U.S. patent application number 09/802523 was filed with the patent office on 2002-04-18 for pharmaceutical compositions comprising metal complexes.
Invention is credited to Abrams, Michael J., Fricker, Simon P., Murrer, Barry A., Vaughan, Owen John.
Application Number | 20020045611 09/802523 |
Document ID | / |
Family ID | 27266826 |
Filed Date | 2002-04-18 |
United States Patent
Application |
20020045611 |
Kind Code |
A1 |
Abrams, Michael J. ; et
al. |
April 18, 2002 |
Pharmaceutical compositions comprising metal complexes
Abstract
New pharmaceutical compositions and pharmaceutical compositions
comprising metal complexes have activity against diseases caused by
or related to overproduction or localized high concentration of
nitric oxide in the body.
Inventors: |
Abrams, Michael J.;
(Glenmore, PA) ; Fricker, Simon P.; (Berkshire,
GB) ; Murrer, Barry A.; (Berkshire, GB) ;
Vaughan, Owen John; (Stockholm, SE) |
Correspondence
Address: |
Laurie A. Axford
Morrision & Foerster LLP
Suite 500
3811 Valley Centre Drive
San Diego
CA
92130-2332
US
|
Family ID: |
27266826 |
Appl. No.: |
09/802523 |
Filed: |
March 9, 2001 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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09802523 |
Mar 9, 2001 |
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09175028 |
Oct 19, 1998 |
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6284752 |
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09175028 |
Oct 19, 1998 |
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08602814 |
Feb 26, 1996 |
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5824673 |
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Current U.S.
Class: |
514/184 ;
424/617 |
Current CPC
Class: |
A61K 31/28 20130101;
A61K 33/24 20130101 |
Class at
Publication: |
514/184 ;
424/617 |
International
Class: |
A61K 033/24; A61K
031/555 |
Foreign Application Data
Date |
Code |
Application Number |
Aug 19, 1994 |
WO |
PCT/GB94/01817 |
Aug 25, 1993 |
GB |
93/17686.5 |
Claims
1. The use of a neutral, anionic or cationic metal complex having
at least one site for coordination with NO, of
formula[M.sub.a(X.sub.bL).sub.cY.su- b.dZ.sub.e].sup.a.+-. formula
I,in the manufacture of a medicament for the attenuation of NO
levels where NO in implicated in disease, where: M is a metal ion
or a mixture of metal ions; X is a cation or a mixture of cations;
L is a ligand, or mixture of ligands each containing at least two
different donor atoms selected from the elements of Group IV, Group
V or Group VI of the Periodic Table; Y is a ligand, or a mixture of
the same or different ligands each containing at least one donor
atom or more than one donor atom, which donor atom is selected from
the elements of Group IV, Group V or Group VI of the Periodic
Table; and Z is a halide or pseudohalide ion or a mixture of halide
ions and pseudohalide ions; a=1-3; b=0-12; c=0-18; d=0-18; e=0-18;
and n=0-10; provided that at least one of c, d and e is 1 or more;
and where c is 0; b is also 0; and where a is 1; c, d and e are not
greater than 9; and where a is 2; c, d and e are not greater than
12.
2. The use of a complex of formula I in the manufacture of a
medicament for the treatment of NO-overproduction related
disease.
3. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 1, wherein M is a first, second or
third row transition metal ion.
4. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 1, wherein M is an Rh, Ru, Os, Mn,
Co, Cr or Re ion.
5. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 4, wherein M is an Rh, Ru or Os
ion.
6. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 5, wherein M is in oxidation state
III.
7. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to any of claims 1-6, wherein X is a mono-,
di- or tri-valent cation.
8. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 7, wherein X is H.sup.+, K.sup.+,
Na.sup.+, NH.sub.4.sup.+ or Ca.sup.2+.
9. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to any of claims 1-8, wherein L is a ligand
containing both nitrogen and oxygen donor atoms.
10. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 9, wherein L is edda, edta, nta,
dipic, pic, dtpa, hedtra, tedta or dtedta.
11. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to any of claims 1-10, wherein Y is a ligand
containing at least one of N, O, S, C or P donor groups.
12. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 11, wherein said N donor group is
ammine, amine, amide, nitrile or nitride or derivatives
thereof.
13. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 11, wherein said O donor group is
carboxylic acid, ester or derivatives thereof, water, oxide,
sulphoxide, hydroxide, acetate, lactate, propionate, oxalate or
maltolate.
14. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 11, wherein said S donor group is
sulphoxide, dialkylsulphide, dialkylcarbamate, dithiocarbamate, or
dithiophosphate.
15. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 11, wherein said C donor group is
carbon monoxide or isocyanide.
16. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 11, wherein said P donor group is
trialkylphosphine.
17. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to any preceding claim, wherein Z is a
halide.
18. The use of a complex of formula I in the manufacture of a
medicament for the attenuation of NO levels where NO is implicated
in disease, according to claim 17, wherein Z is chloride.
19. The use of an optionally hydrated ruthenium complex of
formula[Ru(H.sub.0-6L").sub.1-3Y.sub.0-2Cl.sub.1-4].sup.(0-4).+-.
formula II,where L" is an amide or ester or derivative thereof, or
a polydentate aminocarboxylate ligand, for example edda, tropolone
edta, nta, dipic, pic, dtpa, hedtra, tedta or dtedta or a mixture
of any of these, and Y is as defined above and may for example be
selected from acetylacetone (acac), a .beta.-diketonate, water,
dimethylsulphoxide (dmso), carboxylate, bidentate carboxylate,
catechol, kojic acid, maltol, hydroxide, tropolone, malonic acid,
oxalic acid, 2,3-dihydroxynaphthalene- , squaric acid, acetate, a
sulphate and a glycolate in the manufacture of a medicament for the
attenuation of NO levels where NO is implicated in disease.
20. A pharmaceutical composition containing an optionally hydrated
complex of formula[M.sub.1-3Y.sub.1-18Cl.sub.0-18].sup.(0-6).+-.
formula IIIwherein Y is a sulphur donor ligand.
21. The use of an optionally hydrated complex of ruthenium of
formula[M'".sub.1-3Y'".sub.1-18Cl.sub.0-18].sup.(0-6).+-. formula
IIIwhere M'" is ruthenium and Y'" is an oxygen-donor ligand such as
acetate, lactate, water, oxide, propionate, oxalate or mialtolate
or a combination of these, in the manufacture of a medicament for
the attenuation of NO levels where NO is implicated in disease.
