U.S. patent application number 11/521591 was filed with the patent office on 2007-05-03 for methods of modulating neurotrophin-mediated activity.
This patent application is currently assigned to PainCeptor Pharma Corporation. Invention is credited to Daniel Scott Auld, Kazimierz Babinski, Xilin Cui, Rahul Vohra.
Application Number | 20070099900 11/521591 |
Document ID | / |
Family ID | 37865286 |
Filed Date | 2007-05-03 |
United States Patent
Application |
20070099900 |
Kind Code |
A1 |
Babinski; Kazimierz ; et
al. |
May 3, 2007 |
Methods of modulating neurotrophin-mediated activity
Abstract
Disclosed are compositions which modulate the interaction with
nerve growth factor and precursors thereof with neurotrophic
receptors. Also disclosed are methods of using the compositions of
the invention, including methods of administration.
Inventors: |
Babinski; Kazimierz;
(Dorval, CA) ; Vohra; Rahul; (Kanata, CA) ;
Cui; Xilin; (Orleans, CA) ; Auld; Daniel Scott;
(Montreal, CA) |
Correspondence
Address: |
LAHIVE & COCKFIELD, LLP
ONE POST OFFICE SQUARE
BOSTON
MA
02109-2127
US
|
Assignee: |
PainCeptor Pharma
Corporation
St. Laurent
CA
H4S 2C1
|
Family ID: |
37865286 |
Appl. No.: |
11/521591 |
Filed: |
September 14, 2006 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
60718256 |
Sep 15, 2005 |
|
|
|
Current U.S.
Class: |
514/217.07 ;
514/232.8; 514/253.02; 514/288 |
Current CPC
Class: |
A61P 13/12 20180101;
C07D 401/06 20130101; A61P 29/00 20180101; C07D 209/92 20130101;
A61K 31/403 20130101; A61P 9/10 20180101; A61P 43/00 20180101; C07D
413/06 20130101; A61P 13/10 20180101; C07D 471/16 20130101; A61K
31/454 20130101; A61K 31/5377 20130101; A61P 13/00 20180101; A61K
31/4745 20130101; A61K 31/55 20130101; A61K 31/473 20130101; A61P
25/24 20180101; A61P 3/04 20180101; A61P 25/08 20180101; A61P 3/12
20180101; A61P 9/00 20180101; A61P 11/06 20180101; C07D 471/22
20130101; A61P 1/04 20180101; A61P 13/02 20180101; A61P 25/14
20180101; A61P 25/16 20180101; A61P 9/06 20180101; C07D 491/06
20130101; A61P 17/06 20180101; A61P 27/02 20180101; A61P 1/00
20180101; A61P 17/02 20180101; C07D 471/06 20130101; A61P 17/00
20180101; A61P 25/04 20180101; A61K 31/496 20130101; A61K 31/4375
20130101; A61P 1/02 20180101; C07D 221/14 20130101; A61P 19/00
20180101; A61P 25/00 20180101; A61P 11/00 20180101; A61P 25/18
20180101; C07D 401/04 20130101; A61P 15/00 20180101; A61P 25/28
20180101; A61P 35/00 20180101; A61P 19/02 20180101; A61P 27/06
20180101 |
Class at
Publication: |
514/217.07 ;
514/232.8; 514/253.02; 514/288 |
International
Class: |
A61K 31/55 20060101
A61K031/55; A61K 31/5377 20060101 A61K031/5377; A61K 31/496
20060101 A61K031/496; A61K 31/4745 20060101 A61K031/4745 |
Claims
1. A method of modulating the interaction of a neurotrophin and a
neurotrophin receptor, comprising contacting cells expressing a
neurotrophin receptor with an effective amount of a compound of
Formula 16, ##STR37## and pharmaceutically acceptable salts,
enantiomers, stereoisomers, rotamers, tautomers, diastereomers, or
racemates thereof; wherein R.sup.1, R.sup.1a, R.sup.2, R.sup.3,
R.sup.4 and R.sup.5 are each, independently, selected from the
group consisting of hydrogen, C.sub.1-6-alkyl, C.sub.1-6-alkoxy,
amino, halogen, hydroxyl, acid, cyano, C.sub.1-6-alkyl-sulfonamide,
aryl, heteroaryl, C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl;
wherein the alkyl, C.sub.1-6-alkoxy, amino, aryl, heteroaryl,
cycloalkyl, and heterocycloalkyl groups may be further
independently substituted one or more times with C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, hydroxyl, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl or azepanyl.
2-12. (canceled)
13. The method of claim 1, wherein R.sup.2, R.sup.3, R.sup.4 and
R.sup.5 are hydrogen, R.sup.1 is (CH.sub.2).sub.xR.sup.13, wherein
R.sup.13 is selected from the group consisting of hydrogen,
C.sub.1-6-alkyl, amino, C.sub.1-6-alkoxy, --OH, halogen, acid,
cyano, C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperazinyl, piperidinyl and azepanyl; x is
0, 1, 2, 3 or 4; and R.sup.1a is: ##STR38## wherein R.sup.6,
R.sup.7 and R.sup.12 are each, independently, selected from the
group consisting of hydrogen, C.sub.1-6-alkyl, C.sub.1-6-alkoxy,
amino, halogen, acid, cyano, C.sub.1-6-alkyl-sulfonamide, aryl,
heteroaryl, C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl.
14-17. (canceled)
18. The method of claim 1, wherein the compound is selected from
the group consisting of
4-[7-(3-Carboxy-propyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-benzo[lmn]-
[3,8]phenanthrolin-2-yl]-butyric acid (12),
3-[7-(3-Carboxy-phenyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-benzo[lmn]-
[3,8]phenanthrolin-2-yl]-4-benzoic acid (17),
3-[7-(1-chloro-4-carboxy-phenyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-b-
enzo[lmn][3,8]phenanthrolin-2-yl]-4-chloro-benzoic acid (18),
4-(1,3,6,8-Tetraoxo-7-(4-benzenesulfonic
acid)-3,6,7,8-tetrahydro-1H-benzo[lmn][3,8]phenanthrolin-2-yl)-benzenesul-
fonic acid (54), 3-(1,3,6,8-Tetraoxo-7-(3-benzenesulfonic
acid)-3,6,7,8-tetrahydro-1H-benzo[lmn][3,8]phenanthrolin-2-yl)-benzenesul-
fonic acid (55) and
3-[7-(3-Carboxy-propyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-benzo[lmn]-
[3,8]phenanthrolin-2-yl]-4-chloro-benzoic acid (59).
19. A method of modulating the interaction of a neurotrophin and a
neurotrophin receptor, comprising contacting cells expressing a
neurotrophin receptor with an effective amount of a compound of
Formula 17, ##STR39## and pharmaceutically acceptable salts,
enantiomers, stereoisomers, rotamers, tautomers, diastereomers, or
racemates thereof; wherein R.sup.6, R.sup.7, R.sup.12, R.sup.6a,
R.sup.7a and R.sup.12a are each, independently, selected from the
group consisting of hydrogen, C.sub.1-6-alkyl, C.sub.1-6-alkoxy,
amino, halogen, hydroxyl, acid, cyano, C.sub.1-6-alkyl-sulfonamide,
aryl, heteroaryl, C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, tetrazolyl, sulfone, urea, thiourea, morpholino,
piperidinyl, piperazinyl and azepanyl.
20. The method of claim 19, wherein R.sup.6, R.sup.6a, R.sup.7 and
R.sup.7a are each, independently, selected from the group
consisting of hydrogen, halogen, hydroxyl, trihalomethyl, cyano,
nitro, --N(H)C(O)C.sub.1-C.sub.4-alkyl, --NH.sub.2,
--C(O)N(H)C.sub.1-4alkyl, methoxy, morpholino, piperazinyl and
C.sub.1-C.sub.4-alkyl-ester; and R.sup.12 and R.sup.12a are
hydrogen.
21-23. (canceled)
24. The method of claim 19, wherein the compound is selected from
the group consisting of
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]b-
enzo[4,5]imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid (90),
2-(3-Carboxy-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]benzo[4,5]-
imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid (93),
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]b-
enzo[4,5]imidazo[2,1-b][3,8]phenanthroline-9 (98),
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-9-chloro-be-
nzo[[lmn]benzo[4,5]imidazo[2,1-b][3,8]phenanthroline (99),
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-9-nitro-ben-
zo[lmn]benzo[4,5]imidazo[2,1-b][3,8]phenanthroline (100),
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-9-methoxy-b-
enzo[lmn]benzo[4,5]imidazo[2,1-b][3,8]phenanthroline (101) and
Methyl-2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benz-
o[lmn]benzo[4,5]imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid
ester (102).
25. A method of modulating the interaction of a neurotrophin and a
neurotrophin receptor, comprising contacting cells expressing a
neurotrophin receptor with an effective amount of a compound of
Formula 18, ##STR40## and pharmaceutically acceptable salts,
enantiomers, stereoisomers, rotamers, tautomers, diastereomers, or
racemates thereof; wherein R.sup.6a, R.sup.7a and R.sup.12a are
each, independently, selected from the group consisting of
hydrogen, C.sub.1-6-alkyl, C.sub.1-6-alkoxy, amino, halogen,
hydroxyl, acid, cyano, C.sub.1-6-alkyl-sulfonamide, aryl,
heteroaryl, C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, tetrazolyl, sulfone, urea, thiourea, morpholino,
piperidinyl, piperazinyl and azepanyl; and R.sup.1 is
(CH.sub.2).sub.xR.sup.13, wherein R.sup.13 is selected from the
group consisting of hydrogen, C.sub.1-6-alkyl, amino,
C.sub.1-6-alkoxy, --OH, halogen, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperazinyl, piperidinyl and azepanyl; and x
is 0, 1, 2, 3 or 4.
26. The method of claim 25, wherein R.sup.13 is selected from the
group consisting of --COOH, imidazolyl, --SO.sub.3H, --OSO.sub.3H,
--OH, morpholino, piperazinyl, --PO.sub.3H,
--PO.sub.3C.sub.1-4alkyl and --NO.sub.2.
27. The method of claim 25, wherein x is 3 and R.sup.13 is
COOH.
28. The method of claim 25, wherein R.sup.13 is
C(O)N(R.sup.8)R.sup.9 or N(R.sup.8)R.sup.9, wherein R.sup.8 and
R.sup.9 are each, independently, selected from the group consisting
of --H and --(C.sub.1-4alkyl).sub.0-1G, wherein G is selected from
the group consisting of --COOH, --H, --PO.sub.3H, --SO.sub.3H,
--Br, --Cl, --F, --OC.sub.1-4alkyl, --SC.sub.1-4alkyl, aryl,
--C(O)OC.sub.1-C.sub.6-alkyl, --C(O)C.sub.1-4alkyl-COOH,
C(O)C.sub.1-C.sub.4-alkyl and --C(O)-aryl; or N(R.sup.8)R.sup.9 is
pyrrolyl, tetrazolyl, pyrrolidinyl, pyrrolidin-2-one,
dimethylpyrrolyl, imidazolyl, morpholino.
29. The method of claim 25, wherein R.sup.13 is NH.sub.2 or
N(CH.sub.3).sub.2.
30. The method of claim_25, wherein the compound is
2-(3-Carboxy-propyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]benzo[4,5]-
imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid (95).
31. A method of modulating the interaction of a neurotrophin and a
neurotrophin receptor, comprising contacting cells expressing a
neurotrophin receptor with an effective amount of a compound of
Formulas 19 or 20, ##STR41## and pharmaceutically acceptable salts,
enantiomers, stereoisomers, rotamers, tautomers, diastereomers, or
racemates thereof; wherein R.sup.6a, R.sup.7a, R.sup.8a and
R.sup.9a are each, independently, selected from the group
consisting of hydrogen, C.sub.1-6-alkyl, C.sub.1-6-alkoxy, amino,
halogen, hydroxyl, acid, cyano, C.sub.1-6-alkyl-sulfonamide, aryl,
heteroaryl, C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, tetrazolyl, sulfone, urea, thiourea, morpholino,
piperidinyl, piperazinyl and azepanyl.
32. The method of claim 31, wherein R.sup.6a and R.sup.8a are
hydrogen, and R.sup.7a and R.sup.9a are COONa; or R.sup.6a,
R.sup.7a, R.sup.8a and R.sup.9a are hydrogen.
33. A method of modulating the interaction of a neurotrophin and a
neurotrophin receptor, comprising contacting cells expressing a
neurotrophin receptor with an effective amount of a compound of
Formula 21, ##STR42## and pharmaceutically acceptable salts,
enantiomers, stereoisomers, rotamers, tautomers, diastereomers, or
racemates thereof; wherein R.sup.1, R.sup.8a, and R.sup.9a are
each, independently, selected from the group consisting of a
hydrogen atom, C.sub.1-6-alkyl, C.sub.1-6-alkoxy, amino, halogen,
hydroxyl, acid, cyano, C.sub.1-6-alkyl-sulfonamide, aryl,
heteroaryl, C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl;
wherein the alkyl, C.sub.1-6-alkoxy, amino, aryl, heteroaryl,
cycloalkyl, and heterocycloalkyl groups may be further
independently substituted one or more times with C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, hydroxyl, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl or azepanyl.
34-40. (canceled)
41. A method of modulating the interaction of a neurotrophin and a
neurotrophin receptor, comprising contacting cells expressing a
neurotrophin receptor with an effective amount of a compound of
Formula 22, ##STR43## and pharmaceutically acceptable salts,
enantiomers, stereoisomers, rotamers, tautomers, diastereomers, or
racemates thereof; wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4 and
R.sup.5 are each, independently, selected from the group consisting
of a hydrogen atom, C.sub.1-6-alkyl, C.sub.1-6-alkoxy, amino,
halogen, hydroxyl, acid, cyano, C.sub.1-6-alkyl-sulfonamide, aryl,
heteroaryl, C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl;
wherein the alkyl, C.sub.1-6-alkoxy, amino, aryl, heteroaryl,
cycloalkyl, and heterocycloalkyl groups may be further
independently substituted one or more times with C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, hydroxyl, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl or azepanyl.
42-51. (canceled)
52. The method of claim 41, wherein the compound is
3-(1,3,6,8-Tetraoxo-1,3,6,8-tetrahydro-2-oxa-7-aza-pyren-7-yl)-benzoic
acid (45),
4-Chloro-3-(1,3,6,8-tetraoxo-1,3,6,8-tetrahydro-2-oxa-7-aza-pyren-7-yl)-b-
enzoic acid (58), and
Tetraoxo-3,6,7,8-tetrahydro-1H-2-oxa-pyren-7-yl)-butyric acid
(94).
53-57. (canceled)
58. The method of any one of claims 1, 19, 25, 31, 33 or 41,
wherein the method is used to modulate a neurotrophin-mediated
activity in a subject in need thereof.
59. The method of claim 58, wherein the neurotrophin-mediated
activity is associated with pain.
60. The method of claim 58, wherein the neurotrophin-mediated
activity is associated with an inflammatory disorder.
61. The method of claim 58, wherein the neurotrophin-mediated
activity is associated with a neurological disorder.
62-82. (canceled)
83. A method of treating pain in a subject in need thereof,
comprising administering to the subject an effective amount of a
compound selected from the group consisting of Formula 16, Formula
17, Formula 18, Formula 19, Formula 20, Formula 21 and Formula 22,
or pharmaceutically acceptable salts, enantiomers, stereoisomers,
rotamers, tautomers, diastereomers, or racemates thereof.
84-85. (canceled)
86. A method of treating an inflammatory disorder in a subject in
need thereof, comprising administering to the subject an effective
amount of a compound selected from the group consisting of Formula
16, Formula 17, Formula 18, Formula 19, Formula 20, Formula 21 and
Formula 22 or pharmaceutically acceptable salts, enantiomers,
stereoisomers, rotamers, tautomers, diastereomers, or racemates
thereof.
87. (canceled)
88. A method of treating a neurological disorder in a subject in
need thereof, comprising administering an effective amount of a
compound selected from the group consisting of Formula 16, Formula
17, Formula 18, Formula 19, Formula 20, Formula 21 and Formula 22,
or pharmaceutically acceptable salts, enantiomers, stereoisomers,
rotamers, tautomers, diastereomers, or racemates thereof.
89. (canceled)
90. A method of treating a disease or disorder associated with the
genitourinary and/or gastrointestinal systems of a subject in need
thereof, comprising administering to the subject an effective
amount of a compound selected from the group consisting of Formula
16, Formula 17, Formula 18, Formula 19, Formula 20, Formula 21 and
Formula 22, or pharmaceutically acceptable salts, enantiomers,
stereoisomers, rotamers, tautomers, diastereomers, or racemates
thereof.
91-100. (canceled)
101. A compound of the Formula 16, Formula 17, Formula 18, Formula
19, Formula 20, Formula 21 or Formula 22, or pharmaceutically
acceptable salts, enantiomers, stereoisomers, rotamers, tautomers,
diastereomers, or racemates thereof.
102. The compound of claim 101, wherein the compound is selected
from the group consisting of
4-[7-(3-Carboxy-propyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-benzo[lmn]-
[3,8]phenanthrolin-2-yl]-butyric acid (12),
3-[7-(3-Carboxy-phenyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-benzo[lmn]-
[3,8]phenanthrolin-2-yl]-4-benzoic acid (17),
3-[7-(1-chloro-4-carboxy-phenyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-b-
enzo[lmn][3,8]phenanthrolin-2-yl]-4-chloro-benzoic acid (18),
4-(1,3,6,8-Tetraoxo-7-(4-benzenesulfonic
acid)-3,6,7,8-tetrahydro-1H-benzo[lmn][3,8]phenanthrolin-2-yl)-benzenesul-
fonic acid (54), 3-(1,3,6,8-Tetraoxo-7-(3-benzenesulfonic
acid)-3,6,7,8-tetrahydro-1H-benzo[lmn][3,8]phenanthrolin-2-yl)-benzenesul-
fonic acid (55),
3-[7-(3-Carboxy-propyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-benzo[lmn]-
[3,8]phenanthrolin-2-yl]-4-chloro-benzoic acid (59),
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]b-
enzo[4,5]imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid (90),
2-(3-Carboxy-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]benzo[4,5]-
imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid (93),
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]b-
enzo[4,5]imidazo[2,1-b][3,8]phenanthroline-9 (98),
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-9-chloro-be-
nzo[lmn]benzo[4,5]imidazo[2,1-b][3,8]phenanthroline (99),
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-9-nitro-ben-
zo[lmn]benzo[4,5]imidazo[2,1-b][3,8]phenanthroline (100),
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-9-methoxy-b-
enzo[lmn]benzo[4,5]imidazo[2,1-b][3,8]phenanthroline (101),
Methyl-2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benz-
o[lmn]benzo[4,5]imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid
ester (102),
2-(3-Carboxy-propyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]ben-
zo[4,5]imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid (95),
3-(1,3,6,8-Tetraoxo-1,3,6,8-tetrahydro-2-oxa-7-aza-pyren-7-yl)-benzoic
acid (45),
4-Chloro-3-(1,3,6,8-tetraoxo-1,3,6,8-tetrahydro-2-oxa-7-aza-pyren-7-yl)-b-
enzoic acid (58), and
Tetraoxo-3,6,7,8-tetrahydro-1H-2-oxa-pyren-7-yl)-butyric acid (94).
Description
RELATED APPLICATION
[0001] This application claims priority to U.S. Provisional
Application No. 60/718,256, Attorney Docket No. PCI-030-1, filed
Sep. 15, 2005, entitled "METHODS OF MODULATING
NEUROTROPHIN-MEDIATED ACTIVITY." The contents of any patents,
patent applications, and references cited throughout this
specification are hereby incorporated by reference in their
entireties.
TECHNICAL FIELD
[0002] The present invention relates to compositions which modulate
the interaction of nerve growth factor, and precursors thereof,
with the receptor TrkA, as well as the common neurotrophin receptor
p75.sup.NTR, and methods of use thereof.
BACKGROUND
[0003] The neurotrophins are a family of structurally and
functionally related proteins, including Nerve Growth Factor (NGF),
Brain-Derived Neurotrophic Factor (BDNF), Neurotrophin-3 (NT-3),
Neurotrophin-4/5 (NT-4/5) and Neurotrophin-6 (NT-6). These proteins
promote the survival and differentiation of diverse neuronal
populations in both the peripheral and central nervous systems and
are involved in the pathogenesis of diverse neurological disorders
(Hefti, J. Neurosci. 6:2155-2162 (1986); Hefti and Weiner, Annals
of Neurology 20:275-281 (1986); Levi-Montalcini, EMBO J.
6:1145-1154 (1987); Barde, Neuron 2:1525-1534 (1989); Leibrock et
al., Nature 341:149-152 (1989); Maisonpierre et al., Science
247:1446-1451 (1990); Rosenthal et al., Neuron 4:767-773 (1990);
Hohn et al., Nature 344:339-341 (1990); Gotz et al., Nature
372:266-269 (1994); Maness et al., Neurosci. Biobehav. Rev.
18:143-159 (1994); Dechant et al., Nature Neurosci. 5:1131-1136
(2002)). This broad spectrum of biological activities exerted by
the neurotrophins results from their ability to bind and activate
two structurally unrelated receptor types, the p75 neurotrophin
receptor (p75.sup.NTR) and the three members of the Trk receptor
family of tyrosine kinases (Kaplan et al., Curr. Opin. Cell Biol.
9:213-221 (1997); Friedman et al., Exp. Cell Res. 253:131-142
(1999); Patapoutian et al., Curr. Opin. Neurobiol. 11:272-280
(2001)).
[0004] While NGF was initially studied for its essential role in
neuronal growth and survival, recent reports indicate that this
neurotrophin may also play a role in inflammation and disorders of
the respiratory, the genitourinary and the gastrointestinal
systems. For example, in the gastrointestinal tract, neurotrophins
and neurotrophic factors regulate neuropeptide expression, interact
with immunoregulatory cells and epithelial cells, and regulate
motility during inflammation (Reinshagen, M. et al., Curr. Opin.
Investig. Drugs. 2002; 3(4): 565-568). NGF has been shown to play a
role in bladder overactivity (Lamb, K. et al., J. Pain. 2004; 5(3):
150-156), bladder outlet obstruction (Kim, J. C. et al., BJU Int.
2004; 94(6): 915-918), pancreatic cancer (Shi, X. et al.,
Pancreatology. 2001; 1(5):517-524), and intestinal inflammation
(Lin, A. et al., Exp. Neurol. 2005; 191(2):337-43).
[0005] NGF is synthesized as a larger precursor form (referred to
herein as "proNGF," also known as "preproNGF" or "pro-peptide NGF")
which is then processed by proteolytic cleavages to produce the
mature neurotrophic factor. This prepro region is located at the
amino terminus of the precursor molecule and is needed for proper
folding and secretion of the NGF protein. The mature form of NGF
has arginine residues at its carboxy termini which requires that a
leucine residue be inserted between the naturally occurring
arginine and the hydrophilic spacer. The primary structure of
proNGF has been deduced from the nucleotide sequence of the mouse
NGF cDNA (Scott et al. Nature 302:538 (1983); Ullrich et al. Nature
303:821 (1983)).
[0006] The common neurotrophin receptor p75.sup.NTR is a
transmembrane glycoprotein structurally related to the tumor
necrosis factor and CD-40 receptors (Meakin and Shooter, Trends
Neurosci. 15:323-331 (1992), Ryden and Ibanez, J. Biol. Chem.
271:5623-5627 (1996)). As all neurotrophins bind to p75.sup.NTR
with similar affinities (Rodrigues-Tebar et al., Neuron 4:487-492
(1990); Hallbook et al., Neuron 6:845-858 (1991); Rodrigues-Tebar
et al., EMBO J. 11:917-922 (1992); Ibanez, Trends Biotech.
13:217-227 (1995)), neurotrophin specificity is conventionally
thought to be conferred by the binding selectivity for Trk
receptors which are differentially expressed in different neuronal
populations (Ibanez, Trends Biotech. 13:217-227 (1995)). However,
accumulated experimental data on neurotrophin activity reveal
important functional aspects of p75.sup.NTR (Heldin et al., J.
