U.S. patent application number 09/918292 was filed with the patent office on 2002-06-27 for aryl and heteroaryl fused aminoalkyl-imidazole derivatives.
This patent application is currently assigned to Neurogen Corporation. Invention is credited to DeSimone, Robert W., Hutchison, Alan.
Application Number | 20020082425 09/918292 |
Document ID | / |
Family ID | 26825845 |
Filed Date | 2002-06-27 |
United States Patent
Application |
20020082425 |
Kind Code |
A1 |
DeSimone, Robert W. ; et
al. |
June 27, 2002 |
Aryl and heteroaryl fused aminoalkyl-imidazole derivatives
Abstract
Disclosed are compounds of the formula: 1 or the
pharmaceutically acceptable non-toxic salts thereof wherein X,
R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5, R.sub.6, R.sub.7, A,
B, C, and D are variables defined herein. Such compounds useful in
treatment of obesity and diabetes. The invention also provides
labeled probes for the localization of cellular receptors that are
involved in the modulation of blood glucose levels.
Inventors: |
DeSimone, Robert W.;
(Durham, CT) ; Hutchison, Alan; (Madison,
CT) |
Correspondence
Address: |
Steven J. Sarussi
McDonnell Boehnen Hulbert & Berghoff
32nd Floor
300 S. Wacker Drive
Chicago
IL
60606
US
|
Assignee: |
Neurogen Corporation
|
Family ID: |
26825845 |
Appl. No.: |
09/918292 |
Filed: |
July 30, 2001 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
09918292 |
Jul 30, 2001 |
|
|
|
09539835 |
Mar 31, 2000 |
|
|
|
60127656 |
Apr 2, 1999 |
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Current U.S.
Class: |
546/118 ;
544/277; 548/309.7 |
Current CPC
Class: |
C07D 471/04 20130101;
C07D 235/14 20130101; G01N 2333/72 20130101 |
Class at
Publication: |
546/118 ;
544/277; 548/309.7; 514/263.2; 514/263.1; 514/263.22; 514/303;
514/394 |
International
Class: |
A61K 031/52; C07D
471/02; C07D 473/00; A61K 031/4745 |
Claims
What is claimed is:
1. A compound of the formula: 25or pharmaceutically acceptable
non-toxic salts thereof wherein: R.sub.7 represents H, or
C.sub.1-C.sub.6 alkyl; when R.sub.7 is H, R.sub.1 represents 2-,
3-, or 4-picolyl or benzyl, each of which is optionally mono-, di-,
or trisubstituted independently with halogen, nitro,
trifluoromethyl, cyano, hydroxyl, C.sub.1-C.sub.6 alkoxy, or
C.sub.1-C.sub.6 alkyl; amino, mono or di(C.sub.1-C.sub.6)alkyl-
amino, amino(C.sub.1-C.sub.6)alkyl, or mono- or
di(C.sub.1-C.sub.6)alkylam- ino(C.sub.1-C.sub.6)alkyl, or mono- or
di (C.sub.1-C.sub.6) alkylamino (C.sub.1-C.sub.6) alkoxy;
--O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n is 1,2,3,4,
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or C.sub.1-C.sub.6 alkyl; or NR.sub.8R.sub.9 forms a 5-,
6- or 7-membered heterocycloalkyl ring; SO.sub.2R.sub.8,
NHSO.sub.2R.sub.8, SO.sub.2NHR.sub.8, SO.sub.2NHCOR.sub.8,
CONHSO.sub.2R.sub.8 where R.sub.8 is as defined above;
O(CH.sub.2)n--G where n=1,2,3 or 4 and G represents
SO.sub.2R.sub.8, NHSO.sub.2R.sub.8, SO.sub.2NHR.sub.8,
SO.sub.2NHCOR.sub.8, CONHSO.sub.2R.sub.8, where R.sub.8 is as
defined above; or tetrazole, triazole, imidazole, thiazole,
oxazole, thiophene, or pyridyl; when R.sub.7 represents
C.sub.1-C.sub.6 alkyl, R.sub.1 represents C.sub.1-C.sub.6 alkyl,
cyclopentyl, or cyclopropylmethyl; R.sub.2 represents hydroxy;
C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, each of which is
optionally substituted with amino or mono- or
di(C.sub.1-C.sub.6)alkylamino, C.sub.5-C.sub.7 cycloalkylamino or
C.sub.1-C.sub.7 cycloalkoxy; or O(CH.sub.2).sub.nCO.sub.2R.sub.8
where n=1,2,3 or 4, NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9
or CO.sub.2R.sub.8 where R.sub.8 and R.sub.9 are the same or
different and represent hydrogen or C.sub.1-C.sub.6 alkyl; or
NR.sub.8R.sub.9 forms 5-, 6-, or 7-membered heterocyclic ring;
R.sub.3 represents C.sub.1-C.sub.6 alkyl; R.sub.4 represents
C.sub.1-C.sub.6 alkoxy; or R.sub.4 represents methyl when R.sub.1
and R.sub.7 are lower alkyl; R.sub.5 and R.sub.6 are the same or
different and represent hydrogen or halogen; C.sub.1-C.sub.6 alkyl
or C.sub.1-C.sub.6 alkoxy, the alkyl portion of each being
optionally substituted with amino, mono- or di(C.sub.1-C.sub.6)
alkylamino, or a C.sub.5-C.sub.7 heterocycloalkyl group where the
heteroatom is nitrogen and the nitrogen is attached to the parent
alkyl portion; O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1,2,3,4,
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or straight or branched chain lower alkyl having 1-6
carbon atoms; NR.sub.8R.sub.9 forms a 5-, 6- or 7-membered
heterocyclic ring; X represents a bond, CH.sub.2O, or CH.dbd.CH;
and A, B, C, and D are the same or different and represent CH or N
with the proviso that not more than two of A, B, C and D represent
N.
2. A compound of the formula 26where R.sub.7 is C.sub.1-C.sub.6
alkyl; R.sub.1 represents benzyl optionally mono-, di-, or
trisubstituted independently with halogen, nitro, trifluoromethyl,
cyano, hydroxyl, C.sub.1-C.sub.6 alkoxy, or C.sub.1-C.sub.6 alkyl;
or amino, mono or di(C.sub.1-C.sub.6)alkylamino,
amino(C.sub.1-C.sub.6)alkyl, or mono- or
di(C.sub.1-C.sub.6)alkylamino(C.sub.1-C.sub.6)alkyl, or mono- or di
(C.sub.1-C.sub.6) alkylamino (C.sub.1-C.sub.6) alkoxy; R.sub.2
represents hydroxy; C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6
alkoxy, each of which is optionally substituted with amino or mono-
or di(C.sub.1-C.sub.6)alkyl- amino, C.sub.5-C.sub.7 cycloalkylamino
or C.sub.5-C.sub.7 cycloalkoxy; or O(CH.sub.2).sub.nCO.sub.2R.sub.8
where n=1,2,3 or 4, NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9
or CO.sub.2R.sub.8 where R.sub.8 and R.sub.9 are the same or
different and represent hydrogen or C.sub.1-C.sub.6 alkyl; or
NR.sub.8R.sub.9 forms 5-, 6-, or 7-membered heterocyclic ring;
R.sub.3 represents C.sub.1-C.sub.6 alkyl; R.sub.4 represents
C.sub.1-C.sub.6 alkoxy; or R.sub.5 and R.sub.6 are the same or
different and represent hydrogen or halogen; C.sub.1-C.sub.6 alkyl
or C.sub.1-C.sub.6 alkoxy, the alkyl portion of each being
optionally substituted with amino, mono- or di(C.sub.1-C.sub.6)
alkylamino, or a C.sub.5-C.sub.7 heterocycloalkyl group where the
heteroatom is nitrogen and the nitrogen is attached to the parent
alkyl portion; and X represents a bond or CH.sub.2O.
3. A compound according to claim 2, wherein one of R.sub.5 and
R.sub.6 is hydrogen.
4. A compound according to claim 2, wherein R.sub.4 is methoxy, one
of R.sub.5 and R.sub.6 is hydrogen, and the other of R.sub.5 and
R.sub.6 is alkoxy.
5. A compound according to claim 2, wherein R.sub.7 is
C.sub.4-C.sub.6 alkyl.
6. A compound according to claim 5, wherein X is a bond.
7. A compound according to claim 2, wherein R.sub.1 is benzyl
monosubstituted in the ortho position.
8. A compound according to claim 7, wherein R.sub.7 is
C.sub.4-C.sub.6 alkyl and X is a bond.