22. The use of an optionally hydrated complex of ruthenium of
formula[RuY.sup.IV.sub.1-9Cl.sub.1-9].sup.(0-4).+-. formula IVwhere
Y.sup.IV is a nitrogen-donor ligand such as ammine, ethylenediamine
(en), pyridine (py), 1,10-phenanthroline (phen), 2,2'-bipyridine
(bipy), 1,4,8,11=tetraaza-cyclotetradecane (cyclam),
2,3,7,8,12,13,17,18-octaethy- lporphyrin (oep) or a combination of
these, in the manufacture of a medicament for the attenuation of NO
levels where NO is implicated in disease.
23. The use of an optionally hydrated complex of ruthenium or
osmium of general
formula[M.sub.1-3Y.sub.1-18Cl.sub.0-18].sup.(0-6).+-. formula
Vwhere Y is a combination according to claims 19, 20, 21, or 22 in
the manufacture of a medicament for the attenuation of NO levels
where NO is implicated in disease.
24. The use of an optionally hydrated complex of general
formula[M.sub.1-3Y.sub.1-18Cl.sub.0-18].sup.(0-6).+-.where Y is a
sulphur donor ligand, in the manufacture of a medicament for the
attenuation of NO levels where NO is implicated in disease.
25. A pharmaceutical composition containing an optionally hydrated
ruthenium complex as described in claim 19.
26. A pharmaceutical composition containing an optionally hydrated
complex of ruthenium as described in claim 21, wherein Y'" is an
oxygen donor ligand selected from the group acetate, lactate,
oxide, propionate and maltolate.
27. A pharmaceutical composition containing an optionally hydrated
complex of ruthenium as described in claim 22, wherein Y.sup.IV is
a nitrogen donor ligand selected from the group en, py, phen, bipy,
cyclam or oep.
28. A pharmaceutical composition containing an optionally hydrated
complex of formula[Os(ox)(bipy).sub.2]
29. A pharmaceutical composition containing an optionally hydrated
complex of formula[Ru(acac).sub.2(MeCN).sub.2].sup.+
Description
[0001] This invention relates to new pharmaceutical compositions
and to pharmaceutical compositions having activity against diseases
caused by or related to overproduction or localised high
concentration of nitric oxide in the body.
[0002] Nitric oxide (NO) plays a varied and vital role in the human
body. For example, NO plays a vital role in the control of blood
pressure; it acts as a neurotransmitter; it plays a role in
inhibition of platelet aggregation (important in thrombosis or
blockages of the blood vessels), and in cytostasis (important in
fighting of tumours). Overproduction of NO however, has been
implicated in a number of disease states, including
vascular/pressor diseases such as septic shock, post-ischaemic
cerebral damage, migraine, and dialysis induced renal hypotension;
immunopathologic diseases such as hepatic damage in inflammation
and sepsis, allograft rejection, graft versus host diseases,
diabetes and wound healing; neurodegenerative diseases such as
cerebral ischaemia, trauma, chronic epilepsy, Alzheimer's disease,
Huntington's disease, and AIDS dementia complex; and side effects
of treatment such as restenosis following angioplastic treatment
and secondary hypotension following cytokine therapy.
[0003] Pharmacological modulation of nitric oxide in any of these
disease states should prove extremely beneficial.
[0004] One above-mentioned disease relating to overproduction of
NO, is septic shock. This is precipitated by local septicaemia or
endotoxaemia, (high local levels of bacterial endotoxins). The
result is activation of macrophages, lymphocytes, endothelial cells
and other cell types capable of producing NO, further mediated by
cytokine production by these cells. The activated macrophages
produce excess NO which causes vasodilation of the blood vessels,
and results in local vascular damage and vascular collapse. This
destruction of vascular integrity may be so great that it leads to
the collapse of haemodynamic homeostasis, the end result being
death.
[0005] Current ideas for pharmacological modulation of nitric oxide
in such diseases are based on dealing with the mediators of septic
shock, such as cytokines, endotoxins, and platelet activating
factor (PAF). The approaches include use of antibodies to cytokines
such as tumour necrosis factor (TNF), receptor antagonists such as
interleukin 1, (IL-1), antibodies to lipopolysaccharide (the
endotoxin produced by gram negative bacteria, and PAF antagonists.
All such approaches while challenging a factor mediating septic
shock, do not attempt to deal with the aetiology, or cause, of the
disease. Recent advances in understanding of NO have lead to the
proposal that inhibitors of the NO synthase enzyme, such as
N.sup.G-monomethy-L-arginine (L-NMMA), may be useful in the
treatment of septic shock and other NO overproduction related to
diseases since they inhibit NO production. While these inhibitors
have shown some utility in animal models and preliminary clinical
studies, they have the disadvantage of undesirably inhibiting total
NO synthesis in the body.
[0006] An aim of the present invention is to provide new and
previously indicated pharmaceutical compositions which are able to
modulate NO levels in the body by scavenging, or removing, NO in
situ so that necessary NO synthesis continues while dangerous
excesses are removed. We have found that certain metal complexes
have the ability to carry out this important role.
[0007] Some metal complexes are known in pharmaceutical
compositions for the treatment of diseases of the human body. For
example, certain complexes of platinum and ruthenium have been used
or indicated in the treatment of cancer. Metal complexes have not
however been previously indicated in the treatment of NO
overproduction related diseases.
[0008] This invention provides for the use of a neutral, anionic or
cationic metal complex having at least one site for coordination
with NO, of formula
[M.sub.a(X.sub.bL).sub.cY.sub.dZ.sub.e].sup.a.+-. formula I,
[0009] in the manufacture of a medicament for the attenuation of NO
levels where NO is implicated in disease,
[0010] where:
[0011] M is a metal ion or a mixture of metal ions;
[0012] X is a cation or a mixture of cations;
[0013] L is a ligand, or mixture of ligands each containing at
least two different donor atoms selected from the elements of Group
IV, Group V or Group VI of the Periodic Table;
[0014] Y is a ligand, or a mixture of the same or different ligands
each containing at least one donor atom or more than one donor
atom, which donor atom is selected from the elements of Group IV,
Group V or Group VI of the Periodic Table;
[0015] and
[0016] Z is a halide or pseudohalide ion or a mixture of halide
ions and pseudohalide ions;
[0017] a=1-3; b=0-12; c=0-18; d=0-18; e=0-18; and n=0-10;
[0018] provided that at least one of c, d and e is 1 or more;
[0019] and where c is 0; b is also 0;
[0020] and where a is 1; c, d and e are not greater than 9;
[0021] and where a is 2; c, d and e are not greater than 12.