Biol. Chem. 264:8905-8912 (1989); Jing et al., Neuron 9:1067-1079
(1992); Herrmann et al., Mol. Biol. 4:1205-1216 (1993); Barker and
Shooter, Neuron 13:203-215 (1994); Dobrowsky et al., Science
265:1596-1599 (1994), Matsumoto et al., Cancer Res. 55:1798-1806
(1995); Marchetti et al., Cancer Res. 56:2856-2863 (1996);
Washiyama et al., Amer. J. Path. 148:929-940 (1996)). The common
neurotrophin receptor enhances functions and increases binding
specificity of Trk receptors (Barker and Shooter, Neuron 13:203-215
(1994); Mahadeo et al., J. Biol. Chem. 269:6884-6891 (1994); Chao
and Hempstead, Trends Neurosci. 18:321-326 (1995); Ryden and Ibanz,
J. Biol. Chem. 271:5623-5627 (1996)). In addition, p75.sup.NTR
possesses unique, neurotrophin dependent, Trk-independent signaling
properties which involve ceramide production through activation of
the sphingomyelin cycle (Dobrowsky et al., Science 265:1596-1599
(1994)), apoptosis (cell death) (Cassacia-Bonnefil et al., Nature
383:716-719 (1996)), and activation of the transcription factor
NFKB (Carter et al., Science 272:542-545 (1996)).
[0007] Moreover, while initially studied primarily in neurons,
p75.sup.NTR has also been found to play critical roles in vascular
biology (von Schack et al., Nat. Neurosci. 4:977-978, 2001; Wan et
al., Am. J. Pathol. 157:1247-1258, 2001), glial biology (Bentley et
al., J. Neurosci. 20:7706-7715, 2000; Syroid et al., J. Neurosci.
20:5741-5747, 2000), the immune system (Tokuoka et al., Br. J.
Pharmacol. 134:1580-1586, 2001), and tumor biology (Sakamoti et
al., Oncol. Rep. 8:973-980, 2001; Descamps et al., J. Biol. Chem.
276:17864017870, 2001). For example, p75.sup.NTR has been
demonstrated to participate in human melanoma progression (Herrmann
et al., Mol. Biol. 4:1205-1216 (1993); Marchetti et al., Cancer
Res. 56:2856-2863 (1996)). Furthermore, NGF and NT-3 increase the
production of heparin by 70W melanoma cells, which is associated
with their metastatic potential (Marchetti et al., Cancer Res.
56:2856-2863 (1996)).
[0008] Unlike p75.sup.NTR, the Trk receptors (TrkA, TrkB and TrkC)
exhibit selectivity for specific neurotrophins. (Kaplan et al.,
Science 252:554-558 (1991); Klein et al., Cell 65:189-197 (1991);
Klein et al., Neuron 8:947-956 (1992); Soppet et al., Cell
65:895-903 (1991); Squinto et al., Cell 65:885-893 (1991);
Berkemeier et al., Neuron 7:857-866 (1991); Escandon et al.,
Neurosci. Res. 34:601-613 (1993); Lamballe et al., Cell 66:967-970
(1991)). For example, TrkA primarily binds NGF (Kaplan et al.,
1991; Klein et al., 1991) and has been reported to bind NT-3 (J.
Biol. Chem. 271(10):5623-7, 1996); TrkB binds BDNF and NT-4/5
(Soppet et al., 1991; Squinto et al., 1991; Berkemeier et al.,
1991; Escandon et al., 1993; Lamballe et al., 1991; Klein et al.,
1992; Vale and Shooter, Methods Enzymol. 109:21-39 (1985);
Barbacid, Oncogene 8:2033-2042 (1993)); and TrkC exclusively binds
NT-3 (Lamballe et al., 1991; Vale and Shooter, 1985). This is
particularly evident when the Trk receptors are coexpressed with
the common neurotrophin receptor p75.sup.NTR. (For review see
Meakin and Shooter, 1992; Barbacid, 1993; Chao, 1994; Bradshaw et
al., 1994; Ibanez, 1995).
[0009] Biochemical experiments indicate that neurotrophin receptors
form at least three different types of complexes: homodimers of Trk
receptors, homomeric p75.sup.NTR receptors and mixed complexes of
both Trk and p75.sup.NTR. These complexes may coexist in cells and
may be linked through biochemical equilibria. Functionally, their
signaling has been shown to be independent, synergistic or
antagonistic. The response of a cell to neurotrophins is thus
determined by the quantitative and qualitative composition of its
receptor complement in combination with biochemical equilibria
between pools of active and inactive receptors (Dechant, Cell
Tissue Res. 305:229-238, (2001)), as well as other cellular and
biochemical components downstream of the neurotrophin receptors,
e.g., the availability of proteins, lipids and inorganic molecules
involved in signal transduction.
[0010] Due to the implication of NGF, and its precursor proNGF,
binding to homomeric and heteromeric neurotrophin receptor
complexes in various disease states, especially pain, inflammation,
neurological disorders and disorders of the respiratory,
genitourinary and gastrointestinal systems, a need exists for
pharmaceutical agents and methods of use thereof for modulating the
interactions of NGF with the common neurotrophin receptor
p75.sup.NTR, and the Trk receptor TrkA.
SUMMARY OF THE INVENTION
[0011] There remains a need for new treatments and therapies for
neurotophin-mediated activity, and conditions, diseases and
disorders related to neurotophin-mediated activity. There is also a
need for compounds useful in the treatment or prevention or
amelioration of one or more symptoms of pain, inflammatory
disorders, neurological disorders, respiratory disorders,
genitourinary disorders, and/or gastrointestinal disorders.
Furthermore, there is a need for methods for modulating the
activity of NGF, proNGF, p75.sup.NTR, and/or TrkA, using the
compounds provided herein.
[0012] In one aspect, the invention provides a compound of the
invention of the Formula 16, Formula 17, Formula 18, Formula 19,
Formula 20, Formula 21 and Formula 22, as well as the compounds of
Table 1.
[0013] In another aspect, the invention provides a method of
modulating the interaction of a neurotrophin and a neurotrophin
receptor, comprising contacting cells expressing a neurotrophin
receptor with an effective amount of a compound of the
invention.
[0014] In one embodiment, the neurotrophin is nerve growth factor
and/or precursors thereof. In another embodiment, the neurotrophin
receptor is selected from the group consisting of p75.sup.NTR and
TrkA. In yet another embodiment, the neurotrophin receptor is
p75.sup.NTR. In still another embodiment, the neurotrophin receptor
is TrkA. In another embodiment the compound further modulates the
interaction of NGF and/or proNGF with TrkA.
[0015] In another embodiment, the method is used to modulate a
neurotrophin-mediated activity in a subject in need thereof. In
another embodiment, the neurotrophin-mediated activity is
associated with pain. In still another embodiment, the
neurotrophin-mediated activity is associated with an inflammatory
disorder. In another embodiment, the neurotrophin-mediated activity
is associated with a neurological disorder.
[0016] In another embodiment, the pain treated by the compounds of
the invention is selected from the group consisting of cutaneous
pain, somatic pain, visceral pain and neuropathic pain. In another
embodiment the pain is acute pain or chronic pain.
[0017] In still another embodiment, the cutaneous pain is
associated with injury, disease, disorder or neoplasms of the skin,
subcutaneous tissues and related organs. In another embodiment, the
injury, disease or disorder of the skin, subcutaneous tissues and
related organs is selected from the group consisting of traumas,
cuts, lacerations, punctures, burns, surgical incisions,
infections, psoriasis, eczema, and inflammation (e.g., acute
inflammation).
[0018] In another embodiment, the somatic pain is associated with
an injury, disease, disorder or neoplasms of the musculoskeletal
and connective system. In another embodiment, the injury, disease
or disorder of the musculoskeletal and connective system is
selected from the group consisting of sprains, broken bones,
arthritis, arthralgia, myalgia, chronic lower back pain,
cancer-associated pain, dental pain, fibromyalgia, idiopathic pain
disorder, chronic non-specific pain, post-operative pain, and
referred pain.
[0019] In another embodiment, the visceral pain is associated with
an injury, disease, disorder or neoplasms of the circulatory
system, the respiratory system, the gastrointestinal system, or the
genitourinary system. In one embodiment, the disease or disorder of
the circulatory system treated by the compounds of the invention is
selected from the group consisting of ischaemic heart disease,
angina, acute myocardial infarction, cardiac arrhythmia, phlebitis,
intermittent claudication, varicose veins and hemorrhoids. In one
embodiment, the disease or disorder of the respiratory system
treated by the compounds of the invention is selected from the
group consisting of asthma, Chronic Obstructive Pulmonary Disease
(COPD), respiratory infection, chronic bronchitis and emphysema. In
one embodiment, the disease or disorder of the gastrointestinal
system treated by the compounds of the invention is selected from
the group consisting of gastritis, duodenitis, irritable bowel
syndrome, colitis, Crohn's disease, ulcers and diverticulitis. In
one embodiment, the disease or disorder of the genitourinary system
treated by the compounds of the invention is selected from the
group consisting of cystitis, urinary tract infections,
glomuerulonephritis, polycystic kidney disease, and kidney
stones.
[0020] In another embodiment, the neuropathic pain is associated
with an injury, disease, disorder or neoplasms of the nervous
system. In still another embodiment, the injury, disease or
disorder of the nervous system is selected from the group
consisting of neuralgia, neuropathy, headache, chronic cephalic
pain, phantom limb pain and spinal cord injury.
[0021] In one embodiment, the inflammatory disorder treated by the
compounds of the invention is selected from an inflammatory
disorder of the skin and subcutaneous tissues, the musculoskeletal
and connective tissue system, the respiratory system, the
circulatory system, the genitourinary system, the gastrointestinal
system or the nervous system. In one embodiment, the inflammatory
disorder of the skin and subcutaneous tissues is selected from the
group consisting of psoriasis, dermatitis and eczema. In one
embodiment, the inflammatory disorder of the musculoskeletal and
connective tissue system is selected from the group consisting of
arthritis, gout, myositis, bursitis and synovitis. In one
embodiment, the inflammatory disorder of the respiratory system
treated by the compounds of the invention is selected from the
group consisting of asthma, bronchitis, sinusitis, pharyngitis,
rhinitis and respiratory infections. In another embodiment, the
inflammatory disorder of the circulatory system is selected from
the group consisting of vasculitis, artherosclerosis, phlebitis,
carditis and coronary heart disease. In one embodiment, the
inflammatory disorder of the gastrointestinal system treated by the
compounds of the invention is selected from the group consisting of
inflammatory bowel disorder, ulcerative colitis, Crohn's disease,
diverticulitis, viral infection, bacterial infection, chronic
hepatitis, gingivitis, stomatitis, and gastritis. In one
embodiment, the inflammatory disorder of the genitourinary system
treated by the compounds of the invention is selected from the
group consisting of cystitis, nephritic syndrome,
glomerulonephritis, urinary tract infection, prostatitis,
salpingitis, endometriosis and cystinosis.
[0022] In another embodiment, the neurological disorder treated by
the compounds of the invention is selected from the group
consisting of schizophrenia, bipolar disorder, depression,
Alzheimer's disease, epilepsy, multiple sclerosis, amyotrophic
lateral sclerosis, stroke, cerebral ischemia, neuropathy, retinal
pigment degeneration, glaucoma, cardiac arrhythmia, shingles,
Huntington's chorea, and Parkinson's disease.
[0023] In another aspect, the invention provides a method of
treating pain in a subject in need thereof, comprising
administering to the subject an effective amount of a compound of
the invention. In one embodiment, the pain is selected from the
group consisting of cutaneous pain, somatic pain, visceral pain and
neuropathic pain. In another embodiment, the pain is acute pain,
breakthrough pain or chronic pain.
[0024] In another aspect, the invention provides a method of
treating an inflammatory disorder in a subject in need thereof,
comprising administering to the subject an effective amount of a
compound of the invention. In one embodiment, the inflammatory
disorder is inflammatory disorder of the musculoskeletal and
connective tissue system, the respiratory system, the circulatory
system, the genitourinary system, the gastrointestinal system or
the nervous system.
[0025] In another aspect, the invention provides a method of
treating a neurological disorder in a subject in need thereof,
comprising administering an effective amount of a compound of the
invention. In one embodiment, the neurological disorder is selected
from the group consisting of schizophrenia, bipolar disorder,
depression, Alzheimer's disease, epilepsy, multiple sclerosis,
amyotrophic lateral sclerosis, stroke, cerebral ischemia,
neuropathy, retinal pigment degeneration, glaucoma, cardiac
arrhythmia, Huntington's chorea, and Parkinson's disease.
[0026] In another aspect, the invention provides a method of
treating a disease or disorder associated with the genitourinary
and/or gastrointestinal systems of a subject in need thereof,
comprising administering to the subject an effective amount of a
compound of the invention. In one embodiment, the disease or
disorder of the gastrointestinal system is selected from the group
consisting of gastritis, duodenitis, irritable bowel syndrome,
colitis, Crohn's disease, ulcers and diverticulitis. In another
embodiment the disease or disorder of the genitourinary system is
selected from the group consisting of cystitis, urinary tract
infections, glomuerulonephritis, polycystic kidney disease, kidney
stones and cancers of the genitourinary system.
[0027] In another aspect, the invention provides a method
comprising administering to the subject an additional therapeutic
agent. In one embodiment, the additional therapeutic agent is
selected from the group consisting of an analgesic, an
anti-inflammatory agent, an anesthetic, a corticosteroid, an
anti-convulsant, an antidepressant, an anti-nausea/anti-emetic
agent, an anti-psychiatric agent, a cardiovascular agent and a
cancer therapeutic.
BRIEF DESCRIPTION OF THE DRAWINGS
[0028] FIG. 1 shows a typical Western blot with immunodetection of
phosphorylated Erk 1/2 proteins extracted from NGF-stimulated PC12
cells in the presence or absence of Compound 90, as described in
Example 3. These results demonstrate that Compound 90 inhibits NGF
(5 ng/mL)-induced Erk 1/2 phosphorylation in PC12 cells.
[0029] FIG. 2A shows a dose-response curve for Compound 90 from an
individual NGF binding displacement experiment in PC 12 cells as
described in Example 1. The dose-response curve demonstrates that
Compound 90 effectively blocks NGF binding to cells expressing TrkA
and p75.
[0030] FIG. 2B shows a dose-response curve for Compound 90 from an
individual NGF binding displacement experiment in A875 cells as
described in Example 1. The dose-response curve demonstrates that
Compound 90 effectively blocks NGF binding to cells expressing
p75.
[0031] FIG. 3A shows a binding displacement curve for Compound 90
from an individual NGF binding displacement experiment in HEK_trk
cells as described in Example 1. The dose-response curve
demonstrates that Compound 90 effectively blocks NGF binding to
cells expressing TrkA.
[0032] FIG. 3B shows a neurite outgrowth inhibition curve for
Compound 90 resulting from an experiment as described in Example 4.
The curve demonstrates that Compound 90 effectively inhibits
neurite outgrowth induced by NGF.
[0033] FIG. 4A shows a dose-response curve for Compound 91 from an
individual NGF binding displacement experiment in PC12 cells as
described in Example 1. The dose-response curve demonstrates that
Compound 91 effectively blocks NGF binding to cells expressing TrkA
and p75.
[0034] FIG. 4B shows a typical Western blot with immunodetection of
phosphorylated Erk 1/2 proteins extracted from NGF-stimulated PC12
cells in the presence or absence of Compound 91, as described in
Example 3. These results demonstrate that Compound 91 inhibits
NGF-induced Erk 1/2 phosphorylation in PC12 cells.
[0035] FIG. 5 shows a dose-response curve for Compound 18 from an
individual NGF binding displacement experiment in PC12 cells as
described in Example 1. The dose-response curve demonstrates that
Compound 18 effectively blocks NGF binding to cells expressing TrkA
and p75.
[0036] FIG. 6 shows a phospho-Erk 1/2 blot for Compound 90 as
acquired using the procedure in Example 3, which demonstrates that
Compound 90 inhibits NGF (1 ng/mL)-induced Erk 1/2 phosphorylation
in PC12 cells.
[0037] FIG. 7 illustrates the significant antinociceptive effects
of Compound 91 in the Writhing Test in mice, as described in
Example 6.
DETAILED DESCRIPTION OF THE INVENTION
[0038] The present invention relates to the discovery of compounds
which modulate the interaction of a neurotrophin--either in its
mature (e.g., NGF) or precursor (e.g., proNGF) form--with a
neurotrophin receptor, for example, the common neurotrophin
receptor p75.sup.NTR and/or a Trk receptor. Such compounds are of
use, for example, for modulating the interaction of NGF and/or a
precursor thereof (e.g., proNGF) to p75.sup.NTR, and the compounds
within the invention can also have the ability to modulate the
interaction of NGF and/or proNGF with TrkA. For example, a compound
that modulates the binding of NGF or proNGF to p75.sup.NTR can
further modulate the binding of the neurotrophin to TrkA. Such
compounds can also be used to treat a subject having a condition
with at least one symptom that is directly or indirectly mediated,
at least in part, by the interaction of NGF and/or a precursor
thereof with p75.sup.NTR and/or TrkA.
[0039] Nerve growth factor (also referred to hereinafter as "NGF")
is a prototypic neurotrophin, and is best known for its essential
role during development of peripheral sensory and sympathetic
neurons. NGF is produced as a high-molecular weight precursor
(pro-NGF) that contains a pro-domain linked to the N-terminus which
is cleaved by the endoprotease furin in the trans-Golgi network of
neurons (Mowla et al., J. Biol. Chem. 276:12660-12666, 2001; Mowla
et al., J. Neurosci. 19:2069-2080, 1999). Pro-NGF has been shown to
be induced and secreted after injury to the CNS in an active form
that is capable of triggering cell apoptosis (e.g., of neuronal
cells and oligodendrocytes), and disruption of the interaction of
pro-NGF and p75.sup.NTR has been demonstrated to rescue injured
adult rat corticospinal neurons (e.g., Harrington et al., PNAS USA
101(16):6226-6230, 2004). Mature NGF regulates the phenotype (e.g.,
cell body and dendrite size, gene expression and neurotransmitter
phenotype) of peripheral neurons and certain CNS neurons, notably,
basal forebrain and striatal cholinergic neurons throughout the
life of an animal (Miller et al., Neuron 32:767-770, 2001; Ruberti
et al., J. Neurosci. 20(7):2589, 2000; Chen et al, J. Neurosci.
17(19):7288-96, 1997; Fagan et al., J. Neurosci. 17(20):7644-54,
1997). NGF has been implicated in the pathogenesis of Alzheimer's
disease, epilepsy and pain (Ben Ari and Represa, TINS 13:312-318
(1990); McKee et al., Ann. Neurol. 30:156 (1991); Leven and Mendel,
TINS 16:353-359 (1993); Woolf and Doubell, Current Opinions in
Neurobiol. 4:525-534 (1994); Rashid et al., Proc. Natl. Acad. Sci.
U.S.A. 92:9495-9499 (1995); McMahon et al., Nature Med. 1:774-780
(1995)). The interaction of NGF with its receptors is determined by
distinct sequences within its primary amino acid structure. While
several regions of NGF participate in the NGF/TrkA interaction,
mutation studies suggest that relatively few key residues, namely
those located in the NGF amino and carboxyl termini, are primarily
required for high affinity binding to TrkA.
[0040] Recent results have also shown that NGF may play a role in
inflammation and disorders of the respiratory, genitourinary and
gastrointestinal systems. For example, in the gastrointestinal
tract, neurotrophins and neurotrophic factors regulate neuropeptide
expression, interact with immunoregulatory cells and epithelial
cells, and regulate motility during inflammation (Reinshagen, M. et
al., Curr. Opin. Investig. Drugs. 2002; 3(4): 565-568). NGF has
been shown to be a potentially relevant treatment target for
bladder overactivity (Lamb, K. et al., J. Pain. 2004; 5(3):
150-156). Studies by Kim et al. show that increases in NGF may be
related to the irritative symptoms resulting from correction of
bladder outlet obstruction (BJU Int. 2004; 94(6): 915-918). Shi et
al. have demonstrated that NGF and its receptor are overexpressed
in pancereatic cancer and contribute to its malignant phenotype
(Pancreatology. 2001; 1(5):517-524). The potential of NGF as a
neuroprotective factor in the enteric nervous system has been
demonstrated, as has its role in the mechanism by which intestinal
inflammation can give rise to a permanent imbalance between
excitatory and inhibitory pathways, thus tending to compromise
intestinal function (Maruccio, L. et al. Histol. Histopathol. 2004;
19(2):349-356; Lin, A. et al., Exp. Neurol. 2005; 191(2):337-43).
NGF has also been shown to play an important role in the
development of the rat ovary (Romero, C. et al., 2002;
143(4):1485-1494). Data has further suggested that NGF may play a
role in inflammation, bronchial hyperresponsiveness and airway
remodeling in asthma, and may help us to understand the
neuro-immune cross-talk involved in chronic inflammatory airway
diseases (Frossard, N. et al., Eur. J. Pharmacol. 2004; 500(1-3):
453-465).
[0041] It has been assumed, based on studies in the mouse
submandibular gland that NGF in vivo is largely in the mature form
of NGF, and that mature NGF accounts for the molecule's biological
activity. However, it has been recently shown that proNGF is
abundant in central nervous system tissues whereas mature NGF is
undetectable, suggesting that proNGF may have a function distinct
from its role as a precursor. Moreover, this data suggests that
proNGF may be responsible for some of the biological activity
normally attributed to mature NGF in vivo (Fahnestock, M. et al.,
J. Neurochem. 2004; 89(3):581-592; Fahnestock, M. et al., Prog.
Brain Res. 2004; 146: 107-110). For example, it has been
demonstrated that proNGF levels increase during the preclinical
stage of Alzheimer's disease (Peng, S. et al., J. Neuropathol. Exp.
Neurol. June 2004; 63(6):641-9). Additionally, studies by Beattie
et al. (Neuron Oct. 24, 2002; 36(3): 275-386) have shown that
proNGF plays an important role in eliminating damaged cells by
activating the apoptotic machinery via p75.sup.NTR after spinal
cord injury.
[0042] Based on the above, there is a need for compositions which
modulate the interaction of nerve growth factor, and precursors
thereof, with the receptor TrkA, as well as the common neurotrophin
receptor p75.sup.NTR, and methods of use thereof.
DEFINITIONS
[0043] The term "electronegative atom," as used herein, refers to
an atom which carries a partial or full negative charge in a
particular compound under physiological conditions. The
electronegative atom can be, for example, an oxygen atom, a
nitrogen atom, a sulfur atom or a halogen atom, such as a fluorine,
chlorine, bromine or iodine atom. Preferably, the electronegative
atom is an oxygen atom. The term "electronegative functional
group," as used herein, refers to a functional group which includes
at least one electronegative atom. Electronegative groups include
acid functional groups and other polar functional groups. For
example, suitable electronegative functional groups include, but
are not limited to, carbonyl, thiocarbonyl, ester, imino, amido,
amine, carboxylic acid, sulfonic acid, sulfinic acid, sulfamic
acid, phosphonic acid, boronic acid, sulfate ester, hydroxyl,
mercapto, cyano, cyanate, thiocyanate, isocyanate, isothiocyanate,
carbonate, nitrate and nitro groups. It is to be understood that,
unless otherwise indicated, reference herein to an acidic
functional group also encompasses salts of that functional group in
combination with a suitable cation.
[0044] As used herein, the term "acid" refers to any substituent
that can readily donate a hydrogen ion to another compound.
Particularly preferred acid functional groups include carboxylic
acid, sulfonic acid, sulfinic acid, sulfamic acid, phosphonic acid
and boronic acid functional groups.