9. A compound of the formula: 27wherein: R.sub.7 represents
C.sub.1-C.sub.6 alkyl; R.sub.1 represents C.sub.1-C.sub.6 alkyl,
cyclopentyl, or cyclopropylmethyl; R.sub.2 represents hydroxy;
C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, each of which is
optionally substituted with amino or mono- or
di(C.sub.1-C.sub.6)alkylami- no, C.sub.5-C.sub.7 cycloalkylamino or
C.sub.5-C.sub.7 cycloalkoxy; or O(CH.sub.2).sub.nCO.sub.2R.sub.8
where n=1, 2, 3 or 4 NR.sub.8COR.sub.9, COR.sub.8,
CONR.sub.8R.sub.9 or CO.sub.2R.sub.8 where R.sub.8 and R.sub.9 are
the same or different and represent hydrogen or C.sub.1-C.sub.6
alkyl; or NR.sub.8R.sub.9 forms 5-, 6-, or 7-membered heterocyclic
ring; R.sub.3 represents C.sub.1-C.sub.6 alkyl; R.sub.4 represents
C.sub.1-C.sub.6 alkoxy; or R.sub.4 represents methyl when R.sub.1
and R.sub.7 are lower alkyl; R.sub.5 and R.sub.6 are the same or
different and represent hydrogen or halogen; C.sub.1-C.sub.6 alkyl
or C.sub.1-C.sub.6 alkoxy, the alkyl portion of each being
optionally substituted with amino, mono- or di(C.sub.1-C.sub.6)
alkylamino, or a C.sub.1-C.sub.7 heterocycloalkyl group where the
heteroatom is nitrogen and the nitrogen is attached to the parent
alkyl portion; O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1, 2, 3 or
4 NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or
CO.sub.2R.sub.8 where R.sub.8 and R.sub.9 are the same or different
and represent hydrogen or straight or branched chain lower alkyl
having 1-6 carbon atoms; NR.sub.8R.sub.9 forms a 5-, 6- or
7-membered heterocyclic ring; X represents a bond or CH.sub.2O.
10. A compound according to claim 9 wherein R.sub.3 and R.sub.7 are
methyl.
11. A compound according to claim 9 wherein R.sub.1 is propyl or
cyclopropylmethyl.
12. A compound according to claim 9 wherein R.sub.1 is
cyclopentyl.
13. A compound according to claim 9 wherein X is CH.sub.2O.
14. A compound according to claim 13, wherein R.sub.1 is propyl or
cyclopropylmethyl.
15. A compound according to claim 9, wherein R.sub.1 is propyl or
cyclopropylmethyl and one of R.sub.5 and R.sub.6 is hydrogen.
16. A compound according to claim 9, wherein R.sub.1 is propyl,
R.sub.4 is methoxy or methyl, one of R.sub.5 and R.sub.6 is
hydrogen, and the other of R.sub.5 and R.sub.6 is alkoxy.
17. A compound of the formula: 28wherein: R.sub.7 represents
C.sub.1-C.sub.6 alkyl; R.sub.1 represents C.sub.1-C.sub.6 alkyl,
cyclopentyl, or cyclopropylmethyl; R.sub.2 represents hydroxy;
C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, each of which is
optionally substituted with amino or mono- or
di(C.sub.1-C.sub.6)alkylami- no, C.sub.5-C.sub.7 cycloalkylamino or
C.sub.5-C.sub.7 cycloalkoxy; or O(CH.sub.2).sub.nCO.sub.2R.sub.8
where n=1, 2, 3 or 4 NR.sub.8COR.sub.9, COR.sub.8,
CONR.sub.8R.sub.9 or CO.sub.2R.sub.8 where R.sub.8 and R.sub.9 are
the same or different and represent hydrogen or C.sub.1-C.sub.6
alkyl; or NR.sub.8R.sub.9 forms 5-, 6-, or 7-membered heterocyclic
ring; R.sub.3 represents C.sub.1-C.sub.6 alkyl; R.sub.4 represents
C.sub.1-C.sub.6 alkoxy; or R.sub.4 represents methyl when R.sub.1
and R.sub.7 are lower alkyl; R.sub.5 and R.sub.6 are the same or
different and represent hydrogen or halogen; C.sub.1-C.sub.6 alkyl
or C.sub.1-C.sub.6 alkoxy, the alkyl portion of each being
optionally substituted with amino, mono- or di(C.sub.1-C.sub.6)
alkylamino, or a C.sub.5-C.sub.7 heterocycloalkyl group where the
heteroatom is nitrogen and the nitrogen is attached to the parent
alkyl portion; O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1,2,3 or 4,
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or straight or branched chain lower alkyl having 1-6
carbon atoms; NR.sub.8R.sub.9 forms a 5-, 6- or 7-membered
heterocyclic ring; X represents a bond or CH.sub.2O.
18. A compound according to claim 17 wherein R.sub.3 and R.sub.7
are methyl.
19. A compound according to claim 17 wherein R.sub.1 is propyl or
cyclopropylmethyl.
20. A compound according to claim 17 wherein R.sub.1 is
cyclopentyl.
21. A compound according to claim 17 wherein X is CH.sub.2O.
22. A compound according to claim 21, wherein R.sub.1 is propyl or
cyclopropylmethyl.
23. A compound according to claim 17, wherein R.sub.1 is propyl or
cyclopropylmethyl and one of R.sub.5 and R.sub.6 is hydrogen.
24. A compound according to claim 17, wherein R.sub.1 is propyl,
R.sub.4 is methoxy or methyl, one of R.sub.5 and R.sub.6 is
hydrogen, and the other of R.sub.5 and R.sub.6 is alkoxy.
25. A compound of the formula 29where R.sub.7 is hydrogen or
C.sub.1-C.sub.6 alkyl; R.sub.1 represents benzyl optionally mono-,
di-, or trisubstituted independently with halogen, nitro,
trifluoromethyl, cyano, hydroxyl, C.sub.1-C.sub.6 alkoxy, or
C.sub.1-C.sub.6 alkyl; or amino, mono or
di(C.sub.1-C.sub.6)alkylamino, amino(C.sub.1-C.sub.6)alkyl- , or
mono- or di(C.sub.1-C.sub.6)alkylamino(C.sub.1-C.sub.6)alkyl, or
mono- or di (C.sub.1-C.sub.6) alkylamino (C.sub.1-C.sub.6) alkoxy;
R.sub.2 represents hydroxy; C.sub.1-C.sub.6 alkyl or
C.sub.1-C.sub.6 alkoxy, each of which is optionally substituted
with amino or mono- or di(C.sub.1-C.sub.6)alkylamino,
C.sub.5-C.sub.7 cycloalkylamino or C.sub.1-C.sub.7 cycloalkoxy; or
O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1,2,3 or 4,
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or C.sub.1-C.sub.6 alkyl; or NR.sub.8R.sub.9 forms 5-, 6-,
or 7-membered heterocyclic ring; R.sub.3 represents C.sub.1-C.sub.6
alkyl; R.sub.4 represents C.sub.1-C.sub.6 alkoxy; or R.sub.5 and
R.sub.6 are the same or different and represent hydrogen or
halogen; C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, the alkyl
portion of each being optionally substituted with amino, mono- or
di(C.sub.1-C.sub.6) alkylamino, or a C.sub.5-C.sub.7
heterocycloalkyl group where the heteroatom is nitrogen and the
nitrogen is attached to the parent alkyl portion; and X represents
a bond or CH.sub.2O.
26. A compound according to claim 25, wherein one of R.sub.5 and
R.sub.6 is hydrogen.
27. A compound according to claim 25, wherein R.sub.4 is methoxy,
one of R.sub.5 and R.sub.6 is hydrogen, and the other of R.sub.5
and R.sub.6 is alkoxy.
28. A compound according to claim 25, wherein R.sub.7 is
C.sub.4-C.sub.6 alkyl.
29. A compound according to claim 25, wherein X is a bond.
30. A compound according to claim 25, wherein R.sub.1 is benzyl
monosubstituted in the ortho position.
31. A compound according to claim 30, wherein R.sub.7 is
C.sub.4-C.sub.6 alkyl and X is a bond.
32. A compound according to claim 1, which is
((2,4-dimethoxyphenyl)-N-(3--
methylbutyl)-N-({1-[(2-methylphenyl)methyl]benzimidazol-2-yl}methyl)carbox-
amide
33. A compound according to claim 1, which is
2-(2,3-dimethylphenoxy)-N-me- thyl-N- [(1-propylbenzimidazol-2-yl)
ethyl]acetamide.
34. A compound according to claim 1, which is
2-(2,3-dimethylphenoxy)-N-{[-
1-(cyclopropylmethyl)benzimidazol-2-yl]ethyl}-N-methylacetamide.
35. A compound according to claim 1, which is
(2,4-dimethoxyphenyl)-N-({1--
[(2-chlorophenyl)methyl]benzimidazol-2-yl}methyl)-N-(3-methylbutyl)carboxa-
mide.
36. A compound according to claim 1, which is
(2,4-dimethoxyphenyl)-N-(3-m-
ethylbutyl)-N-({3-[(2-methylphenyl)methyl]imidazolo[5,4-b],pyridin-2-yl}me-
thyl)carboxamide.
37. A compound according to claim 1, which is
2-(2,3-dimethylphenoxy)-N-me-
thyl-N-[(-popylimidazolo[5,4-b]pyridin -2-yl) ethyl]acetamide.
38. A compound according to claim 1, which is
2-(2,3-dimethylphenoxy)-N-{[-
1-(cyclopentyl)-6-chlorobenzimidazol-2-yl]ethyl}-N-methylacetamide
39. A pharmaceutical composition comprising a compound as claimed
in claim 1 and a pharmaceutically acceptable carrier or
excipient.
40. A method of treating diabetes, comprising administering to a
patient in need of such treatment an effective amount of a compound
according to claim 1, or a pharmaceutically acceptable salt
thereof.
41. A method of treating obesity or eating disorders, comprising
administering to a patient in need of such treatment an effective
amount of a compound according to claim 1, or a pharmaceutically
acceptable salt thereof.
42. The use of a compound of claim 1 for the manufacture of a
medicament for the treatment of diabetes or obesity.