[0022] By "complex" in this specification is meant a neutral
complex or anionic or cationic species.
[0023] The term "Group" which is used herein is to be understood as
a vertical column of the periodic table in which elements of each
Group have similar physical and chemical properties. The definition
of the Periodic Table is that credited to Mendeleev; Chambers
Dictionary of Science and Technology, 1974. Published by W & R
Chambers Ltd.
[0024] This invention may also be stated as providing a method of
attenuation of NO levels where NO is implicated in diseases of the
human body, comprising administering a pharmaceutical composition
containing a neutral, anionic or cationic metal complex of formula
I.
[0025] This invention may also provide for the use of a neutral,
anionic or cationic metal complex of formula I in the manufacture
of a medicament for the treatment of NO overproduction related
disease.
[0026] This invention may also be stated as providing a method of
treatment of diseases of the human body resultant of overproduction
of NO in the human body, comprising administering a pharmaceutical
composition containing a neutral, anionic or cationic metal complex
of formula I.
[0027] Where the formula I represents an anionic species, a cation
will also be present. Where formula I represents a cationic
species, an anion will also be present. The metal complexes may be
hydrated.
[0028] Preferably, M is a first, second or third row transition
metal ion. For example, M may be an Rh, Ru, Os, Mn, Co, Cr or Re
ion, and is preferably an Rh, Ru or Os ion.
[0029] Suitably M is in an oxidation state III. We have found
surprisingly that when the metal ion for example ruthenium is in
oxidation state III, the rate at which it binds with NO is
significantly faster than when it is in oxidation state II.
[0030] X may be any cation, such as a mono-, di- or tri-valent
cation. Suitable cations may be H.sup.+, K.sup.+, Na.sup.+,
NH.sub.4.sup.+ or Ca.sup.2+. Conveniently X may be H.sup.+, K.sup.+
or Na.sup.+.
[0031] Preferably, L is a ligand containing both nitrogen and
oxygen donor atoms. Examples of suitable such ligands include
ethylenediamine -N,N'-diacetic acid (edda),
ethylenediaminetetraacetic acid (edta), nitrilotriacetic acid
(nta), dipicolinic acid (dipic), picolinic acid (pic),
diethylenetriaminepentaacetic acid (dtpa), thiobis(ethylenenitrilo-
)tetraacetic acid (tedta), dithioethanebis(ethylenenitrilo)
tetraacetic acid (dtedta) anid
N-(2-hydroxyethyl)ethylenediamine-triacetic acid (hedtra).
[0032] Preferably, Y is a ligand containing nitrogen, oxygen,
sulphur, carbon, or phosphorus donor groups. Suitable nitrogen
donor groups may be for example ammine, amine, nitrile and nitride
or derivations thereof. Suitable oxygen donor groups may be for
example carboxylic acid, ester or derivations thereof, water,
oxide, sulphoxide, hydroxide, acetate, lactate, propionate, oxalate
and maltolate. Suitable sulphur donor groups may be for example
sulphoxide, dialkylsulphide, dithiocarbarnate or dithiophosphate.
Suitable carbon donor groups may be for example carbon monoxide or
isocyanide. Suitable phosphorus donor groups may be for example
triallylphosphine.
[0033] Z may be any halide and is preferably chloride, bromide or
iodide. Most conveniently, Z is chloride.
[0034] Examples of metal complexes for use according to the present
invention include optionally hydrated ruthenium complexes of
formula
[Ru(H.sub.0-6L").sub.1-3Y.sub.0-2Cl.sub.0-4].sup.(0-4).+-. formula
II,
[0035] where L" is an amide or ester or derivative thereof, or a
polydentate aminocarboxylate ligand, for example edta, nta, dipic,
pic, edda, tropolone, dtpa, hedtra, tedta or dtedta or diamide of
edta or dtpa or a mixture of any of these, and Y is as defined
above and may for example be selected from acetylacetone (acac), a
.beta.-diketonate, water, dimethylsulphoxide (dmso), carboxylate,
bidentate carboxylate, catechol, kojic acid, maltol, hydroxide,
tropolone, malonic acid, oxalic acid, 2,3-dihydroxynaphthalene,
squaric acid, acetate, a sulphate and a glycolate. The skilled
addressee will be able to substitute other known ligands at Y and
which will fall within the scope of the inventions. Preparative
methods of tedta, dtedta and diamide of edta and dtpa are described
in the following references respectively:
[0036] P Tse & J E Powell, Inorg Chem, (1985), 24, 2727
[0037] G Schwartzenbach, H Senner, G Anderegg, Helv Chim Acta 1957,
40, 1886
[0038] M S Konings, W C Dow, D B Love, K N Raymond, S C Quay and S
M Rocklage, Inorg Chem (1990), 29, 1488-1491
[0039] P N Turowski, S J Rodgers, R C Scarrow and K N Raymond,
Inorg Chem (1988), 27, 474-481.
[0040] Where the complex of formula II is an anion, a cation will
be required. For example the complexes of formula II are present
in
[0041] K[Ru(Hedta)Cl]2H.sub.2O
[0042] [Ru(H.sub.2edta)(acac)]
[0043] K[Ru(hedtra)Cl]H.sub.2O
[0044] K[Ru(dipic).sub.2]H.sub.2O
[0045] (H.sub.2pic)[RuCl.sub.2(pic).sub.2](Hpic)H.sub.2O
[0046] K[Ru(H.sub.2edta)Cl.sub.2]H.sub.2O
[0047] K[Ru(Hnta).sub.2]1/2H.sub.2O
[0048] K[Ru(H.sub.2dtpa)Cl]H.sub.2O
[0049] [Ru(Hhedtra)acac]H.sub.2O
[0050] [Ru(Hhedtra)trop]
[0051] [Ru(H.sub.3dtpa)Cl]
[0052] Complexes of formula II have not to the best of our
knowledge been previously indicated in any pharmaceutical
composition. Therefore the present invention also provides a
pharmaceutical composition containing an optionally hydrated
ruthenium complex of formula II.