[0045] The term "alkyl" includes saturated aliphatic groups,
including straight-chain alkyl groups (e.g., methyl, ethyl, propyl,
butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, etc.),
branched-chain alkyl groups (isopropyl, tert-butyl, isobutyl,
etc.), cycloalkyl (alicyclic) groups (cyclopropyl, cyclopentyl,
cyclohexyl, cycloheptyl, cyclooctyl), alkyl substituted cycloalkyl
groups, and cycloalkyl substituted alkyl groups. The term "alkyl"
also includes alkenyl groups and alkynyl groups. Furthermore, the
expression "C.sub.x-C.sub.y-alkyl", wherein x is 1-5 and y is 2-10
indicates a particular alkyl group (straight- or branched-chain) of
a particular range of carbons. For example, the expression
C.sub.1-C.sub.4-alkyl includes, but is not limited to, methyl,
ethyl, propyl, butyl, isopropyl, tert-butyl and isobutyl.
[0046] The term alkyl further includes alkyl groups which can
further include oxygen, nitrogen, sulfur or phosphorous atoms
replacing one or more carbons of the hydrocarbon backbone. In an
embodiment, a straight chain or branched chain alkyl has 10 or
fewer carbon atoms in its backbone (e.g., C.sub.1-C.sub.10 for
straight chain, C.sub.3-C.sub.10 for branched chain), and more
preferably 6 or fewer carbons. Likewise, preferred cycloalkyls have
from 4-7 carbon atoms in their ring structure, and more preferably
have 5 or 6 carbons in the ring structure.
[0047] Moreover, alkyl (e.g., methyl, ethyl, propyl, butyl, pentyl,
hexyl, etc.) include both "unsubstituted alkyl" and "substituted
alkyl", the latter of which refers to alkyl moieties having
substituents replacing a hydrogen on one or more carbons of the
hydrocarbon backbone, which allow the molecule to perform its
intended function. The term "substituted" is intended to describe
moieties having substituents replacing a hydrogen on one or more
atoms, e.g. C, O or N, of a molecule. Such substituents can
include, for example, alkenyl, alkynyl, halogen, hydroxyl,
alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy,
aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl,
alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl,
dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate,
phosphonato, phosphinato, amino (including alkyl amino,
dialkylamino, arylamino, diarylamino, and alkylarylamino),
acylamino (including alkylcarbonylamino, arylcarbonylamino,
carbamoyl and ureido), amidino, imino, sulfhydryl, alkylthio,
arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato,
sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido,
heterocyclyl, alkylaryl, morpholino, phenol, benzyl, phenyl,
piperizine, cyclopentane, cyclohexane, pyridine, 5H-tetrazole,
triazole, piperidine, or an aromatic or heteroaromatic moiety.
[0048] Further examples of substituents of the invention, which are
not intended to be is limiting, include moieties selected from
straight or branched alkyl (preferably C.sub.1-C.sub.5), cycloalkyl
(preferably C.sub.3-C.sub.8), alkoxy (preferably C.sub.1-C.sub.6),
thioalkyl (preferably C.sub.1-C.sub.6), alkenyl (preferably
C.sub.2-C.sub.6), alkynyl (preferably C.sub.2-C.sub.6),
heterocyclic, carbocyclic, aryl (e.g., phenyl), aryloxy (e.g.,
phenoxy), aralkyl (e.g., benzyl), aryloxyalkyl (e.g.,
phenyloxyalkyl), arylacetamidoyl, alkylaryl, heteroaralkyl,
alkylcarbonyl and arylcarbonyl or other such acyl group,
heteroarylcarbonyl, or heteroaryl group, (CR'R'').sub.0-3NR'R''
(e.g., --NH.sub.2), (CR'R'').sub.0-3CN (e.g., --CN), --NO.sub.2,
halogen (e.g., --F, --Cl, --Br, or --I),
(CR'R'').sub.0-3C(halogen).sub.3 (e.g., --CF.sub.3),
(CR'R'').sub.0-3CH(halogen).sub.2,
(CR'R'').sub.0-3CH.sub.2(halogen), (CR'R'').sub.0-3CONR'R'',
(CR'R'').sub.0-3(CNH)NR'R'', (CR'R'').sub.0-3S(O).sub.1-2NR'R'',
(CR'R'').sub.0-3CHO, (CR'R'').sub.0-3O(CR'R'').sub.0-3H,
(CR'R'').sub.0-3S(O).sub.0-3R' (e.g., --SO.sub.3H, --OSO.sub.3H),
(CR'R'').sub.0-3O(CR'R'').sub.0-3H (e.g., --CH.sub.2OCH.sub.3 and
--OCH.sub.3), (CR'R'').sub.0-3S(CR'R'').sub.0-3H (e.g., --SH and
--SCH.sub.3), (CR'R'').sub.0-3OH (e.g., --OH),
(CR'R'').sub.0-3COR', (CR'R'').sub.0-3(substituted or unsubstituted
phenyl), (CR'R'').sub.0-3(C.sub.3-C.sub.8 cycloalkyl),
(CR'R'').sub.0-3CO.sub.2R' (e.g., --CO.sub.2H), or
(CR'R'').sub.0-3OR' group, or the side chain of any naturally
occurring amino acid; wherein R' and R'' are each independently
hydrogen, a C.sub.1-C.sub.5 alkyl, C.sub.2-C.sub.5 alkenyl,
C.sub.2-C.sub.5 alkynyl, or aryl group. Such substituents can
include, for example, halogen, hydroxyl, alkylcarbonyloxy,
arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy,
carboxylate, alkylcarbonyl, alkoxycarbonyl, aminocarbonyl,
alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato,
cyano, amino (including alkyl amino, dialkylamino, arylamino,
diarylamino, and alkylarylamino), acylamino (including
alkylcarbonylamino, arylcarbonylamino, carbamoyl and ureido),
amidino, imino, oxime, sulfhydryl, alkylthio, arylthio,
thiocarboxylate, sulfates, sulfonato, sulfamoyl, sulfonamido,
nitro, trifluoromethyl, cyano, azido, heterocyclyl, or an aromatic
or heteroaromatic moiety. In certain embodiments, a carbonyl moiety
(C.dbd.O) may be further derivatized with an oxime moiety, e.g., an
aldehyde moiety may be derivatized as its oxime (--C.dbd.N--OH)
analog. It will be understood by those skilled in the art that the
moieties substituted on the hydrocarbon chain can themselves be
substituted, if appropriate. Cycloalkyls can be further
substituted, e.g., with the substituents described above. An
"aralkyl" moiety is an alkyl substituted with an aryl (e.g.,
phenylmethyl (i.e., benzyl)).
[0049] The term "alkenyl" includes unsaturated aliphatic groups
analogous in length and possible substitution to the alkyls
described above, but which contain at least one double bond.
[0050] For example, the term "alkenyl" includes straight-chain
alkenyl groups (e.g., ethenyl, propenyl, butenyl, pentenyl,
hexenyl, heptenyl, octenyl, nonenyl, decenyl, etc.), branched-chain
alkenyl groups, cycloalkenyl (alicyclic) groups (cyclopropenyl,
cyclopentenyl, cyclohexenyl, cycloheptenyl, cyclooctenyl), alkyl or
alkenyl substituted cycloalkenyl groups, and cycloalkyl or
cycloalkenyl substituted alkenyl groups. The term alkenyl further
includes alkenyl groups that include oxygen, nitrogen, sulfur or
phosphorous atoms replacing one or more carbons of the hydrocarbon
backbone. In certain embodiments, a straight chain or branched
chain alkenyl group has 6 or fewer carbon atoms in its backbone
(e.g., C.sub.2-C.sub.6 for straight chain, C.sub.3-C.sub.6 for
branched chain). Likewise, cycloalkenyl groups may have from 3-8
carbon atoms in their ring structure, and more preferably have 5 or
6 carbons in the ring structure. The term C.sub.2-C.sub.6 includes
alkenyl groups containing 2 to 6 carbon atoms.
[0051] Moreover, the term alkenyl includes both "unsubstituted
alkenyls" and "substituted alkenyls", the latter of which refers to
alkenyl moieties having substituents replacing a hydrogen on one or
more carbons of the hydrocarbon backbone. Such substituents can
include, for example, alkyl groups, alkynyl groups, halogens,
hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy,
aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl,
alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl,
dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate,
phosphonato, phosphinato, cyano, amino (including alkyl amino,
dialkylamino, arylamino, diarylamino, and alkylarylamino),
acylamino (including alkylcarbonylamino, arylcarbonylamino,
carbamoyl and ureido), amidino, imino, sulfhydryl, alkylthio,
arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato,
sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido,
heterocyclyl, alkylaryl, or an aromatic or heteroaromatic
moiety.
[0052] The term "alkynyl" includes unsaturated aliphatic groups
analogous in length and possible substitution to the alkyls
described above, but which contain at least one triple bond.
[0053] For example, the term "alkynyl" includes straight-chain
alkynyl groups (e.g., ethynyl, propynyl, butynyl, pentynyl,
hexynyl, heptynyl, octynyl, nonynyl, decynyl, etc.), branched-chain
alkynyl groups, and cycloalkyl or cycloalkenyl substituted alkynyl
groups. The term alkynyl further includes alkynyl groups that
include oxygen, nitrogen, sulfur or phosphorous atoms replacing one
or more carbons of the hydrocarbon backbone. In certain
embodiments, a straight chain or branched chain alkynyl group has 6
or fewer carbon atoms in its backbone (e.g., C.sub.2-C.sub.6 for
straight chain, C.sub.3-C.sub.6 for branched chain). The term
C.sub.2-C.sub.6 includes alkynyl groups containing 2 to 6 carbon
atoms.
[0054] Moreover, the term alkynyl includes both "unsubstituted
alkynyls" and "substituted alkynyls", the latter of which refers to
alkynyl moieties having substituents replacing a hydrogen on one or
more carbons of the hydrocarbon backbone. Such substituents can
include, for example, alkyl groups, alkynyl groups, halogens,
hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy,
aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl,
alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl,
dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate,
phosphonato, phosphinato, cyano, amino (including alkyl amino,
dialkylamino, arylamino, diarylamino, and alkylarylamino),
acylamino (including alkylcarbonylamino, arylcarbonylamino,
carbamoyl and ureido), amidino, imino, sulfhydryl, alkylthio,
arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato,
sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido,
heterocyclyl, alkylaryl, or an aromatic or heteroaromatic
moiety.
[0055] The term "amine" or "amino" should be understood as being
broadly applied to both a molecule, or a moiety or functional
group, as generally understood in the art, and may be primary,
secondary, or tertiary. The term "amine" or "amino" includes
compounds where a nitrogen atom is covalently bonded to at least
one carbon, hydrogen or heteroatom. The terms include, for example,
but are not limited to, "alkyl amino," "arylamino," "diarylamino,"
"alkylarylamino," "alkylaminoaryl," "arylaminoalkyl,"
"alkaminoalkyl," "amide," "amido," and "aminocarbonyl." The term
"alkyl amino" comprises groups and compounds wherein the nitrogen
is bound to at least one additional alkyl group. The term "dialkyl
amino" includes groups wherein the nitrogen atom is bound to at
least two additional alkyl groups. The term "arylamino" and
"diarylamino" include groups wherein the nitrogen is bound to at
least one or two aryl groups, respectively. The term
"alkylarylamino," "alkylaminoaryl" or "arylaminoalkyl" refers to an
amino group which is bound to at least one alkyl group and at least
one aryl group. The term "alkaminoalkyl" refers to an alkyl,
alkenyl, or alkynyl group bound to a nitrogen atom which is also
bound to an alkyl group.
[0056] The term "amide," "amido" or "aminocarbonyl" includes
compounds or moieties which contain a nitrogen atom which is bound
to the carbon of a carbonyl or a thiocarbonyl group. The term
includes "alkaminocarbonyl" or "alkylaminocarbonyl" groups which
include alkyl, alkenyl, aryl or alkynyl groups bound to an amino
group bound to a carbonyl group. It includes arylaminocarbonyl and
arylcarbonylamino groups which include aryl or heteroaryl moieties
bound to an amino group which is bound to the carbon of a carbonyl
or thiocarbonyl group. The terms "alkylaminocarbonyl,"
"alkenylaminocarbonyl," "alkynylaminocarbonyl,"
"arylaminocarbonyl," "alkylcarbonylamino," "alkenylcarbonylamino,"
"alkynylcarbonylamino," and "arylcarbonylamino" are included in
term "amide." Amides also include urea groups (aminocarbonylamino)
and carbamates (oxycarbonylamino).
[0057] In a particular embodiment of the invention, the term
"amine" or "amino" refers to substituents fo the formulas
N(R.sup.8)R.sup.9, CH.sub.2N(R.sup.8)R.sup.9 and
CH(CH.sub.3)N(R.sup.8)R.sup.9, wherein R.sup.8 and R.sup.9 are
each, independently, selected from the group consisting of --H and
--(C.sub.1-4alkyl).sub.0-1G, wherein G is selected from the group
consisting of --COOH, --H, --PO.sub.3H, --SO.sub.3H, --Br, --Cl,
--F, --O--C.sub.1-4alkyl, --S--C.sub.1-4alkyl, aryl,
--C(O)OC.sub.1-C.sub.6-alkyl, --C(O)C.sub.1-4alkyl-COOH,
--C(O)C.sub.1-C.sub.4-alkyl and --C(O)-aryl;
[0058] or N(R.sup.8)R.sup.9 is pyrrolyl, tetrazolyl, pyrrolidinyl,
pyrrolidinyl-2-one, dimethylpyrrolyl, imidazolyl, morpholino or
##STR1##
[0059] The term "aryl" includes groups, including 5- and 6-membered
single-ring aromatic groups that may include from zero to four
heteroatoms, for example, phenyl, pyrrole, furan, thiophene,
thiazole, isothiaozole, imidazole, triazole, tetrazole, pyrazole,
oxazole, isoxazole, pyridine, pyrazine, pyridazine, and pyrimidine,
and the like. Furthermore, the term "aryl" includes multicyclic
aryl groups, e.g., tricyclic, bicyclic, e.g., naphthalene,
benzoxazole, benzodioxazole, benzothiazole, benzoimidazole,
benzothiophene, methylenedioxyphenyl, quinoline, isoquinoline,
anthryl, phenanthryl, napthridine, indole, benzofuran, purine,
benzofuran, deazapurine, or indolizine. Those aryl groups having
heteroatoms in the ring structure may also be referred to as "aryl
heterocycles", "heterocycles," "heteroaryls" or "heteroaromatics."
The aromatic ring can be substituted at one or more ring positions
with such substituents as described above, as for example, alkyl,
halogen, hydroxyl, alkoxy, alkylcarbonyloxy, arylcarbonyloxy,
alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl,
alkylaminoacarbonyl, aralkylaminocarbonyl, alkenylaminocarbonyl,
alkylcarbonyl, arylcarbonyl, aralkylcarbonyl, alkenylcarbonyl,
alkoxycarbonyl, aminocarbonyl, alkylthiocarbonyl, phosphate,
phosphonato, phosphinato, cyano, amino (including alkyl amino,
dialkylamino, arylamino, diarylamino, and alkylarylamino),
acylamino (including alkylcarbonylamino, arylcarbonylamino,
carbamoyl and ureido), amidino, imino, sulfhydryl, alkylthio,
arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato,
sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido,
heterocyclyl, alkylaryl, or an aromatic or heteroaromatic moiety.
Aryl groups can also be fused or bridged with alicyclic or
heterocyclic rings which are not aromatic so as to form a polycycle
(e.g., tetralin).
[0060] It will be noted that the structures of some of the
compounds of this invention include asymmetric carbon atoms. It is
to be understood accordingly that the isomers arising from such
asymmetry (e.g., all enantiomers and diastereomers) are included
within the scope of this invention. Such isomers can be obtained in
substantially pure form by classical separation techniques and by
stereochemically controlled synthesis. Furthermore, the structures
and other compounds and moieties discussed in this application also
include all tautomers thereof. Compounds described herein may be
obtained through art recognized synthesis strategies.
[0061] Additionally, the phrase "any combination thereof" implies
that any number of the listed functional groups and molecules may
be combined to create a larger molecular architecture. For example,
the terms "phenyl," "carbonyl" (or ".dbd.O"), "--O--," "--OH," and
C.sub.1-6 (i.e., --CH.sub.3 and --CH.sub.2CH.sub.2CH.sub.2--) can
be combined to form a 3-methoxy-4-propoxybenzoic acid substituent;
or the terms "acid" (i.e., --COOH) and C.sub.1-6 (i.e.,
--CH.sub.2CH.sub.3) can be combined to form an ethyl ester
substituent. It is to be understood that when combining functional
groups and molecules to create a larger molecular architecture,
hydrogens can be removed or added, as required to satisfy the
valence of each atom.
[0062] As used herein, the term "pharmaceutically acceptable salts"
refers to salts of the invention prepared from pharmaceutically
acceptable non-toxic acids, including inorganic acids and organic
acids. Suitable non-toxic acids include inorganic and organic acids
such as acetic, benzenesulfonic, benzoic, camphorsulfonic, citric,
ethenesulfonic, fumaric, gluconic, glutamic, hydrobromic,
hydrochloric, isethionic, lactic, maleic, malic, mandelic,
methanesulfonic, mucic, nitric, pamoic, pantothenic, phosphoric,
succinic, sulfuric, tartaric acid, p-toluenesulfonic and the like.
Particularly preferred salts are sodium, lysine and argentine salts
of the compounds of the invention.
[0063] As used herein, the term "neurotrophic factor" or
"neurotrophin" (also referred to herein as "NT") refers to members
of a family of proteins, usually in the form of dimers, which are
structurally homologous to NGF. The term includes the precursors
(pro-neurotrophins, e.g., pro-NGF) and the mature proteins which
include three surface 3-hairpin loops, a p-strand, an internal
reverse turn region, and N- and C-termini. Neurotrophins promote at
least one of the biological activities related to vertebrate neuron
survival, differentiation, and function, as determined using assays
described, for example, in US 2002/0169182A1 and Riopelle et al.,
Can J. of Phys. and Pharm. 60:707 (1982); Harrington et al. PNAS
USA 101(16):6226-6230, (2004)). Neurotrophic factors include, for
example, brain-derived neurotrophic factor (BDNF), NGF,
neurotrophin 3 (NT-3), neurotrophin 4/5 (NT-4/5), and neurotrophin
6 (NT-6) (R. M. Lindsay et al.: TINS, vol. 17, p. 182 (1994) and R.
M. Lindsay: Phil. Trans. R. Soc. Lond. B. vol. 351, p. 365-373
(1996)). In addition, ciliary neurotrophic factor (CNTF),
glia-derived neurotrophic factor (GDNF), glia growth factor (GGF2),
central nerve growth factor (AF-1), hepatocyte growth factor (HGF)
(A. Ebens et al., Neuron, vol. 17, p. 1157-1172 (1996)) can also be
considered as neurotrophic factors. Moreover, biotechnologically
engineered products of the above neurotrophic factors, which are
derived by a partial substitution, an addition, a deletion or a
removal by conventional genetic engineering techniques, are also
included within the scope of the neurotrophic factors of the
present invention as far as such product shows biological
activities of the naturally-occurred neurotrophic factors.
[0064] As used herein, the term "neurotrophin receptor" (also
referred to herein as "NTR") is meant to refer to a receptor which
binds a neurotrophin. In certain embodiments, the neurotrophin
receptor is a member of the tyrosine kinase family of receptors,
generally referred to as the "Trk" receptors or "Trks", which are
expressed on cellular surfaces. The Trk family includes, but is not
limited to, TrkA, TrkB, and TrkC. In a particular embodiment the
neurotrophin receptor is TrkA. In other embodiments, the
neurotrophin receptor is p75.sup.NTR, also called p75 or
low-affinity nerve growth factor receptor or common neurtotrophin
receptor. These receptors may be from any animal species that
expresses neurotrophin receptors (e.g. human, murine, rabbit,
porcine, equine, etc.), and include full length receptors, their
truncated and variant forms, such as those arising by alternate
splicing and/or insertion, and naturally-occurring allelic
variants, as well as functional derivatives of such receptors.
[0065] "Neurotrophin-mediated activity" is a biological activity
that is normally modulated (e.g., inhibited or promoted), either
directly or indirectly, in the presence of a neurotrophin.
Neurotrophin-mediated activities include, for example, neurotrophin
binding to the p75.sup.NTR receptor or neurotrophin binding to one
of the Trk receptors (e.g., TrkA), the ability to promote
neurotrophin receptor dimerization and/or phosphorylation, neuron
survival, neuron differentiation including neuron process formation
and neurite outgrowth, neurotransmission and biochemical changes
such as enzyme induction. A biological activity that is mediated by
a particular neurotrophin, e.g. NGF or pro-NGF, is referred to
herein by reference to that neurotrophin, e.g. NGF-mediated
activity. (It is noted that "NGF-mediated activity" also includes
"proNGF-mediated activity.") To determine the ability of a compound
to inhibit a neurotrophin-mediated activity, conventional in vitro
and in vivo assays can be used. For example, a receptor binding
assay, such as the assay described in US 2002/0169182 A1, can be
used to assess the extent to which a compound inhibits
neurotrophin/receptor binding. Inhibition of neurite survival and
outgrowth can be determined using the in vitro assay described by
Riopelle et al. in the Can. J. of Phys. and Pharm., 1982, 60: 707.
Other examples of in vitro and in vivo assays for use in
determining the ability of a compound to inhibit a
neurotrophin-mediated activity are described in the
"Exemplification of the Invention" section of the application.
[0066] "Neurotransmission," as used herein, is a process by which
small signaling molecules, termed neurotransmitters, are rapidly
passed in a regulated fashion from a neuron to another cell.
Typically, following depolarization associated with an incoming
action potential, a neurotransmitter is secreted from the
presynaptic neuronal terminal. The neurotransmitter then diffuses
across the synaptic cleft to act on specific receptors on the
postsynaptic cell, which is most often a neuron but can also be
another cell type (such as muscle fibers at the neuromuscular
junction). The action of neurotransmitters can either be
excitatory, depolarizing the postsynaptic cell, or inhibitory,
resulting in hyperpolarization. Neurotransmission can be rapidly
increased or decreased by neuromodulators, which typically act
either pre-synaptically or post-synaptically. The neurotrophin
family (notably NGF and BDNF) have been shown to have prominent
neuromodulatory effects on diverse neuronal types (Lohof et al,
Nature. 363(6427):350-3 (1993); Li et al. J Neurosci.
18(24):10231-40. (1998)). BDNF has also been shown to behave like a
neurotransmitter, acting directly on target cells to alter their
excitability by rapidly and directly gating ion certain ion
channels (Rose et al., Bioessays. 26(11):1185-94. (2004)).
[0067] There are several simple fashions in which neurotransmission
can be studied. The release of neurotransmitters from cultured
neurons can be directly quantified using HPLC, radiolabled
neurotransmitters or other methodologies. Neurotransmission can be
estimated by dyes such as FM 1-43, a fluorescent marker of synaptic
vesicle cycling. Moreover, neurotransmission between neurons can be
directly monitored using standard electrophysiological techniques,
as can any direct neurotransmitter-like effects of neurotrophins on
ion channel currents. These various methodologies have been used to
study the effects of neurotrophins, such as BDNF and NGF, on
neurotransmitter release and neurotransmission (Lohof et al.; Li et
al.; Rose et al.).
[0068] The term "contacting" as used herein refers to bringing a
compound of the invention and a target, e.g., NGF, p75.sup.NTR
and/or TrkA together in such a manner that the compound can affect
the activity of the target, either directly; i.e., by interacting
with the target itself, or indirectly; i.e., by interacting with
another target on which the catalytic activity of the target is
dependent. For example, a compound of the invention may effect the
activity of TrkA by contacting (e.g., binding to) TrkA directly, or
by contacting (e.g., binding to) p75.sup.NTR, which may effect the
activity of TrkA. Such "contacting"" can be accomplished "in
vitro," i.e., in a test tube, a petri dish or the like, or "in
vivo," i.e., administered to a subject such as a mouse, rat or
human. In a test tube, contacting may involve only a compound and a
target of interest or it may involve whole cells. Cells may also be
maintained or grown in cell culture dishes and contacted with a
compound in that environment. "Contacting" can refer to a compound
of the invention directly binding to a target, or being in the
vicinity of a target.
[0069] Examples of neurotrophin-mediated activities include, but
are not limited to, pain (e.g., inflammatory pain, acute pain,
chronic malignant pain, chronic nonmalignant pain and neuropathic
pain), inflammatory disorders, diseases and disorders of the
genitourinary and gastrointestinal systems, and neurological
disorders (e.g., neurodegenerative or neuropsychiatric
disorders).
[0070] "Pain" is defined as an unpleasant sensory and emotional
experience associated with actual or potential tissue damage, or
described in terms of such damage (International Association for
the Study of Pain--IASP). Pain is classified most often based on
duration (i.e., acute vs. chronic pain) and/or the underlying
pathophysiology (i.e., nociceptive vs. neuropathic pain).
[0071] Acute pain can be described as an unpleasant experience with
emotional and cognitive, as well as sensory, features that occur in
response to tissue trauma and disease and serves as a defensive
mechanism. Acute pain is usually accompanied by a pathology (e.g.,
trauma, surgery, labor, medical procedures, acute disease states)
and the pain resolves with healing of the underlying injury. Acute
pain is mainly nociceptive, but may also be neuropathic.
[0072] Chronic pain is pain that extends beyond the period of
healing, with levels of identified pathology that often are low and
insufficient to explain the presence, intensity and/or extent of
the pain (American Pain Society--APS). Unlike acute pain, chronic
pain serves no adaptive purpose. Chronic pain may be nociceptive,
neuropathic, or both and caused by injury (e.g., trauma or
surgery), malignant conditions, or a variety of chronic conditions
(e.g., arthritis, fibromyalgia and neuropathy). In some cases,
chronic pain exists de novo with no apparent cause.
[0073] "Nociceptive pain" is pain that results from damage to
tissues and organs. Nociceptive pain is caused by the ongoing
activation of pain receptors in either the surperficial or deep
tissues of the body. Nociceptive pain is further characterized as
"somatic pain", including "cutaneous pain" and "deep somatic pain",
and "visceral pain".
[0074] "Somatic pain" includes "cutaneous pain" and "deep somatic
pain." Cutaneous pain is caused by injury, diseases, disorders or
neoplasms of the skin, subcutaneous tissues and related organs.
Examples of conditions associated with cutaneous pain include, but
are not limited to, cuts, burns, infections, lacerations, as well
as traumatic injury and post-operative or surgical pain (e.g., at
the site of incision).
[0075] "Deep somatic pain" results from injuries, diseases,
disorders or neoplasms of the musculoskeletal tissues, including
ligaments, tendons, bones, blood vessels and connective tissues.
Examples of deep somatic pain or conditions associated with deep
somatic pain include, but are not limited to, sprains, broken
bones, arthralgia, vasculitis, myalgia and myofascial pain.
Arthralgia refers to pain caused by a joint that has been injured
(such as a contusion, break or dislocation) and/or inflamed (e.g.,
arthritis). Vaculitis refers to inflammation of blood vessels with
pain. Myalgia refers to pain originating from the muscles.
Myofascial pain refers to pain stemming from injury or inflammation
of the fascia and/or muscles.
[0076] "Visceral" pain is associated with injury, inflammation,
disease or neoplasms of the body organs and internal cavities,
including but not limited to, the circulatory system, respiratory
system, gastrointestinal system, genitourinary system, immune
system, as well as the ear, nose and throat. Visceral pain can also
be associated with infectious and parasitic diseases that affect
the body organs and tissues. Visceral pain is extremely difficult
to localize, and several injuries to visceral tissue exhibit
"referred" pain, where the sensation is localized to an area
completely unrelated to the site of injury. For example, myocardial
ischaemia (the loss of blood flow to a part of the heart muscle
tissue) is possibly the best known example of referred pain; the
sensation can occur in the upper chest as a restricted feeling, or
as an ache in the left shoulder, arm or even hand. Phantom limb
pain is the sensation of pain from a limb that one no longer has or
no longer gets physical signals from--an experience almost
universally reported by amputees and quadriplegics.
[0077] "Neuropathic pain" or "neurogenic pain" is pain initiated or
caused by a primary lesion, dysfunction or perturbation in the
nervous system. "Neuropathic pain" can occur as a result of trauma,
inflammation, disease or neoplasms of the peripheral nervous system
("peripheral neuropathic pain") and/or the central nervous system
("central pain"). For example, neuropathic pain can be caused by a
nerve or nerves that are irritated, trapped, pinched, severed or
inflamed (neuritis). There are many neuropathic pain syndromes,
such as diabetic neuropathy, trigeminal neuralgia, postherpetic
neuralgia ("shingles"), post-stroke pain, and complex regional pain
syndromes (also called reflex sympathetic dystrophy or "RSD" and
causalgia).
[0078] As used herein, the term "inflammatory disease or disorder"
includes diseases or disorders which are caused, at least in part,
or exacerbated by, inflammation, which is generally characterized
by increased blood flow, edema, activation of immune cells (e.g.,
proliferation, cytokine production, or enhanced phagocytosis),
heat, redness, swelling, pain and loss of function in the affected
tissue and organ. The cause of inflammation may be due to physical
damage, chemical substances, micro-organisms, tissue necrosis,
cancer or other agents. Inflammatory disorders include acute
inflammatory disorders, chronic inflammatory disorders, and
recurrent inflammatory disorders. Acute inflammatory disorders are
generally of relatively short duration, and last for from about a
few minutes to about one to two days, although they may last
several weeks. The main characteristics of acute inflammatory
disorders include increased blood flow, exudation of fluid and
plasma proteins (edema) and emigration of leukocytes, such as
neutrophils. Chronic inflammatory disorders, generally, are of
longer duration, e.g., weeks to months to years or longer, and are
associated histologically with the presence of lymphocytes and
macrophages and with proliferation of blood vessels and connective
tissue. Recurrent inflammatory disorders include disorders which
recur after a period of time or which have periodic episodes. Some
disorders may fall within one or more categories.
[0079] The terms "neurological disorder" and "neurodegenerative
disorder" refer to injuries, diseases and dysfunctions of the
nervous system, including the peripheral nervous system and central
nervous system. Neurological disorders and neurodegenerative
disorders include, but are not limited to, diseases and disorders
that are associated with neurotrophin-mediated biological activity.
Examples of neurological disorders include, but are not limited to,
Alzheimer's disease, epilepsy, cancer, neuromuscular diseases,
multiple sclerosis, amyotrophic lateral sclerosis, stroke, cerebral
ischemia, neuropathy (e.g., chemotherapy-induced neuropathy,
diabetic neuropathy), retinal pigment degeneration, Huntington's
chorea, and Parkinson's disease, and ataxia-telangiectasia.
[0080] As used herein, "neuropathy" is defined as a failure of the
nerves that carry information to and from the brain and spinal cord
resulting in one or more of pain, loss of sensation, and inability
to control muscles. In some cases, the failure of nerves that
control blood vessels, intestines, and other organs results in
abnormal blood pressure, digestion problems, and loss of other
basic body processes. Peripheral neuropathy may involve damage to a
single nerve or nerve group (mononeuropathy) or may affect multiple
nerves (polyneuropathy).
[0081] The term "treated," "treating" or "treatment" includes the
diminishment or alleviation of at least one symptom associated with
the pain, inflammatory disorder, neurological disorder,
genitourinary disorder or gastrointestinal disorder (e.g.,
associated with or caused by neurotrophin mediated activity) being
treated. In certain embodiments, the treatment comprises the
modulation of the interaction of a neurotrophin (e.g., monomer or
dimer) and its receptor by an NT/NTR modulating compound, for
example an NGF/NTR modulating compound, which would in turn
diminish or alleviate at least one symptom directly or indirectly
associated with or caused by the neurotrophin-mediated activity
being treated. For example, treatment can be diminishment of one or
several symptoms of a disorder or complete eradication of a
disorder.
[0082] As used herein, the phrase "therapeutically effective
amount" of the compound is the amount necessary or sufficient to
treat or prevent pain, an inflammatory disorder, a neurological
disorder, a gastrointestinal disorder or a genitourinary disorder,
(e.g., to prevent the various morphological and somatic symptoms of
a neurotrophin-mediated activity). In an example, an effective
amount of the compound is the amount sufficient to alleviate at
least one symptom of the disorder, e.g., pain, inflammation, a
neurological disorder, a gastrointestinal disorder or a
genitourinary disorder, in a subject.
[0083] The term "subject" is intended to include animals, which are
capable of suffering from or afflicted with a
neurotrophin-associated state or neurotrophin-associated disorder,
or any disorder involving, directly or indirectly, neurotrophin
signaling. In another embodiement, a subject is also intended to
include animals, which are capable of suffering from pain, an
inflammatory disorder, a neurological disorder, a respiratory
disorder, a gastrointestinal disorder or a genitourinary disorder.
Examples of subjects include mammals, e.g., humans, dogs, cows,
horses, pigs, sheep, goats, cats, mice, rabbits, rats, and
transgenic non-human animals. In certain embodiments, the subject
is a human, e.g., a human suffering from, at risk of suffering
from, or potentially capable of suffering from pain, inflammation,
a neurological disorder, a gastrointestinal disorder or a
genitourinary disorder (e.g. associated with
neurotrophin-associated activity).
[0084] The language "NT/NTR modulator" refers to compounds that
modulate, i.e., inhibit, promote or otherwise alter the interaction
of a neurotrophin with a neurotrophin receptor. For example,
"NGF/NTR modulator" refers to compounds that modulate, e.g.,
inhibit, promote, or otherwise alter, the interaction of NGF (or
proNGF) with p75.sup.NTR, TrkA, or p75.sup.NTR and TrkA. Examples
of NGF/NTR modulators include compounds of Formulas 16, 17, 18, 19,
20, 21 and 22, including salts thereof, e.g., a pharmaceutically
acceptable salt. Additional examples of NGF/NTR modulators include
compounds of Table 1, or derivatives and fragments thereof,
including salts thereof, e.g., a pharmaceutically acceptable salt.
Compounds of Formulas 16, 17, 18, 19, 20, 21 and 22, as well as the
compounds of Table 1, i.e., the NT/NTR modulators or NGF/NTR
modulators of the invention, are also referred to herein as
"compounds of the invention." In a particular embodiment, the
NGF/NTR modulators of the invention, including the compounds of
Table 1, can be used to treat a disease or disorder associated with
pain, inflammation, neurological disorders, respiratory disorders,
gastrointestinal disorders or genitourinary disorders in a subject
in need thereof. In another embodiment, the compounds of the
invention, including the compounds of Table 1, can be used to treat
an inflammatory disorder in a subject in need thereof.
Modulators of Neurotrophin/Neurotrophin Receptor Interaction
[0085] In one aspect, the present invention provides compounds
which modulate the interaction of a neurotrophin with a
neurotrophin receptor. In certain embodiments, the compounds
modulate the interaction of nerve growth factor (NGF) and/or a
precursor thereof with a neurotrophin receptor (NTR). In other
embodiments the compound modulates the interaction of NGF and/or a
precursor thereof with the p75.sup.NTR receptor. In still other
embodiments, the compound also modulates the interaction of NGF (or
proNGF) with the TrkA receptor. In further embodiments, the
compound modulates the interaction of NGF (or proNGF) with both the
p75.sup.NTR and TrkA receptor.
[0086] In another aspect, the compounds of the invention treat
pain, inflammatory disorders, neurological disorders, respiratory
disorders, gastrointestinal disorders or genitourinary disorders in
a subject in need thereof, comprising administering to the subject
an effective amount of a compound of the invention.
[0087] Without being bound by theory, it is believed that an
electronegative atom of the compound of the invention, e.g, an
NGF/NTR modulator, bears a full or partial negative charge under
physiological conditions and can, therefore, interact
electrostatically with the positively charged side chain of an NGF
lysine residue. Thus, there will be an interaction, such as, for
example, a hydrogen bond, an ion/ion interaction, an ion/dipole
interaction or a dipole/dipole interaction. The hydrophobic region
or moiety of the NGF/NTR modulator can interact with a hydrophobic
region of NGF via a hydrophobic interaction. Without being bound by
theory, it is believed that compounds of the invention can interact
with NGF in such a way as to interfere with, and thereby modulate,
the interaction of NGF and p75mm and/or TrkA. In one embodiment,
the compound of the invention is of the general Formula 16,
##STR2## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4 and R.sup.5 are
each, independently, selected from the group consisting of a
hydrogen atom, --N(H)--, alkyl, alkoxy, amino, halogen, hydroxyl,
acid, cyano, sulfonamide, aryl, heteroaryl, cycloalkyl,
heterocycloalkyl, amide, ester, ether, thioether, alkene, furan,
thiophene, thiazole, nitro, alkene, tetrazole, sulfone, urea,
thiourea, morpholine, piperidine, piperazine, azepane, and any
combination thereof; or a pharmaceutically acceptable salt
thereof.
[0088] In another embodiment of Formula 16, R.sup.1, R.sup.1a,
R.sup.2, R.sup.3, R.sup.4 and R.sup.5 are each, independently,
selected from the group consisting of hydrogen, C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, hydroxyl, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl;
[0089] wherein the alkyl, C.sub.1-6-alkoxy, amino, aryl,
heteroaryl, cycloalkyl, and heterocycloalkyl groups may be further
independently substituted one or more times with C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, hydroxyl, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl or azepanyl.
[0090] In another embodiment of Formula 16, R.sup.2, R.sup.3,
R.sup.4 and R.sup.5 are each, independently, selected from the
group consisting of hydrogen, C.sub.1-C.sub.6-alkyl, halogen,
NO.sub.2, COOH, and N(R.sup.8)R.sup.9, wherein R.sup.8 and R.sup.9
are each, independently, selected from the group consisting of --H
and --C.sub.1-4alkyl).sub.0-1G, wherein G is selected from the
group consisting of --COOH, --PO.sub.3H, --SO.sub.3H, --Br, --Cl,
--F, --OC.sub.1-4alkyl, --SC.sub.1-4alkyl, aryl,
C.sub.1-C.sub.6-alkyl, --C(O)OC.sub.1-C.sub.6-alkyl,
--C(O)C.sub.1-C.sub.4-alkyl and --C(O)-aryl.
[0091] In still another embodiment of Formula 16, R.sup.2, R.sup.3,
R.sup.4 and R.sup.5 are hydrogen, and R.sup.1 and R.sup.1a are
(CH.sub.2).sub.xR.sup.13, wherein R.sup.13 is selected from the
group consisting of hydrogen, C.sub.1-6-alkyl, amino,
C.sub.1-6-alkoxy, --OH, halogen, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperazinyl, piperidinyl and azepanyl; and x
is 0, 1, 2, 3 or 4.
[0092] In yet another embodiment of Formula 16, R.sup.13 is
selected from the group consisting of --COOH, imidazolyl,
--SO.sub.3H, --OSO.sub.3H, --OH, morpholino, piperazinyl,
--PO.sub.3H, --PO.sub.3C.sub.1-4alkyl and --NO.sub.2.
[0093] In another embodiment of Formula 16, R.sup.13 is selected
from the group consisting of --COOH and --COO.sup.-Na.sup.+. In
another embodiment of Formula 16, x is 3 and R.sup.13 is COOH. In
still another embodiment of Formula 16, R.sup.13 is
C(O)N(R.sup.8)R.sup.9 or N(R.sup.8)R.sup.9, wherein R.sup.8 and
R.sup.9 are each, independently, selected from the group consisting
of --H and --C.sub.1-4alkyl).sub.0-1G, wherein G is selected from
the group consisting of --COOH, --H, --PO.sub.3H, --SO.sub.3H,
--Br, --Cl, --F, --OC.sub.1-4alkyl, --SC.sub.1-4alkyl, aryl,
--C(O)OC.sub.1-C.sub.6-alkyl, --C(O)C.sub.1-4alkyl-COOH,
--C(O)C.sub.1-C.sub.4-alkyl and --C(O)-aryl; or N(R.sup.8)R.sup.9
is pyrrolyl, tetrazolyl, pyrrolidinyl, pyrrolidin-2-one,
dimethylpyrrolyl, imidazolyl, morpholino.
[0094] In another embodiment of Formula 16, R.sup.13 is NH.sub.2 or
N(CH.sub.3).sub.2. In still another embodiment of Formula 16,
R.sup.2, R.sup.3, R.sup.4 and R.sup.5 are hydrogen, and R.sup.1 and
R.sup.1a are: ##STR3## wherein
[0095] R.sup.6, R.sup.7 and R.sup.12 are each, independently,
selected from the group consisting of hydrogen, C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl.
[0096] In another embodiment of Formula 16, R.sup.6 and R.sup.7 are
each, independently, selected from the group consisting of
hydrogen, halogen, hydroxyl, trihalomethyl, cyano, nitro,
--N(H)C(O)C.sub.1-C.sub.4-alkyl, --NH.sub.2,
--N(H)C(O)C.sub.1-4alkyl, methoxy, morpholino, piperazinyl and
C.sub.1-C.sub.4-alkyl-ester; and R.sup.12 is hydrogen.
[0097] In another embodiment of Formula 16, R.sup.6 and R.sup.7 are
each, independently, selected from the group consisting of
hydrogen, SO.sub.3H, chloro, fluoro, --OH, --COOH, and
--COO.sup.-Na.sup.+; and R.sup.12 is hydrogen. In still another
embodiment of Formula 17, R.sup.6 and R.sup.12 are hydrogen, and
R.sup.7 is --COOH; or R.sup.6 is chloro, R.sup.12 is hydrogen, and
R.sup.7 is --COOH; or R.sup.6 and R.sup.12 are hydrogen, and
R.sup.7 is SO.sub.3H.
[0098] In another embodiment of Formula 16, R.sup.2, R.sup.3,
R.sup.4 and R.sup.5 are hydrogen, R.sup.1 is
(CH.sub.2).sub.xR.sup.13, wherein
[0099] R.sup.13 is selected from the group consisting of hydrogen,
C.sub.1-6-alkyl, amino, C.sub.1-6-alkoxy, --OH, halogen, acid,
cyano, C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperazinyl, piperidinyl and azepanyl; x is
0, 1, 2, 3 or 4; and
[0100] R.sup.1a is: ##STR4## wherein
[0101] R.sup.6, R.sup.7 and R.sup.12 are each, independently,
selected from the group consisting of hydrogen, C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl.
[0102] In another embodiment of Formula 16, R.sup.13 is
C(O)N(R.sup.8)R.sup.9 or N(R.sup.8)R.sup.9, wherein R.sup.8 and
R.sup.9 are each, independently, selected from the group consisting
of --H and --C.sub.1-4alkyl).sub.0-1G, wherein G is selected from
the group consisting of --COOH, --H, --PO.sub.3H, --SO.sub.3H,
--Br, --Cl, --F, --OC.sub.1-4alkyl, --SC.sub.1-4alkyl, aryl,
--C(O)OC.sub.1-C.sub.6-alkyl, --C(O)C.sub.1-4alkyl-COOH,
--C(O)C.sub.1-C.sub.4-alkyl and --C(O)-aryl; or N(R.sup.8)R.sup.9
is pyrrolyl, tetrazolyl, pyrrolidinyl, pyrrolidin-2-one,
dimethylpyrrolyl, imidazolyl, morpholino. In still another
embodiment, R.sup.13 is NH.sub.2 or N(CH.sub.3).sub.2.
[0103] In another embodiment of Formula 16, R.sup.2, R.sup.3,
R.sup.4 and R.sup.5 are hydrogen, R.sup.1 is
(CH.sub.2).sub.xR.sup.13, wherein R.sup.13 is selected from the
group consisting of --COOH, imidazolyl, --SO.sub.3H, --OSO.sub.3H,
--OH, morpholino, piperazinyl, --PO.sub.3H,
--PO.sub.3C.sub.1-4alkyl and --NO.sub.2;
[0104] x is 0, 1, 2, 3 or 4; and
[0105] R.sup.1a is ##STR5##
[0106] wherein R.sup.6 and R.sup.7 are each, independently,
selected from the group consisting of hydrogen, halogen, hydroxyl,
trihalomethyl, cyano, nitro, --N(H)C(O)C.sub.1-C.sub.4-alkyl,
--NH.sub.2, --N(H)C(O)C.sub.1-4alkyl, methoxy, morpholino,
piperazinyl and C.sub.1-C.sub.4-alkyl-ester; and
[0107] R.sup.12 is hydrogen.
[0108] In another embodiment of Formula 16, x is 3 and R.sup.13 is
COOH, R.sup.6and R.sup.12 are hydrogen, and R.sup.7 is --COOH; or
R.sup.6 is chloro, R.sup.12 is hydrogen, and R.sup.7 is --COOH; or
R.sup.6 and R.sup.12 are hydrogen, and R.sup.7 is SO.sub.3H.
[0109] In another embodiment, the compound of the invention is of
the general Formula 17, ##STR6## wherein R.sup.6, R.sup.7,
R.sup.12, R.sup.6a, R.sup.7a and R.sup.12a are each, independently,
selected from the group consisting of a hydrogen atom, --N(H)--,
alkyl, alkoxy, amino, halogen, hydroxyl, acid, cyano, sulfonamide,
aryl, heteroaryl, cycloalkyl, heterocycloalkyl, amide, ester,
ether, thioether, alkene, furan, thiophene, thiazole, nitro,
alkene, tetrazole, sulfone, urea, thiourea, morpholine, piperidine,
piperazine, azepane, and any combination thereof or a
pharmaceutically acceptable salt thereof.
[0110] In one embodiment of Formula 17, R.sup.6, R.sup.6a, R.sup.7
and R.sup.7a are each, independently, selected from the group
consisting of hydrogen, halogen, hydroxyl, trihalomethyl, cyano,
nitro, --N(H)C(O)C.sub.1-C.sub.4-alkyl, --NH.sub.2,
--C(O)N(H)C.sub.1-4alkyl, methoxy, morpholino, piperazinyl and
C.sub.1-C.sub.4-alkyl-ester; and R.sup.12 and R.sup.12a are
hydrogen.
[0111] In another embodiment of Formula 17, R.sup.6 and R.sup.7 are
each, independently, selected from the group consisting of
hydrogen, chloro, fluoro, --OH, --COOH, and --COO.sup.-Na.sup.+;
R.sup.12 is hydrogen; R.sup.6a is selected from the group
consisting of hydrogen, chloro, fluoro, --OH, --COOH, and
--COO.sup.-Na.sup.+; and R.sup.7a and R.sup.12a are hydrogen.
[0112] In another embodiment of Formula 17, R.sup.6 is COOH,
R.sup.7 is Cl, R.sup.6a is --H, NO.sub.2, OCH.sub.3, COOCH.sub.3 or
Cl, and R.sup.7a, R.sup.12 and R.sup.12a are hydrogen. In still
another embodiment of Formula 17, R.sup.6 and R.sup.6a are COOH,
and R.sup.7, R.sup.7a, R.sup.12 and R.sup.12a are hydrogen.
[0113] In one embodiment, the compound of the invention is of the
general Formula 18, ##STR7##
[0114] and pharmaceutically acceptable salts, enantiomers,
stereoisomers, rotamers, tautomers, diastereomers, or racemates
thereof,
[0115] wherein
[0116] R.sup.6a, R.sup.7a and R.sup.12a are each, independently,
selected from the group consisting of hydrogen, C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, hydroxyl, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, tetrazolyl, sulfone, urea, thiourea, morpholino,
piperidinyl, piperazinyl and azepanyl; and
[0117] R.sup.1 is (CH.sub.2).sub.xR.sup.13, wherein R.sup.13 is
selected from the group consisting of hydrogen, C.sub.1-6-alkyl,
amino, C.sub.1-6-alkoxy, -OH, halogen, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperazinyl, piperidinyl and azepanyl;
and
[0118] x is 0, 1, 2, 3 or 4.
