U.S. patent application number 10/963289 was filed with the patent office on 2005-03-17 for glucagon antagonists/inverse agonists.
This patent application is currently assigned to Novo Nordisk A/S. Invention is credited to Jorgensen, Anker Steen, Madsen, Peter.
Application Number | 20050059611 10/963289 |
Document ID | / |
Family ID | 27222459 |
Filed Date | 2005-03-17 |
United States Patent
Application |
20050059611 |
Kind Code |
A1 |
Jorgensen, Anker Steen ; et
al. |
March 17, 2005 |
Glucagon antagonists/inverse agonists
Abstract
A novel class of compounds, which act to antagonize the action
of the glucagon hormone on the glucagon receptor. Owing to their
antagonizing effect of the glucagon receptor the compounds may be
suitable for the treatment and/or prevention of any diseases and
disorders, wherein a glucagon antagonistic action is beneficial,
such as hyperglycemia, Type 1 diabetes, Type 2 diabetes, disorders
of the lipid metabolism and obesity.
Inventors: |
Jorgensen, Anker Steen;
(Kobenhavn O, DK) ; Madsen, Peter; (Bagsvaerd,
DK) |
Correspondence
Address: |
NOVO NORDISK, INC.
PATENT DEPARTMENT
100 COLLEGE ROAD WEST
PRINCETON
NJ
08540
US
|
Assignee: |
Novo Nordisk A/S
Bagsvaerd
DK
|
Family ID: |
27222459 |
Appl. No.: |
10/963289 |
Filed: |
October 12, 2004 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10963289 |
Oct 12, 2004 |
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09995987 |
Nov 16, 2001 |
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6821960 |
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60252322 |
Nov 20, 2000 |
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Current U.S.
Class: |
514/364 ;
514/11.7; 514/15.7; 514/4.8; 514/563; 514/616; 514/7.3;
514/7.4 |
Current CPC
Class: |
C07C 275/30 20130101;
C07D 319/20 20130101; C07D 209/48 20130101; C07D 271/06 20130101;
C07C 311/47 20130101; C07D 319/08 20130101; C07C 275/42 20130101;
C07C 275/28 20130101; C07C 2601/14 20170501; C07C 317/42 20130101;
C07C 323/44 20130101; C07C 275/36 20130101; C07C 2601/16 20170501;
C07D 333/36 20130101; C07C 275/34 20130101; C07D 257/04
20130101 |
Class at
Publication: |
514/019 ;
514/616; 514/563 |
International
Class: |
A61K 038/04; A61K
031/16; A61K 031/195 |
Foreign Application Data
Date |
Code |
Application Number |
Nov 17, 2000 |
DK |
2000 01733 |
Claims
1. A method of promoting glucagon antagonistic action in a subject
comprising administering to the subject a glucagon antagonist
composition consisting essentially of (a) a compound of the general
formula (I): 48wherein R.sup.2 is hydrogen or C.sub.1-6-alkyl, Z is
arylene or a divalent radical derived from a 5 or 6 membered
heteroaromatic ring containing 1 or 2 heteroatoms selected from
nitrogen, oxygen and sulfur, which may optionally be substituted
with one or two groups R.sup.7 and R.sup.8 selected from halogen,
--CN, --CF.sub.3, --OCF.sub.3, --NO.sub.2, --OR.sup.9,
--NR.sup.9R.sup.10 and C.sub.1-6-alkyl, wherein R.sup.9 and
R.sup.10 independently are hydrogen or C.sub.1-6-alkyl, X is
49wherein r is 0 or 1, q and s independently are 0, 1, 2 or 3,