[0053] Further examples of metal complexes for use according to the
present invention include optionally hydrated complexes of formula
III
[M.sub.1-3Y.sub.1-18Cl.sub.0-18].sup.(0-6).+-. formula III
[0054] Where Y is a sulphur donor ligand. For example, such complex
is present in
[0055] [Ru(mtc).sub.3] (mtc=4-morpholinecarbodithoic acid)
[0056]
Ru(S.sub.2CNCH.sub.2CH.sub.2NMeCH.sub.2CH.sub.2).sub.31/2H.sub.2O
[0057] Complexes of formula III in which Y is a sulphur donor
ligand have not to the best of our knowledge been previously
indicated in any pharmaceutical composition. Therefore, the present
invention also provides a pharmaceutical composition containing an
optionally hydrated complex of formula III wherein Y is a sulphur
donor ligand.
[0058] Yet further examples of metal complexes for use according to
the present invention include optionally hydrated complexes of
ruthenium of formula
[M'".sub.1-3Y'".sub.1-18Cl.sub.0-18].sup.(0-6).+-. formula III
[0059] where M'" is ruthenium and Y'" is an oxygen-donor ligand
such as acetate, lactate, water, oxide, propionate (COEt), oxalate
(ox), or maltolate (maltol) or a combination of these. For example
complexes of formula III are present in
[0060] [RU.sub.3O(OAC).sub.6](OAc)
[0061] [RU.sub.3O(lac).sub.6](lac)
[0062] [Ru.sub.2(OAc).sub.4]NO.sub.3
[0063] [Ru.sub.2(OCOEt).sub.4]NO.sub.3
[0064] K.sub.3[Ru(ox).sub.3]
[0065] [RU.sub.2(OAc).sub.4]Cl
[0066] [Ru(maltol).sub.3]
[0067] Some complexes of formula III have not to the best of our
knowledge been previously indicated in any pharmaceutical
composition. Therefore the present invention also provides a
pharmaceutical composition containing an optionally hydrated
complex of ruthenium of formula III wherein M'" is ruthenium and
Y'" is an oxygen-donor ligand selected from the group acetate,
lactate, oxide, propionate and maltolate.
[0068] Further examples of metal complexes for use according to the
present invention include optionally hydrated complexes of
ruthenium of formula
[RuY.sup.IV.sub.1-9Cl.sub.1-9].sup.(0-4).+-. formula IV
[0069] where Y.sup.IV is a nitrogen-donor ligand such as ammine,
ethylenediamine (en), pyridine (py), 1,10-phenanthroline (phen),
2,2-bipyridine (bipy) or 1,4,8,11-tetra-azacyclotetradecane
(cyclam), 2,3,7,8,12,13,17,18-octaethylporphyrin (oep) or a
combination of these. For example complexes of formula IV are
present in
[0070] [RU(NH.sub.3).sub.5Cl]Cl.sub.2
[0071] [Ru(en).sub.3]I.sub.3
[0072] trans-[RuCl.sub.2(py).sub.4]
[0073] K[Ru(phen)Cl.sub.4]
[0074] [Ru(cyclam)Cl.sub.2]Cl
[0075] K[Ru(bipy)Cl.sub.4]
[0076] [Ru(NH.sub.3).sub.6]Cl.sub.3
[0077] [Ru(NH.sub.3).sub.4Cl.sub.2]Cl
[0078] Ru(oep)Ph
[0079] Some complexes of formula IV have not to the best of our
knowledge been previously indicated in any pharmaceutical
composition. Therefore the present invention also provides a
pharmaceutical composition containing an optionally hydrated
complex of ruthenium of formula IV wherein Y.sup.IV is a
nitrogen-donor ligand selected from the group en, py, phen, bipy,
cyclam and oep. Derivations of these ligands can be prepared by a
skilled addressee and which will fall within the scope of the
inventions.
[0080] Still further examples of metal complexes for use according
to the present invention include optionally hydrated complexes of
ruthenium or osmium of general formula
[M.sub.1-3Y.sup.V.sub.1-18Cl.sub.0-18].sup.(0-6).+-. formula V
[0081] where Y.sup.V is a combination of donor ligands such as are
described hereinabove, for example ammine, dmso, oxalate, bipy,
acac and MeCN. Complexes of formula V are present in for
example
[0082] [Ru(NH.sub.3)(dmso).sub.2Cl.sub.3]
[0083] cis-[Ru(dmso).sub.4Cl.sub.2]
[0084] cis-[Ru(NH.sub.3)(dmso).sub.3Cl.sub.2]
[0085] [Ru(dmso).sub.3Cl.sub.3]
[0086] [Os(ox)(bipy).sub.2]H.sub.2O
[0087] [Ru(acac).sub.2(MeCN).sub.2]CF.sub.3SO.sub.3
[0088] The complex ions of the latter two compounds above have not
to the best of our knowledge been previously indicated in any
pharmaceutical composition. Therefore the present invention also
provides a pharmaceutical composition containing an optionally
hydrated complex of formula [Os(ox)(bipy).sub.2]; and further a
pharmaceutical composition containing an optionally hydrated
complex of formula [Ru(acac).sub.2(MeCN).sub.2].sup.+.
[0089] In use the complexes of the present invention may be
included as an active component in a pharmaceutical composition
containing an optionally hydrated complex of any of formulae I-V,
in admixture with a pharmaceutically acceptable carrier or diluent.
Said pharmaceutical composition may be formulated according to well
known principles, and may be in the form of a solution or
suspension for parenteral administration in single or repeat doses
or be in capsule, tablet, dragee, or other solid composition or as
a solution or suspension for oral administration, or formulated
into pessaries or suppositories, or sustained release forms of any
of the above. The solution or suspension may be administered by a
single or repeat bolus injection or continuous infusion, or any
other desired schedule. Suitable diluents, carriers, excipients and
other components are known. Said pharmaceutical composition may
contain dosages determined in accordance with conventional
pharmacological methods, suitable to provide active complexes in
the dosage range in humans of 1 mg to 10 g per day. Actual required
dosage is largely dependent on where in the body there is the
excess concentration of NO and for how long overproduction
continues or attenuation of NO levels, where NO is implicated in
disease, is required.
[0090] This invention will now be illustrated by Example.
[0091] A number of commercially available compounds, and compounds
prepared by routes known in the literature, containing the
complexes of the present invention were tested in vitro, in vitro
cell culture, and ex-vivo in order to determine ability to
coordinate with NO. The complexes tested were as follows:
1 Example Compound Literature Reference for Preparation 1
K[Ru(Hedta)Cl]2H.sub.2O AA Diamantis & JV Dubrawski,
Inorg.Chem., (1981), 20, 1142-50 2 [Ru(H.sub.2edta)(acac)] AA
Diamantis & JV Dubrawski, Inorg.Chem., (1983), 22, 1934-36 3
K[Ru(hedtra)Cl]H.sub.2O HC Bajaj & R van Eldik,
Inorg.Chem.(1982), 28, 198O-3 4 K[Ru(dipic).sub.2]H.sub.2O NH
Williams & JK Yandell, Aust.J.Chem.(1983), 36(12), 2377-2386 5
(H.sub.2pic)[RuCl.sub.2(pi- c).sub.2](Hpic)H.sub.2O JD Gilbert, D
Rose & G Wilkinson, J.Chem.Soc.(A), (1970), 2765-9 6
K[Ru(H.sub.2edta)Cl.sub.2]H.sub.2- O AA Diamantis & JV
Dubrawski, Inorg.Chem.(1981), 20, 1142-50 7
K[Ru(Hnta).sub.2]1/2H.sub.2O MM Taqui Khan, A Kumar & Z Shirin,
J. Chem. Research (M), (1986), 1001- 1009 8
K[Ru(H.sub.2dtpa)Cl]H.sub.2O MM Taqui Khan, A Kumar & Z Shirin,
J. Chem. Research (M). (1986); 1001- 1009 9
[Ru.sub.3O(lac).sub.6](lac) A Spencer & G Wilkinson, J. Chem.
Soc. Dalton Trans. (1972), 1570-77 10 [Ru.sub.3O(OAc).sub.6](OAc) A
Spencer & G Wilkinson, J.Chem. Soc. Dalton Trans. (1972),
1570-77 11 [Ru.sub.2(OAc).sub.4]NO.sub.3 M Mukaida, T Nomura &
T Ishimori, Bull. Chem. Soc. Japan, (1972), 45, 2143-7 12
[Ru.sub.2(OCOEt).sub.4]NO.sub.3 A Bino, FA Cotton & TR
Felthouse, Inorg. Chem. (1979), 18, 2599-2604 13
K.sub.3[Ru(ox).sub.3] CM Che, SS Kwong, CK Poon, TF Lai & TCW
Mak Inorg. Chem. (1985), 24, 1359-63 14 [Ru.sub.2(OAc).sub.4Cl RW
Mitchell, A Spencer & G Wilkinson J. Chem. Soc. Dalton Trans.,
(1973), 846-54 15 [Ru(NH.sub.3).sub.5Cl]Cl.sub.2 AD Allen, F
Bottomley, RO Harris, VP Reinsahu & CV Senoff J. Amer. Chem.
Soc. (1967), 89, 5595-5599 16 [Ru(en).sub.3]I.sub.3 TJ Meyer &
H Taube Inorg. Chem. (1968), 7, 2369-2379 17
K[RuCL.sub.4(phen)]H.sub.2O BR James & RS McMillan Inorg. Nucl.
Chem. Lett. (1975), 11(12) 837-9 18 [Ru(cyclam)Cl.sub.2]Cl PK Chan,
DA Isabirye & CK Poon Inorg. Chem. (1975), 14, 2579-80 19
K[RuCl.sub.4(bipy)] BR James & RS McMillan Inorg. Nucl. Chem.
Lett. (1975), 11(12), 837-9 20 [RuCl.sub.3(dmso).sub.2(NH.sub.3)]
Patent: International Publication No WO 91/13553 21
[Ru(NH.sub.3).sub.6]Cl.sub.- 3 Matthey Catalogue Sales: Cat No
[190245] 22 cis-[RuCl.sub.2(dmso).sub.4] EA Alessio, G Mestroni, G
Nardin, WM Attia, M Calligaris, G Sava & S Zorget Inorg. Chem.
(1988), 27, 4099-4106 23 cis-[RuCl.sub.2(dmso).sub.3(NH.sub.3)] M
Henn, E Alessio, G Mestroni, M Calligaris & WM Attia lnorg.
Chim. Acta, (1991), 187, 39-50 24 [RuCl.sub.3(dmso).sub.3] E
Alessia, G Balducci, M Calligalis, G Costa, WM Attia & G
Mestroni Inorg. Chem. (1991), 30, 609-618 25 [Ru(mtc).sub.3] AR
Hendrickson, JM Hope & RL Martin J. Chem. Soc. Dalton Trans.
(1976), 20, 2032-9 26 [Ru(maltol).sub.3] WP Griffith & SJ
Greaves Polyhedron, (1988), 7(19) 1973-9 27
[Ru(acac).sub.2(MeCN).sub.2]CF.sub.3SO.sub.3 Y Kasahara, T Hoshino,
K Shimizu & GP Sato Chem. Lett. (1990), 3, 381-4 28
K.sub.2[RuCl.sub.5(H.sub.2O)] Matthey Catalogue Sales: Cat No
[190094] 29 [Os(ox)(bipy).sub.J].H.sub.2O DA Buckingharn, FP Dwyer,
HA Goodwin & AM Sargeson Aust.J.Chem.(1964), 325-336 GM Bryant,
JE Fergusson & HKJ Powell Aust.J.Chem.(1971), 24(2), 257-73 30
[Ru(NH.sub.3).sub.4Cl.sub.2]Cl SD Pell, MM Sherban, V Tramintano
& MJ Clarke Inorg Synth, (1989), 26, 65. 31 [Ru(Hedtra)(dppm)]
MM Taqui Khan, K Venkatasubramanian, Z Shirin, MM Bhadbhade J Chem
Soc Dalt Trans (1992), 885-890 32 Ru(oep)Ph M Ke, SJ Rettig, BR
James and D Dolphin J Chem Soc Chem Commun (1987), 1110
[0092] A number of new compounds were prepared according to the
following protocols. The first four compounds are examples of
ruthenium complexes of formula [Ru(H.sub.0-6L").sub.1-3
Y.sub.0-2Cl.sub.0-4].sup.(0-4).+-. formula II), the subsequent two
are examples of [M.sub.1-3Y.sub.1-8Cl.sub- .0-18].sup.(0-6).+-.