[0119] In one embodiment of Formula 18, R.sup.13 is selected from
the group consisting of --COOH, imidazolyl, --SO.sub.3H,
--OSO.sub.3H, --OH, morpholino, piperazinyl, --PO.sub.3H,
--PO.sub.3C.sub.1-4alkyl and --NO.sub.2. In another embodiment of
Formula 18, x is 3 and R.sup.13 is COOH. In still another
embodiment of Formula 18, R.sup.13 is C(O)N(R.sup.8)R.sup.9 or
N(R.sup.8)R.sup.9, wherein R.sup.8 and R.sup.9 are each,
independently, selected from the group consisting of --H and
--(C.sub.1-4alkyl).sub.0-1G, wherein G is selected from the group
consisting of --COOH, --H, --PO.sub.3H, --SO.sub.3H, --Br, --Cl,
--F, --OC.sub.1-4alkyl, --SC.sub.1-4alkyl, aryl,
--C(O)OC.sub.1-C.sub.6-alkyl, --C(O)C.sub.1-4alkyl-COOH,
--C(O)C.sub.1-C.sub.4-alkyl and --C(O)-aryl; or N(R.sup.8)R.sup.9
is pyrrolyl, tetrazolyl, pyrrolidinyl, pyrrolidin-2-one,
dimethylpyrrolyl, imidazolyl, morpholino. In another embodiment of
Formula 18, R.sup.13 is NH.sub.2 or N(CH.sub.3).sub.2.
[0120] In one embodiment, the compound of the invention is of the
general Formula 19or20, ##STR8##
[0121] and pharmaceutically acceptable salts, enantiomers,
stereoisomers, rotamers, tautomers, diastereomers, or racemates
thereof;
[0122] wherein
[0123] R.sup.6a, R.sup.7a, R.sup.8a and R.sup.9a are each,
independently, selected from the group consisting of hydrogen,
C.sub.1-6-alkyl, C.sub.1-6-alkoxy, amino, halogen, hydroxyl, acid,
cyano, C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, tetrazolyl, sulfone, urea, thiourea, morpholino,
piperidinyl, piperazinyl and azepanyl.
[0124] In one embodiment of Formulas 19 or 20, R.sup.6a and
R.sup.8a are hydrogen, and R.sup.7a and R.sup.9a are COONa; or
R.sup.6a, R.sup.7a, R.sup.8a and R.sup.9a are hydrogen.
[0125] In one embodiment, the compound of the invention is of the
general Formula 21, ##STR9##
[0126] and pharmaceutically acceptable salts, enantiomers,
stereoisomers, rotamers, tautomers, diastereomers, or racemates
thereof;
[0127] wherein
[0128] R.sup.1, R.sup.8a, and R.sup.9a are each, independently,
selected from the group consisting of a hydrogen atom,
C.sub.1-6-alkyl, C.sub.1-6-alkoxy, amino, halogen, hydroxyl, acid,
cyano, C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl;
[0129] wherein the alkyl, C.sub.1-6-alkoxy, amino, aryl,
heteroaryl, cycloalkyl, and heterocycloalkyl groups may be further
independently substituted one or more times with C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, hydroxyl, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl or azepanyl.
[0130] In one embodiment of Formula 21, R.sup.1 is
(CH.sub.2).sub.xR.sup.13, wherein
[0131] R.sup.13 is selected from the group consisting of hydrogen,
C.sub.1-6-alkyl, amino, C.sub.1-6-alkoxy, --OH, halogen, acid,
cyano, C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperazinyl, piperidinyl and azepanyl;
and
[0132] x is 0, 1, 2, 3 or 4.
[0133] In another embodiment of Formula 21, R.sup.13 is
C(O)N(R.sup.8)R.sup.9 or N(R.sup.8)R.sup.9, wherein R.sup.8 and
R.sup.9 are each, independently, selected from the group consisting
of --H and --C.sub.1-4alkyl).sub.0-1G, wherein G is selected from
the group consisting of --COOH, --H, --PO.sub.3H, --SO.sub.3H,
--Br, --Cl, --F, --OC.sub.1-4alkyl, --SC.sub.1-4alkyl, aryl,
--C(O)OC.sub.1-C.sub.6-alkyl, --C(O)C.sub.1-4alkyl-COOH,
--C(O)C.sub.1-C.sub.4-alkyl and --C(O)-aryl;
[0134] or N(R.sup.8)R.sup.9 is pyrrolyl, tetrazolyl, pyrrolidinyl,
pyrrolidin-2-one, dimethylpyrrolyl, imidazolyl, morpholino.
[0135] In another embodiment of Formula 21, R.sup.13 is NH.sub.2 or
N(CH.sub.3).sub.2. In still another embodiment of Formula 21, x is
3, R.sup.13 is COOH, R.sup.8a is H and R.sup.9a is COOH. In yet
another embodiment of Formula 21, R.sup.1 is ##STR10## wherein
R.sup.6, R.sup.7 and R.sup.12 are each, independently, selected
from the group consisting of hydrogen, C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl.
[0136] In yet another embodiment of Formula 21, R.sup.6 and R.sup.7
are each, independently, selected from the group consisting of
hydrogen, halogen, hydroxyl, trihalomethyl, cyano, nitro,
--N(H)C(O)C.sub.1-C.sub.4-alkyl, --NH.sub.2,
--N(H)C(O)C.sub.1-4alkyl, methoxy, morpholino, piperazinyl and
C.sub.1-C.sub.4-alkyl-ester; and R.sup.12 is hydrogen. In still
another embodiment of Formula 21, R.sup.6 is COOH, R.sup.7 is Cl,
R.sup.12 is H, R.sup.8a is H and R.sup.9a is COOH.
[0137] In one embodiment, the compound of the invention is of the
general Formula 22, ##STR11##
[0138] and pharmaceutically acceptable salts, enantiomers,
stereoisomers, rotamers, tautomers, diastereomers, or racemates
thereof;
[0139] wherein
[0140] R.sup.1, R.sup.2, R.sup.3, R.sup.4 and R.sup.5 are each,
independently, selected from the group consisting of a hydrogen
atom, C.sub.1-6-alkyl, C.sub.1-6-alkoxy, amino, halogen, hydroxyl,
acid, cyano, C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl;
[0141] wherein the alkyl, C.sub.1-6-alkoxy, amino, aryl,
heteroaryl, cycloalkyl, and heterocycloalkyl groups may be further
independently substituted one or more times with C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, hydroxyl, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, O--C.sub.1-6-alkyl,
S--C.sub.1-6-alkyl, C.sub.1-6-alkene, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl or azepanyl.
[0142] In one embodiment of Formula 22, R.sup.2, R.sup.3, R.sup.4
and R.sup.5 are hydrogen, and each R.sup.1 is:
(CH.sub.2).sub.xR.sup.13, wherein
[0143] R.sup.13 is selected from the group consisting of hydrogen,
C.sub.1-6-alkyl, amino, C.sub.1-6-alkoxy, --OH, halogen, acid,
cyano, C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperazinyl, piperidinyl and azepanyl;
and
[0144] x is 0, 1, 2, 3 or 4.
[0145] In another embodiment of Formula 22, R.sup.13 is
C(O)N(R.sup.8)R.sup.9 or N(R.sup.8)R.sup.9, wherein R.sup.8 and
R.sup.9 are each, independently, selected from the group consisting
of --H and --(C.sub.1-4alkyl).sub.0-1G, wherein G is selected from
the group consisting of --COOH, --H, --PO.sub.3H, --SO.sub.3H,
--Br, --Cl, --F, --OC.sub.1-4alkyl, --SC.sub.1-4alkyl, aryl,
--C(O)OC.sub.1-C.sub.6-alkyl, --C(O)C.sub.1-4alkyl-COOH,
--C(O)C.sub.1-C.sub.4-alkyl and --C(O)-aryl;
[0146] or N(R.sup.8)R.sup.9 is pyrrolyl, tetrazolyl, pyrrolidinyl,
pyrrolidin-2-one, dimethylpyrrolyl, imidazolyl, morpholino.
[0147] In another embodiment of Formula 22, R.sup.13 is NH.sub.2 or
N(CH.sub.3).sub.2. In still another embodiment of Formula 22,
R.sup.13 is selected from the group consisting of --COOH,
imidazolyl, --SO.sub.3H, --OSO.sub.3H, --OH, morpholino,
piperazinyl, --PO.sub.3H, --PO.sub.3C.sub.1-4alkyl and --NO.sub.2.
In still another embodiment of Formula 22, R.sup.13 is selected
from the group consisting of --COOH and --COO.sup.-Na.sup.+. In yet
another embodiment of Formula 22, x is 3 and R.sup.13 is COOH.
[0148] In still another embodiment of Formula 22, R.sup.2, R.sup.3,
R.sup.4 and R.sup.5 are hydrogen, and each R.sup.1 is: ##STR12##
wherein
[0149] R.sup.6, R.sup.7 and R.sup.12 are each, independently,
selected from the group consisting of hydrogen, C.sub.1-6-alkyl,
C.sub.1-6-alkoxy, amino, halogen, acid, cyano,
C.sub.1-6-alkyl-sulfonamide, aryl, heteroaryl,
C.sub.3-6-cycloalkyl, C.sub.3-6-heterocycloalkyl,
C.sub.1-6-alkyl-amide, C.sub.1-6-alkyl-ester, furanyl, thiophenyl,
thiazolyl, nitro, C.sub.1-6-alkene, tetrazolyl, sulfone, urea,
thiourea, morpholino, piperidinyl, piperazinyl and azepanyl.
[0150] In yet another embodiment of Formula 22, R.sup.6 and R.sup.7
are each, independently, selected from the group consisting of
hydrogen, halogen, hydroxyl, trihalomethyl, cyano, nitro,
--N(H)C(O)C.sub.1-C.sub.4-alkyl, --NH.sub.2,
--N(H)C(O)C.sub.1-4alkyl, methoxy, morpholino, piperazinyl and
C.sub.1-C.sub.4-alkyl-ester; and R.sup.12 is hydrogen.
[0151] In another embodiment of Formula 22, R.sup.6 and R.sup.7 are
each, independently, selected from the group consisting of
hydrogen, chloro, fluoro, --OH, --COOH, and --COO.sup.-Na.sup.+;
and R.sup.12 is hydrogen. In yet another embodiment of Formula 22,
R.sup.6 and R.sup.12 are hydrogen, and R.sup.7 is --COOH; or
R.sup.6 is Cl, R.sup.12 is hydrogen, and R.sup.7 is COOH.
[0152] It is to be understood that all of the compounds of Formulas
16, 17, 18, 19, 20, 21 and 22 described above will further include
double bonds and/or hydrogens between adjacent atoms as required to
satisfy the valence of each atom. That is, double bonds and or
hydrogens are added to provide the following number of total bonds
to each of the following types of atoms: carbon: four bonds;
nitrogen: three bonds; oxygen: two bonds; and sulfur: two-six
bonds.
[0153] In a particular embodiment of the invention, the NGF/NTR
modulator of Formulas 16, 17, 18, 19, 20, 21 and 22 are any one of
the compounds of Table 1 or derivatives and fragments thereof,
including salts thereof, e.g., pharmaceutically acceptable
salts.
[0154] In another embodiment, the invention pertains to the NGF/NTR
modulators of Formulas 16, 17, 18, 19, 20, 21 and 22, including
salts thereof, e.g., pharmaceutically acceptable salts. Particular
embodiments of the invention pertain to the modulating compounds of
Table 1 or derivatives thereof, including salts thereof, e.g.,
pharmaceutically acceptable salts.
[0155] In yet another embodiment, the invention pertains to
pharmaceutical compositions comprising NT/NTR modulating compounds
described herein and a pharmaceutical acceptable carrier.
[0156] In another embodiment, the invention includes any novel
compound or pharmaceutical compositions containing compounds of the
invention described herein. For example, compounds and
pharmaceutical compositions containing compounds set forth herein
(e.g., Table 1) are part of this invention, including salts
thereof, e.g., pharmaceutically acceptable salts.
[0157] The method for acquiring the PC12 data described in Table 1
is described herein in Example 1. Each experiment was done at least
once with an n value of 4 for each data point. (n/a=not acquired)
TABLE-US-00001 TABLE 1 PC12 (TrkA & p75) Single point (% max
binding) Concentration Abfix IC50 Compound Name Structure (.mu.M)
Mean Mean 4-[7-(3-Carboxy-propyl)-1,3,6,8-
tetraoxo-3,6,7,8-tetrahydro-1H- benzo[lmn][3,8]phenanthrolin-2-
yl]-butyric acid (12) ##STR13## n/a n/a ***
3-[7-(3-Carboxy-phenyl)- 1,3,6,8-tetraoxo-3,6,7,8- tetrahydro-1H-
benzo[lmn][3,8]phenanthrolin-2- yl]-4-benzoic acid (17) ##STR14##
n/a n/a *** 3-[7-(1-chloro-4-carboxy-
phenyl)-1,3,6,8-tetraoxo-3,6,7,8- tetrahydro-1H-
benzo[lmn][3,8]phenanthrolin-2- yl]-4-chloro-benzoic acid (18)
##STR15## 50 *** *** 3-(1,3,6,8-Tetraoxo-1,3,6,8-
tetrahydro-2-oxa-7-aza-pyren-7- yl)-benzoic acid (45) ##STR16## n/a
n/a * 4-(1,3,6,8-Tetraoxo-7-(4- benzenesulfonic acid)-3,6,7,8-
tetrahydro-1H- benzo[lmn][3,8]phenanthrolin-2- yl)-benzenesulfonic
acid (54) ##STR17## 50 *** n/a 3-(1,3,6,8-Tetraoxo-7-(3-
benzenesulfonic acid)-3,6,7,8- tetrahydro-1H-
benzo[lmn][3,8]phenanthrolin-2- yl)-benzenesulfonic acid (55)
##STR18## 50 **/*** * 4-Chloro-3-(1,3,6,8-tetraoxo-
1,3,6,8-tetrahydro-2-oxa-7-aza- pyren-7-yl)-benzoic acid (58)
##STR19## 50 *** *** 3-[7-(3-Carboxy-propyl)-1,3,6,8-
tetraoxo-3,6,7,8-tetrahydro-1H- benzo[lmn][3,8]phenanthrolin-2-
yl]-4-chloro-benzoic acid (59) ##STR20## 50 *** ***
2-(5-Carboxy-2-chloro-phenyl)- 1,3,6-trioxo-1,2,3,6-tetrahydro-
benzo[lmn]benzo[4,5]imidazo[2,1- b][3,8]phenanthroline-9-
carboxylic acid (90) ##STR21## 0.2 *** ***
1,3,6-trioxo-1,2,3,6-tetrahydro- benzo[lmn]benzo[4,5]imidazo[2,1-
b][3,8]diphenanthroline-9- carboxylic acid (91) (mixture of
isomers) ##STR22## n/a n/a *** 1,3,6-trioxo-1,2,3,6-tetrahydro-
benzo[lmn]benzo[4,5]imidazo[2,1- b][3,8]diphenanthroline potassium
salt (92) ##STR23## 50 * n/a 2-(3-Carboxy-phenyl)-1,3,6-
trioxo-1,2,3,6-tetrahydro- benzo[lmn]benzo[4,5]imidazo[2,1-
b][3,8]phenanthroline-9- carboxylic acid (93) ##STR24## 50 ** ***
4-(1,3,6,8-Tetraoxo-3,6,7,8- tetrahydro-1H-2-oxa-pyren-7-
yl)-butyric acid (94) ##STR25## 25 *** ***
2-(3-Carboxy-propyl)-1,3,6- trioxo-1,2,3,6-tetrahydro-
benzo[lmn]benzo[4,5]imidazo[2,1- b][3,8]phenanthroline-9-
carboxylic acid (95) ##STR26## 25 * n/a Mixture of two isomers (96)
##STR27## n/a n/a *** Mixture of two isomers (97) ##STR28## n/a n/a
*** 2-(5-Carboxy-2-chloro-phenyl)- 1,3,6-trioxo-1,2,3,6-tetrahydro-
benzo[lmn]benzo[4,5]imidazo[2,1- b][3,8]phenanthroline-9 (98)
##STR29## 25 *** *** 2-(5-Carboxy-2-chloro-phenyl)-
1,3,6-trioxo-1,2,3,6-tetrahydro- 9-chloro-
benzo[lmn]benzo[4,5]imidazo[2,1- b][3,8]phenanthroline (99)
##STR30## 10 *** *** 2-(5-Carboxy-2-chloro-phenyl)-
1,3,6-trioxo-1,2,3,6-tetrahydro- 9-nitro-
benzo[lmn]benzo[4,5]imidazo[2,1- b][3,8]phenanthroline (100)
##STR31## 10 *** *** 2-(5-Carboxy-2-chloro-phenyl)-
1,3,6-trioxo-1,2,3,6-tetrahydro- 9-methoxy-
benzo[lmn]benzo[4,5]imidazo[2,1- b][3,8]phenanthroline (101)
##STR32## 10 ** *** Methyl-2-(5-Carboxy-2-chloro-
phenyl)-1,3,6-trioxo-1,2,3,6- tetrahydro-
benzo[lmn]benzo[4,5]imidazo[2,1- b][3,8]phenanthroline-9-
carboxylic acid ester (102) ##STR33## 10 * *** % max binding Key
IC.sub.50 Key 0 > *** > 30 10 0 > *** > 10 30 > **
> 50 10 > ** > 20 * > 50 * > 20
[0158] In one embodiment of the invention, of
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]b-
enzo[4,5]imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid
(Compound 90) is used to treat pain in a subject.
[0159] In another embodiment of the invention,
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]b-
enzo[4,5]imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid
(Compound 90) is used to treat inflammation in a subject.
[0160] In one embodiment of the invention, of
3-[7-(1-chloro-4-carboxy-phenyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-b-
enzo[lmn][3,8]phenanthrolin-2-yl]-4-chloro-benzoic acid (Compound
18) is used to treat pain in a subject.
[0161] In another embodiment of the invention,
3-[7-(1-chloro-4-carboxy-phenyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-b-
enzo[lmn][3,8]phenanthrolin-2-yl]-4-chloro-benzoic acid (Compound
18) is used to treat inflammation in a subject.
[0162] In one embodiment of the invention, the modulating compounds
of the invention are capable of chemically interacting with NGF,
p75.sup.NTR, and/or TrkA. The language "chemical interaction" is
intended to include, but is not limited to, reversible interactions
such as hydrophobic/hydrophilic, ionic (e.g., coulombic
attraction/repulsion, ion-dipole, charge-transfer), covalent
bonding, Van der Waals, and hydrogen bonding. In certain
embodiments, the chemical interaction is a reversible Michael
addition. In a specific embodiment, the Michael addition involves,
at least in part, the formation of a covalent bond.
[0163] Compounds of the invention can be synthesized according to
standard organic synthesis procedures that are known in the art.
Synthesis procedures for compounds of the invention are described
herein in the Exemplification section.
[0164] Acid addition salts of the compounds of the invention are
most suitably formed from pharmaceutically acceptable acids, and
include for example those formed with inorganic acids, e.g.,
hydrochloric, sulphuric or phosphoric acids and organic acids e.g.
succinic, maleic, acetic or fumaric acid. Other
non-pharmaceutically acceptable salts, e.g., oxalates, may be used
for example in the isolation of the invention, and the compounds of
Table 1 for laboratory use, or for subsequent conversion to a
pharmaceutically acceptable acid addition salt. Also included
within the scope of the invention are solvates and hydrates of the
invention.
[0165] The conversion of a given compound salt to a desired
compound salt is achieved by applying standard techniques, in which
an aqueous solution of the given salt is treated with a solution of
base, e.g., sodium carbonate or potassium hydroxide, to liberate
the free base which is then extracted into an appropriate solvent,
such as ether. The free base is then separated from the aqueous
portion, dried, and treated with the requisite acid to give the
desired salt. Particularly preferred salts are sodium, lysine and
argentine salts of the compounds of the invention.
[0166] In vivo hydrolyzable esters or amides of certain compounds
of the invention can be formed by treating those compounds having a
free hydroxy or amino functionality with the acid chloride of the
desired ester in the presence of a base in an inert solvent such as
methylene chloride or chloroform. Suitable bases include
triethylamine or pyridine. Conversely, compounds of the invention,
and the compounds of Table 1 having a free carboxy group may be
esterified using standard conditions which may include activation
followed by treatment with the desired alcohol in the presence of a
suitable base.
Assays
[0167] The present invention also relates to a method of modulating
the interaction of NGF (or proNGF) with a neurotrophin receptor,
e.g, p75.sup.NTR and/or TrkA. In certain embodiments, the method
comprises contacting NGF and/or a precursor thereof (proNGF) in the
presence of p75.sup.NTR and/or TrkA with a NGF/NTR modulating
amount of a NGF/NTR modulator compound (i.e., a compound of the
invention), thereby modulating the interaction of NGF (and/or
proNGF) with p75.sup.NTR and/or TrkA.
[0168] The methods of the invention can be practiced in vitro, for
example, in a cell culture screening assay to screen compounds
which potentially modulate, directly or indirectly, receptor
function. In such a method, the modulating compound can function by
interacting with and eliminating any function or activity (e.g.,
receptor binding) of NGF and/or a precursor thereof in the sample
or culture. The modulating compounds can also be used to control
NGF activity in neuronal cell culture. In vitro cross-linking
assays for determining the ability of a compound within the scope
of the invention to modulate the interaction of NGF with
p75.sup.NTR and/or TrkA, are well known in the art and described in
the examples herein. Other assays for determining the ability of a
compound to modulate the activity of NGF with its respective
receptors are also readily available to the skilled artisan (see,
Barker et al., Neuron 13(1): 203-215; (1994), Dehant et al.,
Development 119: 545-558 (1993); and US 2002/016982).
[0169] Recombinant and native neurotrophin polypeptides from
different species, including humans, are commercially available
from several sources (e.g., Promega Corporation and R&D
Systems). In addition, neurotrophin polypeptides for use in the
assays described herein can be readily produced by standard
biological techniques or by chemical synthesis. For example, a host
cell transfected with an expression vector containing a nucleotide
sequence encoding the desired neurotrophin can be cultured under
appropriate conditions to allow expression of the peptide to occur.
The secreted peptide can then be isolated according to standard
techniques. Coding polynucleotides, precursors and promoters for a
number of neurotrophins are known, including coding sequences for
neurotrophins of some mammalian species. For example, GenBank
M61176 sets for the coding sequence for BDNF (see also, XM.006027);
BDNF precursor is set forth at BF439589; and a BDNF specific
promoter is set forth at Eo5933. A similar range of coding
sequences for other neurotrophins, including proNGF and mature NGF
(e.g., NCBI ACCESSION NO P01138 and CAA37703), NT-4/5 and NT-3, are
also available through GenBank and other publicly accessible
nucleotide and amino acid sequence databases. Alternatively, the
neurotrophin, e.g., NGF can be obtained by culturing a primary cell
culture or an established cell line that can produce the
neurotrophin, and isolating from the culture broth thereof (e.g.,
culture supernatant, cultured cells).
[0170] The method can also be practiced in vivo, for example, to
modulate one or more processes mediated by the interaction of NGF
(and/or proNGF) with p75.sup.NTR, and/or the interaction of NGF
with TrkA. Animal models for determining the ability of a compound
of the invention to treat a disorder associated with or caused by a
neurotrophin-mediated biological activity (e.g, pain, inflammatory
disorders, respiratory disorders, neurological disorders,
genitourinary disorders and gastrointestinal disorders) are well
known and readily available to the skilled artisan.
[0171] For example, animal models of neuropathic pain based on
injury inflicted to a nerve (mostly the sciatic nerve) are
described in Zeltser et al., 2000, Pain 89:19-24; Bennett et al.,
1988, Pain 33:87-107; Seltzer et al., 1990, Pain 43:205-218; Kim et
al., 1992, Pain 50:355-363; Decosterd et al., 2000, Pain 87:149-158
and DeLeo et al., 1994, Pain 56:9-16. There are also models of
diabetic neuropathy (STZ induced diabetic neuropathy--Courteix et
al., 1994, Pain 57:153-160) and drug induced neuropathies
(vincristine induced neuropathy--Aley et al., 1996, Neuroscience
73: 259-265; oncology-related immunotherapy, anti-GD2
antibodies--Slart et al., 1997, Pain 60:119-125). Acute pain in
humans can be reproduced using in murine animals chemical
stimulation: Martinez et al., Pain 81: 179-186; 1999 (the writhing
test--intraperitoneal acetic acid in mice), Coderre et al., Pain.
1993, 54:43-50 (intraplantar injection of formalin). Other types of
acute pain models are described in Whiteside et al., 2004, Br J
Pharmacol 141:85-91 (the incisional model, a post-surgery model of
pain) and Johanek and Simone, 2004, Pain 109:432-442 (a heat injury
model). An animal model of inflammatory pain using complete
Freund's adjuvant (intraplantar injection) is described in Jasmin
et al., 1998, Pain 75: 367-382. Intracapsular injection of irritant
agents (complete Freund's adjuvant, iodoacetate, capsaicine, urate
crystals, etc.) is used to develop arthritis models in animals
(Fernihough et al., 2004, Pain 112:83-93; Coderre and Wall, 1987,
Pain 28:379-393; Otsuki et al., 1986, Brain Res. 365:235-240). A
stress-induced hyperalgesia model is described in Quintero et al.,
2000, Pharmacology, Biochemistry and Behavior 67:449-458. Further
animal models for pain are considered in an article of Walker et
al. 1999 Molecular Medicine Today 5:319-321, comparing models for
different types of pain, which are acute pain, chronic/inflammatory
pain and chronic/neuropathic pain, on the basis of behavioral
signs. Animal models for depression are described by E.
Tatarczynska et al., Br. J. Pharmacol. 132(7): 1423-1430 (2001) and
P. J. M. Will et al., Trends in Pharmacological Sciences
22(7):331-37 (2001)); models for anxiety are described by D. Treit,
"Animal Models for the Study of Anti-anxiety Agents: A Review,"
Neuroscience & Biobehavioral Reviews 9(2):203-222 (1985).
Additional animal models for pain are also described herein in the
Exemplification section.
[0172] Genitourinary models include methods for reducing the
bladder capacity of test animals by infusing either protamine
sulfate and potassium chloride (See, Chuang, Y. C. et al., Urology
61(3): 664-670, 2003) into the bladder. These methods also include
the use of a well accepted model of for urinary tract disorders
involving the bladder using intravesically administered acetic acid
as described in Sasaki et al. (2002) J. Urol. 168: 1259-64.
Efficacy for treating spinal cord injured patients can be tested
using methods as described in Yoshiyama et al. (1999) Urology 54:
929-33.
[0173] Gastrointestinal models can be found in: Gawad, K. A., et
al., Ambulatory long-term pH monitoring in pigs, Surg Endosc,
(2003); Johnson, S. E. et al., Esophageal Acid Clearance Test in
Healthy Dogs, Can. J. Vet. Res. 53(2): 244-7 (1989); and Cicente,
Y. et al., Esophageal Acid Clearance: More Volume-dependent Than
Motility Dependent in Healthy Piglets, J. Pediatr. Gastroenterol.
Nutr. 35(2): 173-9 (2002). Models for a variety of assays can be
used to assess visceromotor and pain responses to rectal
distension. See, for example, Gunter et al., Physiol. Behav.,
69(3): 379-82 (2000), Depoortere et al., J. Pharmacol. and Exp.
Ther., 294(3): 983-990 (2000), Morteau et al., Fund. Clin.
Pharmacol., 8(6): 553-62 (1994), Gibson et al., Gastroenterology
(Suppl. 1), 120(5): A19-A20 (2001) and Gschossmann et al., Eur. J.
Gastro. Hepat., 14(10): 1067-72 (2002) the entire contents of which
are each incorporated herein by reference.
[0174] Gastrointestinal motility can be assessed based on either
the in vivo recording of mechanical or electrical events associated
intestinal muscle contractions in whole animals or the activity of
isolated gastrointestinal intestinal muscle preparations recorded
in vitro in organ baths (see, for example, Yaun et al., Br. J.
Pharmacol., 112(4):1095-1100 (1994), Jin et al., J. Pharm. Exp.
Ther., 288(1): 93-97 (1999) and Venkova et al., J. Pharm. Exp.
Ther., 300(3): 1046-1052 (2002)). Tatersall et al. and Bountra et
al., European Journal of Pharmacology, 250: (1993) R5 and
249:(1993) R3-R4 and Milano et al., J. Pharmacol. Exp. Ther.,
274(2): 951-961 (1995).
[0175] Animal models for investigating neurological disorders
include, but are not limited to, those described in Morris et al.,
(Learn. Motiv. 1981; 12: 239-60) and Abeliovitch et al., Cell 1993;
75: 1263-71). For example, neurological models for studying spinal
cord injury, are described in Yoshiyama, M. et al., Urology 54(5):
929-933 (1999).
[0176] Further examples of animal models for pain and inflammation
include, but are not limited to the models listed in Table 2.
TABLE-US-00002 TABLE 2 Modality Non-limiting examples of potential
Model Name tested Brief Description clinical indications
(Reference) ACUTE PHASIC PAIN Tail-flick Thermal Tip of tail of
rats is immersed if hot water and time Acute nociceptive pain to
withdrawal from water is measured. Alternatively, (Hardy et al. Am
J Physiol 1957; 189: 1-5.; a radiant heat source is applied to the
tail and time Ben-Bassat et al. Arch Intern to withdrawal is
determined. Analgesic effect is Pharmacodyn Ther 1959; 122:
434-47.) evidenced by a prolongation of the latency period
hot-plate Thermal Rats walk over a heated surface with increasing
Acute nociceptive pain temperature and observed for specific
nociceptive (Woolfe et al. J Pharmacol Exp Ther behavior such paw
licking, jumping. Time to 1944; 80: 300-7.) appearance of such
behavior is measured. Analgesic effects are evidenced by a
prolonged latency. Hargreaves Thermal A focused beam of light is
projected onto a small Acute nociceptive pain Test surface of the
hind leg of a rat with increasing (Yeomans et al. Pain 1994; 59:
85-94.) temperature. Time to withdrawal is measured. Analgesic
effect translates into a prolonged latency Pin Test or Mechanical
An increasing calibrated pressure is applied to the Acute
nociceptive pain Randall paw of rats with a blunt pin. Pressure
intensity is (Green et al. Br J Pharmacol 1951; 6: 572- Selitto
measured. Alternatively increased pressure is 85.; Randall et al.
Arch Int Pharmacodyn applied to the paw using a caliper until pain
Ther 1957; 111: 409-19) threshold is reached and animals withdraw
the paw. HYPERALGESIA MODELS/CHRONIC INFLAMMATORY PAIN MODELS
Hargreaves or Thermal A sensitizing agent (e.g, complete Freund's
Chronic pain associated with tissue Randal & and/or adjuvant
(CFA). carrageenin, turpentine etc.) is inflammation, e.g.
post-surgical pain, Selitto mechanical injected into the paw of
rats creating a local (Hargreaves et al. Pain 1988; 32: 77-
inflammation and sensitivities to mechanical 88.) (Randall &
Selitto) and/or therma (Hargreaves)l Randall L O, Selitto J J. Arch
Int stimulation are measured with comparison to the
Pharmacodyn1957; 3: 409-19. contralateral non-sensitized paw
Yeomans Thermal Rat hind paw in injected with capsaicin, a Chronic
pain associated with tissue model sensitizing agent for small
C-fibers or DMSO, a inflammation, e.g. post-surgical pain
sensitizing agent for A-delta fibers. A radiant heat is (Yeomans et
al. Pain 1994; 59: 85-94.; applied with different gradient to
differentially Otsuki et al. Brain Res 1986; 365: 235- stimulate
C-fibers or A-delta fibers and discriminate 240.) between the
effects mediated by both pathways CHRONIC MALIGNANT PAIN (CANCER
PAIN) Bone Cancer Thermal In this model, osteolytic mouse sarcoma
Bone cancer pain Model and/or NCTC2472 cells are used to induce
bone cancer by (Schwei et al., J. Neurosci. 1999; 19: mechanical
injecting tumor cells into the marrow space of the 10886-10897.)
femur bone and sealing the injection site Cancer Thermal Meth A
sarcoma cells are implanted around the Malignant neuropathic pain
invasion pain and/or sciatic nerve in BALB/c mice and these animals
(Shimoyama et al., Pain 2002; 99: 167- model (CIP) mechanical
develop signs of allodynia and thermal hyperalgesia 174.) as the
tumor grows, compressing the nerve. Spontaneous pain (paw lifting)
is also visible. CHRONIC MALIGNANT PAIN Muscle Pain Thermal
Repeated injections of acidic saline into one Fibromyalgia and/or
gastrocnemius muscle produces bilateral, long- (Sluka et al. Pain
2003; 106: 229-239.) mechanical lasting mechanical hypersensitivity
of the paw (i.e. hyperalgesia) without associated tissue damage
UV-irradiation Thermal Exposure of the rat hind paw to UV
irradiation Inflammatory pain associated with first- and/or
produces highly reliable and persistent allodynia. and
second-degree burns. mechanical Various irradiation periods with
UV-B produce skin (Perkins et al. Pain 1993; 53: 191-197.)
inflammation with different time courses CHRONIC NEUROPATHIC PAIN
Chronic Mostly Loose chronic ligature of the sciatic nerve. Thermal
Clinical Neuropathic pain: nerve Constriction mechanical or
mechanical sensitivities are tested using Von compression and
direct mechanical Injury (CCI) or but aso Frey hairs or the paw
withdrawal test (Hargreaves) neuronal damage might be relevant
Bennett and thermal clinical comparisons Xie model (Bennett &
Xie, Neuropharmacology 1984; 23: 1415-1418.) Chung's Mostly Tight
ligation of one of the two spinal nerves of the Same as above: root
compression model or mechanical sciatic nerve. Thermal or
mechanical sensitivities might be a relevant clinical comparison
Spinal Nerve but aso are tested using Von Frey hairs or the paw
(Kim and Chung, Pain 1990; 41: 235- Ligation thermal withdrawal
test (Hargreaves) 251.) model (SNL)
[0177] Accordingly, an agent identified as described herein (e.g.,
an NGF/NTR modulator) can be used in an animal model to determine
the efficacy, toxicity, or side effects of treatment with such an
agent.
[0178] Accordingly, this invention pertains to uses of novel agents
identified by the above-described screening assays for treatments
as described herein
Pharmaceutical Compositions
[0179] The present invention also provides pharmaceutical
compositions. Such compositions comprise a therapeutically (or
prophylactically) effective amount of a NGF/NTR modulator, and
preferably one or more compounds of the invention described above,
and a pharmaceutically acceptable carrier or excipient. Suitable
pharmaceutically acceptable carriers include, but are not limited
to, saline, buffered saline, dextrose, water, glycerol, ethanol,
and combinations thereof. The carrier and composition can be
sterile. The formulation should suit the mode of
administration.
[0180] The phrase "pharmaceutically acceptable carrier" is art
recognized and includes a pharmaceutically acceptable material,
composition or vehicle, suitable for administering compounds of the
present invention to mammals. The carriers include liquid or solid
filler, diluent, excipient, solvent or encapsulating material,
involved in carrying or transporting the subject agent from one
organ, or portion of the body, to another organ, or portion of the
body. Each carrier must be "acceptable" in the sense of being
compatible with the other ingredients of the formulation and not
injurious to the subject. Some examples of materials which can
serve as pharmaceutically acceptable carriers include: sugars, such
as lactose, glucose and sucrose; starches, such as corn starch and
potato starch; cellulose, and its derivatives, such as sodium
carboxymethyl cellulose, ethyl cellulose and cellulose acetate;
powdered tragacanth; malt; gelatin; talc; excipients, such as cocoa
butter and suppository waxes; oils, such as peanut oil, cottonseed
oil, safflower oil, sesame oil, olive oil, corn oil and soybean
oil; glycols, such as propylene glycol; polyols, such as glycerin,
sorbitol, mannitol and polyethylene glycol; esters, such as ethyl
oleate and ethyl laurate; agar; buffering agents, such as magnesium
hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water;
isotonic saline; Ringer's solution; ethyl alcohol; phosphate buffer
solutions; and other non-toxic compatible substances employed in
pharmaceutical formulations.
[0181] Wetting agents, emulsifiers and lubricants, such as sodium
lauryl sulfate and magnesium stearate, as well as coloring agents,
release agents, coating agents, sweetening, flavoring and perfuming
agents, preservatives and antioxidants can also be present in the
compositions.
[0182] Examples of pharmaceutically acceptable antioxidants
include: water soluble antioxidants, such as ascorbic acid,
cysteine hydrochloride, sodium bisulfate, sodium metabisulfite,
sodium sulfite and the like; oil-soluble antioxidants, such as
ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated
hydroxytoluene (BHT), lecithin, propyl gallate, .alpha.-tocopherol,
and the like; and metal chelating agents, such as citric acid,
ethylenediamine tetraacetic acid (EDTA), sorbitol, tartaric acid,
phosphoric acid, and the like.
[0183] Suitable pharmaceutically acceptable carriers include but
are not limited to water, salt solutions (e.g., NaCl), alcohols,
gum arabic, vegetable oils, benzyl alcohols, polyethylene glycols,
gelatin, carbohydrates such as lactose, amylose or starch,
cyclodextrin, magnesium stearate, talc, silicic acid, viscous
paraffin, perfume oil, fatty acid esters, hydroxymethylcellulose,
polyvinyl pyrolidone, etc. The pharmaceutical preparations can be
sterilized and if desired, mixed with auxiliary agents, e.g.,
lubricants, preservatives, stabilizers, wetting agents,
emulsifiers, salts for influencing osmotic pressure, buffers,
coloring, flavoring and/or aromatic substances and the like which
do not deleteriously react with the active compounds.
[0184] The composition, if desired, can also contain minor amounts
of wetting or emulsifying agents, or pH buffering agents. The
composition can be a liquid solution, suspension, emulsion, tablet,
pill, capsule, sustained release formulation, or powder. The
composition can be formulated as a suppository, with traditional
binders and carriers such as trilycerides. Oral formulation can
include standard carriers such as pharmaceutical grades of
mannitol, lactose, starch, magnesium stearate, polyvinyl
pyrollidone, sodium saccharine, cellulose, magnesium carbonate,
etc.
[0185] The composition can be formulated in accordance with the
routine procedures as a pharmaceutical composition adapted for
intravenous administration to human beings. Typically, compositions
for intravenous administration are solutions in sterile isotonic
aqueous buffer. Where necessary, the composition may also include a
solubilizing agent and a local anesthetic to ease pain at the site
of the injection. Generally, the ingredients are supplied either
separately or mixed together in unit dosage form, for example, as a
dry lyophilized powder or water free concentrate in a hermetically
sealed container such as an ampule or sachet indicating the
quantity of active agent. Where the composition is to be
administered by infusion, it can be dispensed with an infusion
bottle containing sterile pharmaceutical grade water, saline or
dextrose/water. Where the composition is administered by injection,
an ampule of sterile water for injection or saline can be provided
so that the ingredients may be mixed prior to administration.
[0186] The pharmaceutical compositions of the invention can also
include an agent which controls release of the compound of the
invention, thereby providing a timed or sustained relase
composition.
[0187] The present invention also relates to prodrugs of the
compounds disclosed herein, as well as pharmaceutical compositions
comprising such prodrugs. For example, compounds of the invention
which include acid functional groups or hydroxyl groups can also be
prepared and administered as a corresponding ester with a suitable
alcohol or acid. The ester can then be cleaved by endogenous
enzymes within the subject to produce the active agent.
[0188] Formulations of the present invention include those suitable
for oral, nasal, topical, transmucosal, transdermal, buccal,
sublingual, rectal, vaginal and/or parenteral administration. The
formulations may conveniently be presented in unit dosage form and
may be prepared by any methods well known in the art of pharmacy.
The amount of active ingredient that can be combined with a carrier
material to produce a single dosage form will generally be that
amount of the compound that produces a therapeutic effect.
Generally, out of one hundred per cent, this amount will range from
about 1 per cent to about ninety-nine percent of active ingredient,
preferably from about 5 per cent to about 70 per cent, most
preferably from about 10 per cent to about 30 per cent.
[0189] Methods of preparing these formulations or compositions
include the step of bringing into association a compound of the
present invention with the carrier and, optionally, one or more
accessory ingredients. In general, the formulations are prepared by
uniformly and intimately bringing into association a compound of
the present invention with liquid carriers, or finely divided solid
carriers, or both, and then, if necessary, shaping the product.
[0190] Formulations of the invention suitable for oral
administration may be in the form of capsules, cachets, pills,
tablets, lozenges (using a flavored basis, usually sucrose and
acacia or tragacanth), powders, granules, or as a solution or a
suspension in an aqueous or non-aqueous liquid, or as an
oil-in-water or water-in-oil liquid emulsion, or as an elixir or
syrup, or as pastilles (using an inert base, such as gelatin and
glycerin, or sucrose and acacia) and/or as mouth washes and the
like, each containing a predetermined amount of a compound of the
present invention as an active ingredient. A compound of the
present invention may also be administered as a bolus, electuary or
paste.
[0191] In solid dosage forms of the invention for oral
administration (capsules, tablets, pills, dragees, powders,
granules and the like), the active ingredient is mixed with one or
more pharmaceutically acceptable carriers, such as sodium citrate
or dicalcium phosphate, and/or any of the following: fillers or
extenders, such as starches, lactose, sucrose, glucose, mannitol,
and/or silicic acid; binders, such as, for example,
carboxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone,
sucrose and/or acacia; humectants, such as glycerol; disintegrating
agents, such as agar-agar, calcium carbonate, potato or tapioca
starch, alginic acid, certain silicates, and sodium carbonate;
solution retarding agents, such as paraffin; absorption
accelerators, such as quaternary ammonium compounds; wetting
agents, such as, for example, cetyl alcohol and glycerol
monostearate; absorbents, such as kaolin and bentonite clay;
lubricants, such a talc, calcium stearate, magnesium stearate,
solid polyethylene glycols, sodium lauryl sulfate, and mixtures
thereof; and coloring agents. In the case of capsules, tablets and
pills, the pharmaceutical compositions may also comprise buffering
agents. Solid compositions of a similar type may also be employed
as fillers in soft and hard-filled gelatin capsules using such
excipients as lactose or milk sugars, as well as high molecular
weight polyethylene glycols and the like.
[0192] A tablet may be made by compression or molding, optionally
with one or more accessory ingredients. Compressed tablets may be
prepared using binder (for example, gelatin or hydroxypropylmethyl
cellulose), lubricant, inert diluent, preservative, disintegrant
(for example, sodium starch glycolate or cross-linked sodium
carboxymethyl cellulose), surface-active or dispersing agent.
Molded tablets may be made by molding in a suitable machine a
mixture of the powdered compound moistened with an inert liquid
diluent.
[0193] The tablets, and other solid dosage forms of the
pharmaceutical compositions of the present invention, such as
dragees, capsules, pills and granules, may optionally be scored or
prepared with coatings and shells, such as enteric coatings and
other coatings well known in the pharmaceutical-formulating art.
They may also be formulated so as to provide slow or controlled
release of the active ingredient therein using, for example,
hydroxypropylmethyl cellulose in varying proportions to provide the
desired release profile, other polymer matrices, liposomes and/or
microspheres. They may be sterilized by, for example, filtration
through a bacteria-retaining filter, or by incorporating
sterilizing agents in the form of sterile solid compositions that
can be dissolved in sterile water, or some other sterile injectable
medium immediately before use. These compositions may also
optionally contain opacifying agents and may be of a composition
that they release the active ingredient(s) only, or preferentially,
in a certain portion of the gastrointestinal tract, optionally, in
a delayed manner. Examples of embedding compositions that can be
used include polymeric substances and waxes. The active ingredient
can also be in micro-encapsulated form, if appropriate, with one or
more of the above-described excipients.
[0194] Liquid dosage forms for oral administration of the compounds
of the invention include pharmaceutically acceptable emulsions,
microemulsions, solutions, suspensions, syrups and elixirs. In
addition to the active ingredient, the liquid dosage forms may
contain inert diluent commonly used in the art, such as, for
example, water or other solvents, solubilizing agents and
emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl
carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate,
propylene glycol, 1,3-butylene glycol, oils (in particular,
cottonseed, groundnut, corn, germ, olive, castor and sesame oils),
glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fatty
acid esters of sorbitan, and mixtures thereof.
[0195] Besides inert diluents, the oral compositions can also
include adjuvants such as wetting agents, emulsifying and
suspending agents, sweetening, flavoring, coloring, perfuming and
preservative agents.
[0196] Suspensions, in addition to the active compounds, may
contain suspending agents as, for example, ethoxylated isostearyl
alcohols, polyoxyethylene sorbitol and sorbitan esters,
microcrystalline cellulose, aluminum metahydroxide, bentonite,
agar-agar and tragacanth, and mixtures thereof.
[0197] Formulations of the pharmaceutical compositions of the
invention for rectal or vaginal administration may be presented as
a suppository, which may be prepared by mixing one or more
compounds of the invention with one or more suitable nonirritating
excipients or carriers comprising, for example, cocoa butter,
polyethylene glycol, a suppository wax or a salicylate, and which
is solid at room temperature, but liquid at body temperature and,
therefore, will melt in the rectum or vaginal cavity and release
the active compound.
[0198] Formulations of the present invention which are suitable for
vaginal administration also include pessaries, tampons, creams,
gels, pastes, foams or spray formulations containing such carriers
as are known in the art to be appropriate.
[0199] Dosage forms for the topical or transdermal administration
of a compound of this invention include powders, sprays, ointments,
pastes, creams, lotions, gels, solutions, patches and inhalants.
The active compound may be mixed under sterile conditions with a
pharmaceutically acceptable carrier, and with any preservatives,
buffers, or propellants that may be required.
[0200] The ointments, pastes, creams and gels may contain, in
addition to an active compound of this invention, excipients, such
as animal and vegetable fats, oils, waxes, paraffins, starch,
tragacanth, cellulose derivatives, polyethylene glycols, silicones,
bentonites, silicic acid, talc and zinc oxide, or mixtures
thereof.
[0201] Powders and sprays can contain, in addition to a compound of
this invention, excipients such as lactose, talc, silicic acid,
aluminum hydroxide, calcium silicates and polyamide powder, or
mixtures of these substances. Sprays can additionally contain
customary propellants, such as chlorofluorohydrocarbons and
volatile unsubstituted hydrocarbons, such as butane and
propane.
[0202] Transdermal patches have the added advantage of providing
controlled delivery of a compound of the present invention to the
body. Such dosage forms can be made by dissolving or dispersing the
compound in the proper medium. Absorption enhancers can also be
used to increase the flux of the compound across the skin. The rate
of such flux can be controlled by either providing a rate
controlling membrane or dispersing the active compound in a polymer
matrix or gel.
[0203] Ophthalmic formulations, eye ointments, powders, solutions
and the like, are also contemplated as being within the scope of
this invention.
[0204] Pharmaceutical compositions of this invention suitable for
parenteral administration comprise one or more compounds of the
invention in combination with one or more pharmaceutically
acceptable sterile isotonic aqueous or nonaqueous solutions,
dispersions, suspensions or emulsions, or sterile powders which may
be reconstituted into sterile injectable solutions or dispersions
just prior to use, which may contain antioxidants, buffers,
bacteriostats, solutes which render the formulation isotonic with
the blood of the intended recipient or suspending or thickening
agents.
[0205] Examples of suitable aqueous and nonaqueous carriers that
may be employed in the pharmaceutical compositions of the invention
include water, ethanol, polyols (such as glycerol, propylene
glycol, polyethylene glycol, and the like), and suitable mixtures
thereof, vegetable oils, such as olive oil, and injectable organic
esters, such as ethyl oleate. Proper fluidity can be maintained,
for example, by the use of coating materials, such as lecithin, by
the maintenance of the required particle size in the case of
dispersions, and by the use of surfactants.