43. A process for the preparation of a compound of claim 1.
44. A method for localizing receptors in a tissue sample
comprising: contacting the sample with a detectably-labeled
compound of claim 1 under conditions that permit binding of the
compound to the receptors, washing the sample to remove unbound
compound, and detecting the bound compound.
45. The method of claim 44, wherein the receptors modulate blood
glucose levels.
46. The method of claim 44, wherein the receptors are GLP-1
receptors.
47. A packaged pharmaceutical composition comprising the
pharmaceutical composition of claim 39 in a container and
instructions for using the composition to treat a patient suffering
from a disorder responsive to modulation of blood glucose
levels.
48. The packaged pharmaceutical composition of claim 47, wherein
said patient is suffering from obesity or diabetes.
Description
BACKGROUND OF THE INVENTION
[0001] This application claims the benefit of U.S. Provisional
Application no. 60/127,656, filed on Apr. 2, 1999.
[0002] 1. Field of the Invention
[0003] This invention relates to 1-benzylimidazole derivatives, and
more specifically, to the use of such compounds as pharmaceutical
agents, e.g., as modulators of blood glucose levels. This invention
also relates to pharmaceutical compositions comprising such
compounds and to the use of such compounds in treating a variety of
disorders associated with feeding and food metabolism.
Additionally, this invention relates to the use such compounds as
probes for the localization of cellular receptors that are involved
in the modulation of blood glucose levels.
[0004] 2. Background
[0005] Diabetes mellitus is a chronic syndrome of impaired
carbohydrate and fat metabolism resulting from insufficient insulin
secretion and/or target tissue insulin resistance. It occurs in two
major forms: insulin-dependent diabetes mellitus (IDDM, Type 1) and
non-insulin-dependent diabetes mellitus (NIDDM, Type 2). These
forms differ in their etiology, age of onset and treatment. Type 1
is often characterized by onset during childhood and the patients
typically become fully dependent upon exogenous insulin to sustain
life. The disorder is associated with a lack of insulin production
by the pancreatic Islets of Langerhans. The disease is generally
marked by a drastic reduction in the number of insulin secreting
islet beta cells.
[0006] NIDDM usually appears later in life (age 40-60) and is often
associated with obesity. Patients with NIDDM show normal basal
levels of insulin but display an abnormal insulin secretion
response (delayed or reduced) to a glucose load. As the disease
progresses, insulin target tissues show signs of diminished
response to insulin (insulin resistence). Effective treatment of
the disorder is usually obtained by dietary control, with or
without the use of oral hypoglycemic drugs. Sulphonylureas are a
class of hypoglycemic compounds used in the treatment of NIDDM.
These drugs exert their action by causing insulin to be released
from intracellular stores. Care must be taken in the administration
of these agents in order to not induce severe hypoglycemia due to
excessive insulin release. In addition, overdose may deplete
insulin stores to a point requiring administration of exogenous
insulin.
[0007] The discovery that glucose administered via the
gastrointestinal tract provides greater stimulation of insulin
release than a comparable glucose challenge given intravenously led
to the identification of certain gut secreted `incretin` hormones
which augment glucose stimulated insulin secretion, and the
identification of specific cell surface receptors that modulate the
effects of such incretin hormones. Glucagon-like Peptide-1
(7-36)-amide (GLP-1) is one such incretin hormone that is secreted
from gastrointestinal L cells in response to food intake and
increases insulin secretion from pancreatic beta cells (Fehmann, H.
C.; Goke, R. and Goke, B. (1995) Endocr. Rev. 16: 390-410). GLP-1
exerts its actions via binding to a G-protein-linked receptor
expressed in islet .beta.-cells.
[0008] Unlike the sulphonylureas, the effects of GLP-1 are
dependent upon plasma glucose concentration in that the
insulinotropic effects of GLP-1 are abolished at low plasma glucose
levels. In addition to its stimulation of insulin secretion, GLP-1
also increases insulin synthesis (Drucker, D J (1987) Proc. Natl
Acad. Sci USA 84: 3434-3438), inhibits glucagon secretion (Kawai
(1989) Endocrinology 124: 1768-1773) and delays gastric emptying
(Nauck, M A (1995) Gut 37(2): A124). This combination of actions
gives GLP-1 unique potential therapeutic advantages over other
agents presently used to treat non-insulin dependent diabetes
mellitus. In a clinical trial of patients with NIDDM it was found
that subcutaneous administration of GLP-1 could normalize
postprandial glucose levels (Todd et al. (1997) Eur. J. Clin.
Invest. 27: 533-536). Drugs that mimic the action of GLP-1, i.e.
stimulate insulin secretion from pancreatic .beta.-cells, but only
at higher than normal blood glucose levels, are particularly
desirable for us in the treatment of NIDDM. Such drugs may work by
modulating the signal-transducing activity of the GLP-1
receptor.
[0009] In clinical studies GLP-1 has been shown to reduce appetite
and increase satiety in both normal weight and obese subjects (see,
e.g., Christophe J. Ann. N Y Acad. Sci. (1998) 865:323-335 and
Gutwiller, J. P., Am. J. Physio. (1999) 276: R1541-1544). Thus
drugs that modulate the activity of the GLP-1 receptor may be
useful for the treatment of obesity and eating disorders.
[0010] The effect of a compound on blood glucose levels can be
determined in vivo, through the use a glucose tolerance test, in
which the blood glucose levels laboratory animals subjected to a
glucose challenge are monitored in the presence and absence of the
compound. The effects of test compounds on glucose tolerance may be
evaluated in non-diabetic laboratory animals as discussed in Wang
et al., J. Clin. Invest. (1995) 95: 417-421 and Holst, Curr.
Opinion in Endocrinology and Diabetes (1998) 5: 108-115.
[0011] Alternatively, the effects of test compounds on blood
glucose levels may be assessed in an animal model of diabetes,
e.g., streptozotocin (STZ)-induced diabetes. Such assays have been
disclosed by Tancrde et al. (Br. J. Exp. Path. (1983) 64: 117-123),
Junod et al. (J. Clin. Inv. (1969) 48: 2129-2139, Rondu et al. (J.
Med. Chem. (1997) 40:3793-3803), and Maloff and Boyd (Diabetologia
(1986) 29: 295-300).
[0012] In vitro experiments that monitor the interaction of the
compound with GLP-1 receptors may also be used to reliably predict
the effects of a compound on blood glucose levels. In one such
experiment the interaction of compounds with GLP-1 receptors,
expressed either recombinantly or naturally in high abundance in
certain cell lines, may be determined by a cell-based luciferase
screen or by binding experiments measuring competition binding
e.g., with a labeled GLP-1 ligand such as GLP-1 or GLP(7-36)
peptide.
[0013] Receptors that are coupled to the G.sub.s stimulatory
G-protein subunit transduce intracellular signals via the adenylate
cyclase pathway. Stimulation of these receptors with an agonist
typically results in an elevation of cytoplasmic cAMP levels which
can trigger the subsequent transcription of a variety of genes,
generally those with promoters containing binding sites (cAMP
responsive elements--CREs) for the transcription factor, CREB (CRE
binding protein).
[0014] Receptor modulation may be measured via measurement of
transcriptional activation of a firefly luciferase reporter gene.
Such an assay may use a Chinese hamster ovary cell line (CHO-K1)
stably transfected with a GLP-1 receptor (a Gs coupled receptor)
expression plasmid and a luciferase reporter plasmid, wherein
luciferase expression is under the transcriptional control of
multiple CRES. In these cell lines, the GLP1 agonist GLP(7-36)
peptide stimulates luciferase expression in a dose dependent manner
with a potency (EC.sub.50.about.20 pM) similar to the data reported
by Gromada et al. (1995) FEBS Lett. 373: 182-186.
[0015] Compounds are screened by seeding 15,000 cells per well in
opaque multi-well plates. Cells are then incubated overnight in a
tissue culture incubator. Compounds are dispensed to a final
concentration of 4 uM in 1% DMSO. After 6 hours of incubation,
cells are assayed for luciferase activity, which is measured in a
luminometer.
SUMMARY OF THE INVENTION
[0016] This invention provides compounds of Formula I, below. The
invention also provides compounds of Formula I that bind
specifically, and preferably with high affinity, to specific
cellular receptors. Preferably the receptors are cell surface
receptors, and more preferably G-protein coupled receptors. Even
more preferably, the receptors are Secretin-like receptors. Highly
preferred receptors are GLP receptors, and most preferably, the
receptors are GLP-1 receptors. Such compounds are useful in the
treatment of diabetes, especially non-insulin-dependent diabetes
mellitus (Type 2 diabetes), and in the treatment of obesity and
eating disorders.
[0017] The invention further comprises methods of treating patients
suffering from diabetes, especially non-insulin-dependent diabetes
mellitus (Type 2 diabetes), obesity or eating disorders by
administering to a patient in need of such treatment an effective
amount of a compound of the invention. Treatment of human patients,
domesticated companion animals (pets) or livestock animals
suffering from these disorders with an effective amount of a
compound of the invention is encompassed by the invention.
[0018] In a separate aspect, this invention provides compounds that
are useful as probes for the localization of specific receptors.
Preferably these receptors modulate blood glucose levels. Such
receptors are preferably so localized in tissue samples, especially
tissue sections. Such probes are also useful for measuring levels
of such receptors expressed in tissue samples or cell membrane
preparations of tissue samples and for localizing receptors in
living patients (e.g., via PET scanning).