(formula III).
[0093] Preparation of [Ru(Hhedtra)acac].H.sub.2O
[0094] Excess acetylacetone (1 cm.sup.3) was added to an aqueous
solution (5 cm.sup.3) of K[Ru(hedtra)Cl] (0.5 g). The solution
colour changed to violet. The mixture was warmed for 20 minutes
then left to stand at room temperature for 20 minutes. The violet
solution was extracted with chloroform (20 cm.sup.3). The
extraction was repeated twice more. A violet product precipitated
from the aqueous fraction. The product was filtered, washed in
acetone and dried in vacuo, yield 0.1 g (18%).
[0095] Anal. Calc. for C.sub.15H.sub.25O.sub.10N.sub.2Ru: C, 36.43;
H, 5.11; N, 5.70. Found: C, 36.16; H 5.42; N, 5.61%.
[0096] Preparation of [Ru(Hhedtra)trop]2H.sub.2O
[0097] A three-fold excess of tropolone (0.78 g) dissolved in 50:50
water/absolute ethanol (5 cm.sup.3) was added to a warm aqueous
solution of K[Ru(hedtra)Cl] (10 cm.sup.3). The mixture was heated
for 1 hour. On cooling, the dark green mixture was extracted with
3.times.20 cm.sup.3 portions of dichloromethane. On standing, a
dark green product precipitated from the aqueous fraction. The
product was filtered, washed with water (1 cm.sup.3), ether and
dried in vacuo, yield 0.4 g (36%).
[0098] Anal. Calc. for C.sub.17H.sub.22N.sub.2O.sub.9Ru0.2H.sub.2O:
C, 38.13; H, 4.86; N, 5.23. Found: C, 38.55; H, 4.67; N, 5.28%.
[0099] Preparation of [Ru(H.sub.3dtpa)Cl]
[0100] K.sub.2[RuCl.sub.5H.sub.2O].xH.sub.2O (1 g) was suspended in
HClO.sub.4 (15 cm.sup.3, 1 mM) and diethylenetriaminepentaacetic
acid (1.05 g) added. The reaction mixture was heated under reflux
for 1.5 hours forming a yellow/brown solution. On cooling a yellow
product crystallised which was collected by filtration, washed with
90% absolute ethanol/water, diethyl ether and dried in vacuo, yield
0.75 g, 53%.
[0101] Anal. calcd. for C.sub.14H.sub.21N.sub.3O.sub.10ClRu: C,
31.85; H, 3.98; N, 7.96; Cl, 6.73. Found: C, 29.77; H, 3.81; N,
7.36; Cl, 6.64.
[0102] Preparation of K[RuHHBEDCl]3H.sub.2O
[0103] 0.41 g of K.sub.2[RuCl.sub.5]xH.sub.2O was dissolved in
water (20 ml). To this solution was added 1 equivalent (0.39 g) of
N,N'di(2-hydroxy-benzyl)ethylene-diamine N,N-dacetic acid (hbed)
dissolved in water (50 ml) with KOH (0.12 g) and MeOH (1 ml). This
miixture was heated at reflux for 90 minutes. Upon cooling a dark,
insoluble precipitate formed. This material was removed by
filtration and the resulting red-violet solution was taken to
dryness by rotary evaporation. Trituration with water and washing
with acetone yielded 90 mg of a dark solid.
[0104] Anal. calcd. for C.sub.18H.sub.22N.sub.2O.sub.9RuClK: C,
36.89; H, 3.96; N, 4.78; Cl, 6.04. Found: C, 37.09; H, 4.23; N,
4.92; Cl, 6.28.
[0105] Preparation of
Ru(S.sub.2CNCH.sub.2CH.sub.2NMeCH.sub.2CH.sub.2).sub-
.31/2H.sub.2O
[0106] Me.sub.4N[S.sub.2CNCH.sub.2CH.sub.2NMeCH.sub.2CH.sub.2] was
made by the standard method and crystallised from methanol-ether in
71% yield.
[0107] RuCl.sub.3xH.sub.2O, 0.50 g, 2.15 mmol was refluxed in 30 ml
of methanol for 10 minutes and cooled. 1.87 g, 7.50 mmol of
Me.sub.4N[S.sub.2CNCH.sub.2CH.sub.2NMeCH.sub.2CH.sub.2] was added
and the mixture refluxed for 16 hours. After cooling 0.72 g of
crude product was filtered off, dissolved in dichloromethane and
filtered. The filtrate was loaded into 15 cc of basic alumina and
eluted with dichloromethane. Removal of solvent and crystallisation
from dichloromethane with ether by vapour-phase diffusion gave 0.51
g, 0.80 mmol, 37% of brown-black crystals,
Ru(S.sub.2CNCH.sub.2CH.sub.2NMeCH.sub.2CH.sub.2).sub.3{fraction
(1/2)}H.sub.2O.
[0108] Analysis for C.sub.18H.sub.34N.sub.6O.sub.5RuS.sub.6: Calc:
C, 34.00; H, 5.39; N, 13.22; S, 30.25. Found: C, 34.21; H, 5.47; N,
13.12; S, 30.36.
[0109] Preparation of
Ru[S.sub.2P(OC.sub.2H.sub.2OC.sub.2H.sub.4OMe).sub.2- ].sub.3
[0110]
K[S.sub.2P(OC.sub.2H.sub.4OC.sub.2H.sub.4OMe).sub.2].sub.3was made
by standard method and crystallised from methanol in 76% yield.
[0111] RuCl.sub.3xH.sub.2O, 1.00 g, 4.30 mmol was refluxed in 50 ml
of 0.1 N HCl with 1 ml of ethanol for 20 minutes and cooled. To
this solution was added 5.28 g (excess)
K[S.sub.2P(OC.sub.2H.sub.4OC.sub.2H.sub.4OMe).s- ub.2] and the
mixture stirred at 30.degree. C. for 1 hour. the reaction mixture
was extracted with dichloromethane and the solvent removed. The
residue was extracted with ether-hexane and solvents removed. This
residue was crystallised from 25 ml of hot ether by cooling to
-20.degree. C. giving 2.98 of red crystals. 2.41 g of the crude
product was purified by chromatography on 60 cc of silica gel with
5% ethanol in ether. The first band was collected, reduced to
dryness and crystallised from ether by cooling to -20.degree. C.