[0206] These compositions may also contain adjuvants such as
preservatives, wetting agents, emulsifying agents and dispersing
agents. Prevention of the action of microorganisms may be ensured
by the inclusion of various antibacterial and antifungal agents,
for example, paraben, chlorobutanol, phenol sorbic acid, and the
like. It may also be desirable to include isotonic agents, such as
sugars, sodium chloride, and the like into the compositions. In
addition, prolonged absorption of the injectable pharmaceutical
form may be brought about by the inclusion of agents that delay
absorption such as aluminum monostearate and gelatin.
[0207] In some cases, in order to prolong the effect of a drug, it
is desirable to slow the absorption of the drug from subcutaneous
or intramuscular injection. This may be accomplished by the use of
a liquid suspension of crystalline or amorphous material having
poor water solubility. The rate of absorption of the drug then
depends upon its rate of dissolution which, in turn, may depend
upon crystal size and crystalline form. Alternatively, delayed
absorption of a parenterally-administered drug form is accomplished
by dissolving or suspending the drug in an oil vehicle.
[0208] Injectable depot forms are made by forming microencapsule
matrices of the subject compounds in biodegradable polymers such as
polylactide-polyglycolide. Depending on the ratio of drug to
polymer, and the nature of the particular polymer employed, the
rate of drug release can be controlled. Examples of other
biodegradable polymers include poly(orthoesters) and
poly(anhydrides). Depot injectable formulations are also prepared
by entrapping the drug in liposomes or microemulsions that are
compatible with body tissue.
Methods of Administration
[0209] The invention provides a method of treating a condition
mediated by an NGF/NTR interaction in a subject, including, but not
limited to, pain, inflammatory disorders, respiratory disorders,
neurological disorders, gastrointestinal disorders and
genitourinary disorders. The method comprises the step of
administering to the subject a therapeutically effective amount of
a NGF/NTR modulator. The condition to be treated can be any
condition which is mediated, at least in part, by the interaction
of a neurotrophin (e.g. NGF) with a neurotrophin receptor (e.g.,
p75.sup.NTR and TrkA).
[0210] The quantity of a given compound to be administered will be
determined on an individual basis and will be determined, at least
in part, by consideration of the individual's size, the severity of
symptoms to be treated and the result sought. The NGF/NTR
modulators described herein can be administered alone or in a
pharmaceutical composition comprising the modulator, an acceptable
carrier or diluent and, optionally, one or more additional
drugs.
[0211] These compounds may be administered to humans and other
animals for therapy by any suitable route of administration. The
NGF/NTR modulator can be administered subcutaneously,
intravenously, parenterally, intraperitoneally, intradermally,
intramuscularly, topically, enterally (e.g., orally), rectally,
nasally, buccally, sublingually, systemically, vaginally, by
inhalation spray, by drug pump or via an implanted reservoir in
dosage formulations containing conventional non-toxic,
physiologically acceptable carriers or vehicles. The preferred
method of administration is by oral delivery. The form in which it
is administered (e.g., syrup, elixir, capsule, tablet, solution,
foams, emulsion, gel, sol) will depend in part on the route by
which it is administered. For example, for mucosal (e.g., oral
mucosa, rectal mucosa, intestinal mucosa, bronchial mucosa)
administration, nose drops, aerosols, inhalants, nebulizers, eye
drops or suppositories can be used. The compounds and agents of
this invention can be administered together with other biologically
active agents, such as analgesics, e.g., opiates, anti-inflammatory
agents, e.g., NSAIDs, anesthetics and other agents which can
control one or more symptoms or causes of an NTR-mediated
condition.
[0212] In a specific embodiment, it may be desirable to administer
the agents of the invention locally to a localized area in need of
treatment; this may be achieved by, for example, and not by way of
limitation, local infusion during surgery, topical application,
transdermal patches, by injection, by means of a catheter, by means
of a suppository, or by means of an implant, said implant being of
a porous, non-porous, or gelatinous material, including membranes,
such as sialastic membranes or fibers. For example, the agent can
be injected into the joints or the urinary bladder.
[0213] The compounds of the invention can, optionally, be
administered in combination with one or more additional drugs
which, for example, are known for treating and/or alleviating
symptoms of the condition mediated by NGF p75.sup.NTR or TrkA. The
additional drug can be administered simultaneously with the
compound of the invention, or sequentially. For example, the
compounds of the invention can be administered in combination with
at least one of an analgesic, an anti-inflammatory agent, an
anesthetic, a corticosteroid (e.g., dexamethasone, beclomethasone
diproprionate (BDP) treatment), an anti-convulsant, an
antidepressant, an anti-nausea agent, an anti-psychotic agent, a
cardiovascular agent (e.g., a beta-blocker) or a cancer
therapeutic. In certain embodiments, the compounds of the invention
are administered in combination with a pain drug. As used herein
the phrase, "pain drugs" is intended to refer to analgesics,
anti-inflammatory agents, anesthetics, corticosteroids,
antiepileptics, barbiturates, antidepressants, and marijuana.
[0214] The combination treatments mentioned above can be started
prior to, concurrent with, or after the administration of the
compositions of the present invention. Accordingly, the methods of
the invention can further include the step of administering a
second treatment, such as a second treatment for the disease or
disorder or to ameliorate side effects of other treatments. Such
second treatment can include, e.g., anti-inflammatory medication
and any treatment directed toward treating pain. Additionally or
alternatively, further treatment can include administration of
drugs to further treat the disease or to treat a side effect of the
disease or other treatments (e.g., anti-nausea drugs,
anti-inflammatory drugs, anti-depressants, anti-psychiatric drugs,
anti-convulsants, steroids, cardiovascular drugs, and cancer
chemotherapeutics).
[0215] As used herein, an "analgesic" is an agent that relieves
pain without significant impairment of consciousness or sense
perception and may result in the reduction of inflammation as do
corticosteroids, e.g., an anti-inflammatory agent. Analgesics can
be subdivided into NSAIDs (non-steroidal-anti-inflammatory agents),
narcotic analgesics, and non-narcotic analgesics. NSAIDs can be
further subdivided into non-selective COX (cyclooxygenase)
inhibitors, and selective COX2 inhibitors. Opioid analgesics can be
natural, synthetic or semi-synthetic opioid (narcotic) analgesics,
and include for example, morphine, codeine, meperidine, propxyphen,
oxycodone, hydromorphone, heroine, tramadol, and fentanyl.
Non-opioid analgesics (non-narcotic) analgesics include, for
example, acetaminophen, clonidine, NMDA antagonists, and
cannabinoids. Non-selective COX inhibitors include, but are not
limited to acetylsalicylic acid (ASA), ibuprofen, naproxen,
ketoprofen, piroxicam, etodolac, and bromfenac. Selective COX2
inhibitors include, but are not limited to celecoxib, valdecoxib,
parecoxib, and etoricoxib.
[0216] As used herein an "anesthetic" is an agent that interferes
with sense perception near the site of administration, a local
anesthetic, or result in alteration or loss of consciousness, e.g.,
systemic anesthetic agents. Local anesthetics include but are not
limited to lidocaine and buvicaine.
[0217] Non-limiting examples of antiepileptic agents are
carbamazepine, phenytoin and gabapentin. Non-limiting examples of
antidepressants are amitriptyline and desmethylimiprimine.
[0218] Non-limiting examples of anti-inflammatory drugs include
corticosteroids (e.g., hydrocortisone, cortisone, prednisone,
prednisolone, methyl prednisone, triamcinolone, fluprednisolone,
betamethasone and dexamethasone), salicylates, antihistamines and
H.sub.2 receptor antagonists.
[0219] The phrases "parenteral administration" and "administered
parenterally" as used herein mean modes of administration other
than enteral and topical administration, usually by injection, and
include, without limitation, intravenous, intramuscular,
intraarterial, intrathecal, intracapsular, intraorbital,
intracardiac, intradermal, intraperitoneal, transtracheal,
subcutaneous, subcuticular, intraarticular, subcapsular,
subarachnoid, intraspinal and intrasternal injection and
infusion.
[0220] The phrases "systemic administration," "administered
systemically," "peripheral administration" and "administered
peripherally" as used herein mean the administration of a compound,
drug or other material other than directly into the central nervous
system, such that it enters the subject's system and, thus, is
subject to metabolism and other like processes, for example,
subcutaneous administration.
[0221] Regardless of the route of administration selected, the
compounds of the present invention, which may be used in a suitable
hydrated form, and/or the pharmaceutical compositions of the
present invention, are formulated into pharmaceutically acceptable
dosage forms by conventional methods known to those of skill in the
art.
[0222] Actual dosage levels of the active ingredients in the
pharmaceutical compositions of this invention may be varied so as
to obtain an amount of the active ingredient which is effective to
achieve the desired therapeutic response for a particular subject,
composition, and mode of administration, without being toxic to the
subject.
[0223] The selected dosage level will depend upon a variety of
factors including the activity of the particular compound of the
present invention employed, or the ester, salt or amide thereof,
the route of administration, the time of administration, the rate
of excretion of the particular compound being employed, the
duration of the treatment, other drugs, compounds and/or materials
used in combination with the particular compound employed, the age,
sex, weight, condition, general health and prior medical history of
the subject being treated, and like factors well known in the
medical arts.
[0224] A physician or veterinarian having ordinary skill in the art
can readily determine and prescribe the effective amount of the
pharmaceutical composition required. For example, dosages of a
compound of the invention may be determined by deriving
dose-response curves using an animal model for the condition to be
treated. For example, the physician or veterinarian could start
doses of the compounds of the invention employed in the
pharmaceutical composition at levels lower than that required in
order to achieve the desired therapeutic effect and gradually
increase the dosage until the desired effect is achieved.
[0225] In general, a suitable daily dose of a compound of the
invention will be that amount of the compound that is the lowest
dose effective to produce a therapeutic effect. Such an effective
dose will generally depend upon the factors described above.
Generally, intravenous and subcutaneous doses of the compounds of
this invention for a subject, when used for the indicated analgesic
effects, will range from about 0.0001 to about 100 mg per kilogram
of body weight per day, more preferably from about 0.01 to about
100 mg per kg per day, and still more preferably from about 1.0 to
about 50 mg per kg per day. An effective amount is that amount
treats a neurotrophin-associated state or neurotrophin
disorder.
[0226] If desired, the effective daily dose of the active compound
may be administered as two, three, four, five, six or more
sub-doses administered separately at appropriate intervals
throughout the day, optionally, in unit dosage forms.
[0227] While it is possible for a compound of the present invention
to be administered alone, it is preferable to administer the
compound as a pharmaceutical composition.
Methods of Treatment
[0228] The above compounds can be used for administration to a
subject for the modulation of a neurotrophin-mediated activity,
involved in, but not limited to, pain, inflammatory disorders,
neurological disorders, and any abnormal function of cells, organs,
or physiological systems that are modulated, at least in part,
directly or indirectly by a neurotrophin-mediated activity.
Additionally, it is understood that the compounds may also
alleviate or treat one or more additional symptoms of a disease or
disorder discussed herein.
[0229] Accordingly, in one aspect, the compounds of the invention
may be used to treat pain, including acute, chronic, malignant and
non-malignant somatic pain (including cutaneous pain and deep
somatic pain), visceral pain, and neuropathic pain. It is further
understood that the compounds may also alleviate or treat one or
more additional signs or symptoms of pain and sensory deficits
(e.g., hyperalgesia, allodynia, dysesthesia, hyperesthesia,
hyperpathia, paresthesia).
[0230] In some embodiments of this aspect of the invention, the
compounds of the invention may be used to treat somatic or
cutaneous pain associated with injuries, inflammation, diseases and
disorders of the skin, subcutaneous tissues and related organs
including, but not limited to, cuts, burns, lacerations, punctures,
incisions, surgical pain, post-operative pain, orodental surgery,
psoriasis, eczema, dermatitis, and allergies. The compounds of the
invention may also be used to treat somatic pain associated with
malignant and non-malignant neoplasm of the skin, subcutaneous
tissues and related organs (e.g., melanoma, basal cell
carcinoma).
[0231] In other embodiments of this aspect of the invention, the
compounds of the invention may be used to treat deep somatic pain
associated with injuries, inflammation, diseases and disorders of
the musculoskeletal and connective tissues including, but not
limited to, arthralgias, myalgias, fibromyalgias, myofascial pain
syndrome, dental pain, lower back pain, pain during labor and
delivery, surgical pain, post-operative pain, headaches, idiopathic
pain disorder, sprains, bone fractures, bone injury, osteoporosis,
severe burns, gout, arthiritis, osteoarthithis, myositis, and
dorsopathies (e.g., spondylolysis, subluxation, sciatica, and
torticollis). The compounds of the invention may also be used to
treat deep somatic pain associated with malignant and non-malignant
neoplasm of the musculoskeletal and connective tissues (e.g.,
sarcomas, rhabdomyosarcomas, and bone cancer).
[0232] In other embodiments of this aspect of the invention,
compounds of the invention may be used to treat visceral pain
associated with injuries, inflammation, diseases or disorders of
the circulatory system, the respiratory system, the genitourinary
system, the gastrointestinal system and the eye, ear, nose and
throat.
[0233] For example, the compounds of the invention may be used to
treat visceral pain associated with injuries, inflammation and
disorders of the circulatory system including, but are not limited
to, ischaemic diseases, ischaemic heart diseases (e.g., angina
pectoris, acute myocardial infarction, coronary thrombosis,
coronary insufficiency), diseases of the blood and lymphatic
vessels (e.g., peripheral vascular disease, intermittent
claudication, varicose veins, haemorrhoids, embolism or thrombosis
of the veins, phlebitis, thrombophlebitis lymphadenitis,
lymphangitis), and visceral pain associated with malignant and
non-malignant neoplasm of the circulatory system (e.g., lymphomas,
myelomas, Hodgkin's disease).
[0234] In another example, the compounds of the invention may be
used to treat visceral pain associated with injuries, inflammation,
diseases and disorders of the respiratory system including, but are
not limited to, upper respiratory infections (e.g.,
nasopharyngitis, sinusitis, and rhinitis), influenza, pneumoniae
(e.g., bacterial, viral, parasitic and fungal), lower respiratory
infections (e.g., bronchitis, bronchiolitis, tracheobronchitis),
interstitial lung disease, emphysema, bronchiectasis, status
asthmaticus, asthma, pulmonary fibrosis, chronic obstructive
pulmonary diseases (COPD), diseases of the pleura, and visceral
pain associated with malignant and non-malignant neoplasm of the
respiratory system (e.g., small cell carcinoma, lung cancer,
neoplasm of the trachea, of the larynx).
[0235] In another example, the compounds of the invention may be
used to treat visceral pain associated with injuries, inflammation
and disorders of the gastrointestinal system including, but are not
limited to, injuries, inflammation and disorders of the tooth and
oral mucosa (e.g., impacted teeth, dental caries, periodontal
disease, oral aphthae, pulpitis, gingivitis, periodontitis, and
stomatitis), of the oesophagus, stomach and duodenum (e.g., ulcers,
dyspepsia, oesophagitis, gastritis, duodenitis, diverticulitis and
appendicitis), of the intestines (e.g., Crohn's disease, paralytic
ileus, intestinal obstruction, irritable bowel syndrome, neurogenic
bowel, megacolon, inflammatory bowel disease, ulcerative colitis,
and gastroenteritis), of the peritoneum (e.g. peritonitis), of the
liver (e.g., hepatitis, liver necrosis, infarction of liver,
hepatic veno-occlusive diseases), of the gallbladder, biliary tract
and pancreas (e.g., cholelithiasis, cholecystolithiasis,
choledocholithiasis, cholecystitis, and pancreatitis), functional
abdominal pain syndrome (FAPS), gastrointestinal motility
disorders, as well as visceral pain associated with malignant and
non-malignant neoplasm of the gastrointestinal system (e.g.,
neoplasm of the oesophagus, stomach, small intestine, colon, liver
and pancreas).
[0236] In another example, the compounds of the invention may be
used to treat visceral pain associated with injuries, inflammation,
diseases, and disorders of the genitourinary system including, but
are not limited to, injuries, inflammation and disorders of the
kidneys (e.g., nephrolithiasis, glomerulonephritis, nephritis,
interstitial nephritis, pyelitis, pyelonephritis), of the urinay
tract (e.g. include urolithiasis, urethritis, urinary tract
infections), of the bladder (e.g. cystitis, neuropathic bladder,
neurogenic bladder dysfunction, overactive bladder, bladder-neck
obstruction), of the male genital organs (e.g., prostatitis,
orchitis and epididymitis), of the female genital organs (e.g.,
inflammatory pelvic disease, endometriosis, dysmenorrhea, ovarian
cysts), as well as pain associated with malignant and non-malignant
neoplasm of the genitourinary system (e.g., neoplasm of the
bladder, the prostate, the breast, the ovaries).
[0237] In further embodiments of this aspect of the invention,
compounds of the invention may be used to treat neuropathic pain
associated with injuries, inflammation, diseases and disorders of
the nervous system, including the central nervous system and the
peripheral nervous systems. Examples of such injuries,
inflammation, diseases or disorders associated with neuropathic
pain include, but are not limited to, neuropathy (e.g., diabetic
neuropathy, drug-induced neuropathy, radiotherapy-induced
neuropathy), neuritis, radiculopathy, radiculitis,
neurodegenerative diseases (e.g., muscular dystrophy), spinal cord
injury, peripheral nerve injury, nerve injury associated with
cancer, Morton's neuroma, headache (e.g., nonorganic chronic
headache, tension-type headache, cluster headache and migraine),
multiple somatization syndrome, postherpetic neuralgia (shingles),
trigeminal neuralgia complex regional pain syndrome (also known as
causalgia or Reflex Sympathetic Dystrophy), radiculalgia, phantom
limb pain, chronic cephalic pain, nerve trunk pain, somatoform pain
disorder, central pain, non-cardiac chest pain, central post-stroke
pain.
[0238] In another aspect, the compounds of the invention may be
used to treat inflammation associated with injuries, diseases or
disorders of the skin, subcutaneous tissues and related organs, the
musculoskeletal and connective tissue system, the respiratory
system, the circulatory system, the genitourinary system and the
gastrointestinal system.
[0239] In some embodiments of this aspect of the invention,
examples of inflammatory conditions, diseases or disorders of the
skin, subcutaneous tissues and related organs that may be treated
with the compounds of the invention include, but are not limited to
allergies, atopic dermatitis, psoriasis, eczema and dermatitis.
[0240] In other embodiments of this aspect of the invention,
inflammatory conditions, diseases or disorders of the
musculoskeletal and connective tissue system that may be treated
with the compounds of the invention include, but are not limited to
arthritis, osteoarthritis, and myositis.
[0241] In other embodiments of this aspect of the invention,
inflammatory conditions, diseases or disorders of the respiratory
system that may be treated with the compounds of the invention
include, but are not limited to allergies, asthma, rhinitis,
neurogenic inflammation, pulmonary fibrosis, chronic obstructive
pulmonary disease (COPD), adult respiratory distress syndrome,
nasopharyngitis, sinusitis, and bronchitis.
[0242] In still other embodiments of this aspect of the invention,
inflammatory conditions, disease or disorders of the circulatory
system that may be treated with the compounds of the invention
include, but are not limited to, endocarditis, pericarditis,
myocarditis, phlebitis, lymphadenitis and artherosclerosis.
[0243] In further embodiments of this aspect of the invention,
inflammatory conditions, diseases or disorders of thegenitourinary
system that may be treated with the compounds of the invention
include, but are not limited to, inflammation of the kidney (e.g.,
nephritis, interstitial nephritis), of the bladder (e.g.,
cystitis), of the urethra (e.g., urethritis), of the male genital
organs (e.g., prostatitis), and of the female genital organs (e.g.,
inflammatory pelvic disease).
[0244] In further embodiments of this aspect of the invention,
inflammatory conditions, diseases or disorders of the
gastrointestinal system that may be treated with the compounds of
the invention include, but are not limited to, gastritis,
gastroenteritis, colitis (e.g., ulcerative colitis), inflammatory
bowel syndrome, Crohn's disease, cholecystitis, pancreatitis and
appendicitis.
[0245] In still further embodiments of this aspect of the
invention, inflammatory conditions, diseases or disorders that may
be treated with the compounds of the invention, but are not limited
to inflammation associated with microbial infections (e.g.,
bacterial, viral and fungal infections), physical agents (e.g.,
burns, radiation, and trauma), chemical agents (e.g., toxins and
caustic substances), tissue necrosis and various types of
immunologic reactions and autoimmune diseases (e.g., Lupus
erythematosus).
[0246] In another aspect, the compounds of the invention may be
used to treat injuries, diseases or disorders of the nervous system
including, but not limited to neurodegenerative diseases (e.g.,
Alzheimer's disease, Duchenne's disease), epilepsy, multiple
sclerosis, amyotrophic lateral sclerosis, stroke, cerebral
ischemia, neuropathies (e.g., chemotherapy-induced neuropathy,
diabetic neuropathy), retinal pigment degeneration, trauma of the
central nervous system (e.g., spinal cord injury), and cancer of
the nervous system (e.g., neuroblastoma, retinoblastoma, brain
cancer, and glioma), and other certain cancers (e.g., melanoma,
pancreatic cancer).
[0247] In further aspects of the invention, the compounds of the
invention may also be used to treat other disorders of the skin,
subcutaneous tissues and related organs (e.g., hair loss), of the
respiratory system (e.g., asthma), of the circulatory system,
(e.g., cardiac arrhythmias and fibrillation and sympathetic
hyper-innervation), and of the genitourinary system (e.g.,
neurogenic bladder dysfunction and overactive bladder).
[0248] The present invention provides a method for treating a
subject that would benefit from administration of a composition of
the present invention. Any therapeutic indication that would
benefit from a NGF/NTR modulator (i.e., a compound of the
invention) can be treated by the methods of the invention. The
method includes the step of administering to the subject a
composition of the invention, such that the disease or disorder is
treated.
[0249] The invention further provides a method for preventing in a
subject, a disease or disorder which can be treated with
administration of the compositions of the invention. Subjects "at
risk" may or may not have detectable disease, and may or may not
have displayed detectable disease prior to the treatment methods
described herein. "At risk" denotes that an individual who is
determined to be more likely to develop a symptom based on
conventional risk assessment methods or has one or more risk
factors that correlate with development of a disease or disorder
that may be treated according the methods of the invention. For
example, risk factors include family history, medication history,
and history of exposure to an environmental substance which is
known or suspected to increase the risk of disease. Subjects at
risk for a disease or condition which can be treated with the
agents mentioned herein can also be identified by, for example, any
or a combination of diagnostic or prognostic assays known to those
skilled in the art. Administration of a prophylactic agent can
occur prior to the manifestation of symptoms characteristic of the
disease or disorder, such that the disease or disorder is prevented
or, alternatively, delayed in its progression.
EXEMPLIFICATION OF THE INVENTION
[0250] The invention is further illustrated by the following
example, which could be used to examine the
neurotrophin/neurotrophin precursor binding inhibition of the
compounds of the invention. The example should not be construed as
further limiting. The animal models used throughout the Examples
are accepted animal models and the demonstration of efficacy in
these animal models is predictive of efficacy in humans.
Compound Synthesis
Example 1
General Procedure for the Preparation Naphthalimide Derivatives
[0251] To a stirred solution of naphthalic anhydride derivative (1
equiv.) in glacial acetic acid was added, under an N.sub.2
atmosphere, the primary amine (1 equiv.) and sodium acetate (1
equiv.). The reaction mixture was brought to reflux. Refluxing was
continued and the progress of the reaction was monitored by TLC.
Upon completion, if the resultant solution was clear, then the
solvent was evaporated under reduced pressure and the residual
solid was reprecipitated and/or recrystallised from appropriate
solvent(s). If the final mixture contained precipitate then the
mixture was cooled to room temperature and the solid was collected
by filtration, washed with distilled water or dilute acid and
further reprecipitated and/or recrystallised from appropriate
solvent(s). The filtrate was recovered and treated as the clear
solution to obtain further product. The final products were dried
in vacuum for 24-48 hours.