[0019] The invention also comprises a method for altering the
signal-transducing activity of a cell surface GLP1 receptor, said
method comprising exposing cells expressing such a receptor to an
effective amount of a compound of the Formula I, below.
[0020] The invention also provides pharmaceutical compositions
comprising compounds of Formula I, including packaged
pharmaceutical compositions. Packaged pharmaceutical compositions
may include a container and instructions for using the composition
to treat a patient in need thereof. Particulary, the invention
includes packaged pharmaceutical compositions that include a
container and instructions for using the composition to treat a
patient suffering from diabetes, obesity or eating disorders.
[0021] Accordingly, a broad embodiment of the invention is directed
to compounds of Formula I: 2
[0022] or the pharmaceutically acceptable non-toxic salts thereof
wherein:
[0023] R.sub.7 represents H, or C.sub.1-C.sub.6 alkyl;
[0024] when R.sub.7 is H, R.sub.1 represents 2-, 3-, or 4-picolyl
or benzyl, each of which is optionally mono-, di-, or
trisubstituted independently with
[0025] halogen, nitro, trifluoromethyl, cyano, hydroxyl,
C.sub.1-C.sub.6 alkoxy, or C.sub.1-C.sub.6 alkyl;
[0026] amino, mono or di(C.sub.1-C.sub.6)alkylamino,
amino(C.sub.1-C.sub.6)alkyl, or mono- or
di(C.sub.1-C.sub.6)alkylamino(C.- sub.1-C.sub.6)alkyl, or mono- or
di (C.sub.1-C.sub.6) alkylamino (C.sub.1-C.sub.6) alkoxy;
[0027] --O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n is 1, 2, 3 or 4,
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or C.sub.1-C.sub.6 alkyl; or
[0028] NR.sub.8R.sub.9 forms a 5-, 6- or 7-membered
heterocycloalkyl ring;
[0029] SO.sub.2R.sub.8, NHSO.sub.2R.sub.8, SO.sub.2NHR.sub.8,
SO.sub.2NHCOR.sub.8, CONHSO.sub.2R.sub.8 where R.sub.8 is as
defined above;
[0030] O(CH.sub.2).sub.n--G where n=1, 2, 3 or 4 and G represents
SO.sub.2R.sub.8, NHSO.sub.2R.sub.8, SO.sub.2NHR.sub.8,
SO.sub.2NHCOR.sub.8, CONHSO.sub.2R.sub.8, where R.sub.8 is as
defined above; or tetrazole, triazole, imidazole, thiazole, oxazo,
thiophene, or pyridyl;
[0031] when R.sub.7 represents C.sub.1-C.sub.6 alkyl, R.sub.1
represents C.sub.1-C.sub.6 alkyl, cyclopentyl, or
cyclopropylmethyl;
[0032] R.sub.2 represents
[0033] hydroxy;
[0034] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, each of
which is optionally substituted with amino or mono- or
di(C.sub.1-C.sub.6)alkylami- no, C.sub.5-C.sub.7 cycloalkylamino or
C.sub.1-C.sub.7 cycloalkoxy; or
[0035] O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1,2,3,4,
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or
[0036] CO.sub.2R.sub.8 where R.sub.8 and R.sub.9 are the same or
different and represent hydrogen or C.sub.1-C.sub.6 alkyl; or
[0037] NR.sub.8R.sub.9 forms 5-, 6-, or 7-membered heterocyclic
ring;
[0038] R.sub.3 represents C.sub.1-C.sub.6 alkyl;
[0039] R.sub.4 represents C.sub.1-C.sub.6 alkoxy; or
[0040] R.sub.4 represents methyl when R.sub.1 and R.sub.7 are lower
alkyl;
[0041] R.sub.5 and R.sub.6 are the same or different and represent
hydrogen or halogen;
[0042] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, the alkyl
portion of each being optionally substituted with amino, mono- or
di(C.sub.1-C.sub.6) alkylamino, or a C.sub.1-C.sub.7
heterocycloalkyl group where the heteroatom is nitrogen and the
nitrogen is attached to the parent alkyl portion;
[0043] O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1,2,3,4,
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or straight or branched ch- lower alkyl having 1-6 carbon
atoms;
[0044] NR.sub.8R.sub.9 forms a 5-, 6- or 7-membered heterocyclic
ring;
[0045] X represents a bond, CH.sub.2O, or CH.dbd.CH; and
[0046] A, B, C, and D independently represent CH or N with the
proviso that not more than two of A, B, C and D represent N.
[0047] These compounds are useful in the diagnosis and treatment of
obesity and diabetes.
DETAILED DESCRIPTION OF THE INVENTION
[0048] The compounds of the instant invention are represented by
the formula I: 3
[0049] or pharmaceutically acceptable non-toxic salts there
wherein:
[0050] R.sub.7 represents H, or C.sub.1-C.sub.6 alkyl;
[0051] when R.sub.7 is H, R.sub.1 represents 2-, 3-, or 4-picolyl
or benzyl, each of which is optionally mono-, di-, or
trisubstituted independently with
[0052] halogen, nitro, trifluoromethyl, cyano, hydroxyl,
C.sub.1-C.sub.6 alkoxy, or C.sub.1-C.sub.6 alkyl;
[0053] amino, mono or di(C.sub.1-C.sub.6)alkylamino,
amino(C.sub.1-C.sub.6)alkyl, or mono- or
di(C.sub.1-C.sub.6)alkylamino(C.- sub.1-C.sub.6)alkyl, or mono- or
di (C.sub.1-C.sub.6) alkylamino (C.sub.1-C.sub.6) alkoxy;
[0054] --O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n is 1, 2, 3 or 4
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or C.sub.1-C.sub.6 alkyl; or
[0055] NR.sub.8R.sub.9 forms a 5-, 6- or 7-membered
heterocycloalkyl ring;
[0056] SO.sub.2R.sub.8, NHSO.sub.2R.sub.8, SO.sub.2NHR.sub.8,
SO.sub.2NHCOR.sub.8, CONHSO.sub.2R.sub.8 where R.sub.8 is as
defined above;
[0057] O(CH.sub.2).sub.n--G where n=1, 2, 3 or 4 and G represents
SO.sub.2R.sub.8, NHSO.sub.2R.sub.8, SO.sub.2NHR.sub.8,
SO.sub.2NHCOR.sub.8, CONHSO.sub.2R.sub.8, where R.sub.8 is as
defined above; or
[0058] tetrazole, triazole, imidazole, thiazole, oxazole,
thiophene, or pyridyl;
[0059] when R.sub.7 represents C.sub.1-C.sub.6 alkyl, R.sub.1
represents C.sub.1-C.sub.6 alkyl, cyclopentyl, or
cyclopropylmethyl;
[0060] R.sub.2 represents
[0061] hydroxy;
[0062] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, each of
which is optionally substituted with amino or mono- or
di(C.sub.1-C.sub.6)alkylami- no, C.sub.5-C.sub.7 cycloalkylamino or
C.sub.1-C.sub.7 cycloalkoxy; or
[0063] O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1, 2, 3 or 4
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or C.sub.1-C.sub.6 alkyl; or
[0064] NR.sub.8R.sub.9 forms 5-, 6-, or 7-membered heterocyclic
ring;
[0065] R.sub.3 represents C.sub.1-C.sub.6 alkyl;
[0066] R.sub.4 represents C.sub.1-C.sub.6 alkoxy; or
[0067] R.sub.4 represents methyl when R.sub.1 and R.sub.7 are lower
alkyl;
[0068] R.sub.5 and R.sub.6 are the same or different and represent
hydrogen or halogen;
[0069] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, the alkyl
portion of each being optionally substituted with amino, mono- or
di(C.sub.1-C.sub.6) alkylamino, or a C.sub.5-C.sub.7
heterocycloalkyl group where the heteroatom is nitrogen and the
nitrogen is attached to the parent alkyl portion;
[0070] O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1, 2, 3 or 4
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or straight or branched chain lower alkyl having 1-6
carbon atoms;
[0071] NR.sub.8R.sub.9 forms a 5-, 6- or 7-membered heterocyclic
ring;
[0072] X represents a bond, CH.sub.2O, or CHCH; and
[0073] A, B, C, and D are the same or different and represent CH or
N with the proviso that not more than two of A, B, C and D
represent N.
[0074] Preferred compounds of the invention are represented by
Formula II 4
[0075] where
[0076] R.sub.1 represents benzyl optionally mono-, di-, or
trisubstituted independently with
[0077] halogen, nitro, trifluoromethyl, cyano, hydroxyl,
C.sub.1-C.sub.6 alkoxy, or C.sub.1-C.sub.6 alkyl; or
[0078] amino, mono or di(C.sub.1-C.sub.6)alkylamino,
amino(C.sub.1-C.sub.6)alkyl, or mono- or
di(C.sub.1-C.sub.6)alkylamino(C.- sub.1-C.sub.6)alkyl, or mono- or
di (C.sub.1-C.sub.6) alkylamino (C.sub.1-C.sub.6) alkoxy;
[0079] R.sub.2 represents
[0080] hydroxy;
[0081] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, each of
which is optionally substituted with amino or mono- or
di(C.sub.1-C.sub.6)alkylami- no, C.sub.5-C.sub.7 cycloalkylamino or
C.sub.5-C.sub.7 cycloalkoxy; or
[0082] O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1,2,3,4,
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or C.sub.1-C.sub.6 alkyl; or
[0083] NR.sub.8R.sub.9 forms 5-, 6-, or 7-membered heterocyclic
ring;
[0084] R.sub.3 represents C.sub.1-C.sub.6 alkyl;
[0085] R.sub.4 represents C.sub.1-C.sub.6 alkoxy; or
[0086] R.sub.5 and R6 are the same or different and represent
hydrogen or halogen;
[0087] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, the alkyl
portion of each being optionally substituted with amino, mono- or
di(C.sub.1-C.sub.6) alkylamino, or a C.sub.5-C.sub.7
heterocycloalkyl group where the heteroatom is nitrogen and the
nitrogen is attached to the parent alkyl portion; and
[0088] X represents a bond or CH.sub.2O.
[0089] Preferred compounds of Formula II include those where one of
R.sub.5 and R.sub.6 is hydrogen. Other preferred compounds of II
are those where R4 is methoxy, one of R.sub.5 and R.sub.6 is
hydrogen, and the other of R.sub.5 and R.sub.6 is alkoxy. Still
other preferred compounds of II are those where R.sub.7 is
C.sub.4-C.sub.6 alkyl.
[0090] More preferred compounds of Formula II include those where
wherein X is a bond. Other more preferred compounds of II are those
where R.sub.1 is benzyl monosubstituted in the ortho position.
Particularly preferred compounds of II are those where R.sub.7 is
C.sub.4-C.sub.6 alkyl and X is a bond.
[0091] Other preferred compounds of the invention are represented
by Formula III 5
[0092] wherein:
[0093] R.sub.7 represents C.sub.1-C.sub.6 alkyl;
[0094] R.sub.1 represents C.sub.1-C.sub.6 alkyl, cyclopentyl, or
cyclopropylmethyl;
[0095] R.sub.2 represents
[0096] hydroxy;
[0097] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, each of
which is optionally substituted with amino or mono- or
di(C.sub.1-C.sub.6)alkylami- no, C.sub.5-C.sub.7 cycloalkylamino or
C.sub.5-C.sub.7 cycloalkoxy; or
[0098] O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1, 2, 3 or 4
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or C.sub.1-C.sub.6 alkyl; or
[0099] NR.sub.8R.sub.9 forms 5-, 6-, or 7-membered heterocyclic
ring;
[0100] R.sub.3 represents C.sub.1-C.sub.6 alkyl;
[0101] R.sub.4 represents C.sub.1-C.sub.6 alkoxy; or
[0102] R.sub.4 represents methyl when R.sub.1 and R.sub.7 are lower
alkyl;
[0103] R.sub.5 and R.sub.6 are the same or different and represent
hydrogen or halogen;
[0104] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, the alkyl
portion of each being optionally substituted with amino, mono- or
di(C.sub.1-C.sub.6) alkylamino, or a C.sub.1-C.sub.7
heterocycloalkyl group where the heteroatom is nitrogen and the
nitrogen is attached to the parent alkyl portion;
[0105] O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1, 2, 3 or 4
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or straight or branched chain lower alkyl having 1-6
carbon atoms;
[0106] NR.sub.8R.sub.9 forms a 5-, 6- or 7-membered heterocyclic
ring;
[0107] X represents a bond or CH.sub.2O.
[0108] Preferred compounds of Formula III include those where
R.sub.3 and R.sub.7 are methyl. Other preferred compounds of
Formula III are those where R.sub.1 is propyl or cyclopropylmethyl.
Still other preferred compounds of III are those where R.sub.1 is
cyclopentyl.
[0109] Particularly preferred compounds of Formula III include
those where X is CH.sub.2O. Other particularly preferred compounds
of III are those where R.sub.1 is propyl or cyclopropylmethyl.
Still other particularly preferred compounds of Formula III are
those where R.sub.1 is propyl or cyclopropylmethyl and one of
R.sub.5 and R.sub.6 is hydrogen. Other particularly preferred
compounds of Formula III are those where R.sub.1 is propyl, R.sub.4
is methoxy or methyl, one of R.sub.5 and R.sub.6 is hydrogen, and
the other of R.sub.5 and R.sub.6 is alkoxy.
[0110] Other preferred compounds of the invention are represented
by Formula IV 6
[0111] wherein:
[0112] R.sub.7 represents C.sub.1-C.sub.6 alkyl;
[0113] R.sub.1 represents C.sub.1-C.sub.6 alkyl, cyclopentyl, or
cyclopropylmethyl;
[0114] R.sub.2 represents
[0115] hydroxy;
[0116] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, each of
which is optionally substituted with amino or mono- or
di(C.sub.1-C.sub.6)alkylami- no, C.sub.5-C.sub.7 cycloalkylamino or
C.sub.5-C.sub.7 cycloalkoxy; or
[0117] O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1, 2, 3 or 4
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or C.sub.1-C.sub.6 alkyl; or
[0118] NR.sub.8R.sub.9 forms 5-, 6-, or 7-membered heterocyclic
ring;
[0119] R.sub.3 represents C.sub.1-C.sub.6 alkyl;
[0120] R.sub.4 represents C.sub.1-C.sub.6 alkoxy; or
[0121] R.sub.4 can represent methyl when R.sub.1 and R.sub.7 are
lower alkyl;
[0122] R.sub.5 and R.sub.6 are the same or different and represent
hydrogen or halogen;
[0123] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, the alkyl
portion of each being optionally substituted with amino, mono- or
di(C.sub.1-C.sub.6) alkylamino, or a C.sub.5-C.sub.7
heterocycloalkyl group where the heteroatom is nitrogen and the
nitrogen is attached to the parent alkyl portion;
[0124] O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1,2,3,4,
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or straight or branched chain lower alkyl having 1-6
carbon atoms;
[0125] NR.sub.8R.sub.9 forms a 5-, 6- or 7-membered heterocyclic
ring; X represents a bond or CH.sub.2O.
[0126] Preferred compounds of Formula IV are those where R.sub.3
and R.sub.7 are methyl. Other preferred compounds of IV are those
where R.sub.1 is propyl or cyclopropylmethyl. Still other preferred
compounds of Formula IV are those where R.sub.1 is cyclopentyl. Yet
other preferred compounds of IV include those where X is
CH.sub.2O.
[0127] More preferred compounds of IV are those wherein R.sub.1 is
propyl or cyclopropylmethyl. Particularly preferred compounds of
Formula IV are those where R.sub.1 is propyl or cyclopropylmethyl
and one of R.sub.5 and R.sub.6 is hydrogen. Still other
particularly preferred compounds of IV include those where R.sub.1
is propyl, R.sub.4 is methoxy or methyl, one of R.sub.5 and R.sub.6
is hydrogen, and the other of R.sub.5 and R.sub.6 is alkoxy.
[0128] Other preferred compounds of the invention are represented
by Formula V 7
[0129] where
[0130] R.sub.1 represents benzyl optionally mono-, di-, or
trisubstituted independently with
[0131] halogen, nitro, trifluoromethyl, cyano, hydroxyl,
C.sub.1-C.sub.6 alkoxy, or C.sub.1-C.sub.6 alkyl; or
[0132] amino, mono or di(C.sub.1-C.sub.6)alkylamino,
amino(C.sub.1-C.sub.6)alkyl, or mono- or
di(C.sub.1-C.sub.6)alkylamino(C.- sub.1-C.sub.6)alkyl, or mono- or
di (C.sub.1-C.sub.6) alkylamino (C.sub.1-C.sub.6) alkoxy;
[0133] R.sub.2 represents
[0134] hydroxy;
[0135] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, each of
which is optionally substituted with amino or mono- or
di(C.sub.1-C.sub.6)alkylami- no, C.sub.5-C.sub.7 cycloalkylamino or
C.sub.5-C.sub.7 cycloalkoxy; or
[0136] O(CH.sub.2).sub.nCO.sub.2R.sub.8 where n=1, 2, 3 or 4
NR.sub.8COR.sub.9, COR.sub.8, CONR.sub.8R.sub.9 or CO.sub.2R.sub.8
where R.sub.8 and R.sub.9 are the same or different and represent
hydrogen or C.sub.1-C.sub.6 alkyl; or
[0137] NR.sub.8R.sub.9 forms 5-, 6-, or 7-membered heterocyclic
ring;
[0138] R.sub.3 represents C.sub.1-C.sub.6 alkyl;
[0139] R.sub.4 represents C.sub.1-C.sub.6 alkoxy; or
[0140] R.sub.5 and R.sub.6 are the same or different and represent
hydrogen or halogen;
[0141] C.sub.1-C.sub.6 alkyl or C.sub.1-C.sub.6 alkoxy, the alkyl
portion of each being optionally substituted with amino, mono- or
di(C.sub.1-C.sub.6) alkylamino, or a C.sub.5-C.sub.6
heterocycloalkyl group where the heteroatom is nitrogen and the
nitrogen is attached to the parent alkyl portion; and
[0142] X represents a bond or CH.sub.2O.
[0143] Preferred compounds of Formula V are those where one of
R.sub.5 and R.sub.6 is hydrogen. Other preferred compounds of
Formula V are those where R.sub.4 is methoxy, one of R.sub.5 and
R.sub.6 is hydrogen, and the other of R.sub.5 and R.sub.6 is
alkoxy. More preferred compounds of Formula V include those where
R.sub.7 is C.sub.4-C.sub.6 alkyl.
[0144] Particularly preferred compounds of Formula V include those
where X is a bond. Other particularly preferred compounds of
Formula V are those where R.sub.1 is benzyl monosubstituted in the
ortho position. Still other particularly preferred compounds of
Formula V are those where R.sub.7 is C.sub.4-C.sub.6 alkyl and X is
a bond.
[0145] In certain situations, the compounds of Formula I may
contain one or more asymmetric carbon atoms, so that the compounds
can exist in different stereoisomeric forms. These compounds can
be, for example, racemates or optically active forms. In these
situations, the single enantiomers, i.e., optically active forms,
can be obtained by asymmetric synthesis or by resolution of the
racemates. Resolution of the racemates can be accomplished, for
example, by conventional methods such as crystallization in the
presence of a resolving agent, or chromatography, using, for
example a chiral HPLC column.
[0146] Representative compounds of the present invention, which are
encompassed by Formula I, include, but are not limited to the
compounds described in the Examples and their pharmaceutically
acceptable acid addition salts. In addition, if the compound of the
invention is obtained as an acid addition salt, the free base can
be obtained by basifying a solution of the acid salt. Conversely,
if the product is a free base, an addition salt, particularly a
pharmaceutically acceptable addition salt, may be produced by
dissolving the free base in a suitable organic solvent and treating
the solution with an acid, in accordance with conventional
procedures for preparing acid addition salts from base
compounds.
[0147] Non-toxic pharmaceutical salts include salts of acids such
as hydrochloric, phosphoric, hydrobromic, sulfuric, sulfinic,
formic, toluenesulfonic, methanesulfonic, nitric, benzoic, citric,
tartaric, maleic, hydroiodic, alkanoic such as acetic,
HOOC--(CH.sub.2)n--COOH where n is 0-4, and the like. Those skilled
in the art will recognize a wide variety of non-toxic
pharmaceutically acceptable addition salts.
[0148] The present invention also encompasses the acylated prodrugs
of the compounds of Formula I. Those skilled in the art will
recognize various synthetic methodologies which may be employed to
prepare non-toxic pharmaceutically acceptable addition salts and
acylated prodrugs of the compounds encompassed by Formula I.
[0149] By "alkyl" or "lower alkyl" in the present invention is
meant C.sub.1-C.sub.6 alkyl, i.e., straight or branched chain alkyl
groups having 1-6 carbon atoms, such as, for example, methyl,
ethyl, propyl, isopropyl, n-butyl, sec-butyl, tert-butyl, pentyl,
2-pentyl, isopentyl, neopentyl, hexyl, 2-hexyl, 3-hexyl, and
3-methylpentyl. Preferred C.sub.1-C.sub.6 alkyl groups are methyl,
ethyl, propyl, butyl, cyclopropyl or cyclopropylmethyl.
[0150] By "alkoxy" or "lower alkoxy" in the present invention is
meant C.sub.1-C.sub.6 alkoxy, i.e., straight or branched chain
alkoxy groups having 1-6 carbon atoms, such as, for example,
methoxy, ethoxy, propoxy, isopropoxy, n-butoxy, sec-butoxy,
tert-butoxy, pentoxy, 2-pentyl, isopentoxy, neopentoxy, hexoxy,
2-hexoxy, 3-hexoxy, and 3-methylpentoxy.
[0151] By (hetero) cyclic ring is meant a ring that is either
aliphatic or aromatic and optionally contains at least one hetero
atom. Hetero atoms include nitrogen, sulfur, and oxygen. Examples
of such (hetero) cyclic rings are cyclohexyl, cyclopentyl,
piperidinyl, piperazinyl, pyrrolidinyl, morpholinyl, etc.
[0152] By heteroaryl (aromatic heterocycle) in the present
invention is meant one or more aromatic ring systems of 5-, 6-, or
7-membered rings containing at least one and up to four hetero
atoms selected from nitrogen, oxygen, or sulfur. Such heteroaryl
groups include, for example, thienyl, furanyl, thiazolyl,
imidazolyl, (is)oxazolyl, pyridyl, pyrimidinyl, imidazolyl,
(iso)quinolinyl, naphthyridinyl, benzimidazolyl, and
benzoxazolyl.
[0153] As used herein, each alkyl group in a
di(C.sub.1-C.sub.6)alkylamino group is independent of the other.
Specific examples of heteroaryl groups are the following: 8
[0154] wherein
[0155] L is nitrogen or --CR.sup.11;
[0156] T is --NR.sup.19, oxygen, or sulfur;
[0157] R.sup.11 and R.sup.11 are the same or different and are
selected from hydrogen, halogen, hydroxy, C.sub.1-C.sub.6 alkyl,
(C.sub.1-C.sub.6)alkoxy, amino, or mono- or
di(C.sub.1-C.sub.6)alkylamino- ;
[0158] R.sup.12, R.sup.121, and R.sup.13 are the same or different
and are selected from hydrogen, halogen, (C.sub.1-C.sub.6)alkyl,
(C.sub.1-C.sub.6)alkoxy, amino, mono- or
di(C.sub.1-C.sub.6)alkylamino, hydroxy, or trifluoromethyl; and
[0159] R.sup.19 is hydrogen, lower alkyl having 1-6 carbon
atoms.
[0160] The invention includes all possible tautomers and rotamers
of the compounds represented by Formula I.
[0161] By the term "halogen" in the present invention is meant
fluorine, bromine, chlorine, and iodine.
[0162] The compounds of general Formula I may be administered
orally, topically, parenterally, by inhalation or spray or rectally
in dosage unit formulations containing conventional non-toxic
pharmaceutically acceptable carriers, adjuvants and vehicles. The
term parenteral as used herein includes subcutaneous injections,
intravenous, intramuscular intrasternal injection or infusion
techniques. In addition, there is provided a pharmaceutical
formulation comprising a compound of general Formula I and a
pharmaceutically acceptable carrier. One or more compounds of
general Formula I may be present in association with one or more
non-toxic pharmaceutically acceptable carriers and/or diluents
and/or adjuvants and if desired other active ingredients. The
pharmaceutical compositions containing compounds of general Formula
I may be in a form suitable for oral use, for example, as tablets,
troches, lozenges, aqueous or oily suspensions, dispersible powders
or granules, emulsion, hard or soft capsules, or syrups or
elixirs.
[0163] Compositions intended for oral use may be prepared according
to any method known to the art for the manufacture of
pharmaceutical compositions and such compositions may contain one
or more agents selected from the group consisting of sweetening
agents, flavoring agents, coloring agents and preserving agents in
order to provide pharmaceutically elegant and palatable
preparations. Tablets contain the active ingredient in admixture
with non-toxic pharmaceutically acceptable excipients which are
suitable for the manufacture of tablets. These excipients may be
for example, inert diluents, such as calcium carbonate, sodium
carbonate, lactose, calcium phosphate or sodium phosphate;
granulating and disintegrating agents, for example, corn starch, or
alginic acid; binding agents, for example starch, gelatin or
acacia, and lubricating agents, for example magnesium stearate,
stearic acid or talc. The tablets may be uncoated or they may be
coated by known techniques to delay disintegration and absorption
in the gastrointestinal tract and thereby provide a sustained
action over a longer period. For example, a time delay material
such as glyceryl monosterate or glyceryl distearate may be
employed.
[0164] Formulations for oral use may also be presented as hard
gelatin capsules wherein the active ingredient is mixed with an
inert solid diluent, for example, calcium carbonate, calcium
phosphate or kaolin, or as soft gelatin capsules wherein the active
ingredient is mixed with water or an oil medium, for example peanut
oil, liquid paraffin or olive oil.
[0165] Aqueous suspensions contain the active materials in
admixture with excipients suitable for the manufacture of aqueous
suspensions. Such excipients are suspending agents, for example
sodium carboxymethylcellulose, methylcellulose,
hydropropylmethylcellulose, sodium alginate, polyvinylpyrrolidone,
gum tragacanth and gum acacia; dispersing or wetting agents may be
a naturally-occurring phosphatide, for example, lecithin, or
condensation products of an alkylene oxide with fatty acids, for
example polyoxyethylene stearate, or condensation products of
ethylene oxide with long chain aliphatic alcohols, for example
heptadecaethyleneoxycetanol, or condensation products of ethylene
oxide with partial esters derived from fatty acids and a hexitol
such as polyoxyethylene sorbitol monooleate, or condensation
products of ethylene oxide with partial esters derived from fatty
acids and hexitol anhydrides, for example polyethylene sorbitan
monooleate. The aqueous suspensions may also contain one or more
preservatives, for example ethyl, or n-propyl p-hydroxybenzoate,
one or more coloring agents, one or more flavoring agents, and one
or more sweetening agents, such as sucrose or saccharin.
[0166] Oily suspensions may be formulated by suspending the active
ingredients in a vegetable oil, for example arachis oil, olive oil,
sesame oil or coconut oil, or in a mineral oil such as liquid
paraffin. The oily suspensions may contain a thickening agent, for
example beeswax, hard paraffin or cetyl alcohol. Sweetening agents
such as those set forth above, and flavoring agents may be added to
provide palatable oral preparations. These compositions may be
preserved by the addition of an anti-oxidant such as ascorbic
acid.