The yield of red crystals,
Ru(S.sub.2P{OC.sub.2H.sub.4OC.sub.2H.sub.4OMe}.sub.2).sub.3, was
2.16 g, 56%.
[0112] Analysis for C.sub.30H.sub.60O.sub.18P.sub.3RuS.sub.6: Calc:
C, 32.72; H, 6.04; S, 17.47. Found: C, 32.68; H, 6.08; S,
17.16.
[0113] In the in vitro tests, which were carried out in an
atmosphere of argon, each compound (1.times.10.sup.4 moles) was
dissolved in double-distilled deionized and deoxygenated water. The
resulting solution was placed in a three-necked pear-shaped flask
and stirred by a magnetic stirrer at constant speed of 1000 rpm, at
a constant temperature in the range 20.degree. C.-24.degree. C. A
manometer was attached to the flask, and purified, dried nitric
oxide gas (known volume in the range 3-5 cm.sup.3) was introduced
via a septum, using a gas syringe, at atmospheric pressure into the
headspace above the reaction solution. The pressure within the
flask was recorded periodically over a period of one hour.
[0114] A control experiment was carried out according to the above
but without any complex present.
[0115] The recorded pressures in association with the results of
the control experiment were analysed in order to determine the rate
of NO uptake as a function of time for each compound tested.
[0116] On completion of each in vitro test, the reaction solution
was freeze-dried. An infrared spectrum of the freeze-dried product
provided information on metal-NO bond formation.
[0117] In the in vitro cell culture tests, murine (RAW264)
macrophage cell lines, which can be induced to produce nitric
oxide, were seeded, 10.sup.6cells/well, onto 24 well culture plates
of 2 ml volume per well, in Eagles modified minimal essential
medium (MEM) plus 10% foetal bovine serum without phenol red.
[0118] The cells were activated to produce nitric oxide, with 10
.mu.g/ml lipopolysaccharide and 100 units/ml interferon .gamma. for
18 hours. Concurrently, test compounds made up in MEM were added at
non-cytotoxic concentrations.
[0119] Control cells as above, which were activated to produce
nitric oxide as above, but to which no test compound was added,
were used as a measure of the amount of nitric oxide produced by
the cells during the tests.
[0120] Background nitric oxide was assessed by measurement of
nitrate and nitrite in cells which were not activated.
[0121] Cell viability was confirmed by Trypan blue dye exclusion at
the end of the incubation period.
[0122] Nitric oxide was determined by measurement of nitrate and
nitrite in the cell supernatant. These anions are the stable
end-products of reactions of NO in solution. Such reactions may or
may not be catalysed in biological systems. The sum of nitrite and
nitrate concentrations gives the total NO production. Nitrite was
determined using the Griess reaction in which nitrite reacts with
1% sulphanilamide in 5% H.sub.3PO.sub.4/0.1%
naphthylethylenediamine dihydrochloride to form a chromophore
absorbing at 540 nm. Nitrate was determined by reducing nitrate to
nitrite with a bacterial nitrate reductase from Pseudomonas
oleovorans and then measuring nitrite with the Griess reaction. In
the absence of test compounds nitrite concentration plus nitrate
concentration is equal to total nitric oxide production. The effect
of test compounds on available nitric oxide (measured as
nitrite+nitrate) was detemined. The reduction in available nitric
oxide compared with the control level may be taken as an indication
of the degree of binding of NO by the test compounds.
[0123] In the ex vivo tests, segments of rat tail artery (0.8-1.5
cm) were dissected free from normotensive adult Wistar rats. The
arteries were internally perfused with Krebs solution (mM: NaCl
118, KCl 4.7, NaHCO.sub.3 25, NaH.sub.2PO.sub.4 1.15, CaCl.sub.2
2.5, MgCl.sub.2 1.1, glucose 5.6 and gassed with 95% O.sub.2/5%
CO.sub.2 to maintain a pH of 7.4) in a constant flow perfusion
apparatus. A differential pressure transducer located upstream of
the vessel detected changes in back pressure. The rat tail artery
preparation was pre-contracted with 6.5 .mu.M phenylephrine to give
a physiologically normal pressure of 100-120 mm Hg. The
pre-contracted vessels were then perfused with the test compound.
The arteries were perfused with Krebs solution between applications
of test compound to wash out the test compound.
[0124] Pressure changes in the system served to indicate artery
vasoconstriction. The vasoconstriction is a direct result of the
removal of endogenous nitric oxide (edrf) from the endothelial
cells of the rat tail artery.
RESULTS
[0125] The results of the in vitro, in vitro cell culture and
ex-vivo tests were as follows:
[0126] In Vitro Tests
EXAMPLE 1
K[Ru(Hedta)Cl]2H.sub.2O
[0127] A pressure decrease indicated binding of NO to the metal
compound. This is illustrated in FIG. 1.
[0128] The IR spectrum showed a peak at 1897 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 2
[Ru(H.sub.2edta)(acac)]
[0129] The IR spectrum showed a peak at 1896 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 3
K[Ru(hedtra)Cl]H.sub.2O
[0130] A pressure decrease indicated binding of NO to the metal
compound. This is illustrated in FIG. 1.
[0131] The IR spectrum showed a peak at 1889 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 4
K[Ru(dipic).sub.2]H.sub.2O
[0132] A pressure decrease indicated binding of NO to the metal
compound. This is illustrated in FIG. 1.
[0133] The IR spectrum showed a peak at 1915 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 5
(H.sub.2pic)[RuCl.sub.2(pic).sub.2](Hpic)H.sub.2O
[0134] The IR spectrum showed a peak at 1888 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 6
K[Ru(H.sub.2edta)Cl.sub.2]H.sub.2O
[0135] A pressure decrease indicated binding of NO to the metal
compound. This is illustrated in FIG. 1.
[0136] The IR spectrum showed a peak at 1896 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 7
K[Ru(Hnta).sub.2]1/2H.sub.2O
[0137] A pressure decrease indicated binding of NO to the metal
compound. This is illustrated in FIG. 1.