R.sup.11, R.sup.12, R.sup.13 and R.sup.14 independently are
hydrogen or C.sub.1-6-alkyl, D is 50wherein R.sup.15, R.sup.16,
R.sup.17 and R.sup.18 independently are hydrogen, halogen, --CN,
--CH.sub.2CN, --CHF.sub.2, --CF.sub.3, --OCF.sub.3, --OCHF.sub.2,
--OCH.sub.2CF.sub.3, --OCF.sub.2CHF.sub.2, --S(O).sub.2CF.sub.3,
--SCF.sub.3, --NO.sub.2, --OR.sup.21, NR.sub.21R.sup.22,
--SR.sup.21, --NR.sup.21S(O)R.sup.22,
--S(O).sub.2NR.sup.21R.sup.22, --S(O)NR.sup.21R.sup.22,
--S(O)R.sup.21, --S(O).sub.2R.sup.21, --C(O)NR.sup.21R.sup.22,
--OC(O)N.sup.21R.sup.22, --NR.sup.21C(O)R.sup.22,
--CH.sub.2C(O)NR.sup.21R.sup.22, --OCH.sub.2C(O)NR.sup.21R.sup.22,
--CH.sub.2OR.sup.21, --CH.sub.2NR.sup.21R.sup.22, --OC(O)R.sup.21,
--C(O)R.sup.21 or --C(O)OR.sup.21, C.sub.1-6-alkyl,
C.sub.2-6-alkenyl or C.sub.2-6-alkynyl, which may optionally be
substituted with one or more substituents selected from halogen,
--CN, --CF.sub.3, --OCF.sub.3, --NO.sub.2, --OR.sup.21,
--NR.sup.21R.sup.22 and C.sub.1-6-alkyl, C.sub.3-8-cycloalkyl,
C.sub.4-8-cycloalkenyl, heterocyclyl,
C.sub.3-8-cycloalkyl-C.sub.1-6-alkyl,
C.sub.3-8-cyclo-alkyl-C.sub.1-6-alk- oxy, C.sub.3-8-cycloalkyloxy,
C.sub.3-8-cycloalkyl-C.sub.1-6-alkylthio, C.sub.3-8-cycloalkylthio,
C.sub.3-8-cycloalkyl-C.sub.2-6-alkenyl,
C.sub.3-8-cycloalkyl-C.sub.2-6-alkynyl,
C.sub.4-8-cycloalkenyl-C.sub.1-6-- alkyl,
C.sub.4-8-cycloalkenyl-C.sub.2-6-alkenyl,
C.sub.4-8-cycloalkenyl-C.- sub.2-6-alkynyl,
heterocyclyl-C.sub.1-6-alkyl, heterocyclyl-C.sub.2-6-alke- nyl,
heterocyclyl-C.sub.2-6-alkynyl, aryl, aryloxy, aryloxycarbonyl,
aroyl, aryl-C.sub.1-6-alkoxy, aryl-C.sub.1-6-alkyl,
aryl-C.sub.2-6-alkenyl, aryl-C.sub.2-6-alkynyl, heteroaryl,
heteroaryl-C.sub.2-6-alkyl, heteroaryl-C.sub.2-6-alkenyl or
heteroaryl-C.sub.2-6-alkynyl, of which the cyclic moieties
optionally may be substituted with one or more substituents
selected from halogen, --CN, --CF.sub.3, --OCF.sub.3, --NO.sub.2,
--OR.sup.21, --NR.sup.21R.sup.22 and C.sub.1-6-alkyl, wherein
R.sup.21 and R.sup.22 independently are hydrogen, C.sub.1-6-alkyl
or aryl, or R.sup.21 and R.sup.22 when attached to the same
nitrogen atom together with the said nitrogen atom may form a 3 to
8 membered heterocyclic ring optionally containing one or two
further heteroatoms selected from nitrogen, oxygen and sulfur, and
optionally containing one or two double bonds, or two of the groups
R.sup.15 to R.sup.18 when placed in adjacent positions together may
form a bridge
--(CR.sup.23R.sup.24).sub.a--O--(CR.sup.25R.sup.26).sub.c--O--,
wherein a is 0, 1 or 2, c is 1 or 2, R.sup.23, R.sup.24, R.sup.25
and R.sup.26 independently are hydrogen, C.sub.1-6-alkyl or
fluorine, R.sup.19 and R.sup.20 independently are hydrogen,
C.sub.1-6-alkyl, C.sub.3-8-cycloalkyl or
C.sub.3-8-cyclo-alkyl-C.sub.1-6-alkyl, E is 51wherein R.sup.27 and
R.sup.28 independently are hydrogen, halogen, --CN, --CF.sub.3,
--OCF.sub.3, --OR.sup.32, --NR.sup.32R.sup.33, C.sub.1-6-alkyl,
C.sub.3-8-cycloalkyl, C.sub.4-8-cycloalkenyl or aryl, wherein the
cyclic moieties optionally may be substituted with one or more
substituents selected from halogen, --CN, --CF.sub.3, --OCF.sub.3,
--NO.sub.2, --OR.sup.32, --NR.sup.32R.sup.33 and C.sub.1-6-alkyl,
wherein R.sup.32 and R.sup.33 independently are hydrogen or
C.sub.1-6-alkyl, or R.sup.32 and R.sup.33 when attached to the same
nitrogen atom together with the said nitrogen atom may form a 3 to
8 membered heterocyclic ring optionally containing one or two
further heteroatoms selected from nitrogen, oxygen and sulfur, and
optionally containing one or two double bonds, R.sup.2, R.sup.30
and R.sup.3 lindependently are hydrogen, halogen, --CHF.sub.2,
--CF.sub.3, --OCF.sub.3, --OCHF.sub.2, --OCH.sub.2CF.sub.3,
--OCF.sub.2CHF.sub.2, --SCF.sub.3, --OR.sup.34,
--NR.sup.34R.sup.35, --SR.sup.34, --S(O)R.sup.34,
--S(O).sub.2R.sup.34, --C(O)NR.sup.34R.sup.35,
--OC(O)NR.sup.34R.sup.35, --NR.sup.34C(O)R.sup.35,
--OCH.sub.2C(O)NR.sup.34R.sup.35, --C(O)R.sup.34 or
--C(O)OR.sup.34, C.sub.1-6-alkyl, C.sub.2-6-alkenyl or
C.sub.2-6-alkynyl, which may optionally be substituted with one or
more substituents selected from halogen, --CN, --CF.sub.3,
--OCF.sub.3, --NO.sub.2, --OR.sup.34, --NR.sup.34R.sup.35 and
C.sub.1-6-alkyl, C.sub.3-8-cycloalkyl, C.sub.4-8-cycloalkenyl,
heterocyclyl, C.sub.3-8-cycloalkyl-C.sub.1-6-alkyl,
C.sub.3-8-cyclo-alkyl-C.sub.2-6-alk- enyl,
C.sub.3-8-cycloalkyl-C.sub.2-6-alkynyl,
C.sub.4-8-cycloalkenyl-C.sub- .1-6-alkyl,
C.sub.4-8-cycloalkenyl-C.sub.2-6-alkenyl,
C.sub.4-8-cycloalkenyl-C.sub.2-6-alkynyl,
heterocyclyl-C.sub.1-6-alkyl, heterocyclyl-C.sub.2-6-alkenyl,
heterocyclyl-C.sub.2-6-alkynyl, aryl, aryloxy, aroyl,
aryl-C.sub.1-6-alkoxy, aryl-C.sub.1-6-alkyl,
aryl-C.sub.2-6-alkenyl, aryl-C.sub.2-6-alkynyl, heteroaryl,
heteroaryl-C.sub.1-6-alkyl, heteroaryl-C.sub.2-6-alkenyl or
heteroaryl-C.sub.2-6-alkynyl, of which the cyclic moieties
optionally may be substituted with one or more substituents
selected from halogen, --CN, --CF.sub.3, --OCF.sub.3, --NO.sub.2,
--OR.sup.34, --NR.sup.34R.sup.35 and C.sub.1-6-alkyl, wherein
R.sup.34 and R.sup.35 independently are hydrogen, C.sub.1-6-alkyl
or aryl, or R.sup.34 and R.sup.35 when attached to the same
nitrogen atom together with the said nitrogen atom may form a 3 to
8 membered heterocyclic ring optionally containing one or two
further heteroatoms selected from nitrogen, oxygen and sulfur, and
optionally containing one or two double bonds, or two of the groups
R.sup.29, R.sup.30 and R.sup.31 when attached to the same ring
carbon atom or different ring carbon atoms together may form a
radical
--O--(CH.sub.2).sub.t--CR.sup.36R.sup.37--(CH.sub.2).sub.l--O--,
--(CH.sub.2).sub.t--CR.sup.36R.sup.37--(CH.sub.2).sub.l-- or
--S--(CH.sub.2).sub.t--CR.sup.36R.sup.37--(CH.sub.2).sub.l--S--,
wherein t and l independently are 0, 1, 2, 3, 4 or 5, R.sup.36 and
R.sup.37 independently are hydrogen or C.sub.1-6-alkyl; (b) an
optical isomer, geometric isomer, or tautomeric form thereof; (c) a
mixture of any combination of compounds provided by (b); (d) a
pharmaceutically acceptable salt of any compound provided by (a) or
(b); or (e) a combination of any of compounds provided by any of
(a)-(d), in an amount effective for antagonizing glucagon.
2. The method of claim 1, wherein the subject is diagnosed as
having or being at risk of developing a condition for which
glucagon antagonistic action is likely to be therapeutically or
prophylactically beneficial.
3. The method of claim 2, wherein the method comprises
administering about 0.05 mg to about 2000 mg of the glucagon
antagonist composition to the subject per day.
4. The method of claim 3, wherein the method comprises
administering about 0.1 mg to about 1000 mg of the glucagon
antagonist composition to the subject per day.
5. The method of claim 4, wherein the method comprises
administering about 0.5 mg to about 500 mg of the glucagon
antagonist composition to the subject per day.
6. The method of claim 2, wherein the subject is diagnosed as
having hyperglycemia and the method is performed as a treatment for
hyperglycemia.
7. The method of claim 2, wherein the subject is diagnosed as being
at risk of developing hyperglycemia and the method is performed to
prevent the development of hyperglycemia in the subject.
8. The method of claim 1, wherein the composition is administered
in an amount and under conditions such that the subject's blood
glucose level is detectably lowered.
9. The method of claim 8, wherein the subject has been diagnosed as
having a condition wherein the reduction of blood glucose level is
therapeutically desirable.
10. The method of claim 2, wherein the subject is diagnosed as
having impaired glucose tolerance and the method is performed as a
treatment for impaired glucose tolerance.
11. The method of claim 2, wherein the subject is diagnosed as
having Type 2 diabetes and the method is performed as a treatment
for Type 2 diabetes.
12. The method of claim 2, wherein the subject is diagnosed as
being at risk of developing Type 2 diabetes and the method is
performed to prevent the development of Type 2 diabetes in the
subject.
13. The method of claim 2, wherein the subject has been diagnosed
as having impaired glucose tolerance and the method is performed to
prevent a progression from impaired glucose tolerance to Type 2
diabetes in the subject.
14. The method of claim 11, wherein the method is performed to
prevent the progression from non-insulin-requiring Type 2 diabetes
to insulin-requiring Type 2 diabetes.
15. The method of claim 2, wherein the subject has been diagnosed
as having Type 1 diabetes and the method is performed as a
treatment for Type 1 diabetes.
16. The method of claim 11, wherein the method further comprises
administering an antidiabetic agent to the subject, wherein the
glucagon antagonist composition and antidiabetic agent are
administered in amounts and under conditions suitable for treating
diabetes in the subject.
17. The method of claim 1, further comprising administering an
antiobesity agent to said subject.
18. The method of claim 1, further comprising administering an
antihypertensive agent to said subject.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority under 35 U.S.C. 119 of
Danish application PA 2000 01733 filed Nov. 17, 2000, U.S.
application 60/252,322, filed Nov. 20, 2000, and benefit under U.S.
application Ser. No. 09/995,987 filed Nov. 16, 2001, the contents
of which are fully incorporated herein by reference.
FIELD OF THE INVENTION
[0002] The present invention relates to agents that act to
antagonize the action of the glucagon peptide hormone on the
glucagon receptor. More particularly, it relates to glucagon
antagonists or inverse agonists.
BACKGROUND OF THE INVENTION
[0003] Glucagon is a key hormonal agent that, in co-operation with
insulin, mediates homeostatic regulation of the amount of glucose
in the blood. Glucagon primarily acts by stimulating certain cells
(mostly liver cells) to release glucose when blood glucose levels
fall. The action of: glucagon is opposite to that of insulin, which
stimulates cells to take up and store glucose whenever blood
glucose levels rise. Both glucagon and insulin are peptide
hormones.
[0004] Glucagon is produced in the alpha islet cells of the
pancreas and insulin in the beta islet cells. Diabetes mellitus is
a common disorder of glucose metabolism. The disease is
characterized by hyperglycemia and may be classified as Type 1
diabetes, the insulin-dependent form, or Type 2 diabetes, which is
non-insulin-dependent in character. Subjects with Type 1 diabetes
are hyperglycemic and hypoinsulinemic, and the conventional
treatment for this form of the disease is to provide insulin.
However, in some patients with Type 1 or Type 2 diabetes, absolute
or relative elevated glucagon levels have been shown to contribute
to the hyperglycemic state. Both in healthy control animals as well
as in animal models of Type 1 and Type 2 diabetes, removal of
circulating glucagon with selective and specific antibodies has
resulted in reduction of the glycemic level (Brand et al.,
Diabetologia 37, 985 (1994); Diabetes 43, [suppl 1], 172A (1994);
Am. J. Physiol. 269, E469-E477 (1995); Diabetes 44 [suppl 1], 134A
(1995); Diabetes 45, 1076 (1996)). These studies suggest that
glucagon suppression or an action that antagonizes glucagon could
be a useful adjunct to conventional treatment of hyperglycemia in
diabetic patients. The action of glucagon can be suppressed by
providing an antagonist or an inverse agonist, ie substances that
inhibit or prevent glucagon-induced responses. The antagonist can
be peptidic or non-peptidic in nature.
[0005] Native glucagon is a 29 amino acid peptide having the
sequence:
1 His-Ser-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-
Tyr-Leu-Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-
Trp-Leu-Met-Asn-Thr-OH
[0006] Glucagon exerts its action by binding to and activating its
receptor, which is part of the Glucagon-Secretin branch of the
7-transmembrane G-protein coupled receptor family (Jelinek et al.,
Science 259, 1614, (1993)). The receptor functions by activating
the adenylyl cyclase second messenger system and the result is an
increase in cAMP levels. Several publications disclose peptides
that are stated to act as glucagon antagonists. Probably, the most
thoroughly characterized antagonist is
DesHis.sup.1[Glu.sup.9]-glucagon amide (Unson et al., Peptides 10,
1171 (1989); Post et al., Proc. Natl. Acad. Sci. USA 90, 1662
(1993)). Other antagonists are
DesHis.sup.1,Phe.sup.6[Glu.sup.9]-glu- cagon amide (Azizh et al.,
Bioorganic & Medicinal Chem. Left. 16, 1849 (1995)) and
NLeu.sup.9,Ala.sup.11,16-glucagon amide (Unson et al., J. Biol.
Chem. 269 (17), 12548 (1994)).
[0007] Peptide antagonists of peptide hormones are often quite
potent. However, they are generally known not to be orally
available because of degradation by physiological enzymes, and
because of poor distribution in vivo. Therefore, orally available
non-peptide antagonists of peptide hormones are generally
preferred. Among the non-peptide glucagon antagonists, a
quinoxaline derivative, (2-styryl-3-[3-(dimethylamino)prop-
ylmethylamino]-6,7-dichloroquinoxaline was found to displace
glucagon from the rat liver receptor (Collins, J. L. et al.,
Bioorganic and Medicinal Chemistry Letters 2(9):915-918 (1992)). WO
94/14426 (The Wellcome Foundation Limited) discloses use of skyrin,
a natural product comprising a pair of linked 9,10-anthracenedione
groups, and its synthetic analogues, as glucagon antagonists. U.S.
Pat. No. 4,359,474 (Sandoz) discloses the glucagon inhibiting
properties of 1-phenyl pyrazole derivatives. U.S. Pat. No.
4,374,130 (Sandoz) discloses substituted disilacyclohexanes as
glucagon inhibiting agents. WO 98/04528 (Bayer Corporation)
discloses substituted pyridines and biphenyls as glucagon
antagonists. U.S. Pat. No. 5,776,954 (Merck & Co., Inc.)
discloses substituted pyridyl pyrroles as glucagon antagonists and
WO 98/21957, WO 98/22108, WO 98/22109 and U.S. Pat. No. 5,880,139
(Merck & Co., Inc.) disclose 2,4-diaryl-5-pyridylimidazoles as
glucagon antagonists. Furthermore, WO 97/16442 and U.S. Pat. No.
5,837,719 (Merck & Co., Inc.) disclose 2,5-substituted aryl
pyrroles as glucagon antagonists. WO 98/24780, WO 98/24782, WO
99/24404 and WO 99/32448 (Amgen Inc.) disclose substituted
pyrimidinone and pyridone compounds and substituted pyrimidine
compounds, respectively, which are stated to possess glucagon
antagonistic activity. Madsen et al. (J. