U.S. patent application number 11/704421 was filed with the patent office on 2007-09-06 for compounds, compositions and methods for the treatment of inflammatory diseases.
Invention is credited to Joel Cummings, Virginia J. Sanders, Alan D. Snow.
Application Number | 20070208087 11/704421 |
Document ID | / |
Family ID | 38472216 |
Filed Date | 2007-09-06 |
United States Patent
Application |
20070208087 |
Kind Code |
A1 |
Sanders; Virginia J. ; et
al. |
September 6, 2007 |
Compounds, compositions and methods for the treatment of
inflammatory diseases
Abstract
The use of a composition comprising a compound: ##STR1## where R
is a C.sub.1-C.sub.10 alkylene group, in which, when the number of
carbon atoms is at least 2, there are optionally 1 or 2
non-adjacent double bonds; 1 to 3 non-adjacent methylene groups are
optionally replaced by NR.sup.1 (where R.sup.1 is H, alkyl, or
acyl), O, or S; and 1 or 2 methylene groups are optionally replaced
by a carbonyl or hydroxymethylene group for the preparation of a
medicament or a pharmaceutical for the treatment of inflammatory
diseases.
Inventors: |
Sanders; Virginia J.; (San
Francisco, CA) ; Cummings; Joel; (Seattle, WA)
; Snow; Alan D.; (Lynnwood, WA) |
Correspondence
Address: |
PROTEOTECH, INC.
12040 115TH AVE NE
KIRKLAND
WA
98034
US
|
Family ID: |
38472216 |
Appl. No.: |
11/704421 |
Filed: |
February 10, 2007 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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11503400 |
Jan 26, 2007 |
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11704421 |
Feb 10, 2007 |
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10077596 |
Feb 15, 2002 |
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11704421 |
Feb 10, 2007 |
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10053625 |
Nov 2, 2001 |
6929808 |
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10077596 |
Feb 15, 2002 |
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60707567 |
Aug 10, 2005 |
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Current U.S.
Class: |
514/734 |
Current CPC
Class: |
A61P 25/00 20180101;
A61P 1/04 20180101; A61P 3/10 20180101; A61P 19/06 20180101; A61P
17/06 20180101; A61P 19/02 20180101; A61P 1/00 20180101; A61P 29/00
20180101; A61K 31/055 20130101 |
Class at
Publication: |
514/734 |
International
Class: |
A61K 31/055 20060101
A61K031/055 |
Claims
1. Use of a composition for treatment of inflammatory diseases, the
composition comprising a compound ##STR45## where: R is a
C.sub.1-C.sub.10 alkylene group, in which, when the number of
carbon atoms is at least 2, there are optionally 1 or 2
non-adjacent double bonds; 1 to 3 non-adjacent methylene groups are
optionally replaced by NR.sup.1 (where R.sup.1 is H, alkyl, or
acyl), O, or S; and 1 or 2 methylene groups are optionally replaced
by a carbonyl or hydroxymethylene group.
2. The use of the composition of claim 1 where the compound is
selected from the group consisting of;
3,4,3.sup.1,4.sup.1-tetrahydroxybenzoin (compound 1);
3,4,3.sup.1,4.sup.1-tetrahydroxydesoxybenzoin (compound 2);
3,4,3.sup.1,4.sup.1-tetrahydroxydiphenylmethane (compound 3);
1,2-bis(3,4-dihydroxyphenyl)ethane (compound 4);
1,3-bis(3,4-dihydroxyphenyl)propoane (compound 5);
3,4,3.sup.1,4.sup.1-tetrahydroxychalcone (compound 6);
3,5-bis(3,4-dihydroxyphenyl)-1-methyl-2-pyrazoline (compound 7);
4,6-bis(3,4-dihydroxyphenyl)-3-cyano-2-methylpyridine (compound 8);
1,4-bis(3,4-dihydroxybenzyl)piperazine (compound 9);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-N,N.sup.1-dimethylethylenediamine
(compound 10);
2,5-bis(3,4-dihydroxybenzyl)-2,5-diaza[2.2.1]bicycloheptane
(compound 11);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-trans-1,2-diaminocyclohexane
(compound 12);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-trans-1,4-diaminocyclohexane
(compound 13);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-cis-1,3-bis(aminomethyl)cyclohex-
ane (compound 14); N-(3,4-dihydroxybenzyl)proline
3,4-dihydroxybenzylamide (compound 15);
2-(3,4-dihydroxybenzyl)isoquinoline-3-carboxylic acid
3,4-dihydroxyhenethylamide (compound 16);
2,6-bis(3,4-dihydroxybenzyl)cyclohexanone (compound 17);
3,5-bis(3,4-dihydroxybenzyl)-1-methyl-4-piperidinone (compound 18);
2,4-bis(3,4-dihydroxybenzyl)-3-tropinone (compound 19);
tris-(3,4-dihydroxybenzyl)methane (compound 20);
.alpha.-(3,4-dihydroxybenzamido)-3,4-dihydroxycinnamic acid
3,4-dihydroxybenzyl amide (compound 21);
4-(3,4-dihydroxybenzylaminomethylene)-2-(3,4-dihydroxyhenyl)oxazolin-5-on-
e (compound 22); 1,4-bis(3,4-dihydroxybenzoyl)iperazine (compound
23);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-N,N.sup.1-dimethylethylenediamine
(compound 24);
2,5-bis(3,4-dihydroxybenzoyl)-2,5-diaza[2.2.1]bicycloheptane
(compound 25);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-trans-1,2-diaminocyclohexane
(compound 26);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-cis-1,3-bis(aminomethyl)cyclohexane
(compound 27); 3,6-bis(3,4-dihydroxybenzyl)-2,5-diketopiperazine
(compound 28);
3,6-bis(3,4-dihydroxybenzylidene)-1,4-dimethyl-2,5-diketopiperazine
(compound 29); N-(3,4-dihydroxyhenylacetyl)proline
3,4-dihydroxyanilide (compound 30);
2,3-bis(3,4-dihydroxyhenyl)butane (compound 31);
1,3-bis(3,4-dihydroxybenzyl)benzene (compound 32);
1,4-bis(3,4-dihydroxybenzyl)benzene (compound 33);
2,6-bis(3,4-dihydroxybenzyl)yridine (compound 34);
2,5-bis(3,4-dihydroxybenzyl)thiophene (compound 35);
2,3-bis(3,4-dihydroxybenzyl)thiohene (compound 36);
1,2-bis(3,4-dihydroxyphenyl)cyclohexane (compound 37);
1,4-bis(3,4-dihydroxyphenyl)cyclohexane (compound 38);
3,7-bis(3,4-dihydroxyphenyl)bicyclo[3.3.0]octane (compound 39);
2,3-bis(3,4-dihydroxyphenyl)-1,7,7-trimethylgicyclo[2.2.1]heptane
(compound 40); 1,2-bis(3,4-dihydroxyphenoxy)ethane (compound 41);
1,3-bis(3,4-dihydroxyphenoxy)propane (compound 42);
trans-1,2-bis(3,4-dihydroxyphenoxy)-cyclopentane (compound 43);
N-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenoxy)-2-hydroxyropylamine
(compound 44); 3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyanilide
(compound 45); 3,4-dihydroxyphenoxyacetic acid
3,4-dihydroxybenzylamide (compound 46); 3,4-dihydroxyphenoxyacetic
acid 3,4-dihydroxyphenethylamide (compound 47);
3,4-dihydroxybenzoic acid p-(3,4-dihydroxyphenoxy)anilide (compound
48); 3,4-dihydroxybenzoic acid o-(3,4-dihydroxyphenoxy)anilide
(compound 49); 2,6-bis(3,4-dihydroxyphenoxy)pyridine (compound 50);
3,4-dihydroxybenzoic acid 3,4-dihydroxyanilide (compound 51);
3,4-dihydroxybenzoic acid 3,4-dihydroxybenzylamide (compound 52);
3,4-dihydroxybenzoic acid 3,4-dihydroxyphenethylamide (compound
53); 3,4-dihydroxyphenylacetic acid 3,4-dihydroxyanilide (compound
54); 3,4-dihydroxyphenylacetic acid 3,4-dihydroxybenzylamide
(compound 55); 3,4-dihydroxyphenylacetic acid
3,4-dihydroxyphenethylamide (compound 56);
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyanilide
(compound 57); 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxybenzaylamide (compound 58);
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyphenethylamide
(compound 59); 3,4-dihydroxycinnamic acid 3,4-dihydroxyanilide
(compound 60); 3,4-dihydroxycinnamic acid 3,4-dihydroxybenzylamide
(compound 61); 3,4-dihydroxycinnamic acid
3,4-dihydroxyphenethylamide (compound 62); oxalic acid
bis(3,4-dihydroxyanilide) (compound 63); oxalic acid
bis(3,4-dihydroxybenzylamie)(compound 64); oxalic acid
bis(3,4-dihydroxyphenethylamide) (compound 65); succinic acid
bis(3,4-dihydroxyanilide)(compound 66); succinic acid
bis(3,4-dihydroxybenzylamide) (compound 67); succinic acid
bis(3,4-dihydroxyphenethylamide) (compound 68); maleic acid
bis(3,4-dihydroxyanilide) (compound 69); maleic acid
bis(3,4-dihydroxyanilide) (compound 70); fumaric acid
bis(3,4-dihydroxyanilide) (compound 71); fumaric acid
bis(3,4-dihydroxybenzylamide) (compound 72);
bis(3,4-dihydroxybenzyl)amine (compound 73);
N(3,4-dihydroxybenzyl)-3,4-dihydroxyphenethylamine (compound 74);
tris(3,4-dihydroxybenzyl)amine (compound 75);
1,3-bis(3,4-dihydroxyhenzyl)urea (compound 76);
1-(3,4-dihydroxyphenzyl)-3-(3,4-dihydroxybenzyl)urea (compound 77);
1-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenethyl)urea (compound
78); 3-deoxy-3-(3,4-dihydroxybenzyl)aminoepicatechin (compound 79);
3-deoxy-3-(3,4-dihydroxyphenethyl)aminoepicatechin (compound 80);
2,3,6,7-tetrahydroxy-9,10-epoxy-9,10-dihydroacridine (compound 81);
10-aminoanthracene-1,2,7,8-tetraol (compound 82);
acridine-1,2,6,7-tetraol (compound 83);
phenoxazine-2,3,7,8,10-pentaol (compound 84);
dibenzo[c,f][2,7]napthyridine-2,3,10,11-tetraol (compound 85);
6-methyl-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline-2,10,11-trio-
l (compound 86); and methylenedioxy analogs and pharmaceutically
acceptable esters or salts of the compounds.
3. The use of the composition of claim 1 or 2 wherein the
inflammatory disease is selected from the group consisting of
ulcerative colitis, endotoxic shock, rheumatoid arthritis, juvenile
arthritis, osteoarthritis, psoriasis, Crohn's disease, inflammatory
bowel disease, multiple sclerosis, insulin dependent diabetes
mellitus, gout, psoriatic arthritis, reactive arthritis, viral or
post-viral arthritis and ankylosing spondylarthritis.
4. The use of the composition of claim 2 wherein the inflammatory
disease is rheumatoid arthritis.
5. Use of a composition for preparation of a medicament for
treatment of inflammatory diseases, the composition comprising a
compound ##STR46## where: R is a C.sub.1-C.sub.10 alkylene group,
in which, when the number of carbon atoms is at least 2, there are
optionally 1 or 2 non-adjacent double bonds; 1 to 3 non-adjacent
methylene groups are optionally replaced by NR.sup.1 (where R.sup.1
is H, alkyl, or acyl), O, or S; and 1 or 2 methylene groups are
optionally replaced by a carbonyl or hydroxymethylene group.
6. The use of claim 5 where the compounds are selected from the
group consisting of; 3,4,3.sup.1,4.sup.1-tetrahydroxybenzoin
(compound 1); 3,4,3.sup.1,4.sup.1-tetrahydroxydesoxybenzoin
(compound 2); 3,4,3.sup.1,4.sup.1-tetrahydroxydiphenylmethane
(compound 3); 1,2-bis(3,4-dihydroxyphenyl)ethane (compound 4);
1,3-bis(3,4-dihydroxyphenyl)propoane (compound 5);
3,4,3.sup.1,4.sup.1-tetrahydroxychalcone (compound 6);
3,5-bis(3,4-dihydroxyphenyl)-1-methyl-2-pyrazoline (compound 7);
4,6-bis(3,4-dihydroxyphenyl)-3-cyano-2-methylpyridine (compound 8);
1,4-bis(3,4-dihydroxybenzyl)piperazine (compound 9);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-N,N.sup.1-dimethylethylenediamine
(compound 10);
2,5-bis(3,4-dihydroxybenzyl)-2,5-diaza[2.2.1]bicycloheptane
(compound 11);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-trans-1,2-diaminocyclohexane
(compound 12);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-trans-1,4-diaminocyclohexane
(compound 13);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-cis-1,3-bis(aminomethyl)cyclohex-
ane (compound 14); N-(3,4-dihydroxybenzyl)proline
3,4-dihydroxybenzylamide (compound 15);
2-(3,4-dihydroxybenzyl)isoquinoline-3-carboxylic acid
3,4-dihydroxyhenethylamide (compound 16);
2,6-bis(3,4-dihydroxybenzyl)cyclohexanone (compound 17);
3,5-bis(3,4-dihydroxybenzyl)-1-methyl-4-piperidinone (compound 18);
2,4-bis(3,4-dihydroxybenzyl)-3-tropinone (compound 19);
tris-(3,4-dihydroxybenzyl)methane (compound 20);
.alpha.-(3,4-dihydroxybenzamido)-3,4-dihydroxycinnamic acid
3,4-dihydroxybenzyl amide (compound 21);
4-(3,4-dihydroxybenzylaminomethylene)-2-(3,4-dihydroxyhenyl)oxazolin-5-on-
e (compound 22); 1,4-bis(3,4-dihydroxybenzoyl)iperazine (compound
23);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-N,N.sup.1-dimethylethylenediamine
(compound 24);
2,5-bis(3,4-dihydroxybenzoyl)-2,5-diaza[2.2.1]bicycloheptane
(compound 25);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-trans-1,2-diaminocyclohexane
(compound 26);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-cis-1,3-bis(aminomethyl)cyclohexane
(compound 27); 3,6-bis(3,4-dihydroxybenzyl)-2,5-diketopiperazine
(compound 28);
3,6-bis(3,4-dihydroxybenzylidene)-1,4-dimethyl-2,5-diketopiperazine
(compound 29); N-(3,4-dihydroxyhenylacetyl)proline
3,4-dihydroxyanilide (compound 30);
2,3-bis(3,4-dihydroxyhenyl)butane (compound 31);
1,3-bis(3,4-dihydroxybenzyl)benzene (compound 32);
1,4-bis(3,4-dihydroxybenzyl)benzene (compound 33);
2,6-bis(3,4-dihydroxybenzyl)yridine (compound 34);
2,5-bis(3,4-dihydroxybenzyl)thiophene (compound 35);
2,3-bis(3,4-dihydroxybenzyl)thiohene (compound 36);
1,2-bis(3,4-dihydroxyphenyl)cyclohexane (compound 37);
1,4-bis(3,4-dihydroxyphenyl)cyclohexane (compound 38);
3,7-bis(3,4-dihydroxyphenyl)bicyclo[3.3.0]octane (compound 39);
2,3-bis(3,4-dihydroxyphenyl)-1,7,7-trimethylgicyclo[2.2.1]heptane
(compound 40); 1,2-bis(3,4-dihydroxyphenoxy)ethane (compound 41);
1,3-bis(3,4-dihydroxyphenoxy)propane (compound 42);
trans-1,2-bis(3,4-dihydroxyphenoxy)-cyclopentane (compound 43);
N-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenoxy)-2-hydroxyropylamine
(compound 44); 3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyanilide
(compound 45); 3,4-dihydroxyphenoxyacetic acid
3,4-dihydroxybenzylamide (compound 46); 3,4-dihydroxyphenoxyacetic
acid 3,4-dihydroxyphenethylamide (compound 47);
3,4-dihydroxybenzoic acid p-(3,4-dihydroxyphenoxy)anilide (compound
48); 3,4-dihydroxybenzoic acid o-(3,4-dihydroxyphenoxy)anilide
(compound 49); 2,6-bis(3,4-dihydroxyphenoxy)pyridine (compound 50);
3,4-dihydroxybenzoic acid 3,4-dihydroxyanilide (compound 51);
3,4-dihydroxybenzoic acid 3,4-dihydroxybenzylamide (compound 52);
3,4-dihydroxybenzoic acid 3,4-dihydroxyphenethylamide (compound
53); 3,4-dihydroxyphenylacetic acid 3,4-dihydroxyanilide (compound
54); 3,4-dihydroxyphenylacetic acid 3,4-dihydroxybenzylamide
(compound 55); 3,4-dihydroxyphenylacetic acid
3,4-dihydroxyphenethylamide (compound 56);
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyanilide
(compound 57); 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxybenzaylamide (compound 58);
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyphenethylamide
(compound 59); 3,4-dihydroxycinnamic acid 3,4-dihydroxyanilide
(compound 60); 3,4-dihydroxycinnamic acid 3,4-dihydroxybenzylamide
(compound 61); 3,4-dihydroxycinnamic acid
3,4-dihydroxyphenethylamide (compound 62); oxalic acid
bis(3,4-dihydroxyanilide) (compound 63); oxalic acid
bis(3,4-dihydroxybenzylamie)(compound 64); oxalic acid
bis(3,4-dihydroxyphenethylamide) (compound 65); succinic acid
bis(3,4-dihydroxyanilide)(compound 66); succinic acid
bis(3,4-dihydroxybenzylamide) (compound 67); succinic acid
bis(3,4-dihydroxyphenethylamide) (compound 68); maleic acid
bis(3,4-dihydroxyanilide) (compound 69); maleic acid
bis(3,4-dihydroxyanilide) (compound 70); fumaric acid
bis(3,4-dihydroxyanilide) (compound 71); fumaric acid
bis(3,4-dihydroxybenzylamide) (compound 72);
bis(3,4-dihydroxybenzyl)amine (compound 73);
N(3,4-dihydroxybenzyl)-3,4-dihydroxyphenethylamine (compound 74);
tris(3,4-dihydroxybenzyl)amine (compound 75);
1,3-bis(3,4-dihydroxyhenzyl)urea (compound 76);
1-(3,4-dihydroxyphenzyl)-3-(3,4-dihydroxybenzyl)urea (compound 77);
1-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenethyl)urea (compound
78); 3-deoxy-3-(3,4-dihydroxybenzyl)aminoepicatechin (compound 79);
3-deoxy-3-(3,4-dihydroxyphenethyl)aminoepicatechin (compound 80);
2,3,6,7-tetrahydroxy-9,10-epoxy-9,10-dihydroacridine (compound 81);
10-aminoanthracene-1,2,7,8-tetraol (compound 82);
acridine-1,2,6,7-tetraol (compound 83);
phenoxazine-2,3,7,8,10-pentaol (compound 84);
dibenzo[c,f][2,7]napthyridine-2,3,10,11-tetraol (compound 85);
6-methyl-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline-2,10,11-trio-
l (compound 86); and methylenedioxy analogs and pharmaceutically
acceptable esters or salts of the compounds.
7. The use of claims 5 or 6 wherein the inflammatory disease is
selected from the group consisting of ulcerative colitis, endotoxic
shock, rheumatoid arthritis, juvenile arthritis, osteoarthritis,
psoriasis, Crohn's disease, inflammatory bowel disease, multiple
sclerosis, insulin dependent diabetes mellitus, gout, psoriatic
arthritis, reactive arthritis, viral or post-viral arthritis and
ankylosing spondylarthritis.
8. The use of claim 5 wherein the inflammatory disease is
rheumatoid arthritis.
9. Use of a composition for inhibition of the inflammatory cascade,
the composition comprising a compound ##STR47## where: R is a
C.sub.1-C.sub.10 alkylene group, in which, when the number of
carbon atoms is at least 2, there are optionally 1 or 2
non-adjacent double bonds; 1 to 3 non-adjacent methylene groups are
optionally replaced by NR.sup.1 (where R.sup.1 is H, alkyl, or
acyl), O, or S; and 1 or 2 methylene groups are optionally replaced
by a carbonyl or hydroxymethylene group.
10. The use of the composition of claim 9 where the compound is
selected from the group consisting of;
3,4,3.sup.1,4.sup.1-tetrahydroxybenzoin (compound 1);
3,4,3.sup.1,4.sup.1-tetrahydroxydesoxybenzoin (compound 2);
3,4,3.sup.1,4.sup.1-tetrahydroxydiphenylmethane (compound 3);
1,2-bis(3,4-dihydroxyphenyl)ethane (compound 4);
1,3-bis(3,4-dihydroxyphenyl)propoane (compound 5);
3,4,3.sup.1,4.sup.1-tetrahydroxychalcone (compound 6);
3,5-bis(3,4-dihydroxyphenyl)-1-methyl-2-pyrazoline (compound 7);
4,6-bis(3,4-dihydroxyphenyl)-3-cyano-2-methylpyridine (compound 8);
1,4-bis(3,4-dihydroxybenzyl)piperazine (compound 9);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-N,N.sup.1-dimethylethylenediamine
(compound 10);
2,5-bis(3,4-dihydroxybenzyl)-2,5-diaza[2.2.1]bicycloheptane
(compound 11);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-trans-1,2-diaminocyclohexane
(compound 12);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-trans-1,4-diaminocyclohexane
(compound 13);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-cis-1,3-bis(aminomethyl)cyclohex-
ane (compound 14); N-(3,4-dihydroxybenzyl)proline
3,4-dihydroxybenzylamide (compound 15);
2-(3,4-dihydroxybenzyl)isoquinoline-3-carboxylic acid
3,4-dihydroxyhenethylamide (compound 16);
2,6-bis(3,4-dihydroxybenzyl)cyclohexanone (compound 17);
3,5-bis(3,4-dihydroxybenzyl)-1-methyl-4-piperidinone (compound 18);
2,4-bis(3,4-dihydroxybenzyl)-3-tropinone (compound 19);
tris-(3,4-dihydroxybenzyl)methane (compound 20);
.alpha.-(3,4-dihydroxybenzamido)-3,4-dihydroxycinnamic acid
3,4-dihydroxybenzyl amide (compound 21);
4-(3,4-dihydroxybenzylaminomethylene)-2-(3,4-dihydroxyhenyl)oxazolin-5-on-
e (compound 22); 1,4-bis(3,4-dihydroxybenzoyl)iperazine (compound
23);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-N,N.sup.1-dimethylethylenediamine
(compound 24);
2,5-bis(3,4-dihydroxybenzoyl)-2,5-diaza[2.2.1]bicycloheptane
(compound 25);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-trans-1,2-diaminocyclohexane
(compound 26);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-cis-1,3-bis(aminomethyl)cyclohexane
(compound 27); 3,6-bis(3,4-dihydroxybenzyl)-2,5-diketopiperazine
(compound 28);
3,6-bis(3,4-dihydroxybenzylidene)-1,4-dimethyl-2,5-diketopiperazine
(compound 29); N-(3,4-dihydroxyhenylacetyl)proline
3,4-dihydroxyanilide (compound 30);
2,3-bis(3,4-dihydroxyhenyl)butane (compound 31);
1,3-bis(3,4-dihydroxybenzyl)benzene (compound 32);
1,4-bis(3,4-dihydroxybenzyl)benzene (compound 33);
2,6-bis(3,4-dihydroxybenzyl)yridine (compound 34);
2,5-bis(3,4-dihydroxybenzyl)thiophene (compound 35);
2,3-bis(3,4-dihydroxybenzyl)thiohene (compound 36);
1,2-bis(3,4-dihydroxyphenyl)cyclohexane (compound 37);
1,4-bis(3,4-dihydroxyphenyl)cyclohexane (compound 38);
3,7-bis(3,4-dihydroxyphenyl)bicyclo[3.3.0]octane (compound 39);
2,3-bis(3,4-dihydroxyphenyl)-1,7,7-trimethylgicyclo[2.2.1]heptane
(compound 40); 1,2-bis(3,4-dihydroxyphenoxy)ethane (compound 41);
1,3-bis(3,4-dihydroxyphenoxy)propane (compound 42);
trans-1,2-bis(3,4-dihydroxyphenoxy)-cyclopentane (compound 43);
N-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenoxy)-2-hydroxyropylamine
(compound 44); 3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyanilide
(compound 45); 3,4-dihydroxyphenoxyacetic acid
3,4-dihydroxybenzylamide (compound 46); 3,4-dihydroxyphenoxyacetic
acid 3,4-dihydroxyphenethylamide (compound 47);
3,4-dihydroxybenzoic acid p-(3,4-dihydroxyphenoxy)anilide (compound
48); 3,4-dihydroxybenzoic acid o-(3,4-dihydroxyphenoxy)anilide
(compound 49); 2,6-bis(3,4-dihydroxyphenoxy)pyridine (compound 50);
3,4-dihydroxybenzoic acid 3,4-dihydroxyanilide (compound 51);
3,4-dihydroxybenzoic acid 3,4-dihydroxybenzylamide (compound 52);
3,4-dihydroxybenzoic acid 3,4-dihydroxyphenethylamide (compound
53); 3,4-dihydroxyphenylacetic acid 3,4-dihydroxyanilide (compound
54); 3,4-dihydroxyphenylacetic acid 3,4-dihydroxybenzylamide
(compound 55); 3,4-dihydroxyphenylacetic acid
3,4-dihydroxyphenethylamide (compound 56);
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyanilide
(compound 57); 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxybenzaylamide (compound 58);
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyphenethylamide
(compound 59); 3,4-dihydroxycinnamic acid 3,4-dihydroxyanilide
(compound 60); 3,4-dihydroxycinnamic acid 3,4-dihydroxybenzylamide
(compound 61); 3,4-dihydroxycinnamic acid
3,4-dihydroxyphenethylamide (compound 62); oxalic acid
bis(3,4-dihydroxyanilide) (compound 63); oxalic acid
bis(3,4-dihydroxybenzylamie)(compound 64); oxalic acid
bis(3,4-dihydroxyphenethylamide) (compound 65); succinic acid
bis(3,4-dihydroxyanilide)(compound 66); succinic acid
bis(3,4-dihydroxybenzylamide) (compound 67); succinic acid
bis(3,4-dihydroxyphenethylamide) (compound 68); maleic acid
bis(3,4-dihydroxyanilide) (compound 69); maleic acid
bis(3,4-dihydroxyanilide) (compound 70); fumaric acid
bis(3,4-dihydroxyanilide) (compound 71); fumaric acid
bis(3,4-dihydroxybenzylamide) (compound 72);
bis(3,4-dihydroxybenzyl)amine (compound 73);
N(3,4-dihydroxybenzyl)-3,4-dihydroxyphenethylamine (compound 74);
tris(3,4-dihydroxybenzyl)amine (compound 75);
1,3-bis(3,4-dihydroxyhenzyl)urea (compound 76);
1-(3,4-dihydroxyphenzyl)-3-(3,4-dihydroxybenzyl)urea (compound 77);
1-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenethyl)urea (compound
78); 3-deoxy-3-(3,4-dihydroxybenzyl)aminoepicatechin (compound 79);
3-deoxy-3-(3,4-dihydroxyphenethyl)aminoepicatechin (compound 80);
2,3,6,7-tetrahydroxy-9,10-epoxy-9,10-dihydroacridine (compound 81);
10-aminoanthracene-1,2,7,8-tetraol (compound 82);
acridine-1,2,6,7-tetraol (compound 83);
phenoxazine-2,3,7,8,10-pentaol (compound 84);
dibenzo[c,f][2,7]napthyridine-2,3,10,11-tetraol (compound 85);
6-methyl-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline-2,10,11-trio-
l (compound 86); and methylenedioxy analogs and pharmaceutically
acceptable esters or salts of the compounds.
11. Use of a composition for preparation of a medicament for
inhibition of the inflammatory cascade, the composition comprising
a compound ##STR48## where: R is a C.sub.1-C.sub.10 alkylene group,
in which, when the number of carbon atoms is at least 2, there are
optionally 1 or 2 non-adjacent double bonds; 1 to 3 non-adjacent
methylene groups are optionally replaced by NR.sup.1 (where R.sup.1
is H, alkyl, or acyl), O, or S; and 1 or 2 methylene groups are
optionally replaced by a carbonyl or hydroxymethylene group.
12. The use of the composition of claim 11 where the compound is
selected from the group consisting of;
3,4,3.sup.1,4.sup.1-tetrahydroxybenzoin (compound 1);
3,4,3.sup.1,4.sup.1 tetrahydroxydesoxybenzoin (compound 2);
3,4,3.sup.1,4.sup.1-tetrahydroxydiphenylmethane (compound 3);
1,2-bis(3,4-dihydroxyphenyl)ethane (compound 4);
1,3-bis(3,4-dihydroxyphenyl)propoane (compound 5);
3,4,3.sup.1,4.sup.1-tetrahydroxychalcone (compound 6);
3,5-bis(3,4-dihydroxyphenyl)-1-methyl-2-pyrazoline (compound 7);
4,6-bis(3,4-dihydroxyphenyl)-3-cyano-2-methylpyridine (compound 8);
1,4-bis(3,4-dihydroxybenzyl)piperazine (compound 9);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-N,N.sup.1-dimethylethylenediamine
(compound 10);
2,5-bis(3,4-dihydroxybenzyl)-2,5-diaza[2.2.1]bicycloheptane
(compound 11);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-trans-1,2-diaminocyclohexane
(compound 12);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-trans-1,4-diaminocyclohexane
(compound 13);
N,N.sup.1-bis(3,4-dihydroxybenzyl)-cis-1,3-bis(aminomethyl)cyclohex-
ane (compound 14); N-(3,4-dihydroxybenzyl)proline
3,4-dihydroxybenzylamide (compound 15);
2-(3,4-dihydroxybenzyl)isoquinoline-3-carboxylic acid
3,4-dihydroxyhenethylamide (compound 16);
2,6-bis(3,4-dihydroxybenzyl)cyclohexanone (compound 17);
3,5-bis(3,4-dihydroxybenzyl)-1-methyl-4-piperidinone (compound 18);
2,4-bis(3,4-dihydroxybenzyl)-3-tropinone (compound 19);
tris-(3,4-dihydroxybenzyl)methane (compound 20);
.alpha.-(3,4-dihydroxybenzamido)-3,4-dihydroxycinnamic acid
3,4-dihydroxybenzyl amide (compound 21);
4-(3,4-dihydroxybenzylaminomethylene)-2-(3,4-dihydroxyhenyl)oxazolin-5-on-
e (compound 22); 1,4-bis(3,4-dihydroxybenzoyl)iperazine (compound
23);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-N,N.sup.1-dimethylethylenediamine
(compound 24);
2,5-bis(3,4-dihydroxybenzoyl)-2,5-diaza[2.2.1]bicycloheptane
(compound 25);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-trans-1,2-diaminocyclohexane
(compound 26);
N,N.sup.1-bis(3,4-dihydroxybenzoyl)-cis-1,3-bis(aminomethyl)cyclohexane
(compound 27); 3,6-bis(3,4-dihydroxybenzyl)-2,5-diketopiperazine
(compound 28);
3,6-bis(3,4-dihydroxybenzylidene)-1,4-dimethyl-2,5-diketopiperazine
(compound 29); N-(3,4-dihydroxyhenylacetyl)proline
3,4-dihydroxyanilide (compound 30);
2,3-bis(3,4-dihydroxyhenyl)butane (compound 31);
1,3-bis(3,4-dihydroxybenzyl)benzene (compound 32);
1,4-bis(3,4-dihydroxybenzyl)benzene (compound 33);
2,6-bis(3,4-dihydroxybenzyl)yridine (compound 34);
2,5-bis(3,4-dihydroxybenzyl)thiophene (compound 35);
2,3-bis(3,4-dihydroxybenzyl)thiohene (compound 36);
1,2-bis(3,4-dihydroxyphenyl)cyclohexane (compound 37);
1,4-bis(3,4-dihydroxyphenyl)cyclohexane (compound 38);
3,7-bis(3,4-dihydroxyphenyl)bicyclo[3.3.0]octane (compound 39);
2,3-bis(3,4-dihydroxyphenyl)-1,7,7-trimethylgicyclo[2.2.1]heptane
(compound 40); 1,2-bis(3,4-dihydroxyphenoxy)ethane (compound 41);
1,3-bis(3,4-dihydroxyphenoxy)propane (compound 42);
trans-1,2-bis(3,4-dihydroxyphenoxy)-cyclopentane (compound 43);
N-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenoxy)-2-hydroxyropylamine
(compound 44); 3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyanilide
(compound 45); 3,4-dihydroxyphenoxyacetic acid
3,4-dihydroxybenzylamide (compound 46); 3,4-dihydroxyphenoxyacetic
acid 3,4-dihydroxyphenethylamide (compound 47);
3,4-dihydroxybenzoic acid p-(3,4-dihydroxyphenoxy)anilide (compound
48); 3,4-dihydroxybenzoic acid o-(3,4-dihydroxyphenoxy)anilide
(compound 49); 2,6-bis(3,4-dihydroxyphenoxy)pyridine (compound 50);
3,4-dihydroxybenzoic acid 3,4-dihydroxyanilide (compound 51);
3,4-dihydroxybenzoic acid 3,4-dihydroxybenzylamide (compound 52);
3,4-dihydroxybenzoic acid 3,4-dihydroxyphenethylamide (compound
53); 3,4-dihydroxyphenylacetic acid 3,4-dihydroxyanilide (compound
54); 3,4-dihydroxyphenylacetic acid 3,4-dihydroxybenzylamide
(compound 55); 3,4-dihydroxyphenylacetic acid
3,4-dihydroxyphenethylamide (compound 56);
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyanilide
(compound 57); 3-(3,4-dihydroxyphenyl) propionic acid
3,4-dihydroxybenzaylamide (compound 58);
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyphenethylamide
(compound 59); 3,4-dihydroxycinnamic acid 3,4-dihydroxyanilide
(compound 60); 3,4-dihydroxycinnamic acid 3,4-dihydroxybenzylamide
(compound 61); 3,4-dihydroxycinnamic acid
3,4-dihydroxyphenethylamide (compound 62); oxalic acid
bis(3,4-dihydroxyanilide) (compound 63); oxalic acid
bis(3,4-dihydroxybenzylamie)(compound 64); oxalic acid
bis(3,4-dihydroxyphenethylamide) (compound 65); succinic acid
bis(3,4-dihydroxyanilide)(compound 66); succinic acid
bis(3,4-dihydroxybenzylamide) (compound 67); succinic acid
bis(3,4-dihydroxyphenethylamide) (compound 68); maleic acid
bis(3,4-dihydroxyanilide) (compound 69); maleic acid
bis(3,4-dihydroxyanilide) (compound 70); fumaric acid
bis(3,4-dihydroxyanilide) (compound 71); fumaric acid
bis(3,4-dihydroxybenzylamide) (compound 72);
bis(3,4-dihydroxybenzyl)amine (compound 73);
N(3,4-dihydroxybenzyl)-3,4-dihydroxyphenethylamine (compound 74);
tris(3,4-dihydroxybenzyl)amine (compound 75);
1,3-bis(3,4-dihydroxyhenzyl)urea (compound 76);
1-(3,4-dihydroxyphenzyl)-3-(3,4-dihydroxybenzyl)urea (compound 77);
1-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenethyl)urea (compound
78); 3-deoxy-3-(3,4-dihydroxybenzyl)aminoepicatechin (compound 79);
3-deoxy-3-(3,4-dihydroxyphenethyl)aminoepicatechin (compound 80);
2,3,6,7-tetrahydroxy-9,10-epoxy-9,10-dihydroacridine (compound 81);
10-aminoanthracene-1,2,7,8-tetraol (compound 82);
acridine-1,2,6,7-tetraol (compound 83);
phenoxazine-2,3,7,8,10-pentaol (compound 84);
dibenzo[c,f][2,7]napthyridine-2,3,10,11-tetraol (compound 85);
6-methyl-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline-2,10,11-trio-
l (compound 86); and methylenedioxy analogs and pharmaceutically
acceptable esters or salts of the compounds.
Description
[0001] This application is a Continuation-in-Part of U.S.
application Ser. No. 11/503,400 filed Aug. 10, 2006, now abandoned,
which claimed priority to U.S. Provisional Application No.
60/707,567 filed Aug. 10, 2005. This application is also a
Continuation-in-Part of U.S. application Ser. No. 10/077,596 filed
Feb. 15, 2002, which is a Continuation-in-Part of application Ser.
No. 10/053,625, now issued as U.S. Pat. No. 6,929,808 on Aug. 16,
2005.
TECHNICAL FIELD
[0002] This invention relates to bis- and tris-dihydroxyaryl
compounds and their methylenedioxy analogs and pharmaceutically
acceptable esters, their synthesis, pharmaceutical compositions
containing them, and their use in the treatment of inflammatory
diseases such as arthritis, and in the manufacture of medicaments
for such treatment.
BACKGROUND OF THE INVENTION
[0003] Various forms of inflammation are characterized by
activation of macrophages. Macrophages are thought to induce and
maintain inflammatory processes mainly by producing various
products that, by acting on other cells, bring about the
deleterious consequences of inflammation such as the generation of
nitric oxide. For example, macrophages produce cytokines. These
proteins are central mediators in inflammatory processes, such as
the local inflammatory processes characteristic of arthritis or
colitis. Cytokines produced by macrophages are also thought to be
involved in systemic inflammatory processes, such as endotoxic
shock. Macrophage products are more generally involved in
pathophysiological mechanisms, such as plasma extravasation,
inflammatory cell diapedesis, release of toxic free radicals such
as nitric oxide, endothelial injury, and release of tissue
degrading enzymes, which result in tissue injury and, ultimately,
organ failure.
[0004] Tumor necrosis factor alpha (TNF-.alpha.) is a cytokine
associated with macrophage activation. TNF-.alpha. is also thought
to be involved in inducing most of the pathophysiological events
characteristic of inflammation. TNF-.alpha. plays an important role
in regulating inflammation, cellular immune response, and host
defense. TNF-.alpha. is a key cytokine associated with the toxic
effect of lipopolysaccharide (LPS) endotoxin and in the
pathogenesis of septic shock, as evidenced by high serum plasma
levels of TNF-.alpha. after LPS administration to animals or to
human volunteers, or in septic subjects. Administration of
anti-TNF-.alpha. antibodies protects against the lethal effects of
LPS and of live bacteria in a variety of animal models. Moreover,
TNF-.alpha. can be a central target in the treatment of rheumatoid
arthritis. Conversely in diseases such as rheumatoid arthritis,
osteoarthritis, psoriasis, Crohn's disease, inflammatory bowel
disease and other chronic disorders of the immune system, excessive
levels of TNF-.alpha. play a role in the pathophysiology. Indeed,
blocking TNF-.alpha. can halt disease progression and has led to
the search for inhibitors of TNF-.alpha..
[0005] Interleukin-12 (IL-12) is another macrophage product that
has been shown to be involved in the induction of pathology in
several inflammatory diseases. These diseases include autoimmune
diseases such as multiple sclerosis, inflammatory bowel disease,
insulin dependent diabetes mellitus, and rheumatoid arthritis, and
inflammatory states such as septic shock and the generalized
Schwarzman reaction.
[0006] Rheumatoid arthritis is a common rheumatic disease,
affecting more than two million people in the United States alone.
The disease is three times more prevalent in women as in men but
afflicts all races equally. The disease can begin at any age, but
most often starts between the ages of forty and sixty. In some
families, multiple members can be affected, suggesting a genetic
basis for the disorder. The cause of rheumatoid arthritis is
unknown. It is suspected that certain infections or factors in the
environment might trigger the immune system to attack the body's
own tissues, resulting in inflammation in various organs of the
body. Regardless of the exact trigger, the result is an immune
system that is geared up to promote inflammation in the joints and
occasionally other tissues of the body, Lymphocytes are activated
and cytokines, such as TNF-.alpha. and interleukin-1 (IL-1) are
expressed in the inflamed areas.
[0007] The clinical expression of rheumatoid arthritis is
manifested by chronic inflammation of the joints, the tissue
surrounding the joints such as the tendons, ligaments, and muscles,
as well as other organs in the body such as the eyes. The
inflammation process of causes swelling, pain stiffness, and
redness in the joints. In some patients with rheumatoid arthritis,
chronic inflammation leads to the destruction of the cartilage,
bone and ligaments causing deformity of the joints. Rheumatic
diseases can involve other, seemingly unrelated organs as well,
such as eyes, skin and glands. Rheumatic diseases are usually
divided into those that primarily involve joints, known as
arthritis, and those involving other tissues, generally referred to
connective tissue diseases. Arthritis is further subdivided into
inflammatory and non-inflammatory arthritis. The more common types
of inflammatory arthritis are rheumatoid arthritis, gout, psoriatic
arthritis (associated with the skin condition psoriasis), reactive
arthritis, viral or post-viral arthritis (occurring after an
infection), and spondylarthritis which affects the spine as well as
joints.
SUMMARY OF THE INVENTION
[0008] In a first aspect, this invention is bis- and
tris-dihydroxyaryl compounds and their methylenedioxy analogs and
pharmaceutically acceptable esters, and pharmaceutically acceptable
salts thereof. The compounds are useful in the treatment of
inflammatory diseases.
[0009] The compounds are: (1) compounds of the formula: ##STR2##
where: R is a C.sub.1-C.sub.10 alkylene group, in which, when the
number of carbon atoms is at least 2, there are optionally 1 or 2
non-adjacent double bonds; 1 to 3 non-adjacent methylene groups are
optionally replaced by NR' (where R' is H, alkyl, or acyl), O, or
S; and 1 or 2 methylene groups are optionally replaced by a
carbonyl or hydroxymethylene group; and (2) the compounds that are:
3,4,3',4'-tetrahydroxybenzoin (compound 1);
3,4,3',4'-tetrahydroxydesoxybenzoin (compound 2);
3,4,3',4'-tetrahydroxydiphenylmethane (compound 3);
1,2-bis(3,4-dihydroxyphenyl)ethane (compound 4);
1,3-bis(3,4-dihydroxyphenyl)propane (compound 5);
3,4,3',4'-tetrahydroxychalcone (compound 6);
3,5-bis(3,4-dihydroxyphenyl)-1-methyl-2-pyrazoline (compound 7);
4,6-bis(3,4-dihydroxyphenyl)-3-cyano-2-methylpyridine (compound 8);
1,4-bis(3,4-dihydroxybenzyl)piperazine (compound 9);
N,N'-bis(3,4-dihydroxybenzyl)-N,N'-dimethylethylenediamine
(compound 10);
2,5-bis(3,4-dihydroxybenzyl)-2,5-diaza[2.2.1]bicycloheptane
(compound 11);
N,N'-bis(3,4-dihydroxybenzyl)-trans-1,2-diaminocyclohexane
(compound 12);
N,N'-bis(3,4-dihydroxybenzyl)-trans-1,4-diaminocyclohexane
(compound 13);
N,N'-bis(3,4-dihydroxybenzyl)-cis-1,3-bis(aminomethyl)cyclohexane
(compound 14); N-(3,4-dihydroxybenzyl)proline
3,4-dihydroxybenzylamide (compound 15);
2-(3,4-dihydroxybenzyl)isoquinoline-3-carboxylic acid
3,4-dihydroxyphenethylamide (compound 16);
2,6-bis(3,4-dihydroxybenzyl)cyclohexanone (compound 17);
3,5-bis(3,4-dihydroxybenzyl)-1-methyl-4-piperidinone (compound 18);
2,4-bis(3,4-dihydroxybenzyl)-3-tropinone (compound 19);
tris-(3,4-dihydroxybenzyl)methane (compound 20);
.alpha.-(3,4-dihydroxybenzamido)-3,4-dihydroxycinnamic acid
3,4-dihydroxybenzyl amide (compound 21);
4-(3,4-dihydroxybenzylaminomethylene)-2-(3,4-dihydroxyphenyl)oxazolin-5-o-
ne (compound 22); 1,4-bis(3,4-dihydroxybenzoyl)piperazine (compound
23); N,N'-bis(3,4-dihydroxybenzoyl)-N,N'-dimethylethylenediamine
(compound 24);
2,5-bis(3,4-dihydroxybenzoyl)-2,5-diaza[2.2.1]bicycloheptane
(compound 25);
N,N'-bis(3,4-dihydroxybenzoyl)-trans-1,2-diaminocyclohexane
(compound 26);
N,N'-bis(3,4-dihydroxybenzoyl)-cis-1,3-bis(aminomethyl)cyclohexane
(compound 27); 3,6-bis(3,4-dihydroxybenzyl)-2,5-diketopiperazine
(compound 28);
3,6-bis(3,4-dihydroxybenzylidene)-1,4-dimethyl-2,5-diketopiperazine
(compound 29); N-(3,4-dihydroxyphenylacetyl)proline
3,4-dihydroxyanilide (compound 30);
2,3-bis(3,4-dihydroxyphenyl)butane (compound 31);
1,3-bis(3,4-dihydroxybenzyl)benzene (compound 32);
1,4-bis(3,4-dihydroxybenzyl)benzene (compound 33);
2,6-bis(3,4-dihydroxybenzyl)pyridine (compound 34);
2,5-bis(3,4-dihydroxybenzyl)thiophene (compound 35);
2,3-bis(3,4-dihydroxybenzyl)thiophene (compound 36);
1,2-bis(3,4-dihydroxyphenyl)cyclohexane (compound 37);
1,4-bis(3,4-dihydroxyphenyl)cyclohexane (compound 38);
3,7-bis(3,4-dihydroxyphenyl)bicyclo[3.3.0]octane (compound 39);
2,3-bis(3,4-dihydroxyphenyl)-1,7,7-trimethylbicyclo[2.2.1]heptane
(compound 40); 1,2-bis(3,4-dihydroxyphenoxy)ethane (compound 41);
1,3-bis(3,4-dihydroxyphenoxy)propane (compound 42);
trans-1,2-bis(3,4-dihydroxyphenoxy)-cyclopentane (compound 43);
N-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenoxy)-2-hydroxypropylamine
(compound 44); 3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyanilide
(compound 45); 3,4-dihydroxyphenoxyacetic acid
3,4-dihydroxybenzylamide (compound 46); 3,4-dihydroxyphenoxyacetic
acid 3,4-dihydroxyphenethylamide (compound 47);
3,4-dihydroxybenzoic acid p-(3,4-dihydroxyphenoxy)anilide (compound
48); 3,4-dihydroxybenzoic acid o-(3,4-dihydroxyphenoxy)anilide
(compound 49); 2,6-bis(3,4-dihydroxyphenoxy)pyridine (compound 50),
3,4-dihydroxybenzoic acid 3,4-dihydroxyanilide (compound 51);
3,4-dihydroxybenzoic acid 3,4-dihydroxybenzylamide (compound 52);
3,4-dihydroxybenzoic acid 3,4-dihydroxyphenethylamide (compound
53); 3,4-dihydroxyphenylacetic acid 3,4-dihydroxyanilide (compound
54); 3,4-dihydroxyphenylacetic acid 3,4-dihydroxybenzylamide
(compound 55); 3,4-dihydroxyphenylacetic acid
3,4-dihydroxyphenethylamide (compound 56);
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyanilide
(compound 57); 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxybenzylamide (compound 58);
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyphenethylamide
(compound 59); 3,4-dihydroxycinnamic acid 3,4-dihydroxyanilide
(compound 60); 3,4-dihydroxycinnamic acid 3,4-dihydroxybenzylamide
(compound 61); 3,4-dihydroxycinnamic acid
3,4-dihydroxyphenethylamide (compound 62); oxalic acid
bis(3,4-dihydroxyanilide) (compound 63); oxalic acid
bis(3,4-dihydroxybenzylamide) (compound 64); oxalic acid
bis(3,4-dihydroxyphenethylamide) (compound 65); succinic acid
bis(3,4-dihydroxyanilide) (compound 66); succinic acid
bis(3,4-dihydroxybenzylamide) (compound 67); succinic acid
bis(3,4-dihydroxyphenethylamide) (compound 68); maleic acid
bis(3,4-dihydroxyanilide) (compound 69); maleic acid
bis(3,4-dihydroxybenzylamide) (compound 70); fumaric acid
bis(3,4-dihydroxyanilide) (compound 71); fumaric acid
bis(3,4-dihydroxybenzylamide) (compound 72);
bis(3,4-dihydroxybenzyl)amine (compound 73);
N-(3,4-dihydroxybenzyl)-3,4-dihydroxyphenethylamine (compound 74);
tris(3,4-dihydroxybenzyl)amine (compound 75);
1,3-bis(3,4-dihydroxyphenyl)urea (compound 76);
1-(3,4-dihydroxyphenyl)-3-(3,4-dihydroxybenzyl)urea (compound 77);
1-(3,4-dihydroxyphenyl)-3-(3,4-dihydroxyphenethyl)urea (compound
78); 3-deoxy-3-(3,4-dihydroxybenzyl)aminoepicatechin (compound 79);
3-deoxy-3-(3,4-dihydroxyphenethyl)aminoepicatechin (compound 80);
2,3,6,7-tetrahydroxy-9,10-epoxy-9,10-dihydroacridine (compound 81);
10-aminoanthracene-1,2,7,8-tetraol (compound 82);
acridine-1,2,6,7-tetraol (compound 83);
phenoxazine-2,3,7,8,10-pentaol (compound 84);
dibenzo[c,f][2,7]napthyridine-2,3,10,11-tetraol (compound 85); and
6-methyl-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline-2,10,11-t-
riol (compound 86); (3) the methylenedioxy analogs and
pharmaceutically acceptable esters of compounds of (1) and (2); and
(4) the pharmaceutically acceptable salts of the compounds of (1)
to (3).
[0010] In a second aspect, this invention is pharmaceutical
compositions comprising a compound of the first aspect of this
invention and a pharmaceutically acceptable excipient; and
pharmaceutical compositions comprising a pharmaceutically
acceptable excipient and, as the sole active ingredient, a compound
of the first aspect of the invention.
[0011] In a third aspect, this invention is a method of treating
inflammatory diseases such as arthritis in mammals, especially
humans, by administration of a therapeutically effective amount of
a compound of the first aspect of this invention, for example as a
pharmaceutical composition.
[0012] In a fourth aspect, this invention is the use of a compound
of the first aspect of this invention in the manufacture of a
medicament for the treatment of inflammatory diseases such as
arthritis and use of a compound for inhibition of the inflammatory
cascade.
[0013] In a fifth aspect, this invention is a method of treatment
of inflammatory diseases such as arthritis and includes the step of
administering a therapeutically effective amount of a compound of
this invention. Preferably the compound is selected from the groups
described below with respect to their inhibitory activity of
microglial nitric oxide (NO) production and/or TNF-.alpha.
release.
[0014] The various compounds disclosed herein inhibit
lipopolysaccharide(LPS)/interferon gamma (IFN.gamma.) induced NO
release by microglia. This release by the microglia is in response
to LPS/IFN.gamma. pro-inflammatory stimulation. These compounds may
prevent oxidative stress induced cell death in addition to the
inhibition of NO and TNF-.alpha. release by microglia and
subsequent inhibition of the inflammatory cascade.
DETAILED DESCRIPTION OF THE INVENTION
Definitions
[0015] In this application, the following terms shall have the
following meanings, without regard to whether the terms are used
variantly elsewhere in the literature or otherwise in the known
art.
[0016] The compounds of the invention, i.e. the compounds of the
formula shown in the paragraph numbered (1) at the top of page 1 of
the application and the compounds on the list immediately following
and numbered (2) (compounds #1 to #86), are referred to generally
as bis- and tris-dihydroxyaryl compounds, or sometimes just as
"dihydroxyaryl compounds" and sometimes just "compounds". It will
be noted that compound #84 has an additional hydroxy group, but
does have two dihydroxyaryl groups; while compound #86 has only one
dihydroxyaryl group but has an additional phenolic hydroxyl
moiety.
[0017] "Methylenedioxy analogs" refers to the compounds of this
invention in which each of the pairs of adjacent hydroxyl moieties
of the dihydroxyaryl groups have been replaced by methylenedioxy
groups. The methylenedioxy compounds are illustrated and referred
to as compounds #1B to #86B. The methylenedioxy groups also are
convenient intermediate protecting groups for the dihydroxy
moieties and therefore these disclosed compounds are believed to
also serve as effective prodrugs. The methylenedioxy analogs #1B to
#80B are illustrated in Example 3.
[0018] "Pharmaceutically acceptable esters" refers to the compounds
of this invention where the hydroxyl moieties of the dihydroxyaryl
groups of the compounds are esterified with an acid or acids that
result in a pharmaceutically acceptable poly(ester). The compounds
are shown in Example 4 as acetylated, and these acetylated
compounds are illustrated and referred to as compounds #1C to #86C;
but it should be understood that the depiction of acetyl esters in
Example 4 is merely illustrative, and all pharmaceutically
acceptable esters are included within this invention. The ester
groups are expected to serve as intermediate protecting groups for
the hydroxyl moieties and therefore the pharmaceutically acceptable
esters are expected to serve as effective prodrugs for their
underlying bis- and tris-dihydroxyaryl compounds.
[0019] Chemical structures for each of the compounds of this
invention (with the note that the acetates are shown as
representative of the pharmaceutically acceptable esters as a
class) are shown. The names of the compounds are variously IUPAC
names [names derived according to the accepted IUPAC (International
Union of Pure and Applied Chemistry) system established by the
coalition of the Commission on Nomenclature of Organic Chemistry
and the Commission on Physical Organic Chemistry, as can be found
at http://www.chem.qmul.ac.uk/iupac], names derived from IUPAC
names by addition or substitution (for example, by the use of
"3,4-methylenedioxyphenyl" derived from "phenyl" instead of
"benzo[1,3]dioxol-5-yl"), and names derived from the names of
reactants (for example, by the use of "3,4-dihydroxybenzoic acid
3,4-dihydroxyanilide" instead of
"N-(3,4-dihydroxyphenyl)-3,4-dihydroxybenzamide"). However, the
names used are explicitly equated to chemical structures, and are
believed to be readily understood by a person of ordinary skill in
the art.
[0020] "Mammal" includes both humans and non-human mammals, such as
companion animals (cats, dogs, and the like), laboratory animals
(such as mice, rats, guinea pigs, and the like) and farm animals
(cattle, horses, sheep, goats, swine, and the like).
[0021] "Pharmaceutically acceptable excipient" means an excipient
that is conventionally useful in preparing a pharmaceutical
composition that is generally safe, non-toxic, and desirable, and
includes excipients that are acceptable for veterinary use as well
as for human pharmaceutical use. Such excipients may be solid,
liquid, semisolid, or, in the case of an aerosol composition,
gaseous.
[0022] "Pharmaceutically acceptable salt" means a salt that is
pharmaceutically acceptable and have the desired pharmacological
properties. Such salts include salts that may be formed where
acidic protons present in the compounds are capable of reacting
with inorganic or organic bases. Suitable inorganic salts include
those formed with the alkali metals, e.g. sodium and potassium,
magnesium, calcium, and aluminum. Suitable organic salts include
those formed with organic bases such as the amine bases, e.g.
ethanolamine, diethanolamine, triethanolamine, tromethamine,
N-methylglucamine, and the like. Such salts also include acid
addition salts formed with inorganic acids (e.g. hydrochloric and
hydrobromic acids) and organic acids (e.g. acetic acid, citric
acid, maleic acid, and the alkane- and arene-sulfonic acids such as
methanesulfonic acid and benzenesulfonic acid). When there are two
acidic groups present, a pharmaceutically acceptable salt may be a
mono-acid-mono-salt or a di-salt; and similarly where there are
more than two acidic groups present, some or all of such groups can
be salified.
[0023] A "therapeutically effective amount" in general means the
amount that, when administered to a subject or animal for treating
a disease, is sufficient to affect the desired degree of treatment
for the disease. A "therapeutically effective amount" or a
"therapeutically effective dosage" preferably inhibits, reduces,
disrupts, NO or TNF-.alpha. release, or treats a disease associated
with these conditions, such as an inflammatory disease, by at least
20%, more preferably by at least 40%, even more preferably by at
least 60%, and still more preferably by at least 80%, relative to
an untreated subject. Effective amounts of a compound of this
invention or composition thereof for treatment of a mammalian
subject are about 0.1 to about 1000 mg/Kg of body weight of the
subject/day, such as from about 1 to about 100 mg/Kg/day,
especially from about 10 to about 100 mg/Kg/day. A broad range of
disclosed composition dosages are believed to be both safe and
effective.
[0024] "Treating" or "treatment" of a disease includes preventing
the disease from occurring in a mammal that may be predisposed to
the disease but does not yet experience or exhibit symptoms of the
disease (prophylactic treatment), inhibiting the disease (slowing
or arresting its development), providing relief from the symptoms
or side-effects of the disease (including palliative treatment),
and relieving the disease (causing regression of the disease), such
as by disruption of the inflammatory cascade. One such preventive
treatment may be use of the disclosed compounds for the treatment
of inflammatory diseases.
[0025] "A pharmaceutical agent" or "pharmacological agent" or
"pharmaceutical composition" refers to a compound or combination of
compounds used for treatment, preferably in a pure or near pure
form. In the specification, pharmaceutical or pharmacological
agents include the compounds of this invention. The compounds are
desirably purified to 80% homogeneity, and preferably to 90%
homogeneity. Compounds and compositions purified to 99.9%
homogeneity are believed to be advantageous. As a test or
confirmation, a suitable homogeneous compound on HPLC would yield,
what those skilled in the art would identify as a single sharp-peak
band.
Compounds of the Invention
[0026] The compounds of this invention are: (1) compounds of the
formula: ##STR3## where: R is a C.sub.1-C.sub.10 alkylene group, in
which, when the number of carbon atoms is at least 2, there are
optionally 1 or 2 non-adjacent double bonds; 1 to 3 non-adjacent
methylene groups are optionally replaced by NR' (where R' is H,
alkyl, or acyl), O, or S; and 1 or 2 methylene groups are
optionally replaced by a carbonyl or hydroxymethylene group; and
(2) the compounds that are: 3,4,3',4'-tetrahydroxybenzoin;
3,4,3',4'-tetrahydroxydesoxybenzoin;
3,4,3',4'-tetrahydroxydiphenylmethane;
1,2-bis(3,4-dihydroxyphenyl)ethane;
1,3-bis(3,4-dihydroxyphenyl)propane;
3,4,3',4'-tetrahydroxychalcone;
3,5-bis(3,4-dihydroxyphenyl)-1-methyl-2-pyrazoline;
4,6-bis(3,4-dihydroxyphenyl)-3-cyano-2-methylpyridine;
1,4-bis(3,4-dihydroxybenzyl)piperazine;
N,N'-bis(3,4-dihydroxybenzyl)-N,N'-dimethylethylenediamine;
2,5-bis(3,4-dihydroxybenzyl)-2,5-diaza[2.2.1]bicycloheptane;
N,N'-bis(3,4-dihydroxybenzyl)-trans-1,2-diaminocyclohexane;
N,N'-bis(3,4-dihydroxybenzyl)-trans-1,4-diaminocyclohexane;
N,N'-bis(3,4-dihydroxybenzyl)-cis-1,3-bis(aminomethyl)cyclohexane;
N-(3,4-dihydroxybenzyl)proline 3,4-dihydroxybenzylamide;
2-(3,4-dihydroxybenzyl)isoquinoline-3-carboxylic acid
3,4-dihydroxyphenethylamide;
2,6-bis(3,4-dihydroxybenzyl)cyclohexanone;
3,5-bis(3,4-dihydroxybenzyl)-1-methyl-4-piperidinone;
2,4-bis(3,4-dihydroxybenzyl)-3-tropinone;
tris(3,4-dihydroxybenzyl)methane;
.alpha.-(3,4-dihydroxybenzamido)-3,4-dihydroxycinnamic acid
3,4-dihydroxybenzyl amide;
4-(3,4-dihydroxybenzylaminomethylene)-2-(3,4-dihydroxyphenyl)oxazolin-5-o-
ne; 1,4-bis(3,4-dihydroxybenzoyl)piperazine;
N,N'-bis(3,4-dihydroxybenzoyl)-N,N'-dimethylethylenediamine;
2,5-bis(3,4-dihydroxybenzoyl)-2,5-diaza[2.2.1]bicycloheptane;
N,N'-bis(3,4-dihydroxybenzoyl)-trans-1,2-diaminocyclohexane;
N,N'-bis(3,4-dihydroxybenzoyl)-cis-1,3-bis(aminomethyl)cyclohexane;
3,6-bis(3,4-dihydroxybenzyl)-2,5-diketopiperazine;
3,6-bis(3,4-dihydroxybenzylidene)-1,4-dimethyl-2,5-diketopiperazine;
N-(3,4-dihydroxyphenylacetyl)proline-3,4-dihydroxyanilide;
2,3-bis(3,4-dihydroxyphenyl)butane;
1,3-bis(3,4-dihydroxybenzyl)benzene;
1,4-bis(3,4-dihydroxybenzyl)benzene;
2,6-bis(3,4-dihydroxybenzyl)pyridine;
2,5-bis(3,4-dihydroxybenzyl)thiophene;
2,3-bis(3,4-dihydroxybenzyl)thiophene;
1,2-bis(3,4-dihydroxyphenyl)cyclohexane;
1,4-bis(3,4-dihydroxyphenyl)cyclohexane;
3,7-bis(3,4-dihydroxyphenyl)bicyclo[3.3.0]octane;
2,3-bis(3,4-dihydroxyphenyl)-1,7,7-trimethyl-bicyclo[2.2.1]heptane;
1,2-bis(3,4-dihydroxyphenoxy)ethane;
1,3-bis(3,4-dihydroxyphenoxy)propane;
trans-1,2-bis(3,4-dihydroxyphenoxy)cyclopentane;
N-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenoxy)-2-hydroxypropylamine;
3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyanilide;
3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxybenzylamide;
3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyphenethylamide;
3,4-dihydroxybenzoic acid p-(3,4-dihydroxyphenoxy)anilide;
3,4-dihydroxybenzoic acid o-(3,4-dihydroxyphenoxy)anilide;
2,6-bis(3,4-dihydroxyphenoxy)pyridine; 3,4-dihydroxybenzoic acid
3,4-dihydroxyanilide; 3,4-dihydroxybenzoic acid
3,4-dihydroxybenzylamide; 3,4-dihydroxybenzoic acid
3,4-dihydroxyphenethylamide; 3,4-dihydroxyphenyl acetic acid
3,4-dihydroxyanilide; 3,4-dihydroxyphenylacetic acid
3,4-dihydroxybenzylamide; 3,4-dihydroxyphenylacetic acid
3,4-dihydroxyphenethylamide; 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxyanilide; 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxybenzylamide; 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxyphenethylamide; 3,4-dihydroxycinnamic acid
3,4-dihydroxyanilide; 3,4-dihydroxycinnamic acid
3,4-dihydroxybenzylamide; 3,4-dihydroxycinnamic acid
3,4-dihydroxyphenethylamide; oxalic acid bis(3,4-dihydroxyanilide);
oxalic acid bis(3,4-dihydroxybenzylamide); oxalic acid
bis(3,4-dihydroxyphenethylamide); succinic acid
bis(3,4-dihydroxyanilide); succinic acid
bis(3,4-dihydroxybenzylamide); succinic acid
bis(3,4-dihydroxyphenethylamide); maleic acid
bis(3,4-dihydroxyanilide); maleic acid
bis(3,4-dihydroxybenzylamide); fumaric acid
bis(3,4-dihydroxyanilide); fumaric acid
bis(3,4-dihydroxybenzylamide); bis(3,4-dihydroxybenzyl)amine;
N-(3,4-dihydroxybenzyl)-3,4-dihydroxyphenethylamine;
tris(3,4-dihydroxybenzyl)amine; 1,3-bis(3,4-dihydroxyphenyl)urea;
1-(3,4-dihydroxyphenyl)-3-(3,4-dihydroxybenzyl)urea;
1-(3,4-dihydroxyphenyl)-3-(3,4-dihydroxyphenethyl)urea;
3-deoxy-3-(3,4-dihydroxybenzyl)aminoepicatechin;
3-deoxy-3-(3,4-dihydroxyphenethyl)aminoepicatechin;
2,3,6,7-tetrahydroxy-9,10-epoxy-9,10-dihydroacridine;
10-aminoanthracene-1,2,7,8-tetraol; acridine-1,2,6,7-tetraol;
phenoxazine-2,3,7,8,10-pentaol;
dibenzo[c,f][2,7]napthyridine-2,3,10,11-tetraol; and
6-methyl-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline-2,10,11-triol;
(3) the methylenedioxy analogs and pharmaceutically acceptable
esters of the compounds of (1) and (2); and (4) the
pharmaceutically acceptable salts of the compounds of (1) to
(3).
[0027] Within the compounds of this invention, a first group of
compounds is the compounds selected from the group consisting of:
(1) compounds of the formula: ##STR4## where: R is a
C.sub.1-C.sub.10, especially a C.sub.1-6, alkylene group, in which,
when the number of carbon atoms is at least 2, there are optionally
1 or 2 non-adjacent double bonds; 1 to 3 non-adjacent methylene
groups are optionally replaced by NR' (where R' is H, C.sub.1-3
alkyl, or C.sub.2-4 acyl), O, or S, especially NH or N--CH.sub.3;
and 1 or 2 methylene groups are optionally replaced by a carbonyl
or hydroxymethylene group; (2) the methylenedioxy analogs and
pharmaceutically acceptable tetraesters thereof; and (3) the
pharmaceutically acceptable salts of the compounds of (1) and
(2).
[0028] Within this first group, a subgroup of compounds is the
group of compounds selected from the group consisting of: (1)
compounds of the formula: ##STR5## where: R is a C.sub.2-C.sub.10,
especially a C.sub.2-6, alkylene group, in which there is
optionally 1 double bond; and 1 or 2 non-adjacent ethylene groups
are replaced by --C(O)NR'-- or --NR'C(O)-- (where R' is H or lower
alkyl); (2) the methylenedioxy analogs and pharmaceutically
acceptable tetraesters thereof; and (3) the pharmaceutically
acceptable salts of compounds of (1) and (2).
[0029] Within the compounds of this invention, a second group of
compounds is:
(1) the compounds that are:
[0030] 3,4,3',4'-tetrahydroxybenzoin;
3,4,3',4'-tetrahydroxydesoxybenzoin;
3,4,3',4'-tetrahydroxydiphenylmethane;
1,2-bis(3,4-dihydroxyphenyl)ethane;
1,3-bis(3,4-dihydroxyphenyl)propane;
3,4,3',4'-tetrahydroxychalcone;
3,5-bis(3,4-dihydroxyphenyl)-1-methyl-2-pyrazoline;
4,6-bis(3,4-dihydroxyphenyl)-3-cyano-2-methylpyridine;
1,4-bis(3,4-dihydroxybenzyl)piperazine;
N,N'-bis(3,4-dihydroxybenzyl)-N,N'-dimethylethylenediamine;
2,5-bis(3,4-dihydroxybenzyl)-2,5-diaza[2.2.1]bicycloheptane;
N,N'-bis(3,4-dihydroxybenzyl)-trans-1,2-diaminocyclohexane;
N,N'-bis(3,4-dihydroxybenzyl)-trans-1,4-diaminocyclohexane;
N,N'-bis(3,4-dihydroxybenzyl)-cis-1,3-bis(aminomethyl)cyclohexane;
N-(3,4-dihydroxybenzyl)proline 3,4-dihydroxybenzylamide;
2-(3,4-dihydroxybenzyl)isoquinoline-3-carboxylic acid
3,4-dihydroxyphenethylamide;
2,6-bis(3,4-dihydroxybenzyl)cyclohexanone;
3,5-bis(3,4-dihydroxybenzyl)-1-methyl-4-piperidinone;
2,4-bis(3,4-dihydroxybenzyl)-3-tropinone;
tris(3,4-dihydroxybenzyl)methane;
.alpha.-(3,4-dihydroxybenzamido)-3,4-dihydroxycinnamic acid
3,4-dihydroxybenzyl amide;
4-(3,4-dihydroxybenzylaminomethylene)-2-(3,4-dihydroxyphenyl)oxazolin-5-o-
ne; 1,4-bis(3,4-dihydroxybenzoyl)piperazine;
N,N'-bis(3,4-dihydroxybenzoyl)-N,N'-dimethylethylenediamine;
2,5-bis(3,4-dihydroxybenzoyl)-2,5-diaza[2.2.1]bicycloheptane;
N,N'-bis(3,4-dihydroxybenzoyl)-trans-1,2-diaminocyclohexane;
N,N'-bis(3,4-dihydroxybenzoyl)-cis-1,3-bis(aminomethyl)cyclohexane;
3,6-bis(3,4-dihydroxybenzyl)-2,5-diketopiperazine;
3,6-bis(3,4-dihydroxybenzylidene)-1,4-dimethyl-2,5-diketopiperazine;
N-(3,4-dihydroxyphenylacetyl)proline-3,4-dihydroxyanilide;
2,3-bis(3,4-dihydroxyphenyl)butane;
1,3-bis(3,4-dihydroxybenzyl)benzene;
1,4-bis(3,4-dihydroxybenzyl)benzene;
2,6-bis(3,4-dihydroxybenzyl)pyridine;
2,5-bis(3,4-dihydroxybenzyl)thiophene;
2,3-bis(3,4-dihydroxybenzyl)thiophene;
1,2-bis(3,4-dihydroxyphenyl)cyclohexane;
1,4-bis(3,4-dihydroxyphenyl)cyclohexane;
3,7-bis(3,4-dihydroxyphenyl)bicyclo[3.3.0]octane;
2,3-bis(3,4-dihydroxyphenyl)-1,7,7-trimethyl-bicyclo[2.2.1]heptane;
1,2-bis(3,4-dihydroxyphenoxy)ethane;
1,3-bis(3,4-dihydroxyphenoxy)propane;
trans-1,2-bis(3,4-dihydroxyphenoxy)cyclopentane;
N-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenoxy)-2-hydroxypropylamine;
3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyanilide;
3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxybenzylamide;
3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyphenethylamide;
3,4-dihydroxybenzoic acid p-(3,4-dihydroxyphenoxy)anilide;
3,4-dihydroxybenzoic acid o-(3,4-dihydroxyphenoxy)anilide;
2,6-bis(3,4-dihydroxyphenoxy)pyridine; 3,4-dihydroxybenzoic acid
3,4-dihydroxyanilide; 3,4-dihydroxybenzoic acid
3,4-dihydroxybenzylamide; 3,4-dihydroxybenzoic acid
3,4-dihydroxyphenethylamide; 3,4-dihydroxyphenyl acetic acid
3,4-dihydroxyanilide; 3,4-dihydroxyphenylacetic acid
3,4-dihydroxybenzylamide; 3,4-dihydroxyphenylacetic acid
3,4-dihydroxyphenethylamide; 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxyanilide; 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxybenzylamide; 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxyphenethylamide; 3,4-dihydroxycinnamic acid
3,4-dihydroxyanilide; 3,4-dihydroxycinnamic acid
3,4-dihydroxybenzylamide; 3,4-dihydroxycinnamic acid
3,4-dihydroxyphenethylamide; oxalic acid bis(3,4-dihydroxyanilide);
oxalic acid bis(3,4-dihydroxybenzylamide); oxalic acid
bis(3,4-dihydroxyphenethylamide); succinic acid
bis(3,4-dihydroxyanilide); succinic acid
bis(3,4-dihydroxybenzylamide); succinic acid
bis(3,4-dihydroxyphenethylamide); maleic acid
bis(3,4-dihydroxyanilide); maleic acid
bis(3,4-dihydroxybenzylamide); fumaric acid
bis(3,4-dihydroxyanilide); fumaric acid
bis(3,4-dihydroxybenzylamide); bis(3,4-dihydroxybenzyl)amine;
N-(3,4-dihydroxybenzyl)-3,4-dihydroxyphenethylamine;
tris(3,4-dihydroxybenzyl)amine; 1,3-bis(3,4-dihydroxyphenyl)urea;
1-(3,4-dihydroxyphenyl)-3-(3,4-dihydroxybenzyl)urea;
1-(3,4-dihydroxyphenyl)-3-(3,4-dihydroxyphenethyl)urea;
3-deoxy-3-(3,4-dihydroxybenzyl)aminoepicatechin;
3-deoxy-3-(3,4-dihydroxyphenethyl)aminoepicatechin;
2,3,6,7-tetrahydroxy-9,10-epoxy-9,10-dihydroacridine;
10-aminoanthracene-1,2,7,8-tetraol; acridine-1,2,6,7-tetraol;
phenoxazine-2,3,7,8,10-pentaol;
dibenzo[c,f][2,7]napthyridine-2,3,10,11-tetraol; and
6-methyl-5,6,6a,7-tetrahydro-4H-dibenzo[de,g]quinoline-2,10,11-triol;
(2) the methylenedioxy analogs and pharmaceutically acceptable
esters thereof; and
(3) the pharmaceutically acceptable salts of the compounds of (1)
and (2).
[0031] Within this second group, a subgroup of compounds is:
(1) the compounds that are:
[0032] 3,4,3',4'-tetrahydroxybenzoin;
3,4,3',4'-tetrahydroxydesoxybenzoin;
3,4,3',4'-tetrahydroxydiphenylmethane;
1,2-bis(3,4-dihydroxyphenyl)ethane;
1,3-bis(3,4-dihydroxyphenyl)propane;
3,4,3',4'-tetrahydroxychalcone;
3,5-bis(3,4-dihydroxyphenyl)-1-methyl-2-pyrazoline;
4,6-bis(3,4-dihydroxyphenyl)-3-cyano-2-methylpyridine;
1,4-bis(3,4-dihydroxybenzyl)piperazine;
N,N'-bis(3,4-dihydroxybenzyl)-N,N'-dimethylethylenediamine;
2,5-bis(3,4-dihydroxybenzyl)-2,5-diaza[2.2.1]bicycloheptane;
N,N'-bis(3,4-dihydroxybenzyl)-trans-1,2-diaminocyclohexane;
N,N'-bis(3,4-dihydroxybenzyl)-trans-1,4-diaminocyclohexane;
N,N'-bis(3,4-dihydroxybenzyl)-cis-1,3-bis(aminomethyl)cyclohexane;
N-(3,4-dihydroxybenzyl)proline 3,4-dihydroxybenzylamide;
2-(3,4-dihydroxybenzyl)isoquinoline-3-carboxylic acid
3,4-dihydroxyphenethylamide;
2,6-bis(3,4-dihydroxybenzyl)cyclohexanone;
3,5-bis(3,4-dihydroxybenzyl)-1-methyl-4-piperidinone;
2,4-bis(3,4-dihydroxybenzyl)-3-tropinone;
tris(3,4-dihydroxybenzyl)methane;
.alpha.-(3,4-dihydroxybenzamido)-3,4-dihydroxycinnamic acid
3,4-dihydroxybenzyl amide;
4-(3,4-dihydroxybenzylaminomethylene)-2-(3,4-dihydroxyphenyl)oxazolin-5-o-
ne; 1,4-bis(3,4-dihydroxybenzoyl)piperazine;
N,N'-bis(3,4-dihydroxybenzoyl)-N,N'-dimethylethylenediamine;
2,5-bis(3,4-dihydroxybenzoyl)-2,5-diaza[2.2.1]bicycloheptane;
N,N'-bis(3,4-dihydroxybenzoyl)-trans-1,2-diaminocyclohexane;
N,N'-bis(3,4-dihydroxybenzoyl)-cis-1,3-bis(aminomethyl)cyclohexane;
3,6-bis(3,4-dihydroxybenzyl)-2,5-diketopiperazine;
3,6-bis(3,4-dihydroxybenzylidene)-1,4-dimethyl-2,5-diketopiperazine;
N-(3,4-dihydroxyphenylacetyl)proline-3,4-dihydroxyanilide;
2,3-bis(3,4-dihydroxyphenyl)butane;
1,3-bis(3,4-dihydroxybenzyl)benzene;
1,4-bis(3,4-dihydroxybenzyl)benzene;
2,6-bis(3,4-dihydroxybenzyl)pyridine;
2,5-bis(3,4-dihydroxybenzyl)thiophene;
2,3-bis(3,4-dihydroxybenzyl)thiophene;
1,2-bis(3,4-dihydroxyphenyl)cyclohexane;
1,4-bis(3,4-dihydroxyphenyl)cyclohexane;
3,7-bis(3,4-dihydroxyphenyl)bicyclo[3.3.0]octane;
2,3-bis(3,4-dihydroxyphenyl)-1,7,7-trimethyl-bicyclo[2.2.1]heptane;
1,2-bis(3,4-dihydroxyphenoxy)ethane;
1,3-bis(3,4-dihydroxyphenoxy)propane;
trans-1,2-bis(3,4-dihydroxyphenoxy)cyclopentane;
N-(3,4-dihydroxybenzyl)-3-(3,4-dihydroxyphenoxy)-2-hydroxypropylamine;
3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyanilide;
3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxybenzylamide;
3,4-dihydroxyphenoxyacetic acid 3,4-dihydroxyphenethylamide;
3,4-dihydroxybenzoic acid p-(3,4-dihydroxyphenoxy)anilide;
3,4-dihydroxybenzoic acid o-(3,4-dihydroxyphenoxy)anilide;
2,6-bis(3,4-dihydroxyphenoxy)pyridine; 3,4-dihydroxybenzoic acid
3,4-dihydroxyanilide; 3,4-dihydroxybenzoic acid
3,4-dihydroxybenzylamide; 3,4-dihydroxybenzoic acid
3,4-dihydroxyphenethylamide; 3,4-dihydroxyphenyl acetic acid
3,4-dihydroxyanilide; 3,4-dihydroxyphenylacetic acid
3,4-dihydroxybenzylamide; 3,4-dihydroxyphenylacetic acid
3,4-dihydroxyphenethylamide; 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxyanilide; 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxybenzylamide; 3-(3,4-dihydroxyphenyl)propionic acid
3,4-dihydroxyphenethylamide; 3,4-dihydroxycinnamic acid
3,4-dihydroxyanilide; 3,4-dihydroxycinnamic acid
3,4-dihydroxybenzylamide; 3,4-dihydroxycinnamic acid
3,4-dihydroxyphenethylamide; oxalic acid bis(3,4-dihydroxyanilide);
oxalic acid bis(3,4-dihydroxybenzylamide); oxalic acid
bis(3,4-dihydroxyphenethylamide); succinic acid
bis(3,4-dihydroxyanilide); succinic acid
bis(3,4-dihydroxybenzylamide); succinic acid
bis(3,4-dihydroxyphenethylamide); maleic acid
bis(3,4-dihydroxyanilide); maleic acid
bis(3,4-dihydroxybenzylamide); fumaric acid
bis(3,4-dihydroxyanilide); fumaric acid
bis(3,4-dihydroxybenzylamide); bis(3,4-dihydroxybenzyl)amine;
N-(3,4-dihydroxybenzyl)-3,4-dihydroxyphenethylamine;
tris(3,4-dihydroxybenzyl)amine; 1,3-bis(3,4-dihydroxyphenyl)urea;
1-(3,4-dihydroxyphenyl)-3-(3,4-dihydroxybenzyl)urea;
1-(3,4-dihydroxyphenyl)-3-(3,4-dihydroxyphenethyl)urea;
3-deoxy-3-(3,4-dihydroxybenzyl)aminoepicatechin; and
3-deoxy-3-(3,4-dihydroxyphenethyl)aminoepicatechin;
(2) the methylenedioxy analogs and pharmaceutically acceptable
esters thereof; and
(3) the pharmaceutically acceptable salts of the compounds of (1)
and (2).
[0033] Within this subgroup, a further subgroup is:
(1) the compounds that are:
[0034] 3,4,3',4'-tetrahydroxybenzoin;
3,4,3',4'-tetrahydroxydiphenylmethane;
1,2-bis(3,4-dihydroxyphenyl)ethane;
4,6-bis(3,4-dihydroxyphenyl)-3-cyano-2-methylpyridine;
1,4-bis(3,4-dihydroxybenzyl)piperazine;
N,N'-bis(3,4-dihydroxybenzyl)-trans-1,2-diaminocyclohexane;
2,4-bis(3,4-dihydroxybenzyl)-3-tropinone;
.alpha.-(3,4-dihydroxybenzamido)-3,4-dihydroxycinnamic acid
3,4-dihydroxybenzyl amide; 1,4-bis(3,4-dihydroxybenzoyl)piperazine;
N,N'-bis(3,4-dihydroxybenzoyl)-trans-1,2-diaminocyclohexane;
3,4-dihydroxybenzoic acid 3,4-dihydroxyanilide;
3,4-dihydroxybenzoic acid 3,4-dihydroxybenzylamide;
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxyanilide;
3-(3,4-dihydroxyphenyl)propionic acid 3,4-dihydroxybenzylamide;
3,4-dihydroxycinnamic acid 3,4-dihydroxybenzylamide; oxalic acid
bis(3,4-dihydroxyanilide); succinic acid bis(3,4-dihydroxyanilide);
succinic acid bis(3,4-dihydroxybenzylamide);
bis(3,4-dihydroxybenzyl)amine; tris(3,4-dihydroxybenzyl)amine;
1,3-bis(3,4-dihydroxyphenyl)urea; and
1-(3,4-dihydroxyphenyl)-3-(3,4-dihydroxyphenethyl)urea;
(2) the methylenedioxy analogs and pharmaceutically acceptable
esters thereof; and
(3) the pharmaceutically acceptable salts of the compounds of (1)
and (2).
[0035] Within each of these groups and subgroups, there are
especially the compounds of the invention that are the bis- and
tris(dihydroxyaryl) compounds (i.e. the compounds of the formula or
of the list) and compound #86, and their pharmaceutically
acceptable salts.
Synthesis of the Compounds of the Invention
[0036] The compounds of this invention may be prepared by methods
generally known to the person of ordinary skill in the art, having
regard to that knowledge and the disclosure of this
application.
[0037] The starting materials and reagents used in preparing these
compounds are either available from commercial suppliers such as
the Aldrich Chemical Company (Milwaukee, Wis.), Bachem (Torrance,
Calif.), Sigma (St. Louis, Mo.), or Lancaster Synthesis Inc.
(Windham, N.H.) or are prepared by methods well known to a person
of ordinary skill in the art, following procedures described in
such references as Fieser and Fieser's Reagents for Organic
Synthesis, vols. 1-17, John Wiley and Sons, New York, N.Y., 1991;
Rodd's Chemistry of Carbon Compounds, vols. 1-5 and supps.,
Elsevier Science Publishers, 1989; Organic Reactions, vols. 1-40,
John Wiley and Sons, New York, N.Y., 1991; March J.: Advanced
Organic Chemistry, 4th ed., John Wiley and Sons, New York, N.Y.;
and Larock: Comprehensive Organic Transformations, VCH Publishers,
New York, 1989.
[0038] In most cases, protective groups for the hydroxy groups are
introduced and finally removed. Suitable protective groups are
described in Greene et al., Protective Groups in Organic Synthesis,
Second Edition, John Wiley and Sons, New York, 1991. A preferred
protective group is the methylenedioxy group, as seen in many of
Examples 1-23, and a wide variety of methylenedioxyphenyl compounds
(such as 3,4-methylenedioxyacetophenone, 3,4-methylenedioxyaniline,
3,4-methylenedioxybenzaldehyde, 3,4-methylenedioxybenzoic acid,
3,4-methylenedioxybenzonitrile, 3,4-methylenedioxybenzoic acid,
3,4-methylenedioxybenzoyl chloride, 3,4-methylenedioxycinnamic
acid, 3,4-methylenedioxynitrobenzene, 3,4-methylenedioxyphenol,
3,4-methylenedioxyphenylacetic acid,
3,4-methylenedioxyphenylacetonitrile, 3,4-methylenedioxyphenyl
isocyanate, 3,4-methylenedioxyphenylmagnesium bromide, and
3,4-methylenedioxyphenylmethanol) are commercially available. Other
protecting groups, such as the benzyl and methoxymethyl groups, may
also be used.
[0039] Other starting materials or early intermediates may be
prepared by elaboration of the materials listed above, for example,
by methods well known to a person of ordinary skill in the art.
[0040] The starting materials, intermediates, and compounds of this
invention may be isolated and purified using conventional
techniques, including precipitation, filtration, distillation,
crystallization, chromatography, and the like. The compounds may be
characterized using conventional methods, including physical
constants and spectroscopic methods.
Pharmacology and Utility
[0041] The compounds of this invention, either as the dihydroxyaryl
compounds per se, or as the methylenedioxy analogs or
pharmaceutically acceptable esters (once de-protected either in the
body or in vitro), act to inhibit or prevent microglial NO and/or
TNF-.alpha.. Their activity can be measured in vitro by methods
such as those discussed herein.
[0042] Compounds of special interest for treating inflammatory
diseases such as arthritis are selected from the group consisting
of
(1) the compounds that are:
[0043] 3,4,3',4'-tetrahydroxybenzoin;
3,4,3',4'-tetrahydroxydiphenylmethane;
1,2-bis(3,4-dihydroxyphenyl)ethane;
N,N'-bis(3,4-dihydroxybenzyl)-trans-1,2-diaminocyclohexane;
.alpha.-(3,4-dihydroxybenzamido)-3,4-dihydroxycinnamic acid
3,4-dihydroxybenzylamide; 3,4-dihydroxybenzoic acid
3,4-dihydroxyanilide; bis(3,4-dihydroxybenzyl)amine;
1,3-bis(3,4-dihydroxyphenyl)urea; and
1-(3,4-dihydroxyphenyl)-3-(3,4-dihydroxyphenethyl)urea;
(2) the methylenedioxy analogs and pharmaceutically acceptable
esters thereof; and
(3) the pharmaceutically acceptable salts of the compounds of (1)
and (2).
[0044] Especially of interest are the compounds of (1) above and
their pharmaceutically acceptable salts.
Pharmaceutical Compositions and Administration
[0045] In general, compounds of the invention will be administered
in therapeutically effective amounts by any of the usual modes
known in the art, either singly or in combination with at least one
other compound of this invention and/or at least one other
conventional therapeutic agent for the disease being treated. A
therapeutically effective amount may vary widely depending on the
disease, its severity, the age and relative health of the animal
being treated, the potency of the compound(s), and other factors.
As anti-inflammatory agents, therapeutically effective amounts of
compounds of this invention may range from 0.1-1000 mg/Kg body
weight/day, such as from 1-100 mg/Kg/day; for example, 10-100
mg/Kg/day. A person of ordinary skill in the art will be
conventionally able, and without undue experimentation, having
regard to that skill and to this disclosure, to determine a
therapeutically effective amount of a compound for the treatment of
inflammatory diseases such as arthritis.
[0046] Preferred compositions will contain a compound of this
invention that is at least substantially pure. In general "pure"
means better than 95% pure, and "substantially pure" means a
compound synthesized such that the compound, as made as available
for consideration into a therapeutic dosage, has only those
impurities that can not readily nor reasonably be removed by
conventional purification processes.
[0047] In general, the compounds of this invention will be
administered as pharmaceutical compositions by one of the following
routes: oral, topical, systemic (e.g. transdermal, intranasal, or
by suppository), or parenteral (e.g. intramuscular, subcutaneous,
or intravenous injection). Compositions may take the form of
tablets, pills, capsules, semisolids, powders, sustained release
formulations, solutions, suspensions, elixirs, aerosols, or any
other appropriate compositions; and comprise at least one compound
of this invention in combination with at least one pharmaceutically
acceptable excipient. Suitable excipients are well known to persons
of ordinary skill in the art, and they, and the methods of
formulating the compositions, may be found in such standard
references as Remington: The Science and Practice of Pharmacy, A.
Gennaro, ed., 20th edition, Lippincott, Williams & Wilkins,
Philadelphia, Pa. Suitable liquid carriers, especially for
injectable solutions, include water, aqueous saline solution,
aqueous dextrose solution, and glycols.
[0048] In particular, the compound(s)--optimally only one such
compound is administered in any particular dosage form--can be
administered, orally, for example, as tablets, troches, lozenges,
aqueous or oily suspension, dispersible powders or granules,
emulsions, hard or soft capsules, or syrups or elixirs.
Compositions intended for oral use may be prepared according to any
method known in the art for the manufacture of pharmaceutical
compositions and such compositions may contain one or more agents
selected from the group consisting of sweetening agents, flavoring
agents, coloring agents and preserving agents in order to provide
pharmaceutically elegant and palatable preparations.
[0049] Tablets contain the compound in admixture with non-toxic
pharmaceutically acceptable excipients that are suitable for the
manufacture of tablets. These excipients may be for example, inert
diluents, such as calcium carbonate, sodium carbonate, lactose,
calcium phosphate or sodium phosphate; granulating and
disintegrating agents, for example, maize starch or alginic acid;
binding agents, for example, maize starch, gelatin or acacia, and
lubricating agents, for example, magnesium stearate or stearic acid
or tale. The tablets may be uncoated or they may be coated by known
techniques to delay disintegration and absorption in the
gastrointestinal tract and thereby provide a sustained action over
a longer period. For example, a time delay material such as
glycerol monostearate or glycerol distearate may be employed.
Formulations for oral use may also be presented as hard gelatin
capsules wherein the compound is mixed with an inert solid diluent,
for example, calcium carbonate, calcium phosphate or kaolin, or as
soft gelatin capsules wherein the active ingredient is mixed with
water or an oil medium, for example, peanut oil, liquid paraffin or
olive oil.
[0050] Aqueous suspensions contain the compound in admixture with
excipients suitable for the manufacture of aqueous suspensions.
Such excipients are suspending agents, for example, sodium
carboxymethylcellulose, methylcellulose, hydroxypropylmethyl
cellulose, sodium alginate, polyvinylpyrrolidone, gum tragacanth
and gum acacia; dispersing or wetting agents may be naturally
occurring phosphatides, for example lecithin, or condensation
products of an alkylene oxide with fatty acids, for example
polyoxyethylene stearate, or condensation products of ethylene
oxide with long chain aliphatic alcohols, for example,
heptadecaethyleneoxycetanol, or condensation products of ethylene
oxide with partial esters derived from fatty acids such as hexitol
such as polyoxyethylene sorbitol monooleate, or condensation
products of ethylene oxide with partial esters from fatty acids and
a hexitol anhydrides, for example, polyethylene sorbitan
monooleate. The aqueous suspensions may also contain one or more
preservatives, for example, ethyl or n-propyl p-hydroxybenzoate,
one or more coloring agents, one or more flavoring agents, or one
or more sweetening agents, such as sucrose or saccharin.
[0051] Oily suspensions may be formulated by suspending the
compound in a vegetable oil, for example arachis oil, olive oil,
sesame oil, or coconut oil or in a mineral oil such as liquid
paraffin. The oily suspensions may contain a thickening agent, for
example beeswax, hard paraffin or cetyl alcohol. Sweetening agents,
such as those set forth below, and flavoring agents may be added to
provide a palatable oral preparation. These compositions may be
preserved by the addition of an antioxidant such as ascorbic acid.
Dispersible powders and granules suitable for preparation of an
aqueous suspension by the addition of water provide the active
ingredient in admixture with a dispersing or wetting agent, a
suspending agent and one or more preservatives. Suitable dispersing
or wetting agents and suspending agents are exemplified by those
already described above. Additional excipients, for example
sweetening, flavoring and agents, may also be present.
[0052] The compounds of the invention may also be in the form of
oil-in-water emulsions. The oily phase may be a vegetable oil, for
example olive oil or arachis oils, or a mineral oil, for example
liquid paraffin or mixtures of these. Suitable emulsifying agents
may be naturally-occurring gums, for example gum acacia or gum
tragacanth, naturally occurring phosphatides, for example soy bean,
lecithin, and occurring phosphatides, for example soy bean,
lecithin, and esters or partial esters derived from fatty acids and
hexitol anhydrides, for example sorbitan monooleate, and
condensation products of the said partial esters with ethylene
oxide, for example polyoxyethylene sorbitan monooleate. The
emulsion may also contain sweetening and flavoring agents. Syrups
and elixirs may be formulated with sweetening agents, for example,
glycerol, sorbitol or sucrose. Such formulations may also contain a
demulcent, a preservative and flavoring and coloring agents.
[0053] The compounds of the invention can also be administered by
injection or infusion, either subcutaneously or intravenously, or
intramuscularly, or intrasternally, or intranasally, or by infusion
techniques in the form of sterile injectable or oleaginous
suspension. The compound may be in the form of a sterile injectable
aqueous or oleaginous suspensions. These suspensions may be
formulated according to the known art using suitable dispersing of
wetting agents and suspending agents that have been described
above. The sterile injectable preparation may also be a sterile
injectable solution or suspension in a non-toxic
parenterally-acceptable diluent or solvent for example, as a
solution in 1,3-butanediol. Among the acceptable vehicles and
solvents that may be employed are water, Ringer's solution and
isotonic sodium chloride solution. In addition, sterile, fixed oils
are conventionally employed as a solvent or suspending medium. For
this purpose any bland fixed oils may be conventionally employed
including synthetic mono- or diglycerides. In addition fatty acids
such as oleic acid find use in the preparation of injectables.
Dosage regimens can be adjusted to provide the optimum therapeutic
response. For example, several divided dosages may be administered
daily or the dosage may be proportionally reduced as indicated by
the exigencies of the therapeutic situation.
[0054] It is especially advantageous to formulate the compounds in
dosage unit form for ease of administration and uniformity of
dosage. Dosage unit form as used herein refers to physically
discrete units suited as unitary dosages for the subjects to be
treated; each containing a therapeutically effective quantity of
the compound and at least one pharmaceutical excipient. A drug
product will comprise a dosage unit form within a container that is
labelled or accompanied by a label indicating the intended method
of treatment, such as the treatment of an inflammatory disease such
as arthritis.
Sustained Release Formulations
[0055] The invention also includes the use of sustained release
formulations to deliver the compounds of the present invention to
the desired target (i.e. systemic organs) at high circulating
levels (between 10.sup.-9 and 10.sup.-4 M) are also disclosed.
[0056] It is understood that the compound levels are maintained
over a certain period of time as is desired and can be easily
determined by one skilled in the art using this disclosure and
compounds of the invention. In a preferred embodiment, the
invention includes a unique feature of administration comprising a
sustained release formulation so that a constant level of
therapeutic compound is maintained between 10.sup.-8 and 10.sup.-6M
between 48 to 96 hours in the sera.
[0057] Such sustained and/or timed release formulations may be made
by sustained release means of delivery devices that are well known
to those of ordinary skill in the art, such as those described in
U.S. Pat. Nos. 3,845,770; 3,916,899; 3,536,809; 3,598,123;
4,008,719; 4,710,384; 5,674,533; 5,059,595; 5,591,767; 5,120,548;
5,073,543; 5,639,476; 5,354,556 and 5,733,566, the disclosures of
which are each incorporated herein by reference. These
pharmaceutical compositions can be used to provide slow or
sustained release of one or more of the active compounds using, for
example, hydroxypropylmethyl cellulose, other polymer matrices,
gels, permeable membranes, osmotic systems, multilayer coatings,
microparticles, liposomes, microspheres, or the like. Suitable
sustained release formulations known to those skilled in the art,
including those described herein may be readily selected for use
with the pharmaceutical compositions of the invention. Thus, single
unit dosage forms suitable for oral administration, such as, but
not limited to, tablets, capsules, gelcaps, caplets, powders and
the like, that are adapted for sustained release are encompassed by
the present invention.
[0058] In a preferred embodiment, the sustained release formulation
contains active compound such as, but not limited to,
microcrystalline cellulose, maltodextrin, ethylcellulose, and
magnesium stearate. As described above, all known methods for
encapsulation which are compatible with properties of the disclosed
compounds are encompassed by this invention. The sustained release
formulation is encapsulated by coating particles or granules of the
pharmaceutical composition of the invention with varying thickness
of slowly soluble polymers or by microencapsulation. In a preferred
embodiment, the sustained release formulation is encapsulated with
a coating material of varying thickness (e.g. about 1 micron to 200
microns) that allow the dissolution of the pharmaceutical
composition about 48 hours to about 72 hours after administration
to a mammal. In another embodiment, the coating material is a
food-approved additive.
[0059] In another embodiment, the sustained release formulation is
a matrix dissolution device that is prepared by compressing the
drug with a slowly soluble polymer carrier into a tablet. In one
preferred embodiment, the coated particles have a size range
between about 0.1 to about 300 microns, as disclosed in U.S. Pat.
Nos. 4,710,384 and 5,354,556, which are incorporated herein by
reference in their entireties. Each of the particles is in the form
of a micromatrix, with the active ingredient uniformly distributed
throughout the polymer.
[0060] Sustained release formulations such as those described in
U.S. Pat. No. 4,710,384, which is incorporated herein by reference
in its entirety, having a relatively high percentage of plasticizer
in the coating in order to permit sufficient flexibility to prevent
substantial breakage during compression are disclosed. The specific
amount of plasticizer varies depending on the nature of the coating
and the particular plasticizer used. The amount may be readily
determined empirically by testing the release characteristics of
the tablets formed. If the medicament is released too quickly, then
more plasticizer is used. Release characteristics are also a
function of the thickness of the coating. When substantial amounts
of plasticizer are used, the sustained release capacity of the
coating diminishes. Thus, the thickness of the coating may be
increased slightly to make up for an increase in the amount of
plasticizer. Generally, the plasticizer in such an embodiment will
be present in an amount of about 15 to 30% of the sustained release
material in the coating, preferably 20 to 25%, and the amount of
coating will be from 10 to 25% of the weight of the active
material. Preferably 15 to 20%. Any conventional pharmaceutically
acceptable plasticizer may be incorporated into the coating.
[0061] The compounds of the invention can be formulated as a
sustained and/or timed release formulation. All sustained release
pharmaceutical products have a common goal of improving drug
therapy over that achieved by their non-sustained counterparts.
Ideally, the use of an optimally designed sustained release
preparation in medical treatment is characterized by a minimum of
drug substance being employed to cure or control the condition.
Advantages of sustained release formulations may include: 1)
extended activity of the composition, 2) reduced dosage frequency,
and 3) increased patient compliance. In addition, sustained release
formulations can be used to affect the time of onset of action or
other characteristics, such as blood levels of the composition, and
thus can affect the occurrence of side effects.
[0062] The sustained release formulations of the invention are
designed to initially release an amount of the therapeutic
composition that promptly produces the desired therapeutic effect,
and gradually and continually release of other amounts of
compositions to maintain this level of therapeutic effect over an
extended period of time. In order to maintain this constant level
in the body, the therapeutic composition must be released from the
dosage form at a rate that will replace the composition being
metabolized and excreted from the body.
[0063] The sustained release of an active ingredient may be
stimulated by various inducers, for example pH, temperature,
enzymes, water, or other physiological conditions or compounds. The
term "sustained release component" in the context of the present
invention is defined herein as a compound or compounds, including,
but not limited to, polymers, polymer matrices, gels, permeable
membranes, liposomes, microspheres, or the like, or a combination
thereof, that facilitates the sustained release of the active
ingredient.
[0064] If the complex is water-soluble, it may be formulated in an
appropriate buffer, for example, phosphate buffered saline, or
other physiologically compatible solutions. Alternatively, if the
resulting complex has poor solubility in aqueous solvents, then it
may be formulated with a non-ionic surfactant such as Tween, or
polyethylene glycol. Thus, the compounds and their physiologically
solvents may be formulated for administration by inhalation or
insufflation (either through the mouth or the nose) or oral,
buccal, parenteral, or rectal administration, as examples.
[0065] Preparations for oral administration may be suitably
formulated to give controlled release of the active compound. In a
preferred embodiment, the compounds of the present invention are
formulated as controlled release powders of discrete microparticles
that can be readily formulated in liquid form. The sustained
release powder comprises particles containing an active ingredient
and optionally, an excipient with at least one non-toxic
polymer.
[0066] The powder can be dispersed or suspended in a liquid vehicle
and will maintain its sustained release characteristics for a
useful period of time. These dispersions or suspensions have both
chemical stability and stability in terms of dissolution rate. The
powder may contain an excipient comprising a polymer, which may be
soluble, insoluble, permeable, impermeable, or biodegradable. The
polymers may be polymers or copolymers. The polymer may be a
natural or synthetic polymer. Natural polymers include polypeptides
(e.g., zein), polysaccharides (e.g., cellulose), and alginic acid.
Representative synthetic polymers include those described, but not
limited to, those described in column 3, lines 33-45 of U.S. Pat.
No. 5,354,556, which is incorporated by reference in its entirety.
Particularly suitable polymers include those described, but not
limited to those described in column 3, line 46-column 4, line 8 of
U.S. Pat. No. 5,354,556 which is incorporated by reference in its
entirety.
[0067] The sustained release compounds of the invention may be
formulated for parenteral administration, e.g., by intramuscular
injections or implants for subcutaneous tissues and various body
cavities and transdermal devices. In one embodiment, intramuscular
injections are formulated as aqueous or oil suspensions. In an
aqueous suspension, the sustained release effect is due to, in
part, a reduction in solubility of the active compound upon
complexation or a decrease in dissolution rate. A similar approach
is taken with oil suspensions and solutions, wherein the release
rate of an active compound is determined by partitioning of the
active compound out of the oil into the surrounding aqueous medium.
Only active compounds which are oil soluble and have the desired
partition characteristics are suitable. Oils that may be used for
intramuscular injection include, but are not limited to, sesame,
olive, arachis, maize, almond, soybean, cottonseed and castor
oil.
[0068] A highly developed form of drug delivery that imparts
sustained release over periods of time ranging from days to years
is to implant a drug-bearing polymeric device subcutaneously or in
various body cavities. The polymer material used in an implant,
which must be biocompatible and nontoxic, include but are not
limited to hydrogels, silicones, polyethylenes, ethylene-vinyl
acetate copolymers, or biodegradable polymers.
General Experimental Procedures
[0069] All solvents were distilled before use and were removed by
rotary evaporation at temperatures up to 35.degree. C. Octadecyl
functionalized silica gel (C18) was used for reversed-phase (RP)
flash chromatography, and Merck silica gel 60, 200-400 mesh, 40-63
.mu.m, was used for silica gel flash chromatography. Thin layer
chromatography (TLC) was carried out using Merck DC-plastikfolien
Kieselgel 60 F.sub.254, first visualized with a UV lamp, and then
by dipping in a vanillin solution (1% vanillin, 1% H.sub.2SO.sub.4
in ethanol), and heating. Optical rotations were measured on a
Perkin-Elmer 241 polarimeter. Mass spectra were recorded on a
Kratos MS-80 instrument. NMR spectra, at 25.degree. C., were
recorded at 500 or 300 MHz for .sup.1H and 125 or 75 MHz for
.sup.13C on Varian INOVA-500 or VXR-300 spectrometers. Chemical
shifts are given in ppm on the delta scale referenced to the
solvent peaks CHCl.sub.3 at 7.25 and CDCl.sub.3 at 77.0 ppm,
(CH.sub.3).sub.2CO at 2.15 and (CD.sub.3).sub.2CO at 30.5 ppm, or
CH.sub.3OD at 3.30 and CD.sub.3OD at 39.0 ppm.
HPLC Conditions
[0070] The analytical HPLC equipment consisted of a Waters 717
autosampler, 600 pump and controller, and a 2487 UV detector
controlled by Omega software. Samples were analyzed by using an
RP-18 semi-preparative column (Phenomenex Prodigy 5 mm C18 100A,
250.times.4.6 mm) with a guard column (Phenomenex SecurityGuard
cartridge containing a C18 ODS 4.times.3 mm, 5 mm column) fitted at
30.degree. C. Samples (5 ml) were analyzed using a mobile phase
flow rate of 5.0 ml/min, with UV detection at 280 nm.
TABLE-US-00001 Method 1 Time (minutes) CH.sub.3CN H.sub.2O
containing 0.1% TFA 0 11 89 20 11 89 30 100 0 31 11 89 40 11 89
[0071] TABLE-US-00002 Method 2 CH.sub.3CN/H.sub.2O (95:5) Time
(minutes) containing 0.1% TFA H.sub.2O containing 0.1% TFA 0 11 89
20 11 89 30 100 0 31 11 89 40 11 89
[0072] The following non-limiting Examples are given by way of
illustration only and are not considered a limitation of this
invention, many apparent variations of which are possible without
departing from the spirit or scope thereof.
EXAMPLE 1
Bis- and Tris-Dihydroxyaryl Compounds of the Invention
[0073] This Example describes bis- and tris(dihydroxyaryl)
compounds that serve as potent inhibitors of inflammation and in
particular the release of NO and TNF-.alpha. from microglial cells.
A common structural motif that is present in all of the compounds
disclosed herein is the presence of two or three dihydroxyaryl
groups. These compounds are generally indicated on succeeding pages
and identified variously herein by simple number. ##STR6## ##STR7##
##STR8## ##STR9## ##STR10## ##STR11## ##STR12## ##STR13## ##STR14##
##STR15## ##STR16## ##STR17## ##STR18## ##STR19##
EXAMPLE 2
Compounds of the Invention with Rigid Scaffolds
[0074] This Example illustrates six further compounds of this
invention; compounds #81, 82, 83, 84, 85, and 86. These compounds
have relatively rigid scaffold structures. ##STR20##
EXAMPLE 3
Methylenedioxy Analogs
[0075] A strategy for the delivery of the dihydroxyaryl compounds
of this invention to improve and/or cause more favorable metabolism
and bioavailability characteristics involves the protection of the
hydroxy groups of the dihydroxyaryl compounds with methylenedioxy
groups. This strategy is exemplified in the 80 structures shown
below, and is equally applicable to protect the dihydroxyaryl
groups of compounds #81-86. Methylenedioxy analogs represent
intermediate hydroxy protecting structures that are made to
successfully complete the synthesis of the dihydroxyaryl compounds
described in the invention. These closed-ring compounds also tend
to be more stable, and hydrophobic (water insoluble), and less
likely to be altered or degraded due to the oxidation that could
occur if hydroxyl groups were present. In addition, these compounds
make good prodrugs for delivery. Hydrophobic compounds that are
lipid soluble tend to be attractive compounds for delivery since
they are usually able to penetrate the blood-brain-barrier.
[0076] The methylenedioxy analogs are generally available as
intermediates in the synthesis of the corresponding dihydroxyaryl
compounds. These compounds are expected to be efficacious once the
methylenedioxy structures are cleaved to yield hydroxyl groups.
Conversion of the hydroxyl groups to methylenedioxy derivatives
also yields prodrugs that are believed to improve toxicity (i.e.
being less toxic), metabolism (since the OH groups will be less
likely to be altered by methylation, glucuronidation and
sulfation), and bioavailability. In this prodrug concept, it is
believed that the prodrug conversion takes place in the plasma
(following its protection through the gut), and closer to its
appropriate target tissue (systemic organs). Enzymes in the blood
and appropriate tissues are believed to be able to cleave the
methylenedioxy group on these analogs to yield the dihydroxy
structures to achieve the observed efficacy. ##STR21## ##STR22##
##STR23## ##STR24## ##STR25## ##STR26## ##STR27## ##STR28##
##STR29## ##STR30## ##STR31## ##STR32##
EXAMPLE 4
Acetylated Compounds
[0077] Another potential strategy for the delivery of the bis- and
tris-dihydroxyaryl compounds of this invention to improve and/or
cause more favorable metabolism and bioavailability
characteristics, involves methods of protecting the hydroxy groups
as their pharmaceutically acceptable esters. Ester groups replacing
the hydroxy groups also tend to make the compounds more stable, and
less likely to be altered or degraded due to oxidation of the
hydroxyl groups.
[0078] The compound table below illustrates the acetyl esters of
the 86 dihydroxyaryl compounds previously presented herein, and are
presented below in which the OH groups are replaced by acetyl
groups. The illustration of acetyl esters here is merely exemplary
for the class of pharmaceutically acceptable esters that are part
of the compounds of this invention and may be prepared by analogous
methods. Other compounds disclosed herein also form
pharmaceutically acceptable esters in the same manner, and these
compounds, though not illustrated in the compound table below, are
also compounds of this invention.
[0079] These compounds are expected to be efficacious once the
ester linkages are cleaved (by enzymes in the plasma or in the
brain tissue), and the hydroxyl groups are regenerated. Replacement
of the hydroxyl groups with ester groups will yield prodrugs that
are believed to improve toxicity (i.e. being less toxic),
metabolism (since the OH groups will be less likely to be altered
by methylation, glucuronidation and sulfation), and
bioavailability. In this prodrug concept, it is believed that the
prodrug conversion takes place in the plasma (following its
protection through the gut), and closer to its appropriate target
tissue. Enzymes in the blood and appropriate tissues are believed
to be able to cleave the ester linkages on these pharmaceutically
acceptable esters to yield the dihydroxy structures important for
the observed efficacy.
[0080] The pharmaceutically acceptable esters of compounds #1
through #86 are prepared by methods well known to persons of
ordinary skill in the art, such as by reaction of the dihydroxyaryl
compounds with pharmaceutically acceptable acids, especially in
activated form (such as the acyl halides) and/or in the presence of
reagents facilitating esterification (such as an acidic catalyst)
and/or under conditions favoring esterification (such as by
conducting the reaction under conditions where the water formed in
the esterification is removed, e.g. by distillation). Methods of
esterification of phenolic hydroxyl groups are well known to
persons of ordinary skill in the art.
[0081] Suitable acids for the formation of pharmaceutically
acceptable esters are the C.sub.2-6 alkanoic acids (acetic acid,
propionic acid, and the like), benzoic acid, arylalkanoic acids
(phenylacetic acid, and the like); though many other acids are
suitable for the formulation of pharmaceutically acceptable esters,
and a person of ordinary skill in the art will have no difficulty
in choosing a suitable acid. ##STR33## ##STR34## ##STR35##
##STR36## ##STR37## ##STR38## ##STR39## ##STR40## ##STR41##
##STR42## ##STR43## ##STR44##
EXAMPLE 5
Pharmaceutical Formulations
Compositions of Compounds of this Invention.
[0082] The compounds of this invention, as mentioned previously,
are desirably administered in the form of pharmaceutical
compositions. Suitable pharmaceutical compositions, and the method
of preparing them, are well-known to persons of ordinary skill in
the art and are described in such treatises as Remington: The
Science and Practice of Pharmacy, A. Gennaro, ed., 20th edition,
Lippincott, Williams & Wilkins, Philadelphia, Pa.
[0083] Representative compositions are as follows:
Oral Tablet Formulation
[0084] An oral tablet formulation of a compound of this invention
is prepared as follows: TABLE-US-00003 % w/w Compound of this
invention 10.0 Magnesium stearate 0.5 Starch 2.0
Hydroxypropylmethylcellulose 1.0 Microcrystalline cellulose
86.5
[0085] The ingredients are mixed to homogeneity, then granulated
with the aid of water, and the granulates dried. The granulate is
then compressed into tablets sized to give a suitable dose of the
compound. The tablet is optionally coated by applying a suspension
of a film forming agent (e.g. hydroxypropylmethylcellulose),
pigment (e.g. titanium dioxide), and plasticizer (e.g. diethyl
phthalate), and drying the film by evaporation of the solvent. The
film coat may comprise, for example, 2-6% of the tablet weight.
Oral Capsule Formulation
[0086] The granulate from the previous section of this Example is
filled into hard gelatin capsules of a size suitable to the
intended dose. The capsule is banded for sealing, if desired.
Softgel Formulation
[0087] A softgel formulation is prepared as follows: TABLE-US-00004
% w/w Compound of this invention 20.0 Polyethylene glycol 400
80.0
[0088] The compound is dissolved or dispersed in the polyethylene
glycol, and a thickening agent added if required. A quantity of the
formulation sufficient to provide the desired dose of the compound
is then filled into softgels.
Parenteral Formulation
[0089] A parenteral formulation is prepared as follows:
TABLE-US-00005 % w/w Compound of this invention 1.0 Normal saline
99.0
[0090] The compound is dissolved in the saline, and the resulting
solution is sterilized and filled into vials, ampoules, and
prefilled syringes, as appropriate.
Controlled-Release Oral Formulation
[0091] A sustained release formulation may be prepared by the
method of U.S. Pat. No. 4,710,384, as follows:
[0092] One Kg of a compound of this invention is coated in a
modified Uni-Glatt powder coater with Dow Type 10 ethyl cellulose.
The spraying solution is an 8% solution of the ethyl cellulose in
90% acetone to 10% ethanol. Castor oil is added as plasticizer in
an amount equal to 20% of the ethyl cellulose present. The spraying
conditions are as follows: 1) speed, 1 liter/hour; 2) flap, 10-15%;
3) inlet temperature, 50.degree. C., 4) outlet temperature,
30.degree. C., 5) percent of coating, 17%. The coated compound is
sieved to particle sizes between 74 and 210 microns. Attention is
paid to ensure a good mix of particles of different sizes within
that range. Four hundred mg of the coated particles are mixed with
100 mg of starch and the mixture is compressed in a hand press to
1.5 tons to produce a 500 mg controlled release tablet.
EXAMPLE 6
NO Release Inhibition by Bis- and Tris-Dihydroxyaryl Compounds
[0093] The purpose of this experiment was to screen compounds 3, 4,
21, 26, 51, 52, 66, and 78 for their ability to inhibit Nitric
oxide release from microglia. Previous experiments demonstrated the
ability of some compounds to inhibit NO release from LPS/IFN.gamma.
stimulated microglia. EOC 13.31 microglia cells were plated into 96
well plates for 24 hours. Confluent cells were treated with 50
.mu.M of the bis- and tris-dihydroxyaryl compounds either with or
without stimulation at 1 pg/ml LPS, and 1 ng/ml IFN.gamma.. NO
release was measured at 24 hours using the Griess reaction which is
a spectrophotometric measure of NO release (reagents from Promega).
TABLE-US-00006 TABLE 1 Inhibition of NO release from microglia
exposed to LPS/IFN.gamma. treated with bis- and tris-dihydroxyaryl
compounds Compound Only OD Values uM Avg Std. Dev Media control
0.052 0.06 0.053 0.8750 1.4186 1.0000 1.0979 0.284692 3 0.053 0.054
0.053 1.0000 1.1250 1.0000 1.0417 0.072169 4 0.052 0.058 0.051
0.8750 0.9729 0.7500 0.8660 0.111704 21 0.045 0.046 0.043 0.0000
0.1250 -0.2500 -0.0417 0.190941 26 0.045 0.047 0.043 0.0000 0.2500
-0.2500 0.0000 0.25 51 0.056 0.056 0.053 1.3750 1.3750 1.0000
1.2500 0.216506 52 0.044 0.047 0.043 -0.1250 0.2500 -0.2500 -0.0417
0.260208 66 0.052 0.054 0.049 0.8750 1.1250 0.5000 0.8333 0.314576
78 0.054 0.056 0.053 1.1250 1.3750 1.0000 1.1667 0.190941 L-NAME*
0.067 0.053 0.048 2.9786 1.0000 0.3750 1.4512 1.359164 Compound + %
LPS/IFN.gamma. inhibition Media control 0.192 0.208 17.6020 19.6429
18.6224 1.443075 3 0.094 0.137 0.145 6.0450 10.8259 11.7188 9.5299
3.050835 48.8% 4 0.048 0.055 0.057 0.3750 1.2500 1.5000 1.0417
0.590727 94.4% 21 0.159 0.175 0.189 13.3929 15.4337 17.2194 15.3486
1.914682 17.6% 26 0.169 0.168 0.174 14.6684 14.5408 15.3061 14.8384
0.410019 20.3% 51 0.209 0.218 0.221 19.7704 20.9184 21.3010 20.6633
0.796556 -11.0% 52 0.18 0.174 0.191 16.0714 15.3061 17.4745 16.2840
1.099704 12.6% 66 0.165 0.178 0.183 14.1582 15.8163 16.4541 15.4762
1.18515 16.9% 78 0.251 0.178 0.176 25.000 15.8163 15.5612 18.7925
5.377351 -0.9% L-NAME* 0.104 0.099 0.104 7.1429 6.5848 7.1429
6.9568 0.322182 62.6% *L-NAME (nitro-L-arginine methyl ester) is a
positive control for NO inhibition
[0094] NO was not detected in cells not exposed to LPS/IFN.gamma..
Exposure of cells only to 1 mg/ml LPS and 1 ng/ml IFN.gamma.
induced NO release to 18.6 uM. Concurrent treatment with bis- and
tris-dihydroxyaryl compound inhibited NO release as set out in
table 2. Bis- and tris-dihydroxyaryl compounds 3, 4 and 26 were
very effective inhibitors of NO release by LPS/IFN.gamma.
stimulated microglial cells. Specifically compound 4 caused a
significant, (p<0.001) 94.4% inhibition. TABLE-US-00007 TABLE 2
Summary of Inhibition of NO release from microglia exposed to
LPS/IFN.gamma. treated with bis- and tris-dihydroxyaryl compounds
Compound % Inhibition 3 48.8% 4 94.4% 21 17.6% 26 20.3% 51 -11% 52
12.6% 66 16.9% 78 -0.9% L-name (+control) 62.6%
EXAMPLE 7
Inhibition of NO Release by Bis- and Tris-Dihydroxyaryl
Compounds
[0095] In this experiment, additional bis- and tris-dihydroxyaryl
compounds were screened for their ability to inhibit nitric oxide
release from microglial cells. Microglial cells stimulated with
LPS/IFN.gamma. release nitric oxide. Previous experiments suggest
some bis- and tris-dihydroxyaryl compounds block NO release. This
is a continuation of screening experiments. EOC 13.31 microglial
cells were plated into 96 well plates. At 24 hours and at
confluency, cells were treated with 10 .mu.g/ml LPS/10 ng/ml
IFN.gamma.+50 uM bis- and tris-dihydroxyaryl compounds. NO release
was measured by using the Griess reaction at 48 hours and is shown
in Table 3. TABLE-US-00008 TABLE 3 NO release from microglia Avg. %
Std. Inhibi- Std. Compound .mu.M NO release Avg Dev. % inhibition
tion Dev. Untreated 0.085 0.08 0.074 4.545 3.835 2.964 3.782 0.792
67 0.117 0.152 0.104 9.455 14.910 7.372 10.579 3.893 65.01 44.82
72.72 60.85 14.4 63 0.205 0.211 0.205 22.892 23.795 22.892 23.193
0.522 15.27 11.93 15.27 14.16 1.93 61 0.189 0.196 0.198 20.482
21.536 21.837 21.285 0.712 24.19 20.29 19.18 21.22 2.63 23 0.215
0.202 0.208 24.398 22.440 23.343 23.394 0.980 9.70 16.95 13.60
13.42 3.63 19 0.226 0.225 0.222 26.087 25.932 25.466 25.828 0.323
3.45 4.02 5.75 4.41 1.20 9 0.216 0.233 0.233 24.548 27.174 27.174
26.299 1.516 9.14 -0.57 -0.57 2.66 5.61 8 0.216 0.218 0.228 24.548
24.849 26.398 25.265 0.992 9.14 8.03 2.30 6.49 3.67 1 0.199 0.203
0.196 21.988 22.590 21.536 22.038 0.529 18.62 16.39 20.29 18.43
1.96 L-NAME* 0.067 0.07 0.072 1.872 2.340 2.652 2.288 0.393 93.07
91.34 90.18 91.53 1.45 LPS 0.239 0.228 0.229 28.106 26.398 26.553
27.019 0.945 Untreated 0.092 0.078 0.09 5.540 3.551 5.256 4.782
1.076 77 0.219 0.211 0.215 25.000 23.795 24.398 24.398 0.602 9.89
14.23 12.06 12.06 2.17 58 0.236 0.219 0.233 27.640 25.000 27.174
26.605 1.409 0.37 9.89 2.05 4.10 5.08 75 0.217 0.192 0.215 24.699
20.934 24.398 23.343 2.092 10.97 24.54 12.06 15.86 7.54 57 0.226
0.207 0.199 26.087 23.193 21.988 23.756 2.107 5.97 16.40 20.74
14.37 7.59 12 0.176 0.173 0.188 18.524 18.072 20.331 18.976 1.195
33.23 34.86 26.72 31.60 4.31 76 0.214 0.202 0.216 24.247 22.440
24.548 23.745 1.140 12.60 19.12 11.52 14.41 4.11 73 0.064 0.06
0.085 1.393 0.964 4.545 2.301 1.956 94.98 96.52 83.62 91.71 7.05 85
0.22 0.192 0.214 25.155 20.934 24.247 23.445 2.222 9.33 24.54
12.660 15.49 8.01 3 0.064 0.055 0.062 1.393 0.429 1.179 1.000 0.506
94.98 98.46 95.75 96.40 1.83 LPS 0.243 0.224 0.243 28.727 25.776
28.727 27.743 1.703 *Positive control
[0096] From this experiment, 3, 12, 61, 67 and 73 were compounds
that demonstrated the most dramatic effect. L-NAME treated cells
appeared exactly like cells that weren't exposed to LPS/IFN.gamma..
Compounds 3, 12, 61, 67 and 73 were effective inhibitors of NO
release by LPS/IFN.gamma. stimulated microglia.
EXAMPLE 8
Dose Response Inhibition of NO Release from Microglia by Bis- and
Tris-Dihydroxyaryl Compounds
[0097] This experiment was designed to determine dose response
inhibition of NO release from microglia by bis- and
tris-dihydroxyaryl compounds when stimulated with LPS/IFN.gamma..
Earlier experiments demonstrated that some bis- and
tris-dihydroxyaryl compounds are good inhibitors of NO release from
microglia. This experiment is designed to determine a dose response
for various bis- and tris-dihydroxyaryl compounds. Microglial cells
were plated into 96 well plates at 1.times.10.sup.5 cells/ml. At 48
hours, media was exchanged for media containing 10 ng/ml IFN.gamma.
and 10 .mu.g/ml LPS and the following compounds 3, 4, 21, 26 and
the positive control L-NAME were used. The doses tested were: 0.75
.mu.M, 1.56 .mu.M, 3.125 .mu.M, 6.25 .mu.M, 12.5 .mu.M, 25 .mu.M,
50 .mu.M, 100 .mu.M and 200 .mu.M. Nitric oxide release was
measured after 48 hours using the Griess reaction. TABLE-US-00009
TABLE 4 Dose response Avg % Std. % inhibition Inhibi- Std. Dose
.mu.M NO release Avg Dev % inhibition tion Dev Compound 3 LPS 0.178
0.181 0.177 19.688 20.156 19.531 19.792 0.325 0.75 0.2 0.183 0.187
23.125 20.469 21.094 21.563 1.389 -16.84 -3.42 -6.58 -8.947 7.017
1.56 0.188 0.174 0.174 21.250 19.063 19.063 19.792 1.263 -7.37 3.68
3.68 0.000 6.381 3.125 0.186 0.175 0.172 20.938 19.219 18.750
19.635 1.152 -5.79 2.89 5.26 0.789 5.819 6.25 0.177 0.165 0.166
19.531 17.656 17.813 18.333 1.040 1.32 10.79 10.00 7.368 5.257 12.5
0.17 0.166 0.175 18.438 17.813 19.219 18.490 0.705 6.84 10.00 2.89
6.579 3.560 25 0.164 0.15 0.166 17.500 15.313 17.813 16.875 1.362
11.58 22.63 10.00 14.737 6.882 50 0.119 0.117 0.12 10.469 10.156
10.625 10.417 0.239 47.11 48.68 46.32 47.368 1.206 100 0.1 0.097
0.102 7.500 7.031 7.813 7.448 0.393 62.11 64.47 60.53 62.368 1.987
200 0.095 0.092 0.094 6.719 6.250 6.563 6.510 0.239 66.05 68.42
66.84 67.105 1.206 Compound 4 LPS 0.21 0.203 0.21 24.688 23.594
24.688 24.323 0.631 0.75 0.229 0.22 0.222 27.931 26.379 26.724
27.011 0.815 -14.83 -8.45 -9.87 -11.054 3.350 1.56 0.214 0.197 0.2
25.345 22.656 23.125 23.709 1.436 -4.20 6.85 4.93 2.525 5.905 3.125
0.215 0.188 0.191 25.517 21.250 21.719 22.829 2.340 -4.91 12.63
10.71 6.143 9.621 6.25 0.167 0.142 0.145 17.969 14.063 14.531
15.521 2.133 26.12 42.18 40.26 36.188 8.769 12.5 0.083 0.071 0.073
5.125 3.625 3.875 4,208 0.804 78.93 85.10 84.07 82.698 3.304 25
0.071 0.07 0.064 3.625 3.500 2.947 3.357 0.361 85.10 85.61 87.89
86.197 1.484 50 0.071 0.067 0.067 3.625 3.125 3.125 3.292 0.289
85.10 87.15 87.15 86.467 1.187 100 0.078 0.075 0.077 4.500 4.125
4.375 4.333 0.191 81.50 83.04 82.01 82.184 0.785 200 0.092 0.086
0.092 6.250 5.500 6.250 6.000 0.433 74.30 77.39 74.30 75.332 1.780
Compound 21 LPS 0.178 0.181 0.177 19.688 20.156 19.531 19.792 0.325
0.75 0.195 0.199 0.201 22.344 22.969 23.281 22.865 0.477 -12.89
-16.05 -17.63 -15.526 2.412 1.56 0.181 0.189 0.191 20.156 21.406
21.719 21.094 0.827 -1.84 -8.16 -9.74 -6.579 4.178 3.125 0.186
0.184 0.189 20.938 20.625 21.406 20.990 0.393 -5.79 -4.21 -8.16
-6.053 1.987 6.25 0.183 0.175 0.185 20.469 19.219 20.781 20.156
0.827 -3.42 2.89 -.5.00 -1.842 4.178 12.5 0.161 0.172 0.184 17.031
18.750 20.625 18.802 1.797 13.95 5.26 -4.21 5.000 9.082 25 0.179
0.169 0.177 19.844 18.281 19.531 19.219 0.827 -0.26 7.63 1.32 2.895
4.178 50 0.161 0.158 0.165 17.031 16.563 17.656 17.083 0.549 13.95
16.32 10.79 13.684 2.773 100 0.136 0.136 0.136 13.125 13.125 13.125
13.125 0.000 33.68 33.68 33.68 33.684 0.000 200 0.087 0.093 0.092
5.625 6,406 6..250 6.094 0.413 71.58 67.63 68.42 69.211 2.089
Compound 26 LPS 0.21 0.203 0.21 24.688 23.594 24.688 24.323 0.631
0.75 0.21 0.206 0.215 24.688 24.063 25.517 24.756 0.730 -1.50 1.07
-4.91 -1.780 3.000 1.56 0.198 0.193 0.204 22.813 22.031 23.750
22.865 0.861 6.21 9.42 2.36 5.996 3.538 3.125 0.197 0.195 0.204
22.656 22.344 23.750 22.917 0.738 6.85 8.14 2.36 5.782 3.036 6.25
0.208 0.202 0.211 24.375 23.438 24.844 24.219 0.716 -0.21 3.64
-2.14 0.428 2.944 12.5 0.192 0.192 0.192 21.875 21.875 21.875
21.875 0.000 10.06 10.06 10.06 10.064 0.000 25 0.197 0.187 0.192
22.656 21.094 21.875 21.875 0.781 6.85 13.28 10.06 10.064 3.212 50
0.175 0.177 0.179 19.219 19.531 19.844 19.531 0.313 20.99 19.70
18.42 19.700 1.285 100 0.154 0.181 0.178 15.938 20.156 19.688
18.594 2.312 34.48 17.13 19.06 23.555 9.507 200 0.134 0.136 0.145
12.813 13.125 14.531 13.490 0.916 47.32 46.04 40.26 44.540 3.764
Positive Control L-name LPS 0.195 0.195 0.194 22.344 22.344 22.188
22.292 0.090 0.75 0.198 0.201 0.209 22.813 23.281 24.531 23.542
0.888 -2.34 -4.44 -10.05 -5.607 3.986 1.56 0.19 0.198 0.199 21.563
22.813 22.969 22.448 0.771 3.27 -2.34 -3.04 -0.701 3.458 3.125
0.187 0.19 0.193 21.094 21.563 21.031 21.563 0.469 5.37 3.27 1.17
3.271 2.103 6.25 0.179 0.174 01.79 19.844 19.063 19.844 19.583
0.451 10.98 14.49 10.98 12.150 2.023 12.5 0.155 0.159 0.161 16.094
16.719 17.031 16.615 0.477 27.80 25.00 23.60 25.467 2.141 25 0.143
0.132 0.142 14.219 12.500 14.063 13.594 0.950 36.21 43.93 36.92
39.019 4.264 50 0.115 0.112 0.112 9.844 9.375 9.375 9.531 0.271
55.84 57.94 57.94 57.243 1.214 100 0.092 0.088 0.096 6.250 5.750
6.875 6.292 0.564 71.96 74.21 69.16 71.776 2.529 200 0.079 0.077
0.078 4.625 4.375 4.500 4.500 0.125 79.25 80.37 79.81 79.813
0.561
[0098] Compounds 3, 4, 21, and to a lesser extent 26 were effective
inhibitors of NO release. At the highest concentration tested (200
.mu.M), compounds 3, 4, and 21 inhibited NO release by 67%, 75%,
and 69% respectively.
EXAMPLE 9
Dose Response Inhibition of NO Release from Microglia by Bis- and
Tris-Dihydroxyaryl Compounds
[0099] This experiment was designed to determine the dose response
inhibition of NO release from microglia by bis- and
tris-dihydroxyaryl compounds when stimulated with LPS/IFN.gamma..
Earlier experiments have demonostrated that some bis- and
tris-dihydroxyaryl compounds are good inhibitors of NO release from
microglia. This experiment is designed to determine a dose response
for various effective bis- and tris-dihydroxyaryl compounds.
Microglial cells were plated into 96 well plates at
1.times.10.sup.5 cells/ml. At 48 hours, media was exchanged for
media containing 10 ng/ml IFN.gamma. and 10 .mu.g/ml LPS and the
following compounds 3, 4, 21, 26 and the positive control L-NAME.