[0167] Dispersible powders and granules suitable for preparation of
an aqueous suspension by the addition of water provide the active
ingredient in admixture with a dispersing or wetting agent,
suspending agent and one or more preservatives. Suitable dispersing
or wetting agents and suspending agents are exemplified by those
already mentioned above. Additional excipients, for example
sweetening, flavoring and coloring agents, may also be present.
[0168] Pharmaceutical compositions of the invention may also be in
the form of oil-in-water emulsions. The oily phase may be a
vegetable oil, for example olive oil or arachis oil, or a mineral
oil, for example liquid paraffin or mixtures of these. Suitable
emulsifying agents may be naturally-occurring gums, for example gum
acacia or gum tragacanth, naturally-occurring phosphatides, for
example soy bean, lecithin, and esters or partial esters derived
from fatty acids and hexitol, anhydrides, for example sorbitan
monoleate, and condensation products of the said partial esters
with ethylene oxide, for example polyoxyethylene sorbitan
monoleate. The emulsions may also contain sweetening and flavoring
agents.
[0169] Syrups and elixirs may be formulated with sweetening agents,
for example glycerol, propylene glycol, sorbitol or sucrose. Such
formulations may also contain a demulcent, a preservative and
flavoring and coloring agents. The pharmaceutical compositions may
be in the form of a sterile injectable aqueous or oleaginous
suspension. This suspension may be formulated according to the
known art using those suitable dispersing or wetting agents and
suspending agents which have been mentioned above. The sterile
injectable preparation may also be sterile injectable solution or
suspension in a non-toxic parentally acceptable diluent solvent,
for example as a solution in 1,3-butanediol. Among the acceptable
vehicles and solvents that may be employed are water, Ringer's
solution and isotonic sodium chloride solution. In addition,
sterile, fixed oils are conventionally employed as a solvent or
suspending medium. For this purpose any bland fixed oil may be
employed including synthetic mono- or diglycerides. In addition,
fatty acids such as oleic acid find use in the preparation of
injectables.
[0170] The compounds of general Formula I may also be administered
in the form of suppositories for rectal administration of the drug.
These compositions can be prepared by mixing the drug with a
suitable non-irritating excipient which is solid at ordinary
temperatures but liquid at the rectal temperature and will
therefore melt in the rectum to release the drug. Such materials
are cocoa butter and polyethylene glycols.
[0171] Compounds of general Formula I may be administered
parenterally in a sterile medium. The drug, depending on the
vehicle and concentration used, can either be suspended or
dissolved in the vehicle. Advantageously, adjuvants such as local
anesthetics, preservatives and buffering agents can be dissolved in
the vehicle.
[0172] Dosage levels of the order of from about 0.1 mg to about 140
mg per kilogram of body weight per day are useful in the treatment
of the above-indicated conditions (about 0.5 mg to about 7 g per
patient per day). The amount of active ingredient that may be
combined with the carrier materials to produce a single dosage form
will vary depending upon the host treated and the particular mode
of administration. Dosage unit forms will generally contain between
from about 1 mg to about 500 mg of an active ingredient.
[0173] Frequency of dosage may also vary depending on the compound
used and the particular disease treated. However, for treatment of
most disorders, a dosage regimen of 4 times daily or less is
preferred. For the treatment of obesity or diabetes, a dosage
regimen of 1 or 2 times daily is particularly preferred.
[0174] It will be understood, however, that the specific dose level
for any particular patient will depend upon a variety of factors
including the activity of the specific compound employed, the age,
body weight, general health, sex, diet, time of administration,
route of administration, and rate of excretion, drug combination
and the severity of the particular disease undergoing therapy.
[0175] Preferred compounds of the invention will have certain
pharmacological properties. Such properties include, but are not
limited to oral bioavailability, low toxicity, low serum protein
binding and desirable in vitro and in vivo half-lifes. Penetration
of the blood brain barrier for compounds used to treat CNS
disorders is necessary, while low brain levels of compounds used to
treat periphereal disorders are often preferred.
[0176] Assays may be used to predict these desirable
pharmacological properties. Assays used to predict bioavailability
include transport across human intestinal cell monolayers,
including Caco-2 cell monolayers. Toxicity to cultured hepatocyctes
may be used to predict compound toxicity. Penetration of the blood
brain barrier of a compound in humans may be predicted from the
brain levels of the compound in laboratory animals given the
compound intravenously.
[0177] Serum protein binding may be predicted from albumin binding
assays. Such assays are described in a review by Oravcov, et al.
(Journal of Chromatography B (1996) volume 677, pages 1-27).
[0178] Compound half-life is inversely proportional to the
frequency of dosage of a compound. In vitro half-lifes of compounds
may be predicted from assays of microsomal half-life as described
by Kuhnz and Gieschen (Drug Metabolism and Disposition, (1998)
volume 26, pages 1120-1127).
[0179] The present invention also pertains to packaged
pharmaceutical compositions for treating disorders responsive to
GLP receptor modulation, e.g., treatment obesity or diabetes. The
packaged pharmaceutical compositions include a container holding a
therapeutically effective amount of at least one compound of
Formula I supra and instructions for using the treating disorder
responsive to GLP receptor modulation in the patient.
[0180] The disclosures of all articles and references mentioned in
this application, including patents, are incorporated herein by
reference.
[0181] Compounds of the invention can be prepared using the
reactions depicted in Schemes I to VII. The numbers appearing below
or adjacent the chemical structures in these schemes refer to
intermediates and are not to be confused with the compound numbers
found in the examples. 9 10 11 12 13 14 15
[0182] Those having skill in the art will recognize that starting
materials may be varied and additional steps employed to produce
compounds encompassed by the present invention, as demonstrated by
the following examples.
[0183] The following examples illustrate the general procedures for
the preparation of compounds of the invention using the reactions
outlined above in Schemes I-VII. These examples are not to be
construed as limiting the invention in scope or spirit to the
specific procedures and compounds described in them.
EXAMPLE 1
General Procedure for the Preparation of Chloromethylbenzimidazoles
as Outlined in Scheme I
[0184] 1. Imidate Hydrochloride:
[0185] A solution of 150 mL (2.37 mole) of chloroacetonitrile, 139
mL (2.37 mole) of ethanol in 1,200 mL of dry benzene is cooled to
0.degree. C. in an ice/ethanol bath. Dry HCl gas is bubbled through
the vigorously stirred solution for approximately 30 min. while the
internal temperature is maintained below 10.degree. C. The solution
is allowed to stand at rt. overnight. The resulting solid is
filtered and washed with 2L of dry ether and allowed to air dry to
afford 328 g (88%) of imidate hydrochloride.
[0186] 2.2-(chloromethyl)-1-[(2-methylphenyl)methyl]benzimidazole:
16
[0187] A solution of 27 g (0.13 mole) of (2-aminophenyl)
(2-methylphenyl)amine in 200 mL of anhydrous CHCl.sub.3 is treated
with 30.81 g (0.19 mole) of imidate at room temperature. The
heterogeneous reaction mixture is allowed to stir for 1 hr. at room
temperature at which time no starting material is detectable by
TLC. 100 mL of saturated NaHCO.sub.3 is added and extracted
3.times.100 mL of CH.sub.2Cl.sub.2. The extracts are dried over
anhydrous Na.sub.2SO.sub.4, the solvent removed in vacuo, and the
residue chromatgraphed (SiO.sub.2) with 50% ethyl acetate/hexane to
afford 22 9 (62%) of 2-(chloromethyl)-1-[(2-methy- lphenyl)methyl]
benzimidazole. Mass Spec M.sup.+271.
[0188] 3.2-(chloroethyl)-1-propylbenzimidazole 17
[0189] A solution of 8 9 (0.053 mole) of (2-aminophenyl)propylamine
in 50 mL of anhydrous DMF is treated with 9.0 g (0.056 mole) of
2-chloro-1-ethoxypropanimine hydrochloride at 80.degree. C. for 16
hr. The reaction mixture is cooled to room temperature diluted with
200 mL of ethyl acetate and washed 3.times.100 mL water,
1.times.100 mL brine, organic extracts are dried over anhydrous
Na.sub.2SO.sub.4, the solvent removed in vacuo, and the residue
chromatgraphed (SiO.sub.2) with 30% ethyl acetate/hexane to afford
3 g (28%) of 2-(chloroethyl)-1-propylbenzi- midazole.
EXAMPLE 2
General Procedure for the Preparation of Benzimidazoles as Shown in
Scheme II
[0190]
((2,4-dimethoxyphenyl)-N-(3-methylbutyl)-N-({1-[(2-methylphenyl)met-
hyl]benzimidazol-2-yl}methyl)carboxamide 18
[0191] A solution of 5.4 mmole
2-(chloromethyl)-1-[(2-methylphenyl)methyl] benzimidazole in 20 mL
of dry Acetonitrile is treated with 10 mL of isoamylamine for 16 hr
at room temperature. The solvent is removed in vacuo and the
residue is partitioned between 30 mL of ethyl acetate and 10 mL of
1 N NaOH. The ethyl acetate layer is dried over anhydrous
Na.sub.2SO.sub.4 and solvent removed in vacuo to afford 1.6 g 92%
({1-[(2-methylphenyl)methyl]benzimidazol-2-yl}methyl)(3-methylbutyl)
amine. 2,4-dimethoxybenzoylchloride 1.5 eq is treated with 1.0 eq
of
({1-[2-methylphenyl)methyl]benzimidazol-2-yl}methyl)(3-methylbutyl)
amine in dichloromethane at room temperature for 1 hr. The reaction
is quenched with 1 N NaOH and partitioned between dichloromethane
and water. The organic layer is dried with Na.sub.2SO.sub.4 and the
solvent removed in vacuo. The residue is chromatographed
(SiO.sub.2) with ethyl acetate to afford 95% of
(2,4-dimethoxyphenyl)-N-({1-[(2-methylphenyl)methyl]benzimi-
dazol-2-yl}methyl)-N-(3-methylbutyl)carboxamide (Compound 1). Mass
Spec M.sup.+ 486.