[0138] The IR spectrum showed a peak at 1889 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 8
K[Ru(H.sub.2dtpa)Cl]H.sub.2O
[0139] A pressure decrease indicated binding of NO to the metal
compound. This is illustrated in FIG. 1.
[0140] The IR spectrum showed a peak at 1905 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 9
[Ru.sub.3O(lac).sub.6](lac)
[0141] The IR spectrum showed a peak at 1884 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 10
[Ru.sub.3(OAc).sub.6](OAc)
[0142] The IR spectrum showed a peak at 1877 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 11
[RU.sub.2(OAc).sub.4]NO.sub.3
[0143] The IR spectrum showed a peak at 1891 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 12
[Ru(OCOEt).sub.4]NO.sub.3
[0144] The IR spectrum showed a peak at 1891 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 13
K.sub.3[Ru(ox).sub.3]
[0145] The IR spectrum showed a peak at 1889 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 14
[Ru.sub.2(OAc).sub.4]Cl
[0146] The IR spectrum showed a peak at 1895 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 15
[Ru(NH.sub.3).sub.5Cl]Cl.sub.2
[0147] The IR spectrum showed two peaks at 1909 cm.sup.-1 and 1928
cm.sup.-1, indicating the presence of a Ru--NO bond.
EXAMPLE 16
[Ru(en).sub.3]I.sub.3
[0148] The IR spectrum showed a peak at 1906 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 17
K[RuCl.sub.4(phen)]H.sub.2O
[0149] The IR spectrum showed a peak at 1904 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 18
[Ru(cyclam)Cl.sub.2]Cl
[0150] The IR spectrum showed a peak at 1895 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 19
K[RuCl.sub.4(bipy)]
[0151] The IR spectrum showed a peak at 1885 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 20
[RuCl.sub.3(dmso).sub.2(NH.sub.3)]
[0152] The IR spectrum showed a peak at 1889 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 21
[Ru(NH.sub.3).sub.6]Cl.sub.3
[0153] The IR spectrum showed a peak at 1918 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 22
cis-[RuCl.sub.2(dmso).sub.4]
[0154] The IR spectrum showed a peak at 1881 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 23
cis-[RuCl.sub.2(dmso).sub.3(NH.sub.3)]
[0155] The IR spectrum showed a peak at 1893 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 24
[RuCl.sub.3(dmso).sub.3]
[0156] The IR spectrum showed a peak at 1880 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 25
[Ru(mtc).sub.3]
[0157] The IR spectrum showed a peak at 1862 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 26
[Ru(maltol).sub.3]
[0158] The IR spectrum showed a peak at 1866 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 27
[Ru(acac).sub.2(MeCN).sub.2]CF.sub.3SO.sub.3
[0159] The IR spectrum showed a peak at 1899 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 28
K.sub.2[RuCl.sub.5(H.sub.2O)]
[0160] The IR spectrum showed a peak at 1903 cm.sup.-1, indicating
the presence of a Ru--NO bond.
EXAMPLE 29
[Os(ox)(bipy).sub.2]H.sub.2O
[0161] The IR spectrum showed a peak at 1894 cm.sup.-1, indicating
the presence of a Os--NO bond.
[0162] In Vitro Cell Culture Tests
[0163] Results are shown in Table 1 and FIG. 2.
EXAMPLE 1
K[Ru(Hedta)Cl]2H.sub.2O
[0164] Available nitric oxide was reduced in a dose-dependent
fashion with a maximum reduction of 75% at a concentration of 100
.mu.M.
EXAMPLE 2
[Ru(H.sub.2edta)(acac)]
[0165] Available nitric oxide was reduced by 82% at 100 .mu.M test
compound.
EXAMPLE 3
K[Ru(Hedtra)Cl]H.sub.2O
[0166] Available nitric oxide was reduced by 42% at 100 .mu.M.
EXAMPLE 6
K[Ru(H.sub.2edta)Cl.sub.2]H.sub.2O
[0167] Available nitric oxide was reduced by 77% at 100 .mu.M test
compound.
EXMAPLE 14
[Ru.sub.2(OAc).sub.4]Cl
[0168] Available nitric oxide was reduced by 47% at 100 .mu.M.
EXAMPLE 15
[Ru(NH.sub.3).sub.5Cl]Cl.sub.2
[0169] Available nitric oxide was reduced by 86% at 100 .mu.M test
compound.
EXAMPLE 26
[Ru(maltolato).sub.3]
[0170] Available nitric oxide was reduced by 71% at 100 .mu.M.
2 TABLE 1 % Decrease of Available Nitric Oxide Example 1 25 .mu.M
12 50 .mu.M 23 100 .mu.M 75 Example 2 100 .mu.M 82 Example 3 100
.mu.M 42 Example 6 100 .mu.M 77 Example 14 100 .mu.M 47 Example 15
100 .mu.M 86 Example 26 100 .mu.M 71
[0171] Ex-Vivo Tests
EXAMPLE 2
[0172] Application of test compound resulted in a dose-dependent
vasoconstriction at 10 .mu.M and 100 .mu.M. This effect was
reversible by washout with Krebs solution.
EXAMPLE 3
[0173] Application of test compound resulted in a dose-dependent
vasoconstriction at 10 .mu.M and 100 .mu.M. This effect was
reversible by washout with Krebs solution.
EXAMPLE 14
[0174] Application of test compound resulted in a dose-dependent
vasoconstriction at 10 .mu.M and 100 .mu.M. This effect was
reversible by washout with Krebs solution.
EXAMPLE 15
[0175] Application of test compound resulted in a dose-dependent
vasoconstriction at 10 .mu.M and 100 .mu.M. This effect was
reversible by washout with Krebs solution.
EXAMPLE 26
[0176] Application of test compound resulted in a dose-dependent
vasoconstrietion at 10 .mu.M and 100 .mu.M and 1000 .mu.M. This
effect was reversible by washout with Krebs solution.
3 TABLE 2 % Vasoconstriction Example 2 10 .mu.M 20 100 .mu.M 69
Example 3 10 .mu.M 17 100 .mu.M 59 Example 14 10 .mu.M 11 100 .mu.M
40 Example 15 10 .mu.M 16 100 .mu.M 86 Example 26 10 .mu.M 10 100
.mu.M 18 1000 .mu.M 25
* * * * *