The doses tested were: 0.5 .mu.M, 1.0 .mu.M, 5.0 .mu.M, 10 .mu.M,
25 .mu.M, 50 .mu.M, 100 .mu.M, 250 .mu.M, and 500 .mu.M. Nitric
oxide release was measured after 48 hours using the Griess
reaction. TABLE-US-00010 TABLE 5 Dose Response Avg % Std. Dose
.mu.M NO release Avg % inhibition Inhibition Dev No LPS 0.058 0.059
0.062 1.500 1.600 2.000 1.70 Com- pound 21 LPS 0.18 0.22 0.184
17.917 23.472 18.472 16.22 21.77 16.77 18.25 0.5 0.193 0.196 0.198
19.722 20.139 20.417 18.02 18.44 18.72 1.27% -1.01% -2.54% -0.76%
0.02 1 0..2 0.191 0.189 20.694 19.444 19.167 18.99 17.74 17.47
-4.06% 2.79% 4.31% 1.01% 0.04 5 0.197 0.195 0.189 20.278 20.000
19.167 18.58 18.30 17.47 -1.78% -0.25% 4.31% 0.76% 0.03 10 0.199
0.191 0.199 20.556 19.444 20.556 18.86 17.74 18.86 -3.30% 2.79%
-3.30% -1.27% 0.04 25 0.193 0.187 0.185 19.722 18.889 18.611 18.02
17.19 16.91 1.27% 5.83% 7.36% 4.82% 0.03 50 0.184 0.172 0.176
18.472 16.806 17.361 16.77 15.11 15.66 8.12% 17.25% 14.20% 13.19%
0.05 100 0.156 0.142 0.147 14.583 12.639 13.333 12.88 10.94 11.63
29.42% 40.07% 36.27% 35.25% 0.05 250 0.086 0.077 0.085 5.163 3.940
5.027 3.46 2.24 3.33 81.03% 87.73% 81.77% 83.51% 0.04 500 0.09 0.08
0.091 5.707 4.348 5.842 4.01 2.65 4.14 78.05% 85.49% 77.31% 80.28%
0.05 Com- pound 26 LPS 0.18 0.22 0.184 17.917 23.472 18.472 16.22
21.77 16.77 18.25 0.5 0.207 0.19 0.203 21.667 19.306 21.111 19.97
17.61 19.41 -9.38% 3.55% -6.34% -4.06% 0.07 1 0.193 0.188 0.201
19.722 19.028 20.833 18.02 17.33 19.13 1.27% 5.07% -4.82% 0.51%
0.05 5 0.195 0.206 0.2 20.000 21.528 20.694 18.30 19.83 18.99
-0.25% -8.62% -4.06% -4.31% 0.04 10 0.196 0.192 0.2 20.139 19.583
20.694 18.44 17.88 18.99 -1.01% 2.03% -4.06% -1.01% 0.03 25 0.182
0.183 0.192 18.194 18.333 19.583 16.49 16.63 17.88 9.64% 8.88%
2.03% 6.85% 0.04 50 0.183 0.192 0.183 18.333 19.583 18.333 16.63
17.88 16.63 8.88% 2.03% 8.88% 6.59% 0.04 100 0.172 0.172 0.176
16.806 16.806 17.361 15.11 15.11 15.66 17.25% 17.25% 14.20% 16.23%
0.02 250 0.147 0.146 0.152 13.333 13.194 14.028 11.63 11.49 12.33
36.27% 37.03% 32.46% 35.25% 0.02 500 0.062 0.058 0.061 2.000 1.500
1.867 0.30 -0.20 0.17 98.36% 101.10% 99.09% 99.51% 0.01 Com- pound
67 LPS 0.23 0.181 0.179 24.861 18.056 17.778 23.16 16.36 16.08
18.53 0.5 0.206 0.197 0.198 21.528 20.278 20.417 19.83 18.58 18.72
-7.00% -0.25% -1.00% -2.75% 0.04 1 0.206 0.19 0.188 21.528 19.306
19.028 19.83 17.61 17.33 -7.00% 5.00% 6.50% 1.50% 0.07 5 0.205
0.192 0.196 21.389 19.583 20.139 19.69 17.88 18.44 -6.25% 3.50%
0.50% -0.75% 0.05 10 0.216 0.21 0.215 22.917 22.083 22.778 21.22
20.38 21.08 -14.49% -9.99% -13.74% -12.74% 0.02 25 0.209 0.199
0.204 21.944 20.556 21.250 20.24 18.86 19.55 -9.24% -1.75% -5.50%
-5.50% 0.04 50 0.214 0.203 0.21 22.639 21.111 22.083 20.94 19.41
20.38 -12.99% -4.75% -9.99% -9.24% 0.04 100 0.18 0.156 0.19 17.917
14.583 19.306 16.22 12.88 17.61 12.49% 30.48% 5.00% 15.99% 0.13 250
0.136 0.136 0.14 11.835 11.835 12.367 10.14 10.14 10.67 45.31%
45.31% 42.44% 44.35% 0.02 500 0.094 0.09 0.092 6.250 5.707 5.978
4.55 4.01 4.28 75.45% 78.38% 76.91% 76.91% 0.01 Com- pound 73 LPS
0.23 0.181 0.179 24.861 18.056 17.778 23.16 16.36 16.08 18.53 0.5
0.176 0.182 0.181 17.361 18.194 18.056 15.66 16.49 16.36 15.49%
10.99% 11.74% 12.74% 0.02 1 0.173 0.172 0.179 16.944 16.806 17.778
15.25 15.11 16.08 17.74% 18.49% 13.25% 16.49% 0.03 5 0.195 0.192
0.197 20.000 19.583 20.278 18.30 17.88 18.58 1.25% 3.50% -0.25%
1.50% 0.02 10 0.187 0.198 0.198 18.889 20.417 20.417 17.19 18.72
18.72 7.24% -1.00% -1.00% 1.75% 0.05 25 0.189 0.196 0.202 19.167
20.139 20.972 17.47 18.44 19.27 5.75% 0.50% -4.00% 0.75% 0.05 50
0.191 0.194 0.204 19.444 19.861 21.250 17.74 18.16 19.55 4.25%
2.00% -5.50% 0.25% 0.05 100 0.155 0.145 0.158 14.444 13.056 14.861
12.74 11.36 13.16 31.23% 38.72% 28.98% 32.98% 0.05 250 0.134 0.129
0.145 11.569 10.904 13.056 9.87 9.20 11.36 46.74% 50.33% 38.72%
45.27% 0.06 500 0.098 0.096 0.101 6.782 6.516 7.181 5.08 4.82 5.48
72.58% 74.01% 70.42% 72.34% 0.02 L-name LPS 0.205 0.193 0.196
21.389 19.722 20.139 19.69 18.02 18.44 18.72 0.5 0.168 0.162 0.164
16.250 15.417 15.694 14.55 13.72 13.99 22.26% 26.71% 25.23% 24.74%
0.02 1 0.156 0.161 0.159 14.583 15.278 15.000 12.88 13.58 13.30
31.17% 27.46% 28.94% 29.19% 0.02 5 0.139 0.144 0.154 12.234 12.917
14.306 10.53 11.22 12.61 43.72% 40.07% 32.65% 38.81% 0.06 10 0.143
0.144 0.145 12.778 12.917 13.056 11.08 11.22 11.36 40.81% 40.07%
39.33% 40.07% 0.01 25 0.105 0.109 0.113 7.713 8.245 8.777 6.01 6.54
7.08 67.87% 65.03% 62.19% 65.03% 0.03 50 0.085 0.085 0.092 5.027
5.027 5.978 3.33 3.33 4.28 82.22% 82.22% 77.14% 80.53% 0.03 100
0.083 0.084 0.09 4.755 4.891 5.707 3.06 3.19 4.01 83.68% 82.95%
78.59% 81.74% 0.03 250 0.067 0.067 0.071 2.667 2.667 3.125 0.97
0.97 1.43 94.84% 94.84% 92.39% 94.02% 0.01 500 0.063 0.064 0.066
2.133 21.33 2.533 0.43 0.57 0.83 97.68% 96.97% 95.55% 96.73%
0.01
[0100] All of the compounds tested were effective inhibitors of NO
release. At the highest concentration tested (500 .mu.M, compounds
26, 67, and 73 inhibited NO release by 99%, 76%, and 72%
respectively. Compound 21 was most effective at the 250 .mu.M
dose.
EXAMPLE 10
Inhibition of NO Release from Microglia Following Pretreatment with
Bis- and Tris-Dihydroxyaryl Compounds
[0101] This experiment was designed to determine if pretreatment
for 24 hours with bis- and tris-dihydroxyaryl compounds changes NO
inhibition previously observed with concurrent exposure of bis- and
tris-dihydroxyaryl compounds. Previous screens demonstrated
significant inhibition by different bis- and tris-dihydroxyaryl
compounds of LPS/IFN.gamma. induced nitric oxide release by
microglial cells. This study seeks to determine if pretreatment
with bis- and tris-dihydroxyaryl compounds increases NO release
inhibition. EOC 13.31 microglia cells were plated at
1.times.10.sup.6 cells/ml in 96 well plates. After 48 hours,
confluent cells were exposed to 50 .mu.M compound (stock 10 mM in
DMSO) prepared in fresh cell culture media. After 24 hours, 10
.mu.g/ml LPS and 10 ng/ml IFN.gamma. was added to wells directly
without a media change. Cells were incubated for a further 48 hours
and nitric oxide release was measured using the Griess Reaction.
TABLE-US-00011 TABLE 6 Pretreatment and inhibition of NO release
Avg % 50 uM Inhibi- Std. Compound .mu.M NO release % inhibition
tion Dev. 4 0.066 0.058 0.057 0.000 0.000 0.000 100.00% 100.00%
100.00% 100.00% 0.00 73 0.072 0.066 0.065 0.000 0.000 0.000 100.00%
100.00% 100.00% 100.00% 0.00 57 0.093 0.107 0.094 0.000 0.000 0.000
100.00% 100.00% 100.00% 100.00% 0.00 77 0.110 0.110 0.136 0.120
0.120 1.625 99.39% 99.39% 91.78% 96.85% 0.04 85 0.117 0.127 0.130
0.540 1.140 1.320 97.27% 94.23% 93.32% 94.94% 0.02 21 0.166 0.160
0.158 3.500 3.125 2.933 82.29% 84.17% 85.16% 83.88% 0.01 19-2 0.135
0.161 0.195 1.620 3.188 5.313 91.80% 83.87% 73.11% 82.93% 0.09 63
0.167 0.167 0.166 3.563 3.563 3.500 81.97% 81.97% 82.29% 82.08%
0.00 52 0.172 0.169 0.179 3.875 3.688 4.313 80.39% 81.34% 78.17%
79.97% 0.02 3 0.206 0.163 0.153 6.000 3.313 2.621 69.63% 83.24%
86.74% 79.87% 0.09 76 0.187 0.177 0.191 4.813 4.188 5.063 75.64%
78.81% 74.38% 76.28% 0.02 58 0.180 0.205 0.206 4.375 5.938 6.000
77.86% 69.95% 69.63% 72.48% 0.05 67 0.242 0.214 0.204 8.125 6.375
5.875 58.88% 67.74% 70.27% 65.63% 0.06 78 0.245 0.210 0.206 8.313
6.125 6.000 57.93% 69.00% 69.63% 65.52% 0.07 66 0.241 0.222 0.210
8.063 6.875 6.250 59.20% 65.21% 68.37% 64.26% 0.05 51-S1 0.264
0.219 0.231 9.500 6.688 7.438 51.92% 66.15% 62.36% 60.14% 0.07 75
0.237 0.248 0.234 7.813 8.500 7.625 60.46% 56.98% 61.41% 59.62%
0.02 51 0.277 0.241 0.235 10.313 8.063 7.688 47.81% 59.20% 61.09%
56.03% 0.07 L-NAME* 0.081 0.091 0.098 0.000 0.000 0.000 100.00%
100.00% 100.00% 100.00% 0.00 LPS/IFN 0.425 0.457 0.392 19.781
21.843 17.655 19.759 * Positive control
[0102] Pretreatment with bis- and tris-dihydroxyaryl compounds
significantly inhibited NO release. All of the compounds tested
showed inhibition of NO release by at least 55%. Specifically,
compounds 4, 73, 57, 77, 85, 21, 19, 63, 52, 3, and 76 showed
greater than 75% inhibition of NO release following
pretreatment.
EXAMPLE 11
TNF-.alpha. Inhibition by Bis- and Tris-Dihydroxyaryl Compounds
[0103] This experiment was designed to determine if bis- and
tris-dihydroxyaryl compounds inhibit TNF-.alpha. release from
microglia immunostimulated with LPS and IFN.gamma.. Previous
experiements demonstrated signficant TNF-.alpha. release from
microglia stimulated with LPS and IFN.gamma.. Also, bis- and
tris-dihydroxyaryl compound's ability to inhibit NO release
strongly supports their interference with the inflammatory cascade
and utlimately inhibition of inflammation. Microglia cells were
plated at 1.times.10.sup.5 cells/ml into 96 well plates. At day
three, they were treated with 50 .mu.M bis- and tris-dihydroxyaryl
compound for 1 hour and then exposed to 10 .mu.g/ml LPS+10 ng/ml
IFN.gamma. for 24 hours. 10 .mu.l of media was removed and stored
at -80.degree. until TNF-.alpha. ELISA assays were performed.
Samples were diluted 1:50 before running the TNF-.alpha. ELISA (kit
commercially available from Sigma). TABLE-US-00012 TABLE 7
TNF-.alpha. release from microglia treated with bis- and
tris-dihydroxyaryl compounds and 10 ug LPS and 10 ng IFN.gamma. Avg
% 50 uM Inhibi- Std. Compound .mu.M NO release % inhibition tion
Dev No LPS 0.258 0.239 0.221 264.815 230.903 199.653 51-1 0.880
0.883 0.909 1571.111 1577.778 1635.556 -12.94% -13.42% -17.57%
-14.64% 0.03 51-4 0.907 0.870 0.925 1631.111 1548.889 1671.111
-17.25% -11.34% -20.13% -16.24% 0.04 51-S1 0.867 0.882 0.865
1542.222 1575.556 1537.778 -10.86% -13.26% -10.54% -11.55% 0.01
51-S3 0.913 0.857 0.808 1644.444 1520.000 1411.111 -18.21% -9.27%
-1.44% -9.64% 0.08 51-S4 0.841 0.763 0.802 1484.444 1311.111
1397.778 -6.71% 5.75% -0.48% -0.48% 0.06 51-S5 0.432 0.454 0.448
598.739 644.958 632.353 56.96% 53.64% 54.54% 55.05% 0.02 51-S6
0.831 0.791 0.780 1462.222 1373.333 1348.889 -5.11% 1.28% 3.04%
-0.27% 0.04 51-S7 0.923 0.862 0.866 1666.667 1531.111 1540.000
-19.81% -10.06% -10.70% -13.53% 0.05 51-S8 0.901 0.903 1.003
1617.778 1622.222 1844.444 -16.29% -16.61% -32.59% -21.83% 0.09
51-S9 0.471 0.558 0.513 680.672 863.445 768.908 51.07% 37.93%
44.73% 44.58% 0.07 3-2 0.593 0.627 0.623 936.975 1008.889 1000.000
32.65% 27.48% 28.11% 29.41% 0.03 4-3 0.177 0.185 0.199 123.214
137.153 161.458 91.14% 90.14% 88.39% 89.89% 0.01 73 0.996 1.142
1.121 1828.889 2153.333 2106.667 -31.47% -54.79% -51.44% -45.90%
0.13 67 0.887 0.908 0.971 1586.667 1633.333 1773.333 -14.06%
-17.41% -27.48% -19.65% 0.07 12 0.641 0.733 0.700 1040.000 1244.444
1171.111 25.24% 10.54% 15.81% 17.20% 0.07 LPS 0.756 0.802 0.839
1295.556 1397.778 1480.000 1391.111 No LPS 0.132 0.143 0.137 45.313
62.500 53.125 61 0.885 0.827 0.802 1582.222 1453.333 1397.778
-17.04% -7.51% -3.40% -9.32 007 26 0.914 0.880 0.857 1646.667
1571.111 1520.000 -21.81% -16.22% -12.44% -16.82 0.05 1 0.885 0.892
0.882 1582.222 1597.778 1575.556 -17.04% -18.19% -16.55% -17.26
0.01 21-2 0.924 0.904 0.939 1668.889 1624.444 1702.222 -23.45%
-20.16% -25.92% -23.18 0.03 66-2 0.889 0.838 0.867 1591.111 1477.78
1542.222 -17.70% -9.32% -14.08% -13.70 0.04 75 0.798 0.746 0.766
1388.889 1273.333 1317.778 -2.74% 5.81% 2.52% 1.86 0.04 85 0.897
0.884 0.869 1608.889 1580.000 1546.667 -19.01% -16.88% -14.41%
-16.77 0.02 76-2 0.849 0.840 0.834 1502.222 1482.222 1468.889
-11.12% -9.64% -8.66% -9.81 0.01 57 0.802 0.793 0.820 1397.778
1377.778 1437.778 -3.40% -1.92% -6.36% -3.89 0.02 63 0.825 0.842
0.850 1448.889 1486.667 1504.444 -7.18% -9.97% -11.29% -9.48 0.02
23 0.878 0.890 0.926 1566.667 1593.333 1673.333 -15.89% -17.96%
-23.78% -19.18 0.04 52 0.943 0.968 0.856 1711.111 1766.667 1517.778
-26.58% -30.69% -12.27% -23.18 0.10 77-1 0.756 0.788 0.753 1295.556
1366.667 1288.889 4.16% -1.10% 4.66% 2.58 0.03 8-1 0.169 0.174
0.152 108.929 117.857 78.571 91.94% 91.28% 94.19% 92.47 0.02 19-2
0.730 0.708 0.734 1237.778 1188.889 1246.667 8.44% 12.05% 7.78%
9.42 0.02 58 0.862 0.854 0.858 1531.111 1513.333 1522.222 -13.26%
-11.95% -12.60% -12.60 0.01 LPS 0.793 0.799 0.752 1377.778 1391.111
1286.667 1351.852 *S numbers indicate analogs and numerals after
dashes indicate synthetic batch numbers.
[0104] Dexamethasone, the positive control, demonstrated 72.4%
inhibition. Some of the bis- and tris-dihydroxyaryl compounds like
51-S5 (55% inhibition), 51-S9 (44% inhibition), 4-3 (89%
inhibition) and 8-1 (92% inhibition) were good inhibitors of
TNF-.alpha. release indicating that the compounds are also good
inhibitors of the inflammatory cascade.
EXAMPLE 12
Ranking of NO Release Inhibition
[0105] The bis- and tris-dihydroxyaryl compounds were ranked in
order of effectiveness for the inhibition of NO release as set out
in Table 8. TABLE-US-00013 TABLE 8 Ranking of effectiveness of
compounds for the inhibition of NO release. Compound % Inhibition
at 50 .mu.M 4 94 8 92 67 61 12 32 3 30 61 21 26 20 1 18 21 18 66 17
75 16 85 16 76 14 57 14 63 14 23 13 52 13 77 12 19 4 58 4 9 3 73 2
78 -1 L-NAME 92
[0106] The present invention is not limited in scope by the
specific embodiments described herein. Indeed, various
modifications of the invention in addition to those described will
become apparent to those skilled in the art from the foregoing
descriptions. Such modifications are intended to fall within the
scope of the appended claims. Various publications are cited
herein, the disclosures of which are incorporated by reference in
their entireties.
* * * * *
References