pharmaceutically acceptable salt
thereof;
[0025] (d) BIIB 513, or a pharmaceutically acceptable salt
thereof;
[0026] (e) TY-1 2533, or a pharmaceutically acceptable salt
thereof; and
[0027] (f) SM-15681, or a pharmaceutically acceptable salt
thereof.
[0028] Especially preferred Formula (I) compounds are those
compounds selected from the group consisting of:
[0029]
[1-(2-chlorophenyl)-5-methyl-1H-pyrazole-4-carbonyl]guanidine;
[0030]
[5-methyl-1-(2-trifluoromethylphenyl)-1H-pyrazole-4-carbonyl]guanid-
ine;
[0031] [5-ethyl-1-phenyl-1H-pyrazole-4-carbonyl]guanidine;
[0032]
[5-cyclopropyl-1-(2-trifluoromethylphenyl)-1H-pyrazole-4-carbonyl]g-
uanidine;
[0033]
[5-cyclopropyl-1-phenyl-1H-pyrazole-4-carbonyl]guanidine;
[0034]
[5-cyclopropyl-1-(2,6-dichlorophenyl)-1H-pyrazole-4-carbonyl]guanid-
ine;
[0035] [5-methyl-1-(quinoli
n-6-yl)-1H-pyrazole-4-carbonyl]guanidine;
[0036]
[5-methyl-1-(naphthalen-1-yl)-1H-pyrazole-4-carbonyl]guanidine;
[0037]
[5-cyclopropyl-1-(quinolin-5-yl)-1H-pyrazole-4-carbonyl]guanidine;
[0038]
[5-cyclopropyl-1-(quinolin-8-yl)-1H-pyrazole-4-carbonyl]guanidine;
[0039] [3-methyl-1-phenyl-1H-pyrazole-4-carbonyl]guanidine;
[0040]
[3-methyl-1-(naphthalen-1-yl)-1H-pyrazole-4-carbonyl]guanidine;
[0041]
[3-methyl-1-(isoquinolin-5-yl)-1H-pyrazole-4-carbonyl]guanidine;
[0042] [2-methyl-5-phenyl-2H-pyrazole-3-carbonyl]guanidine;
[0043]
[2-methyl-5-(naphthalen-1-yl)-2H-pyrazole-3-carbonyl]guanidine;
[0044]
[5-methyl-2-phenyl-2H-1,2,3-triazole-4-carbonyl]guanidine;
[0045]
[5-methyl-2-(3-methoxyphenyl)-2H-1,2,3-triazole-4-carbonyl]guanidin-
e;
[0046]
[2-(3-bromophenyl)-5-methyl-2H-1,2,3-triazole-4-carbonyl]guanidine;
[0047]
[2-(naphthalen-1-yl)-5-methyl-2H-1,2,3-triazole-4-carbonyl]guanidin-
e;
[0048] [2-(isoquinolin-5-yl)-5-methyl -2H-1,2,3-triazole-4-
carbonyl]guanidine;
[0049] [5-methyl-2-(quinolin-5-yl)-2H-
1,2,3-triazole-4-carbonyl]guanidine- ;
[0050]
[1-(naphthalen-1-yl)-5-cyclopropyl-2H-pyrazole-4-carbonyl]guanidine-
;
[0051]
[1-(2-chloro-4-methylsulfonylphenyl)-5-cyclopropyl-1H-pyrazole-4-ca-
rbonyl]guanidine;
[0052]
[1-(2-chlorophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanidine;
[0053]
[1-(2-trifluoromethyl-4-fluorophenyl)-5-cyclopropyl-1H-pyrazole-4-c-
arbonyl]guanidine;
[0054]
[1-(2-bromophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanidine;
[0055]
[1-(2-fluorophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanidine;
[0056]
[1-(2-chloro-5-methoxyphenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]-
guanidine;
[0057]
[1-(2-chloro-4-methylaminosulfonylphenyl)-5-cyclopropyl-1H-pyrazole-
-4-carbonyl]guanidine;
[0058]
[1-(2,5-dichlorophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanid-
ine;
[0059]
[1-(2,3-dichlorophenyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanid-
ine;
[0060]
[1-(2-chloro-5-aminocarbonylphenyl)-5-cyclopropyl-1H-pyrazole-4-car-
bonyl]guanidine;
[0061]
[1-(2-chloro-5-aminosulfonylphenyl)-5-cyclopropyl-1H-pyrazole-4-car-
bonyl]guanidine;
[0062]
[1-(2-fluoro-6-trifluoromethylphenyl)-5-cyclopropyl-1H-pyrazole-4-c-
arbonyl]guanidine;
[0063]
[1-(2-chloro-5-methylsulfonylphenyl)-5-cyclopropyl-1H-pyrazole-4-ca-
rbonyl]guanidine;
[0064]
[1-(2-chloro-5-dimethylaminosulfonylphenyl)-5-cyclopropyl-1H-pyrazo-
le-4-carbonyl]guanidine;
[0065]
[1-(2-trifluoromethyl-4-chlorophenyl)-5-cyclopropyl-1H-pyrazole-4-c-
arbonyl]guanidine;
[0066]
[1-(8-bromoquinolin-5-yI)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guan-
idine;
[0067]
[1-(6-chloroquinolin-5-yI)-5-cyclopropyl-1H-pyrazole4-carbonyl]guan-
idine;
[0068]
[1-(indazol-7-yl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanidine;
[0069]
[1-(benzimidazol-5-yl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanidi-
ne;
[0070]
[1-(1-isoquinolyl)-5-cyclopropyl-1H-pyrazole-4-carbonyl]guanidine;
[0071]
[5-cyclopropyl-1-(4-quinolinyl)-1H-pyrazole-4-carbonyl]guanidine;
[0072]
[1-(indazol-6-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[0073]
[1-(indazol-5-yI)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[0074]
[1-(benzimidazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[0075]
[1-(1-methylbenzimidazol-6-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guani-
dine
[0076]
[1-(5-quinolinyl)-5-n-propyl-1H-pyrazole-4-carbonyl]guanidine;
[0077]
[1-(5-quinolinyl)-5-isopropyl-1H-pyrazole-4-carbonyl]guanidine;
[0078]
[5-ethyl-1-(6-quinolinyl)-1H-pyrazole4-carbonyl]guanidine;
[0079]
[1-(2-methylbenzimidazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guani-
dine;
[0080] [1-(1
,4-benzodioxan-6-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine-
;
[0081]
[1-(benzotriazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[0082]
[1-(3-chloroindazol-5-yl)-5-ethyl-1H-pyrazole-4-carbonyl]guanidine;
[0083]
[1-(5-quinolinyl)-5-butyl-1H-pyrazole-4-carbonyl]guanidine;
[0084] [5-propyl-1-(6-quinolinyl)-1H-pyrazole-4-carbonyl]g
uanidine;
[0085]
[5-isopropyl-1-(6-quinolinyl)-1H-pyrazole-4-carbonyl]guanidine;
[0086]
[1-(indazol-7-yl)-3-methyl-1H-pyrazole-4-carbonyl]guanidine;
[0087]
[1-(2,1,3-benzothiadiazol-4-yl)-3-methyl-1H-pyrazole-4-carbonyl]gua-
nidine;
[0088] and
[0089]
[3-methyl-1-(quinolin-5-yl)-1H-pyrazole-4-carbonyl]guanidine; the
prodrugs thereof, and the pharmaceutically acceptable salts of the
compounds, and the prodrugs.
[0090] The compounds of Formula (I), the prodrugs thereof, and the
pharmaceutically acceptable salts of the compounds and prodrugs,
may be prepared as disclosed in the aforementioned,
commonly-assigned PCT International Application Publication No. WO
99/43663, the disclosure of which is incorporated herein by
reference.
[0091] The preferred NHE-1 inhibitor cariporide, i.e.
N-(aminoiminomethyl)-4-(1-methylethyl)-3-(methylsulfonyl)-benzamide,
may be prepared as disclosed in U.S. Pat. No. 5,591,754, the
disclosure of which is incorporated herein by reference. The
preferred NHE-1 inhibitor eniporide, i.e.