Example 2
3-[7-(1-chloro-4-carboxy-phenyl)-1,3,6,8-tetraoxo-3,6,7,8-tetrahydro-1H-be-
nzo[lmn][3,8]phenanthrolin-2-yl]-4-chloro-benzoic acid (Compound
18)
[0252] ##STR34##
[0253] The compound was prepared from 1,4,5,8-naphthalene
tetracarboxylic dianhydride (1.34 g, 5 mmol) and
3-amino-4-chlorobenzoic acid (1.71 g, 10 mmol) in 100 mL glacial
acetic acid using the general procedure shown in Example 1.
[0254] .sup.1H NMR (d6-DMSO, 400 MHz): 7.89 (d, J=8.4 Hz, 2H), 8.12
(dd, J=8.4 Hz, 2.0 Hz, 2H), 8.30 (d, J=2.0 Hz, 2H), 8.82 (s, 4H),
13.40 (bs, 2H);
[0255] .sup.13C NMR (dept): 131.1, 132.0, 132.3, 133.1;
[0256] MS (ES-) m/z: 574.2 (M, 100%).
Example 3
2-(5-Carboxy-2-chloro-phenyl)-1,3,6-trioxo-1,2,3,6-tetrahydro-benzo[lmn]be-
nzo[4,5]imidazo[2,1-b][3,8]phenanthroline-9-carboxylic acid
(Compound 90)
[0257] ##STR35##
[0258] The intermediate compound was prepared from
1,4,5,8-naphthalene tetracarboxylic dianhydride (1.34 g, 5 mmol)
and 3-amino-4-chlorobenzoic acid (0.86 g, 5 mmol) in 100 mL glacial
acetic acid using the general procedure from Example 1.
[0259] .sup.1H NMR (d6-DMSO, 400 MHz): 7.89 (d, J=8.4 Hz, 1H), 8.12
(dd, J=8.4 Hz, 2.0 Hz, 1H), 8.30 (d, J=2.0 Hz, 1H), 8.78 (s,
4H);
[0260] MS (ES-) m/z: 420.1 (M-1, 100%).
[0261] Compound 90 was prepared from the intermediate (Example 1;
0.42 g, 1 mmol) and 3,4-diamino benzoic acid (0.15 g, 1 mmol) in 20
mL glacial acetic acid using the general procedure from Example
1.
[0262] .sup.1H NMR (d6-DMSO, 400 MHz): 7.88 (d, J=4.4 Hz, 1H), 7.90
(d, J=4.0 Hz, 1H), 8.01 (dd, J=8.4 Hz, 1.6 Hz, 1H), 8.11 (dd, J=8.4
Hz, 2.0 Hz, 1H), 8.34 (d, J=2.0 Hz, 1H), 8.73-8.79 (m, 2H),
8.85-8.91 (m, 3H), 13.29 (bs, 2H);
[0263] .sup.13C NMR (dept): 117.4, 121.0, 127.8, 127.9, 131.0,
131.8, 131.9, 132.2, 132.3, 133.2;
[0264] MS (ES-) m/z: 536.2 (M-1, 100%).
Example 4
1,3,6-trioxo-1,2,3,6-tetrahydrobenzo[lmn]benzo[4,5]imidazo[2,1b][3,8]di
phenan-throline-9-carboxylic acid (Compound 91)
[0265] ##STR36##
[0266] The compound was prepared from 1,4,5,8-naphthalene
tetracarboxylic dianhydride (1.34 g, 5 mmol) and 3,4-diaminobenzoic
acid (10 mmol) in 100 mL glacial acetic acid using the general
procedure shown in Example 1.
Biological Activity
Materials and Methods
Cell Culture
[0267] All cells were incubated at 37.degree. C. in 5% CO2. PC12
cells were maintained in RPMI-1640 medium supplemented with 10%
fetal bovine serum (FBS). A875 human melanoma cells were maintained
in DMEM with 10% FBS. HEK 293 cells were stably transfected with
human TrkA plasmid (see below) and maintained in DMEM with 10% FBS
and G418 (600 .mu.g/mL) for selection.
Cell Transfections:
[0268] Expression of human TrkA in HEK cells was achieved by
transfecting the cells with the TrkA expression vector using
Lipofectamine (Invitrogen) reagent. HEK cells were plated in 100 mm
Petri dishes at a concentration of 10.sup.6 cells per dish. The
next day, a solution of DNA diluted in 1 mL OptiMEM (Invitrogen)
per dish was prepared and incubated for 15 min at room temperature.
Concurrently, 42 .mu.l of Lipofectamine reagent was prepared in
OPtiMEM (1 mL per dish) and incubated for 15 min at room
temperature. The DNA and lipofectamine reagent solutions were then
mixed together and incubated for a further 15 minutes. During this
30 min of incubation, the cells were rinsed twice with OptiMEM. The
DNA-lipofectamine solution in OptiMEM was then added to the dish,
which was then placed in the incubator (37.degree. C.; 5% CO.sub.2)
for 3 h. This solution was then aspirated and the cells were rinsed
with DMEM. From this point, cells were grown in DMEM+FBS (10%),
their normal growth medium. For stable cell lines, the culture
medium contained G418 (600 .mu.g/mL) a selection agent for
maintenance of TrkA expression in the cells. The presence of TrkA
was confirmed with .sup.125I-NGF binding (see below) and Western
blots (see below) labeled with TrkA specific antisera.
Example 1
NGF Binding
[0269] NGF binding was evaluated using methods familiar to those
who are skilled in the art. Briefly, cells expressing one or both
NGF receptors (PC12: TrkA+p75; A875: p75 alone; HEK_TrkA: TrkA
alone) were harvested by replacing the medium with
calcium-magnesium free balanced salt (Grey's) solution and
incubating at 37.degree. C. for 15 min. For NGF binding, cells were
resuspended at a concentration of 2.times.10.sup.6 cells/mL in
HEPES-Krebs-Ringer (HKR) buffer (10 mM HEPES; 125 mM NaCl; 4.8 mM
KCl; 1.3 mM CaCl.sub.2; 1.2 mM MgSO.sub.4; 1.2 mM KH.sub.2PO.sub.4;
1 mg/ml BSA; 1 mg/ml glucose; pH 7.4) and exposed to .sup.125I-NGF
(.about.0.1 nM) in the presence or absence of varying
concentrations of the compound. Non-specific binding was determined
for reference by incubating .sup.125I-NGF with an excess of
non-radioactive NGF in the abence of compound. Following a two-hour
incubation period at 4.degree. C., .sup.125I-NGF bound to the cells
was quantified in a gamma radiation counter following separation
from unbound NGF by filtration or centrifugation through glycerol
(10% in HKR). Inhibition of binding was calculated as a percent of
the specific binding (calculated as the differential between
.sup.125I-NGF binding in the absence and presence of an excess of
non-radioactive NGF without compound). Dose-response inhibition
curves were typically generated with seven concentrations of a
given compound, with three replicates for each concentration. For
most compounds, multiple dose-response curves were generated.
[0270] The IC.sub.50 data shown in Table 1 were acquired using the
procedure described herein using PC12 cells. Examples of IC.sub.50
calculation curves from individual experiments using this procedure
are shown in FIGS. 2A, 2B, 3A, 4A, and 5, in which compounds 90, 91
and 18 are shown to effectively block NGF binding to TrkA and/or
p75.
Example 2
NGF Crosslinking to Receptors
[0271] NGF binding to TrkA and p75 was qualitatively evaluated
following chemical cross-linking, and separation of proteins
according to molecular weight with SDS-PAGE. PC12 (for p75 and TrkA
binding), HEK_TrkA (for TrkA only) and A875 (for p75 only) cells
were recovered using Grey's solution, pelleted by centrifugation,
and suspended in HKR. In a total volume of 1 mL, 2.times.10.sup.6
cells/mL were incubated, rotating, with .about.0.1 nM
.sup.125I-NGF, with or without compound, for 2 h at 4.degree. C. At
the conclusion of the binding reaction, a 20 .mu.L volume of
BS.sup.3 (Bis[sulfosuccinimidyl]suberate) crosslinker was added for
a final concentration 0.4 mM and incubated, rocking, for an
additional 30 min at room temperature. Cells were washed twice in
HKR. Following centrifugation, the pellets were solubilized
directly in SDS sample buffer and heated for 10 min at 95.degree.
C. All samples were electrophoresed on a 6% SDS-PAGE gel, which was
then dried and autoradiographed. Bands at the appropriate molecular
weights for p75-NGF conjugates and TrkA-NGF conjugates were
visualized by exposing the dried gel to film overnight (BioMax,
Kodak) overnight. Modulatory effects of the compounds on NGF
binding were determined by variations in band intensity. Reduced
binding of NGF to its receptors was represented by lighter
bands.
[0272] Results from this qualitative evaluation are in accordance
with the quantitative data from the binding assay of Example 1.
Example 3
Erk Phosphorylation
[0273] This assay is useful for establishing that the compounds of
the invention are functional NGF antagonists, not receptor agonists
(an agonist could conceivably block NGF binding but actually
activate the receptor). Erk 1/2 is a kinase activated down stream
of TrkA and is a well studied member of the NGF-induced signal
transduction cascade.
[0274] PC12 cells expressing TrkA and p75 were acutely exposed to 5
ng/mL NGF (15 min; 37.degree. C.; 5% CO.sub.2) that was
pre-incubated (30 min; room temperature) with or without the
compounds. Cells were lysed in Laemmli sample buffer (for SDS-PAGE)
or a lysis buffer containing Triton X-100 (for ELISA). Following
SDS-PAGE, proteins were electroblotted onto nitrocellulose and
immunoprobed for phosphorylated Erk 1 and 2. Blocking and primary
antibody incubations of immunoblots were performed in Tris-buffered
saline-Tween (10 mM Tris, pH 8.0, 150 mM NaCl, and 0.2% Tween 20)
supplemented with 5% (w/v) bovine serum albumin (BSA); secondary
antibody incubations were performed in 5% (w/v) dried skim milk
powder. Immunoreactive bands were detected by
chemiluminescence.
[0275] Representative examples of a phosphoryated Erk 1/2 blot for
Compound 90 and 91 are shown in FIGS. 1 and 4A, which demonstrate
that Compounds 90 and 91 inhibit NGF-induced Erk 1/2
phosphorylation in PC12 cells. FIG. 6 shows an additional
phosphorylated Erk 1/2 blot for Compound 90 as acquired using this
procedure, which demonstrates that Compound 90 inhibits NGF-induced
Erk 1/2 phosphorylation in PC12 cells.
[0276] To detect phosphorylated Erk 1/2 by ELISA, a kit from
R&D (Minneapolis, Minn.) Systems is used. Briefly, cells are
lysed in buffer (1 mM EDTA, 0.5% Triton X-100, 5 mM NaF, 1 M urea,
10 .mu.g/mL, Leupeptin, 10 .mu.g/mL Pepstatin, 100 .mu.M PMSF, 3
.mu.g/mL Aprotinin, 2.5 mM sodium pyrophosphate, 1 mM sodium
orthovanadate in PBS, pH 7.2-7.4). Lysates are incubated overnight
at 4.degree. C. in ELISA plates coated with an anti-Erk 1/2 capture
antibody. Immobilized Erk 1/2 is then exposed to a biotinylated
detection antibody specific for phosphorylated Erk. The quantity of
phosphoryated Erk 1/2 is quantified colorometrically with a
standard HRP-streptavidin reaction.
Example 4
Neurite Outgrowth
[0277] This assay was run as a further functional marker of NGF
antagonism and takes advantage of the differentiation of PC12 cells
(neurite outgrowth) induced by NGF. Cultures of PC12 cells were
grown on Terasaki plates pre-coated with poly-D-lysine. Cells were
exposed to NGF (1-50 (preferably 5) ng/ml) to induce neurite
outgrowth as described elsewhere [L A Greene & A S Tischler,
Establishment of a noradrenergic clonal line of rat adrenal
pheochromocytoma cells which respond to nerve growth factor, Proc
Natl Acad Sci USA. July 1976; 73(7): 2424-2428]. In addition to
NGF, cells were exposed to varying concentration of the compounds
or vehicle. Following 4 days of exposure to NGF in the presence or
absence of compound, neurite outgrowth was quantified. A neurite
was scored if its caliber from origin to terminal was approximately
the same and the length was equal to or greater than 1.5 the cell
body diameter. The number of neurite bearing cells per total number
of viable cells was calculated for each condition; the number of
cells with neurites in the presence of NGF (without compound
exposure) was considered to represent maximal (100%) outgrowth, to
which the inhibitory effect of compounds of the invention on
NGF-induced outgrowth was compared.
[0278] Results from this experiment are shown in FIG. 3B, which
demonstrates that compound 90 effectively inhibits neurite
outgrowth induced by 5 ng/mL NGF in a dose-dependent fashion. The
x-axis represents the log concentration in M.
Example 5
Formalin Model--Model of Acute Tonic Pain
[0279] Male Sprague-Dawley rats are given a subcutaneous injection
of 50 .mu.l of 2.5% formalin into the plantar surface of one
hindpaw using a 27 gauge syringe needle, and nociceptive responses
are measured for the next 60 min. The compound or vehicle was
administered (i.p. or s.c.) 15-45 minutes prior to the formalin
injection. A nociceptive score was determined for each 5 min. block
by measuring the amount of time spent in each of four behavioral
categories: 0, the injected hindpaw is indistinguishable from that
of the contralateral paw; 1, the injected paw has little or no
weight placed on it; 2, the injected paw is elevated and is not in
contact with any surface; 3, the injected paw is licked, bitten, or
shaken. A weighted nociceptive score, ranging from 0 to 3 is
calculated by multiplying the time spent in each category by the
category weight, summing these products, and dividing by the total
time for each 5 min block of time (Coderre et al., Pain 1993; 54:
43). On the basis of the resulting response patterns, 2 phases of
nociceptive behaviour are identified and scored: first phase (P1;
0-5 min) and second phase (P2; 11-40 min).
[0280] Statistical analysis is performed using Prism.TM. 4.01
software package (GraphPad, San Diego, Calif., USA). The difference
in response levels between treatment groups and control vehicle
group is analyzed using an ANOVA followed by Bonferroni's method
for post-hoc pair-wise comparisons. A p value <0.05 is
considered to be significant.
Example 6
Writhing Test
[0281] The acetic acid abdominal contraction test (Martinez et al.,
Pain 1999; 81: 179) was used to determine the efficacy of different
compounds in a model of acute visceral chemical nociception. Mice
were given an intraperitoneal (i.p.) injection of 200 .mu.l of 0.6%
acetic acid in saline and observed for the number of abdominal
contractions (writhing) they exhibited in the 30 minutes following
the injection. Compound or vehicle was administered (i.p. or s.c.)
15-45 minutes prior to the acetic acid injection. Each episode of
writhing is characterized by a lengthwise stretching of the torso
with a concomitant arching of the back.
[0282] Statistical analysis was performed using Prism.TM. 4.01
software package (GraphPad, San Diego, Calif., USA). The difference
in response levels between treatment groups and control vehicle
group was analyzed using an ANOVA followed by Bonferroni's method
for post-hoc pair-wise comparisons. A p value <0.05 was
considered to be significant
[0283] Results from this experiment are shown in FIG. 7. Animals
were treated with vehicle (VEH) or Compound 91 (5 mg/kg i.p.; [CMP
1] and 50 mg/kg i.p. [CMP 2])) 10-20 minutes before the
intraperitoneal injection of acetic acid, as described in this
example. Compound 91 displayed a significant antinociceptive effect
(** p<0.01, compared to vehicle treated group) as indicated by
the attenuation of the chemically-induced abdominal contractions
(from 58.+-.4 [n=6] to 16.+-.9 [n=6]).
Example 7
CFA Model--Model of Chronic Nociceptive (Inflammatory Pain)
Pain
[0284] Injection of complete Freunds adjuvant (CFA) in the hindpaw
of the rat has been shown to produce a long-lasting inflammatory
condition, which is associated with behavioural hyperalgesia and
allodynia at the injection site (Hylden et al., Pain 1989; 37:
229). Rats (body weight 200-250 g) receive a s.c. injection of CFA
(50% in saline, 100 .mu.l, Sigma) into the plantar surface of the
hindpaw under brief halothane anaesthesia. After 24 h, animals are
treated with vehicle or compound (s.c. or i.p.) and, after
different time-points following the treatment (e.g., 30, 60 or 90
min.), they are tested for hindpaw weight bearing responses, as
assessed using an Incapacitance Tester (e.g., Linton
Instrumentation, UK), (Zhu et al., 2005). The instrument
incorporates a dual channel scale that separately measures the
weight of the animal distributed to each hindpaw. While normal rats
distribute their body weight equally between the two hindpaws
(50-50), the discrepancy of weight distribution between an injured
and non-injured paw is a natural reflection of the discomfort level
in the injured paw (nocifensive behavior). The rats are placed in a
plastic chamber designed so that each hindpaw rests on a separate
transducer pad. The averager is set to record the load on the
transducer over 5 s time period and two numbers displayed represent
the distribution of the rat's body weight on each paw in grams (g).
For each rat, three readings from each paw are taken and then
averaged. Side-to-side weight bearing difference is calculated as
the average of the absolute value of the difference between two
hindpaws from three trials (right paw reading--left paw
reading).
Example 8
Seltzer or Partial Nerve Ligation (PNL) Model (Neuropathic Pain
Model)
[0285] The Partial Nerve Ligation (PNL) model (Seltzer et al., Pain
1990; 43:205) is used to induce chronic neuropathic pain. Male
Sprague-Dawley rats are anesthetized with isoflurane, and injury to
the sciatic nerve is achieved by tying a tight ligature around 1/3
to 1/2 of the sciatic nerve just proximal to the nerve's
trifurcation in the mid-thigh. Animals develop thermal and
mechanical hyperalgesia and allodynia, as well as a long-lasting
spontaneous pain or dysesthesia after the nerve injury.
[0286] Mechanical allodynia testing: Pre- and post-injury baselines
as well as post-treatment values (vehicle or compound-treated
animals) for mechanical allodynia are evaluated using Von Frey
filaments (Stoelting, Wood Dale, Ill., USA) with varying stiffness.
Animals are placed on a perforated metallic platform and allowed to
acclimate to their surroundings for a minimum of 30 minutes before
testing. The mean and standard error of the mean (SEM) are
determined for each paw in each treatment group
(ipsilateral/injured paw and contralateral/non-injured paw). Since
this stimulus is normally not considered painful, significant
injury-induced increases in responsiveness (i.e., lower response
thresholds) in this test are interpreted as a measure of mechanical
allodynia. Effects of compounds are evaluated two weeks after the
injury. Compound or vehicle is administered (i.p. or s.c.) and
effects on mechanical allodynia are measured at different time
points after dosing (e.g., 30, 60 and 90 min.).
[0287] Data analysis: Statistical analyses are conducted using
Prism.TM. 4.01 (GraphPad, San Diego, Calif., USA). Mechanical
hypersensitivity of the injured paw is determined by comparing
contralateral to ipsilateral paw values within the vehicle group at
each time point. Effect of vehicle (VEH) and compound are
determined by comparing the post-injury baseline (BL) to post
treatment values using RM-ANOVA or one-way ANOVA followed by
Bonferroni's method for post-hoc pair-wise comparisons (e.g.,
vehicle vs compound).
Example 9
Decosterd Model or Spared Nerve Injury Model (SNI) (Neuropathic
Pain Model)
[0288] The Spared Nerve Injury (SNI) model (Decosterd et al., Pain
2000; 87: 149) is used to induce chronic neuropathic pain. Male
Sprague-Dawley rats are anesthetized with isoflurane, and two of
the three terminal branches of the sciatic nerve (tibial and common
peroneal nerves) are transected, leaving the remaining sural nerve
intact. Animals develop thermal and mechanical hyperalgesia and
allodynia, as well as a long-lasting spontaneous pain or
dysesthesia the last month after the nerve injury.
[0289] Mechanical allodynia testing: Pre- and post-injury baselines
as well as post-treatment values (vehicle or compound-treated
animals) for mechanical allodynia are evaluated using Von Frey
filaments (Stoelting, Wood Dale, Ill., USA) with varying stiffness.
Animals are placed on a perforated metallic platform and allowed to
acclimate to their surroundings for a minimum of 30 minutes before
testing. The mean and standard error of the mean (SEM) are
determined for each paw in each treatment group
(ipsilateral/injured paw and contralateral/non-injured paw). Since
this stimulus is normally not considered painful, significant
injury-induced increases in responsiveness (i.e. lower response
thresholds) in this test are interpreted as a measure of mechanical
allodynia. Effects of compounds are evaluated two weeks after the
injury. Compound or vehicle is administered (i.p. or s.c.) and
effects on mechanical allodynia were measured at different time
points after dosing (e.g., 30, 60 and 90 min.).
[0290] Data analysis: Statistical analyses is conducted using
Prism.TM. 4.01 (GraphPad, San Diego, Calif., USA). Mechanical
hypersensitivity of the injured paw is determined by comparing
contralateral to ipsilateral paw values within the vehicle group at
each time point. Effect of vehicle (VEH) and compound are
determined by comparing the post-injury baseline (BL) to post
treatment values using RM-ANOVA or one-way ANOVA followed by
Bonferroni's method for post-hoc pair-wise comparisons (e.g.
vehicle vs compound).
Example 10
Chung or Spinal Nerve Ligation (SNL) Mode (Neuropathic Pain
Model)
[0291] The Spinal Nerve Ligation (SNL) model (Kim and Chung, Pain
1992; 50: 355) is used to induce chronic neuropathic pain. Male
Sprague-Dawley rats (Harlan, Indianapolis, Ind., USA) are
anesthetized with isoflurane, the left L5 transverse process is
removed, and the L5 and L6 spinal nerves aree tightly ligated with
6-0 silk suture. The wound is then closed with internal sutures and
external staples.
[0292] Mechanical allodynia testing: Pre- and post-injury baselines
as well as post-treatment values (vehicle or compound-treated
animals) for non-noxious mechanical sensitivity are evaluated using
8 Semmes-Weinstein filaments (Stoelting, Wood Dale, Ill., USA) with
varying stiffness according to the up-down method (Chaplan et al.,
J Neurosci Methods 1994; 53: 55). Animals are placed on a
perforated metallic platform and allowed to acclimate to their
surroundings for a minimum of 30 minutes before testing. The mean
and standard error of the mean (SEM) are determined for each paw in
each treatment group. Since this stimulus is normally not
considered painful, significant injury-induced increases in
responsiveness (i.e., lower response thresholds) in this test are
interpreted as a measure of mechanical allodynia. Effects of
compounds are evaluated two weeks after the injury. Compound or
vehicle is administered (i.p. or s.c.) and effects on mechanical
allodynia are measured at different time points after dosing (e.g.,
30, 60 and 90 min.). Injections are performed by a separate
experimenter who was not involved in testing the animals.
[0293] Data analysis: Statistical analyses is conducted using
Prism.TM. 4.01 (GraphPad, San Diego, Calif., USA). Mechanical
hypersensitivity of the injured paw is determined by comparing
contralateral to ipsilateral paw values within the vehicle group at
each time point. Effect of vehicle (VEH) and compound are
determined by comparing the post-injury baseline (BL) to post
treatment values using RM-ANOVA or one-way ANOVA followed by
Bonferroni's method for post-hoc pair-wise comparisons (e.g.
vehicle vs. compound).
EQUIVALENTS
[0294] Those skilled in the art will recognize, or be able to
ascertain using no more than routine experimentation, many
equivalents to the specific embodiments of the invention described
herein. Such equivalents are intended to be encompassed by the
following claims.
INCORPORATION BY REFERENCE
[0295] The entire contents of all patents, published patent
applications and other references cited herein are hereby expressly
incorporated herein in their entireties by reference. The entire
contents of copending applications attorney docket number PCI-030,
filed on even date herewith, titled "METHODS OF MODULATING
NEUROTROPHIN-MEDIATED ACTIVITY"; and attorney docket number
PCI-048, filed on even date herewith, titled "METHODS OF MODULATING
NEUROTROPHIN-MEDIATED ACTIVITY" are expressly incorporated herein,
in their entirety, as applied to the compounds of the present
invention.
* * * * *