[0192] Following the above procedures, compounds 11, 12 and 15, 16
are prepared starting from 2-(chloromethyl)imidazolo[5,4-b]pyridine
and 2-(chloromethyl)imidazolo[5,4-c]pyridine respectivly. (Cleve,
G: Gibian, H.; Hoyer, G.; Rahtz, D.; Schroeder, E.; Schulz, G.
Justus Liebigs Ann. Chem. 1971, 747, 158-171).
EXAMPLE 3
[0193] The following compounds are prepared essentially according
to the procedure described in Examples 1-2, and as shown in Schemes
I-VII:
[0194] (a) 2- (2,3-dimethylphenoxy)-N-methyl-N-
[(1-propylbenzimidazol-2-y- l)ethyl]acetamide M.sup.+ 381 amu.
(Compound 2) 19
[0195] (b)
2-(2,3-dimethylphenoxy)-N-{[1-(cyclopropylmethyl)benzimidazol-2-
-yl]ethyl}-N-methylacetamide M.sup.+ 393 amu. (Compound 3) 20
[0196] (c)
(2,4-dimethoxyphenyl)-N-({-[(2-chlorophenyl)methyl]benzimidazol-
-2-yl}methyl)-N-(3-methylbutyl)carboxamide M.sup.+ 507 amu.
(Compound 4) 21
[0197] (d)
(2,4-dimethoxyphenyl)-N-(3-methylbutyl)-N-({3-[(2-methylphenyl)
methyl]imidazolo[5,4-b]pyridin-2-yl}methyl)carboxamide M.sup.+ 488
amu. (Compound 5) 22
[0198] (e) 2-
(2,3-dimethylphenoxy)-N-methyl-N-[(3-propylimidazolo[5,4-b]p-
yridin-2-yl)ethyl] acetamide M.sup.+ 381 amu. (Compound 6) 23
[0199] (f)
2-(2,3-dimethylphenoxy)-N-{[1-(cyclopentyl)-6-chlorobenzimidazo-
l-2-yl]ethyl}-N-methylacetamide M.sup.+ 441 amu. (Compound 7)
24
EXAMPLE 4
[0200] Assay for Glucagon-Like Peptide Receptor Activation
[0201] The following assay may be used to quantitate the effects of
compounds on GLP-1 activity.
[0202] Glucagon-like peptide 1 receptor activation is carried out
by measuring the second messenger cyclic adenosine monophosphate
(cAMP). The host cell for such studies can be any that endogenously
expresses the Glp-1 receptor, such as the Rin M5F or HIT--TI5
insulinomas, or a cell line that expresses the recombinant form of
that receptor. For this purpose, the appropriate cells are plated
in the either 24 or 96 well plates and the cells are grown to a 75
to 95% level of confluence. Cells are usually plated 24 to 48 hours
prior to assay. Immediately prior to the receptor activation study,
the cells are rinsed with a phosphate buffered saline solution, and
cells are incubated with from 1 to 10 mM isobutylmethylxanthine,
(IBMX). The purpose of IBMX is to inhibit the enzyme cAMP
phosphodiesterase, which breaks down cAMP. The use of IBMX allows
one to more easily detect the ability of a hormone or drug to
activate the Glp-1 receptor. In a typical assay, either the Glp-1
peptide or drug is added directly to the 24 or 96 well plate, and
is incubated with the cells for up to 60 minutes at 37.degree. C.
After the desired time, the receptor activation event is terminated
by the addition of hydrochloric acid to the cells, which also lyses
the cells and liberates the cAMP that has accumulated within the
cells. This cellular extract is harvested and neutralized with
sodium hydroxide, and the cells lysates are cleared by
microcentrifugation. The cell extract is then analyzed in a cAMP
radioimmunoassay, such as that commercially available from NEN Life
Science Products or Amersham. The amount of cAMP generated per well
of cells treated with hormone or drug can be compared to that
observed without the addition of such agent, to obtain an index of
receptor activation. Dose-response curves are also performed to
obtain the level of potency and efficacy of any test compound.
[0203] In the described assay, preferred compounds of the invention
will have 5% or greater stimulation with respect to GLP-1.
EXAMPLE 5
Glucose Tolerance Test
[0204] Following overnight fasting adult male (200-300 g)
Sprague-Dawley rats are injected orally with either vehicle or
glucose solution in a given concentration. Following thirty-five
minutes of resting in their home cages, the animals are brought
back into the laboratory and restrained using a BRAINTREE
SCIENTIFIC adjustable restrainer. Within five minutes of restraint,
one of the lateral tail veins is catheterized, and the animals are
given intravenous (iv) injection of either glucagon-like
polypeptide 1 (GLP1) or test compound. Five minutes after iv
injection, the animals are euthanized by decapitation, and trunk
blood is collected in tubes containing EDTA. The plasma levels of
insulin and glucose measured using appropriate radio-immunoassay
(RIA) kits.
EXAMPLE 6
Streptozocin-Induced Diabetes Glucose Tolerance Test
[0205] Streptozotocin is an antibiotic extracted from Streptomyces
achromogenes, which when injected into animals, causes pancreatic
.beta.-cell degranulation and necrosis. To achieve mild necrosis of
pancreatic .beta.-cells, which induces a state of diabetes without
affecting normal development and weight gain, a 35 mg/kg/5 ml dose
STZ is injected intraperitoneally (ip) into a group of healthy,
naive animals. The control group animals receive 0.1 N citrate
buffer (vehicle, 5 ml/kg, 10=16). Five days after ip injections
(day5), the diabetic symptoms are assessed via the following test
of glucose tolerance. All animals receive an oral injection of 3
g/kg/10 ml glucose solution between 3:00 and 5:00 PM. Forty minutes
later, their blood glucose levels are measured using the LIFE SCAN
ONE TOUCH glucose monitoring system. Animals are restrained and a
blood sample is taken from a lateral tail vein. Animals that show
substantially higher blood glucose levels (100 to 250% higher than
non-STZ treated animals, normally 2/3 of STZ treated animals) are
used to assess the effects of test compounds in this animal model
of diabetes. On day 7, following overnight fast, the animals are
subjected to glucose tolerance test using a procedure identical to
that described above. The test compound is injected iv or orally
and IV injection of GLP-1 is used as a positive control.
EXAMPLE 7
[0206] Preparation of Radiolabeled Probe Compounds of the
Invention
[0207] The compounds of the invention are prepared as radiolabeled
probes by carrying out their synthesis using precursors comprising
at least one atom that is a radioisotope. The radioisotope is
preferably selected from of at least one of carbon (preferably
.sup.14C), hydrogen (preferably .sup.3H), sulfur (preferably
.sup.35S), or iodine (preferably .sup.125I). Such radiolabeled
probes are conveniently synthesized by a radioisotope supplier
specializing in custom synthesis of radiolabeled probe compounds.
Such suppliers include Amersham Corporation, Arlington Heights,
Ill.; Cambridge Isotope Laboratories, Inc. Andover, Mass.; SRI
International, Menlo Park, Calif.; Wizard Laboratories, West
Sacramento, Calif.; ChemSyn Laboratories, Lexena, Kans.; American
Radiolabeled Chemicals, Inc., St. Louis, Mo.; and Moravek
Biochemicals Inc., Brea, Calif.
[0208] Tritium labeled probe compounds are also conveniently
prepared catalytically via platinum-catalyzed exchange in tritiated
acetic acid, acid-catalyzed exchange in tritiated trifluoroacetic
acid, or heterogeneous-catalyzed exchange with tritium gas. Such
preparations are also conveniently carried out as a custom
radiolabeling by any of the suppliers listed in the preceding
paragraph using the compound of the invention as substrate. In
addition, certain precursors may be subjected to tritium-halogen
exchange with tritium gas, tritium gas reduction of unsaturated
bonds, or reduction using sodium borotritide, as appropriate.
EXAMPLE 8
Receptor Autoradiography
[0209] Receptor autoradiography (receptor mapping) is carried out
in vitro as described by Kuhar in sections 8.1.1 to 8.1.9 of
Current Protocols in Pharmacology (1998) John Wiley & Sons, New
York, using radiolabeled compounds of the invention prepared as
described in the preceding Example.
[0210] The invention and the manner and process of making and using
it, are now described in such full, clear, concise and exact terms
as to enable any person skilled in the art to which it pertains, to
make and use the same. It is to be understood that the foregoing
describes preferred embodiments of the present invention and that
modifications may be made therein without departing from the spirit
or scope of the present invention as set forth in the claims. To
particularly point out and distinctly claim the subject matter
regarded as invention, the following claims conclude this
specification.
* * * * *