N-(aminoiminomethyl)-2-methyl-5-(methylsulfonyl)-4-(1H-py-
rrol-1-yl)-benzamide, may be prepared as disclosed in U.S. Pat. No.
5,753,680, the disclosure of which is incorporated herein by
reference. The preferred NHE-1 inhibitor BIIB-513, i.e.
N-(aminoiminomethyl)-4-(4-(2-
-furanylcarbonyl)-1-piperazinyl)-3-(methylsulfonyl)-benzamide, may
be prepared as disclosed in U.S. Pat. No. 6,114,335, the disclosure
of which is incorporated herein by reference. The preferred NHE-1
inhibitor TY-12533, i.e.
6,7,8,9-tetrahydro-2-methyl-5H-cyclohepta[b]pyridine-3-car-
bonylguanidine, may be prepared as disclosed in PCT International
Application Publication No. WO 98/39300, the disclosure of which is
incorporated herein by reference. The preferred NHE-1 inhibitor
SM-15681, i.e.
N-(aminoiminomethyl)-1-methyl-1H-indole-2-carboxamide, may be
prepared as disclosed in EPO 0 708 091, the disclosure of which is
incorporated herein by reference.
[0092] The ability of a compound to function as an NHE-1 inhibitor
may be determined according to the protocols described in detail
hereinbelow.
[0093] A complement modulator refers generally to an agent that
modulates, i.e., regulates or inhibits, certain thermolabile
substances, normally present in the serum, that are destructive to
certain bacteria and other cells sensitized by a specific
complement-fixing antibody known as antibody C. Antibody C
comprises a group of at least 20 disparate serum proteins, the
activity of which is affected by a series of interactions resulting
in enzymatic cleavages which can follow at least two distinct
pathways. The complement pathways contribute to myocardial
ischemia-reperfusion injury in vivo, ostensibly via a combination
of mechanisms including, but not limited to, stimulation of
cytokine release from various cell types, expression of adhesion
molecules, and neutrophil infiltration, of which all mechanisms
directly elicit cell death. Accordingly, it is believed that the
administration of a combination of an NHE-1 inhibitor and a
complement modulator will afford greater protection from tissue
damage resulting from ischemia than either agent administered
alone.
[0094] Although any complement modulator may be employed in the
methods and pharmaceutical compositions of the instant invention,
it is generally preferred that such complement modulator be
selected from the group consisting of a C5a inhibitor, preferably
L-747981, or a pharmaceutically acceptable salt thereof, a soluble
complement receptor type 1 (sCR1) inhibitor, or an analog thereof,
preferably amidinophenylpyruvic acid (APPA), and a Cl esterase
inhibitor.
[0095] The preferred C5a inhibitor L-747981, the chemical structure
of which is shown hereinbelow, and the pharmaceutically acceptable
salts thereof, may be prepared according to known synthetic
methods. 2
[0096] The ability of L-747981 to selectively bind to the C5a
receptor is disclosed in Flanagan, K. L., et al., ACS, 210th
Chicago: MEDI 085 (1995). The preferred soluble complement receptor
type 1 (sCR1) inhibitor amidinophenylpyruvic acid (APPA), may be
obtained commercially.
[0097] The ability of a compound to inhibit C5a may be determined
according to the methodology set forth in Vakeva, A. P., et al.,
Circulation, 97 (22), 2259-67 (1998). The ability of a compound to
inhibit complement activation at the sCR1 receptor may be
determined according to the protocols described by Rittershaus,
C.W., J. Biol. Chem., 274 (16), 11237-44 (1999). The ability of a
compound to inhibit Cl esterase may be determined according to the
methodology disclosed in Benny, A. G., et al., Haematologia,
22(3),189-93 (1989).
[0098] Generally, the term metabolic modulator refers generally to
any agent that serves to modulate, i.e., regulate, stimulate, or
inhibit, one or more metabolic pathways. With respect to the
methods and pharmaceutical compositions of the present invention,
metabolic modulators such as pyruvate dehydrogenase kinase
inhibitors, for example, dichloroacetate (DCA), activate the
myocardial dehydrogenase (PDC) complex, thus increasing glucose
oxidation and decreasing fatty acid oxidation in the ischemic
myocardium. Accordingly, reduction of ischemic tissue damage by
treatment with a combination of an NHE-1 inhibitor and a metabolic
modulator should elicit additional cardioprotective benefit.
[0099] Although any metabolic modulator may be employed in the
methods and pharmaceutical compositions of the instant invention,
it is generally preferred that such modulator be selected from the
group consisting of a pyruvate dehydrogenase complex
up-regulator/activator, preferably dichloroacetate (DCA), a
pyruvate dehydrogenase kinase inhibitor, preferably DCA, a malonyl
CoA decarboxylase inhibitor, an acetyl CoA carboxylase activator, a
partial fatty acid oxidation (pFOX) inhibitor, preferably
ranolazine or trimetazidine, a 5' AMP-activated protein kinase
(AMPK) inhibitor, a carnitine palmitoyl transferase inhibitor,
preferably etomoxir, and a fatty acid CoA synthase inhibitor,
preferably triascin C. The preferred pFOX inhibitors ranolazine and
trimetazidine may be prepared as disclosed in U.S. Pat. Nos.
4,567,264, and 4,663,325 respectively, which patents are
incorporated herein in their entirety by reference. The preferred
carnitine palmitoyl transferase inhibitor etomoxir may be prepared
as disclosed in U.S. Pat. No. 4,337,267, the disclosure of which is
incorporated herein by reference. The preferred fatty acid CoA
synthase inhibitor triascin C, i.e.
(2E,4E,7E)-undecatrienal-nitrosohydrazone, may be prepared as
disclosed in U.S. Pat. No. 4,297,096, the disclosure of which is
incorporated herein by reference.
[0100] The ability of a compound to function as a pyruvate
dehydrogenase complex up-regulator/activator, a pyruvate
dehydrogenase kinase inhibitor, or a malonyl CoA decarboxylase
inhibitor, may be determined according to the methodologies
disclosed in Stanley, W. C., et al., J. Mol. Cell. Cardiol., 28,
905-914 (1996). The ability of a compound to function as an acetyl
CoA carboxylase activator may be determined according to the
protocols described in Belke, D. D., et al., Biochem. Biophys.
Acta, 1391 (1), 25-36 (1998). The ability of a compound to function
as a partial fatty acid oxidation (pFOX) inhibitor may be
determined according to the procedures of McCormack J. G., et al.,
J. Appl. Physiol., 81/2, 905-910 (1996), or Merrill G. F., et al.,
Am. J. Physiol., 273, E1107-1112 (1997). The ability of a compound
to function as a carnitine palmitoyl transferase inhibitor may be
determined according to the methodology disclosed in Kudo, et al.,
J. Biol. Chem., 270, 17513-17520 (1995). The ability of a compound
to function as a 5' AMP-activated protein kinase (AMPK) inhibitor
may be determined according to Haystead T. A., et al., Eur. J.
Biochem., 187, 199-205 (1990). The ability of a compound to
function as a fatty acid CoA synthase inhibitor may be determined
according to the following systems.
[0101] Oxidase/Catalase Coupling System: 3
[0102] A reduction in absorbance at 620 nm indicates fatty acid CoA
synthase inhibition.
[0103] Generally, an anti-apoptotic agent inhibits apoptosis, i.e.,
programmed cell death, the process by which certain cells
self-destruct by fragmentation into membrane-bound particles which
are subsequently phagocytized by other cells, for example,
macrophages. Because apoptosis is known to occur during myocardial
ischemia-reperfusion injury, it is believed that enzymatic
inhibition of the apoptotic cascade with an anti-apoptic agent, in
combination with treatment with an NHE-1 inhibitor, will confer
greater protection from tissue damage resulting from ischemia than
either agent administered alone.
[0104] Although any anti-apoptotic agent may be employed in the
methods and pharmaceutical compositions of the present invention,
it is generally preferred that such anti-apoptotic agent comprise a
caspase inhibitor. The term caspase inhibitor refers to any agent
that inhibits the activity of caspases, a salient family of enzymes
involved in the induction of apoptosis in mammalian cells, for
example, excessive apoptosis of cardiac myocytes during
reperfusion, and neuronal cells during ischemia. Generally
preferred caspase inhibitors, useful in the methods and
pharmaceutical compositions of the instant invention, comprise
those compounds selected from the group consisting of a compound of
structural Formula (II) 4
[0105] wherein
[0106] R.sub.1 and R.sub.2, together with the nitrogen to which
they are attached, form a 4- to 7-membered ring; R.sub.3 and
R.sub.4, together with the nitrogen atom to which they are
attached, form a 4- to 7-membered ring; and R is benzoyl, or
(C.sub.1-6)alkyl; and
[0107] a compound of structural Formula (III) 5
[0108] wherein
[0109] R.sub.1 is hydrogen, or (C.sub.1-4)alkyl; R.sub.2 is
(C.sub.1-10)alkyl, optionally substituted with
aryl(C.sub.1-4)alkyl, optionally substituted
heteroaryl(C.sub.1-4)alkyl, optionally substituted
(C.sub.3-7)cycloalkyl, or R.sub.1 and R.sub.2, together with the
nitrogen to which they are attached, form a 3- to 10-membered ring
which optionally contains an additional heteroatom selected from
oxygen, nitrogen, or sulfur; R.sub.3 and R.sub.4 are
(C.sub.1-6)alkyl, hydrogen, nitro, or halogen; and R.sub.5 is
(C.sub.1-6)alkyl, hydrogen, arylalkyl, or heteroarylalkyl.
[0110] The preferred caspase inhibitors of structural Formula (II)
may be prepared as described in PCT International Application
Publication No. WO 99/65451. The preferred caspase inhibitors of
structural Formula (III) may be prepared as described in PCT
International Application Publication No. WO 99/06367.
[0111] The ability of a compound to inhibit apoptosis may be
determined according to the methodology of Wu, J. C., et al.,
Methods, 17 (4), 320-8 (1999). The ability of an agent to function
as a caspase inhibitor may be determined according to the
methodologies disclosed in the aforementioned PCT International
Application Publication Nos. WO 99/06367 and WO 99-65451.
[0112] The term nitric oxide synthase-related agent, as employed
within the context of the instant invention, refers generally to
any agent that regulates, i.e., inhibits, promotes, or enhances,
the enzymatic formation of nitric oxide (NO) free-radical, a known
mediator of cell-to-cell communication and potent vasodilator,
which free-radical is produced by the nitric oxide
synthase-catalyzed reaction of L-arginine with 2 O.sub.2 and 1.5
NADPH. Because certain agents increase the expression of inducible
nitric oxide synthase (iNOS, i.e., Type 3), the nitric oxide
produced by this enzyme, or that generated by nitric oxide donors,
is currently believed to be cardioprotective. Furthermore, NO
produced acutely by endothelial NOS (Type 2), or neuronal NOS (Type
1) during ischemia-reperfusion injury is also believed to be
cardioprotective. For detailed discussions of the cardioprotective
effects of NO produced by nitric oxide donors, see, for example,
Takano, H., et al., Circ. Res., 83, 73-84 (1998) and Pabla, R., et
al., Heart Circ. Physiol. 38, H1113-1121 (1995). In direct
contrast, it has been further disclosed that the inhibition of NO,
through the activity of NOS inhibitors, also affords protection
against ischemic injury. See, for example, Depre, C., et al.,
Circulation, 92, 1911-1918 (1995) and Woolfson, R. G., et al.,
Circulation, 91, 1545-1551 (1995). Accordingly, it is believed that
treatment with a nitric oxide synthase-related agent, in
combination with an NHE-1 inhibitor, will confer greater protection
from tissue damage resulting from ischemia than either agent
administered alone.
[0113] Although any nitric oxide synthase-related agent may be
employed in the methods and pharmaceutical compositions of the
present invention, it is generally preferred that such agent be
selected from the group consisting of monophosphoryl lipid A, or an
analog thereof, preferably RC-552 (MPL-C) or ONO-4007, a nitric
oxide donor, preferably nipride, and a nitric oxide synthase
inhibitor, preferably aminoguanidine or N(G)-monomethyl-L-arginine
(L-NMMA).
[0114] The preferred monophosphoryl lipid A analog ONO-4077, i.e.
(S)-2-deoxy-2-((1-oxo-3-((1-oxo-9-phenylnonyl)oxy)-tetradecyl)amino)-,
3-benzenenonanoate 4-(hydrogen sulfate)-D-glucose, may be prepared
as disclosed in U.S. Pat. Nos. 5,294,723 and 5,733,927, the
disclosures of which are incorporated herein by reference. The
preferred nitric oxide donor nipride, i.e., sodium
nitrosylpentacyanoferrate (II), may be prepared as described in
Playfair, L., Proc. Roy. Soc. London 5, 846 (1849). The preferred
nitric oxide synthase inhibitor aminoguanidine may be prepared as
described in Smith, G. B. L., et al., J. Am. Chem. Soc., 57, 2730
(1935). The preferred nitric oxide inhibitor
N(G)-monomethyl-L-arginine (L-NMMA) may be obtained from commercial
sources.
[0115] The ability of an agent to function as a nitric oxide donor,
nitric oxide synthase activator, or nitric oxide synthase inhibitor
may be determined according to the methods disclosed in Rees, D.
D., et al., British Journal of Pharmacology, 101, 746-52 (1990),
and Archer, S., FASEB Journal, 7, 349-60 (1999).
[0116] In the practice of the methods and pharmaceutical
compositions of the invention, the NHE-1 inhibitor may further be
employed in combination with an enzyme/protein modulator selected
from the group consisting of a protein kinase C .epsilon.
activator, an endothelin converting enzyme inhibitor, preferably
S-17162, a tissue-activated fibrinolytic (TAFI), a
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor,
preferably KB-R7943, and a poly (ADP ribose) synthetase (PARS/PARP)
inhibitor, preferably 3-aminobenzamide, or a compound, or a
pharmaceutically acceptable salt, prodrug, active metabolite, or
solvate thereof, of Formula (IV) 6
[0117] wherein
[0118] R.sup.1 is H, halogen, cyano, an optionally substituted
alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or
heteroaryl group;
[0119] or --C(O)--R.sup.10, where R.sup.10 is hydrogen, an
optionally substituted alkyl, alkenyl, alkynyl, cycloalkyl,
heterocycloalkyl, aryl, or heteroaryl group;
[0120] or --OR.sup.100 or NR.sup.100R.sup.110, where R.sup.100 and
R.sup.110 are each independently H or an optionally substituted
alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or
heteroaryl group;
[0121] R.sup.2 is H or alkyl;
[0122] R.sup.3 is H or alkyl;
[0123] R.sup.4 is H, halogen or alkyl;
[0124] X is O or S; and
[0125] Y is (CR.sup.5R.sup.6)(CR.sup.7R.sup.8)n or
N.dbd.C(R.sup.5), where n is 0 or 1; R.sup.5 and R.sup.6 are each
independently H or an optionally substituted alkyl, alkenyl,
alkynyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl group;
and R.sup.7 and R.sup.8 are each independently H or an optionally
substituted alky, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,
aryl, or heteroaryl group.
[0126] The term protein kinase C .epsilon. refers to the
cytoplasmic, calcium-activated family of kinases that are believed
to be downstream mediators of, inter alia, the ischemic
pre-conditioning pathway. Accordingly, because activators of this
enzyme are cardioprotective, reduction of ischemic tissue damage by
treatment with a combination of an NHE-1 inhibitor and a protein
kinase C .epsilon. activator should provide additional
cardioprotective benefit. Endothelin is known to contribute to
myocardial ischemia-reperfusion injury and, therefore, by
preventing endothelin production with an endothelin converting
enzyme inhibitor, reduction of ischemic tissue damage by treatment
with a combination of an NHE-1 inhibitor and an endothelin
converting enzyme inhibitor should provide additional
cardioprotective advantages. Tissue-activated fibrinolytic
inhibitors are known to be useful in the treatment of both deep
venous thrombosis (DVT) and acute coronary syndrome (ACS), a
syndrome that embraces, inter alia, ischemic attack. Accordingly,
reduction of ischemic tissue damage by treatment with a combination
of an NHE-1 inhibitor and a TAFI inhibitor should elicit additional
cardioprotective benefit. During cardiac reperfusion,
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) increases
intracellular calcium levels, due to the increased intracellular
sodium levels resulting from NHE-1 activity, which leads to
contracture, arrhythmias, and cellular death. Thus, inhibition of
NCX-1 is cardioprotective and, therefore, reduction of ischemic
tissue damage by treatment with a combination of an NHE-1 inhibitor
and an NCX-1 inhibitor should elicit additional cardioprotective
benefit. PARS/PARP is a DNA repair enzyme that is activated during
myocardial ischemia/reperfusion injury in response to DNA single
strand breaks. The enzyme consumes intracellular NAD.sup.+and ATP
pools, and slows the rate of glycolysis and mitochondrial
respiration which can contribute to, or directly cause,
cardiomyocyte dysfunction and/or death. Accordingly, PARS/PARP
inhibitors are believed to be cardioprotective and, therefore,
reduction of ischemic tissue damage by treatment with a combination
of an NHE-1 inhibitor and a PARS/PARP inhibitor should elicit
additional cardioprotective benefit.
[0127] The preferred endothelin converting enzyme inhibitor
S-17162, i.e. N-(2,3-dihydroxypropylphosphonyl-(S)-Leu-(S)-Trp-OH,
disodium salt, may be prepared as disclosed in U.S. Pat. Nos.
5,481,030, 5,591,728, and 5,608,045, the disclosures of which are
hereby incorporated by reference in their entirety. The preferred
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor KB-R7943,
i.e., (2-(2-(4-(4-nitrobenzyloxy)-phenyl)-eth- yl)-isothiourea
methanesulfonate), may be prepared as described in PCT
International Application Publication No. WO 97/09306. The
preferred poly (ADP ribose) synthetase (PARS/PARP) inhibitor
3-aminobenzamide is available commercially. The preferred poly (ADP
ribose) synthetase (PARS/PARP) inhibitors of Formula (IV) may be
prepared as disclosed in PCT International Application Publication
No. WO 2000/42040.
[0128] The ability of an agent to function as a protein kinase C
.epsilon. activator may be determined according to the protocol
disclosed in Bowling, N., et al., Circulation, 99, 384-91 (1999).
The ability of an agent to function as an endothelin converting
enzyme inhibitor may be determined according to Fassina, G., et
al., Peptide Res., 6, 73-78 (1993). The ability of an agent to
function as a tissue-activated fibrinolytic inhibitor (TAFI)
inhibitor may be determined according to Bajzar, L., et al., J.
Biol. Chem., 270, 14477-14484 (1995). The ability of an agent to
function as a Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1)
inhibitor may be determined according to the methodology described
by Iwamoto, T., et al., Am. J. Physiol., 275, C423-430 (1998). The
ability of an agent to function as a poly (ADP ribose) synthetase
(PARS/PARP) inhibitor may be determined according to the
methodologies of Decker, P., et al., Clinical Cancer Research, 5,
1169-1172 (1999).
[0129] Generally, the NHE-1 inhibitor can be administered to a
mammal at dosage levels in the range of from about 0.001 to about
100 mg/kg body weight per day. For a normal adult human having a
body weight of about 70 kg, a dosage in the range of from about
0.01 to about 50 mg/kg body weight is typically preferred. However,
some variability in these general dosage ranges may be required
depending upon the age and weight of the mammal being treated, the
intended route of administration, the particular agent being
administered, and the like. The determination of dosage ranges and
optimal dosages for a particular mammal is within the ability of
one of ordinary skill in the art having the benefit of the instant
disclosure.
[0130] The dosage of the complement modulator, metabolic modulator,
anti-apoptotic agent, nitric oxide synthase-related agent, or
enzyme/protein modulator will also be generally dependent upon a
number of factors including the health of the mammal being treated,
the extent of treatment desired, the nature and kind of concurrent
therapy, if any, and the frequency of treatment and the nature of
the effect desired. In general, dosage ranges of complement
modulators, metabolic modulators, anti-apoptotic agents, nitric
oxide synthase-related agents, and enzyme/protein modulators range
from about 0.001 to about 250 mg/kg body weight per day. For a
normal adult human having a body weight of about 70 kg, a dosage in
the range of from about 0.1 to about 25 mg/kg body weight is
typically preferred. However, some variability in this general
dosage range may be required depending upon the age and weight of
the subject being treated, the intended route of administration,
the particular agent being administered and the like. The
determination of dosage ranges and optimal dosages for a particular
mammal is also well within the ability of one of ordinary skill in
the art having the benefit of the instant disclosure.
[0131] According to the methods of the invention, the combination
of the NHE-1 inhibitor, and the second compound selected from the
group consisting of a complement modulator, a metabolic modulator,
an anti-apoptotic agent, a nitric oxide synthase-related agent, and
an enzyme/protein modulator selected from the group consisting of a
protein kinase C activator, an endothelin converting enzyme
inhibitor, a tissue-activated fibrinolytic inhibitor (TAFI), a
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor, and a
poly (ADP ribose) synthetase (PARS/PARP) inhibitor is administered
to a mammal in need of treatment therewith, preferably in the form
of a pharmaceutical composition. In the combination aspect of the
invention, the NHE-1 inhibitor, and the second compound selected
from the group consisting of a complement modulator, a metabolic
modulator, an anti-apoptotic agent, a nitric oxide synthase-related
agent, and an enzyme/protein modulator selected from the group
consisting of a protein kinase C activator, an endothelin
converting enzyme inhibitor, a tissue-activated fibrinolytic
inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1)
inhibitor, and a poly (ADP ribose) synthetase (PARS/PARP) inhibitor
may be administered either separately or in the pharmaceutical
composition comprising both. It is generally preferred that such
administration be oral. However, if the subject being treated is
unable to swallow, or oral administration is otherwise impaired or
undesirable, parenteral or transdermal administration will be
appropriate.
[0132] According to the methods of the invention, when the NHE-1
inhibitor, and the second compound selected from the group
consisting of a complement modulator, a metabolic modulator, an
anti-apoptotic agent, a nitric oxide synthase-related agent, and an
enzyme/protein modulator selected from the group consisting of a
protein kinase C activator, an endothelin converting enzyme
inhibitor, a tissue-activated fibrinolytic inhibitor (TAFI), a
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor, and a
poly (ADP ribose) synthetase (PARS/PARP) inhibitor are administered
together, such administration can be sequential in time or
simultaneous with the simultaneous method being generally
preferred. For sequential administration, the NHE-1 inhibitor, and
the second compound selected from the group consisting of a
complement modulator, a metabolic modulator, an anti-apoptotic
agent, a nitric oxide synthase-related agent, and an enzyme/protein
modulator selected from the group consisting of a protein kinase C
activator, an endothelin converting enzyme inhibitor, a
tissue-activated fibrinolytic inhibitor (TAFI), a
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor, and a
poly (ADP ribose) synthetase (PARS/PARP) inhibitor can be
administered in any order. It is generally preferred that such
administration be oral. It is especially preferred that such
administration be oral and simultaneous. When the NHE-1 inhibitor,
and the second compound selected from the group consisting of a
complement modulator, a metabolic modulator, an anti-apoptotic
agent, a nitric oxide synthase-related agent, and an enzyme/protein
modulator selected from the group consisting of a protein kinase C
activator, an endothelin converting enzyme inhibitor, a
tissue-activated fibrinolytic inhibitor (TAFI), a
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor, and a
poly (ADP ribose) synthetase (PARS/PARP) inhibitor are administered
sequentially, the administration of each can be by the same or by
different methods.
[0133] According to the methods of the invention, the combination
of the NHE-1 inhibitor, and the second compound selected from the
group consisting of a complement modulator, a metabolic modulator,
an anti-apoptotic agent, a nitric oxide synthase-related agent, and
an enzyme/protein modulator selected from the group consisting of a
protein kinase C activator, an endothelin converting enzyme
inhibitor, a tissue-activated fibrinolytic inhibitor (TAFI), a
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor, and a
poly (ADP ribose) synthetase (PARS/PARP) inhibitor is preferably
administered in the form of a pharmaceutical composition comprising
a pharmaceutically acceptable carrier, vehicle, or diluent.
Accordingly, the combination of the NHE-1 inhibitor, and the second
compound selected from the group consisting of a complement
modulator, a metabolic modulator, an anti-apoptotic agent, a nitric
oxide synthase-related agent, and an enzyme/protein modulator
selected from the group consisting of a protein kinase C activator,
an endothelin converting enzyme inhibitor, a tissue-activated
fibrinolytic inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger
isoform-1 (NCX-1) inhibitor, and a poly (ADP ribose) synthetase
(PARS/PARP) inhibitor can be administered to a patient separately
or together in any conventional oral, rectal, transdermal,
parenteral, (for example, intravenous, intramuscular, or
subcutaneous) intracisternal, intravaginal, intraperitoneal,
intravesical, local (for example, powder, ointment or drop), or
buccal, or nasal, dosage form .
[0134] Compositions suitable for parenteral injection may comprise
pharmaceutically acceptable sterile aqueous or nonaqueous
solutions, dispersions, suspensions, or emulsions, and sterile
powders for reconstitution into sterile injectable solutions or
dispersions. Examples of suitable aqueous and nonaqueous carriers,
diluents, solvents, or vehicles include water, ethanol, polyols
(propylene glycol, polyethylene glycol, glycerol, and the like),
suitable mixtures thereof, vegetable oils (such as olive oil) and
injectable organic esters such as ethyl oleate. Proper fluidity can
be maintained, for example, by the use of a coating such as
lecithin, by the maintenance of the required particle size in the
case of dispersions, and by the use of surfactants.
[0135] These compositions may also contain adjuvants such as
preserving, wetting, emulsifying, and dispersing agents. Prevention
of microorganism contamination of the compositions can be
accomplished with various antibacterial and antifungal agents, for
example, parabens, chlorobutanol, phenol, sorbic acid, and the
like. It may also be desirable to include isotonic agents, for
example, sugars, sodium chloride, and the like. Prolonged
absorption of injectable pharmaceutical compositions can be brought
about by the use of agents capable of delaying absorption, for
example, aluminum monostearate and gelatin.
[0136] Solid dosage forms for oral administration include capsules,
tablets, powders, and granules. In such solid dosage forms, the
active compound is admixed with at least one inert customary
pharmaceutical excipient (or carrier) such as sodium citrate or
dicalcium phosphate or (a) fillers or extenders, as for example,
starches, lactose, sucrose, mannitol, and silicic acid; (b)
binders, as for example, carboxymethylcellulose, alginates,
gelatin, polyvinylpyrrolidone, sucrose, and acacia; (c) humectants,
as for example, glycerol; (d) disintegrating agents, as for
example, agar-agar, calcium carbonate, potato or tapioca starch,
alginic acid, certain complex silicates, and sodium carbonate; (e)
solution retarders, as for example, paraffin; (f) absorption
accelerators, as for example, quaternary ammonium compounds; (g)
wetting agents, as for example, cetyl alcohol and glycerol
monostearate; (h) adsorbents, as for example, kaolin and bentonite;
and/or (i) lubricants, as for example, talc, calcium stearate,
magnesium stearate, solid polyethylene glycols, sodium lauryl
sulfate, or mixtures thereof. In the case of capsules and tablets,
the dosage forms may also comprise buffering agents.
[0137] Solid compositions of a similar type may also be used as
fillers in soft or hard filled gelatin capsules using such
excipients as lactose or milk sugar, as well as high molecular
weight polyethylene glycols, and the like.
[0138] Solid dosage forms such as tablets, dragees, capsules, and
granules can be prepared with coatings and shells, such as enteric
coatings and others well known in the art. They may also contain
opacifying agents, and can also be of such composition that they
release the active compound or compounds in a delayed manner.
Examples of embedding compositions that can be used are polymeric
substances and waxes. The active compounds can also be in
micro-encapsulated form, if appropriate, with one or more of the
above-mentioned excipients.
[0139] Liquid dosage forms for oral administration include
pharmaceutically acceptable emulsions, solutions, suspensions,
syrups, and elixirs. In addition to the active compounds, the
liquid dosage form may contain inert diluents commonly used in the
art, such as water or other solvents, solubilizing agents and
emulsifiers, as for example, ethyl alcohol, isopropyl alcohol,
ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate,
propylene glycol, 1,3-butylene glycol, dimethylformamide, oils, in
particular, cottonseed oil, groundnut oil, corn germ oil, olive
oil, castor oil, and sesame seed oil, glycerol, tetrahydrofurfuryl
alcohol, polyethylene glycols and fatty acid esters of sorbitan, or
mixtures of these substances, and the like.
[0140] Besides such inert diluents, the composition can also
include adjuvants, such as wetting agents, emulsifying and
suspending agents, sweetening, flavoring, and perfuming agents.
[0141] Suspensions, in addition to the active compound, may further
comprise suspending agents, as for example, ethoxylated isostearyl
alcohols, polyoxyethylene sorbitol and sorbitan esters,
microcrystalline cellulose, aluminum metahydroxide, bentonite,
agar-agar, and tragacanth, or mixtures of these substances, and the
like.
[0142] Compositions for rectal administration preferably comprise
suppositories, which can be prepared by mixing a compound of the
present invention with suitable non-irritating excipients or
carriers such as cocoa butter, polyethylene glycol or a suppository
wax, which are solid at ordinary room temperature, but liquid at
body temperature, and therefore, melt in the rectal cavity thereby
releasing the active component.
[0143] Dosage forms for topical administration of the NHE-1
inhibitor, and the second compound selected from the group
consisting of a complement modulator, a metabolic modulator, an
anti-apoptotic agent, a nitric oxide synthase-related agent, and an
enzyme/protein modulator selected from the group consisting of a
protein kinase C activator, an endothelin converting enzyme
inhibitor, a tissue-activated fibrinolytic inhibitor (TAFI), a
Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1) inhibitor, and a
poly (ADP ribose) synthetase (PARS/PARP) inhibitor may comprise
ointments, powders, sprays and inhalants. The active agent or
agents are admixed under sterile condition with a pharmaceutically
acceptable carrier, and any preservatives, buffers, or propellants
that may be required.
[0144] The combinations and pharmaceutical compositions comprising
the combinations of the present invention are useful as
therapeutants or prophylactic agents for diseases caused or
aggravated by the acceleration of the sodium/hydrogen (Na+/H+)
exchange transport system, for example, cardiovascular diseases
[e.g., arteriosclerosis, hypertension, arrhythmia (e.g. ischemic
arrhythmia, arrhythmia due to myocardial infarction, myocardial
stunning, myocardial dysfunction, arrhythmia after PTCA or after
thrombolysis, etc.), angina pectoris, cardiac hypertrophy,
myocardial infarction, heart failure (e.g., congestive heart
failure, acute heart failure, cardiac hypertrophy, etc.),
restenosis after PTCA, PTCI, shock (e.g., hemorrhagic shock,
endotoxin shock, etc.)], renal diseases (e.g., diabetes mellitus,
diabetic nephropathy, ischemic acute renal failure, etc.) organ
disorders associated with ischemia or ischemic reperfusion [(e.g.,
heart muscle ischemic reperfusion associated disorders, acute renal
failure, or disorders induced by surgical treatment such as
coronary artery bypass grafting (CABG) surgeries, vascular
surgeries, organ transplantation, non-cardiac surgeries or
percutaneous transluminal coronary angioplasty (PTCA)],
cerebrovascular diseases (e.g., ischemic stroke, hemorrhagic
stroke, etc.), cerebro ischemic disorders (e.g., disorders
associated with cerebral infarction, disorders caused after
cerebral apoplexy as sequelae, or cerebral edema. The combinations
and pharmaceutical compositions of this invention can also be used
as an agent for myocardial protection during coronary artery bypass
grafting (CABG) surgeries, vascular surgeries, percutaneous
transluminal coronary angioplasty (PTCA), PTCI, organ
transplantation, or non-cardiac surgeries.
[0145] Preferably, the combinations and pharmaceutical compositions
of this invention can be used to for myocardial protection before,
during, or after coronary artery bypass grafting (CABG) surgeries,
vascular surgeries, percutaneous transluminal coronary angioplasty
(PTCA), organ transplantation, or non-cardiac surgeries.
[0146] Preferably, the combinations and pharmaceutical compositions
of this invention can be used for myocardial protection in patients
presenting with ongoing cardiac (acute coronary syndromes, e.g.,
myocardial infarction or unstable angina) or cerebral ischemic
events (e.g., stroke).
[0147] Preferably, the combinations and pharmaceutical compositions
of this invention can be used for chronic myocardial protection in
patients with diagnosed coronary heart disease (e.g., previous
myocardial infarction or unstable angina) or patients who are at
high risk for myocardial infarction (age greater than 65 and two or
more risk factors for coronary heart disease).
[0148] The utility of the combinations and pharmaceutical
compositions of the present invention as medical agents in the
treatment of diseases, such as are detailed herein in mammals
(e.g., humans) for example, in patients presenting with ongoing
cardiac or cerebral ischemic events, or chronic cardioprotection in
patients with diagnosed coronary heart disease, or at risk for
coronary heart disease, cardiac dysfunction or myocardial stunning
is demonstrated by the activity of the combinations and
pharmaceutical compositions of this invention in conventional
preclinical cardioprotection assays [see, for example, the in vivo
assay in Klein, H. et al., Circulation 92:912-917 (1995); the
isolated heart assay in Scholz, W. et al., Cardiovascular Research
29:260-268 (1995); the antiarrhythmic assay in Yasutake M. et al.,
Am. J. Physiol., 36:H2430-H2440 (1994); the NMR assay in Kolke et
al., J. Thorac. Cardiovasc. Surg. 112: 765-775 (1996)] and the
additional in vitro and in vivo assays described below. Such assays
may also provide a means whereby the activities of the combinations
and pharmaceutical compositions of this invention can be compared
with the activities of other known compounds. The results of these
comparisons are useful for determining dosage levels in mammals,
including humans, for the treatment of such diseases.
[0149] Since the present invention relates to methods of reducing
tissue damage resulting from ischemia with a combination of active
ingredients that may be administered separately, the invention
further relates to combining separate pharmaceutical compositions
in kit form. The kit, according to the instant invention, comprises
an amount of a sodium-hydrogen exchanger type-1 (NHE-1) inhibitor,
and a pharmaceutically acceptable carrier, vehicle, or diluent in a
first unit dosage form; an amount of a second compound selected
from the group consisting of (a) a complement modulator, (b) a
metabolic modulator, (c) an anti-apoptotic agent, (d) a nitric
oxide synthase-related agent, and (e) an enzyme/protein modulator
selected from the group consisting of a protein kinase C activator,
an endothelin converting enzyme inhibitor, a tissue-activated
fibrinolytic inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger
isoform-1 (NCX-1) inhibitor, and a poly (ADP ribose) synthetase
(PARS/PARP) inhibitor, and a pharmaceutically acceptable carrier,
vehicle, or diluent in a second unit dosage form; and a container.
The container is employed to contain the separate components and
may comprise, for example, a divided bottle or a divided foil
packet, however, the separate compositions may also be contained
within a single, undivided container. Normally, the kit will also
include directions for the administration of the separate
components. The kit form is particularly advantageous when the
separate components are administered in different dosage forms
(e.g., oral and parenteral), are administered at different dosage
levels, or when titration of the individual components of the
combination is desired by the prescribing physician.
[0150] One example of such a kit is a so-called blister pack.
Blister packs are well known in the packaging industry and are used
widely for the packaging of pharmaceutical unit dosage forms
(tablets, capsules, and the like). Blister packs generally comprise
a sheet of relatively rigid material covered with a foil of a
preferably transparent plastic material. During the packaging
process, recesses (e.g., blisters) are formed in the plastic foil.
The recesses generally conform to the size and shape of the tablets
or capsules to be contained therein. Next, the tablets or capsules
are placed in the recesses and the sheet of relatively rigid
material is sealed against the plastic foil at the face of the foil
which is opposite from the direction in which the recesses were
formed. As a result, the tablets or capsules are captively retained
and sealed inside the recesses between the plastic foil and the
sheet. Preferably, the strength of the sheet is such that the
tablets or capsules may be removed from the blister pack by the
application of manual pressure on the outer surface of the recesses
whereby an opening is formed in the sheet at the place of the
recess. The tablet or capsule can then be removed through the
formed opening.
[0151] It is further desirable to provide a memory aid on the pack,
for example, in the form of numbers or similar indicia proximate to
the tablets or capsules whereby the indicia correspond to the days
of the regimen which the dosage form so specified is to be
ingested. An additional example of such a memory aid is a calendar
printed on the pack, for example, as follows: "First Week, Monday,
Tuesday, . . . etc . . . Second Week, Monday, Tuesday, . . . " etc.
In light of the instant disclosure, other variations will be
readily apparent to one of ordinary skill in the art. A "daily
dose" can be a single tablet or capsule, or multiple tablets or
capsules, or tablets or capsules to be ingested on a given day.
Also, a daily dose of the sodium-hydrogen exchanger type 1 (NHE-1)
inhibitor can consist of a single tablet or capsule, while a daily
dose of the second compound, selected from the group consisting of
(a) a compliment modulator, (b) a metabolic modulator, (c) an
anti-apoptotic agent, (d) a nitric oxide synthase-related agent,
and (e) an enzyme/protein modulator selected from the group
consisting of a protein kinase C activator, an endothelin
converting enzyme inhibitor, a tissue-activated fibrinolytic
inhibitor (TAFI), a Na.sup.+/Ca.sup.+2 exchanger isoform-1 (NCX-1)
inhibitor, and a poly (ADP ribose) synthetase (PARS/PARP)
inhibitor, can consist of multiple tablets or capsules, and
vice-versa. The memory aid should reflect this.
[0152] In another specific embodiment of the invention, a pack
designed to dispense the daily doses one at a time in the order of
their intended use is provided. Preferably, the pack is equipped
with a memory aid, so as to further facilitate compliance with the
dosage regimen. An example of such a memory aid comprises a
mechanical counter that indicates the number of daily doses to be
dispensed. Another example of such a memory aid comprises a
battery-powered micro-chip memory coupled with a liquid crystal
readout, or audible reminder signal which, for example, reads out
the date and time that the last daily dose has been taken and/or
reminds the patient when the next dose is to be taken.
Measurement of Human NHE-1 Inhibitory Activity
[0153] Methodologies for measurement of human NHE-1 activity and
inhibitor potency are predicated on those published by Watson et
al., Am. J. Physiol., 24:G229-G238, 1991), where NHE-mediated
recovery of intracellular pH is measured following intracellular
acidification. Thus, fibroblasts stably expressing human NHE-1
(Counillon, L. et al., Mol. Pharmacol., 44:1041-1045 (1993) are
plated onto collagen coated 96 well plates (50,000/well) and grown
to confluence in growth media (DMEM high glucose, 10% fetal bovine
serum, 50 u/ml penicillin and streptomycin). Confluent plates are
incubated for 30 min at 37.degree. C. with the pH sensitive
fluorescent probe BCECF (5 .mu.M; Molecular Probes, Eugene, Oreg.).
BCECF loaded cells are incubated for 30 min at 37.degree. C. in
acid loading media (70 mM choline chloride, 50 mM NHCl.sub.4, 5 mM
KCl, 1 mM MgCl.sub.2, 1.8 mM CaCl.sub.2, 5 mM glucose, 10 mM HEPES,
pH 7.5), and then placed in a Fluorescent Imaging Plate Reader
(Molecular Devices, CA). BCECF fluorescence is monitored using
excitation and emission wavelengths of 485 nM and 525 nM,
respectively. Intracellular acidification is initiated via rapid
replacement of acid loading media with recovery media (120 mM NaCl,
5 mM KCl, 1 mM MgCl.sub.2, 1.8 mM CaCl.sub.2, 5 mM glucose, 10 mM
HEPES, pH 7.5) .+-.test compound, and NHE-mediated recovery of
intracellular pH is monitored as the subsequent time-dependent
increase BCECF fluorescence. The potency of human NHE-1 inhibitors
is calculated as the concentration that reduces recovery of
intracellular pH by 50% (IC.sub.50).
[0154] As background information, it is noted that brief periods of
myocardial ischemia followed by coronary artery reperfusion
protects the heart from subsequent severe myocardial ischemia
(Murry et al., Circulation 74:1124-1136, 1986).
[0155] The therapeutic effects of the combinations and
pharmaceutical compositions of this invention in preventing heart
tissue damage resulting from an ischemic insult can be demonstrated
in vitro along lines presented in Tracey, et al. (Cardiovasc. Res.,
33: 410-415, 1997), as described specifically herein.
Cardioprotection, as indicated by a reduction in infarcted
myocardium, can be induced pharmacologically using adenosine
receptor agonists in isolated, retrogradely perfused rabbit hearts
as an in vitro model of myocardial ischemic preconditioning
(Tracey, et al., Cardiovasc. Res., 33: 410-415, 1997). The in vitro
test described below demonstrates that the combinations and
pharmaceutical compositions of the invention can also
pharmacologically induce cardioprotection, i.e., reduced myocardial
infarct size, when administered to a rabbit isolated heart. The
effects of the combinations and pharmaceutical compositions are
compared to ischemic preconditioning and the A1 adenosine agonist,
PIA (N.sup.6-1-(phenyl-2R-isopropyl)adenosi- ne), that has been
shown to pharmacologically induce cardioprotection in the rabbit
isolated heart (Tracey, et aL, Cardiovasc. Res., 33: 410-415,
1997). The exact methodology is described below.
[0156] The protocol used for these experiments closely follows that
described by Tracey, et aL, Cardiovasc. Res., 33: 410-415, 1997.
Male New Zealand White rabbits (3-4 kg) are anesthetized with
sodium pentobarbital (30 mg/kg, i.v.). After deep anesthesia is
achieved (determined by the absence of an ocular blink reflex) the
animal is intubated and ventilated with 100% O.sub.2 using a
positive pressure ventilator. A left thoracotomy is performed, the
heart exposed, and a snare (2-0 silk) is placed loosely around a
prominent branch of the left coronary artery, approximately 2/3 of
the distance towards the apex of the heart. The heart is removed
from the chest and rapidly (<30 sec) mounted on a Langendorff
apparatus. The heart is retrogradely perfused in a
non-recirculating manner with a modified Krebs solution (NaCl 118.5
mM, KCl 4.7 mM, Mg SO.sub.4 1.2 mM, KH.sub.2PO.sub.4 1.2 mM,
NaHCO.sub.3 24.8 mM, CaCl.sub.2 2.5 mM, and glucose 10 mM), at a
constant pressure of 80 mmHg and a temperature of 37.degree. C.
Perfusate pH is maintained at 7.4-7.5 by bubbling with 95%
O.sub.2/5% CO.sub.2. Heart temperature is tightly controlled by
using heated reservoirs for the physiological solution and water
jacketing around both the perfusion tubing and the isolated heart.
Heart rate and left ventricular pressures are determined via a
latex balloon which is inserted in the left ventricle and connected
by stainless steel tubing to a pressure transducer. The
intraventricular balloon is inflated to provide a systolic pressure
of 80-100 mmHg, and a diastolic pressure .ltoreq.10 mm Hg. Total
coronary flow is also continuously monitored using an in-line flow
probe and normalized for heart weight.
[0157] The heart is allowed to equilibrate for 30 min, over which
time the heart must show stable left ventricular pressures within
the parameters outlined above. If the heart rate falls below 180
bpm at any time prior to the 30 min. period of regional ischemia,
the heart is paced at about 200 bpm for the remainder of the
experiment. Ischemic preconditioning is induced by total cessation
of cardiac perfusion (global ischemia) for 5 min, followed by
reperfusion for 10 min. The regional ischemia is provided by
tightening the snare around the coronary artery branch. Following
the 30 min regional ischemia, the snare is released and the heart
reperfused for an additional 120 min.
[0158] Pharmacological cardioprotection is induced by infusing the
combination or pharmaceutical composition at predetermined
concentrations, starting 30 min prior to the 30 min regional
ischemia, and continuing until the end of the 120 min reperfusion
period. Hearts which receive test combinations or pharmaceutical
compositions do not undergo the period of ischemic preconditioning.
A reference compound, PIA (25 nM) is perfused through hearts (which
do not receive the test combination or pharmaceutical composition)
for a 5 min period which ends 10 min before the 30 min regional
ischemia.
[0159] At the end of the 120 min reperfusion period, the coronary
artery snare is tightened, and a 0.5% suspension of fluorescent
zinc cadmium sulfate particles (1-10 .mu.m) Duke Scientific
Corp.(Palo Alto, Calif.) is perfused through the heart; this stains
all of the myocardium, except that area-at-risk for infarct
development (area-at-risk). The heart is removed from the
Langendorff apparatus, blotted dry, wrapped in aluminum foil and
stored overnight at -20.degree. C. The next day, the heart is
sliced into 2 mm transverse sections from the apex to the top of
the ventricles. The slices are stained with 1% triphenyl
tetrazolium chloride (TTC) in phosphate-buffered saline for 20 min
at 37.degree. C. Since TTC reacts with living tissue (containing
NAD-dependent dehydrogenases), this stain differentiates between
living (red stained) tissue, and dead tissue (unstained infarcted
tissue). The infarcted area (no stain) and the area-at-risk (no
fluorescent particles) are calculated for each slice of left
ventricle using a precalibrated image analyzer. To normalize the
ischemic injury for differences in the area-at-risk between hearts,
the data is expressed as the ratio of infarct area vs. area-at-risk
(%IA/AAR). All data are expressed as mean.+-.SE and compared
statistically using a Mann-Whitney non-parametric test with a
Bonferroni correction for multiple comparisons. Significance is
considered as p<0.05.
[0160] The therapeutic effects of the combinations and
pharmaceutical compositions of this invention in preventing heart
tissue damage otherwise resulting from an ischemic insult can also
be demonstrated in vivo along lines presented in Liu et al.
(Circulation, 84: 350-356, 1991) as described specifically herein.
This in vivo assay tests the cardioprotection of the combinations
and pharmaceutical compositions relative to the control group which
receives saline vehicle. Cardioprotection, as indicated by a
reduction in infarcted myocardium, can be induced pharmacologically
using intravenously administered adenosine receptor agonists in
intact, anesthetized rabbits studied as an in situ model of
myocardial ischemic preconditioning (Liu et al., Circulation
84:350-356, 1991). The in vivo assay tests whether combinations and
pharmaceutical compositions can pharmacologically induce
cardioprotection, i.e., reduced myocardial infarct size, when
parenterally administered to intact, anesthetized rabbits. The
effects of the combinations and pharmaceutical compositions of this
invention can be compared to ischemic preconditioning using the A1
adenosine agonist, N.sup.6-1-(phenyl-2R-isopropyl) adenosine (PIA)
that has been shown to pharmacologically induce cardioprotection in
intact anesthetized rabbits studied in situ (Liu et al.,
Circulation 84:350-356, 1991). The methodology is described
below.
[0161] Surgery: New Zealand White male rabbits (3-4 kg) are
anesthetized with sodium pentobarbital (30 mg/kg, i.v.). A
tracheotomy is performed via a ventral midline cervical incision
and the rabbits are ventilated with 100% oxygen using a positive
pressure ventilator. Catheters are placed in the left jugular vein
for drug administration and in the left carotid artery for blood
pressure measurements. The hearts are then exposed through a left
thoracotomy and a snare (00 silk) placed around a prominent branch
of the left coronary artery. Ischemia is induced by pulling the
snare tight and clamping it in place. Releasing the snare allows
the affected area to reperfuse. Myocardial ischemia is evidenced by
regional cyanosis; reperfusion is evidenced by reactive
hyperemia.
[0162] Protocol: Once arterial pressure and heart rate have been
stable for at least 30 min. the test is started. Ischemic
preconditioning is induced by occluding the coronary artery for 5
min followed by a 10 min. reperfusion. Pharmacological
preconditioning is induced by infusing test combination or
pharmaceutical composition over, for example 5 min. and allowing 10
min. before further intervention or by infusing the adenosine
agonist, PIA (0.25 mg/kg). Following ischemic preconditioning,
pharmacological preconditioning or no conditioning (unconditioned,
vehicle control) the artery is occluded for 30 minutes and then
reperfused for two hours to induce myocardial infarction. The
combination or pharmaceutical composition and PIA are dissolved in
saline or other suitable vehicle and delivered at 1 to 5 mg/kg,
respectively.
[0163] Staining (Liu et al., Circulation 84:350-356, 1991): At the
end of the 2 hour reperfusion period, the hearts are quickly
removed, hung on a Langendorff apparatus, and flushed for 1 minute
with normal saline heated to body temperature (38.degree. C.). The
silk suture used as the snare is then tied tightly to reocclude the
artery and a 0.5% suspension of fluorescent zinc cadmium sulphate
particles (1-10 pm) Duke Scientific Corp. (Palo Alto, CA) is
infused with the perfusate to stain all of the myocardium except
the area at risk (nonfluorescent ventricle). The hearts are then
quickly frozen and stored overnight at -20.degree. C. On the
following day, the hearts are cut into 2 mm slices and stained with
1% triphenyl tetrazolium chloride (TTC). Since TTC reacts with
living tissue, this stain differentiates between living (red
stained) tissue, and dead tissue (unstained infarcted tissue). The
infarcted area (no stain) and the area at risk (no fluorescent
particles) are calculated for each slice of left ventricle using a
pre-calibrated image analyzer. To normalize the ischemic injury for
differences in the area at risk between hearts, the data is
expressed as the ratio of infarct area vs. area at risk (%IA/AAR).
All data are expressed as Mean.+-.SEM and compared statistically
using single factor ANOVA or Mann Whitney non-parametric test.
Significance is considered as p<0.05.
[0164] The combinations and pharmaceutical compositions of this
invention can be tested for their utility in reducing or preventing
ischemic injury in non-cardiac tissues, for example, the brain, or
the liver, utilizing procedures reported in the scientific
literature. The combinations and pharmaceutical compositions of
this invention in such tests can be administered by the preferred
route and vehicle of administration and at the preferred time of
administration either prior to the ischemic episode, during the
ischemic episode, following the ischemic episode (reperfusion
period) or during any of the below-mentioned experimental
stages.
[0165] The benefit of the combinations and pharmaceutical
compositions of the invention in reducing ischemic brain damage can
be demonstrated, for example, in mammals using the method of Park,
et al (Ann. Neurol. 1988;24:543-551). According to the procedure of
Park, et al., adult male Sprague Dawley rats are anesthetized
initially with 2% halothane, and thereafter by mechanical
ventilation with a nitrous oxide-oxygen mixture (70%:30%)
containing 0.5-1% halothane. A tracheostomy is then performed. The
stroke volume of the ventilator is adjusted to maintain arterial
carbon dioxide tension at approximately 35 mm Hg and adequate
arterial oxygenation (PaO.sub.2>90 mm Hg). Body temperature can
be monitored by a rectal thermometer, and the animals can be
maintained normothermic, if necessary, by external heating. The
animals next undergo subtemporal craniectomy to expose the main
trunk of the left middle cerebral artery (MCA) under an operating
microscope, and the exposed artery is occluded with microbipolar
coagulation to generate large ischemic lesions in the cerebral
cortex and basal ganglia. After three hours of MCA occlusion, the
rats are deeply anesthetized with 2% halothane and a thoracotomy is
performed to infuse heparinized saline into the left ventricle. The
effluent is collected via an incision of the right atrium. The
saline washout is followed by approximately 200 ml of a 40%
formaldehyde, glacial acetic acid and absolute methanol solution
(FAM; 1:1:8, v/v/v), then the animals are decapitated and the head
is stored in fixative for 24 hours. The brain is then removed,
dissected, embedded in paraffin wax, and sectioned (approximately
100 sections 0.2 mm per brain). The sections are then stained with
hematoxylin-eosin or with a combination of cresyl violet and Luxol
fast blue, and examined by light microscopy to identify and
quantitate the ischemic damage using a precalibrated image
analyzer. The ischemic volumes and areas are expressed in absolute
units (mm.sup.3 and mm.sup.2) and as a percentage of the total
region examined. The effect of the combinations and pharmaceutical
compositions of this invention to reduce ischemic brain damage
induced by MCA occlusion is noted based on a reduction in the area
or volume of relative or absolute ischemic damage in the brain
sections from the rats in the treatment group compared to brain
sections from rats in a placebo-treated control group.
[0166] Other methods which could alternatively be utilized to
demonstrate the benefit of the combinations and pharmaceutical
compositions of the invention in reducing ischemic brain damage
include those described by Nakayama, et al. in Neurology
1988,38:1667-1673; Memezawa, et al. in Stroke 1992,23:552-559;
Folbergrova, et al. in Proc. Nat1. Acad. Sci 1995,92:5057-5059; and
Gotti, et al. in Brain Res. 1990,522:290-307.
[0167] The benefit of the combinations and pharmaceutical
compositions of this invention to reduce ischemic liver damage can
be demonstrated, for example, in mammals using the method of
Yokoyama, et al. (Am. J. Physiol. 1990;258:G564-G570). According to
the procedure of Yokoyama, et al., fasted adult male Sprague Dawley
rats are anesthetized with sodium pentobarbital (40 mg/kg i.p.),
then the animals are tracheotomized and mechanically ventilated
with room air. The liver is extirpated and placed in an
environmental chamber maintained at constant temperature
(37.degree. C.), then perfused through the portal vein at a
constant pressure of 15 cm H.sub.2O with a modified,
hemoglobin-free Krebs-Henseleit buffer (in mM: 118 NaCl, 4.7 KCl,
27 NaHCO.sub.3, 2.5 CaCl.sub.2, 1.2 MgSO.sub.4, 1.2
KH.sub.2PO.sub.4, 0.05 EDTA, and 11 mM glucose, plus 300 U
heparin). The pH of the perfusate is maintained at 7.4 by gassing
the buffer with 95% O.sub.2-5% CO.sub.2. Each liver is perfused at
a flow rate of 20 ml/min in a single-pass manner for a 30 min
washout and equilibration period (preischemic period), followed by
a 2 hour period of global ischemia, and then a 2 hour period of
reperfusion under conditions identical to the preischemic period.
Aliquots (20 ml) of the perfusate are collected during the
preischemic period, immediately after the occlusive ischemic
period, and every 30 min of the 2 hour reperfusion period. The
perfusate samples are assayed for the appearance of hepatocellular
enzymes, for example, aspartate amino-transferase (AST), alanine
amino-transferase (ALT), and lactate dehydrogenase (LDH), which are
taken to quantitatively reflect the degree of ischemic liver tissue
damage during the procedure. AST, ALT, and LDH activities in the
perfusate can be determined by several methods, for example, by the
reflectometry method using an automatic Kodak Ektachem 500 analyzer
reported by Nakano, et al. (Hepatology 1995;22:539-545). The effect
of the combinations and pharmaceutical compositions of this
invention in reducing ischemic liver damage induced by occlusion is
noted based on a reduction in the release of hepatocellular enzymes
immediately following the occlusive period and/or during the
postischemic-reperfusion period in the perfused livers from the
rats in the treatment group compared to perfused livers from rats
in a placebo-treated control group.
[0168] Other methods which may be utilized to demonstrate the
benefits of the compositions and methods of this invention in
reducing ischemic liver damage include those described by Nakano,
et al. (Hepatology 1995;22:539-545).
* * * * *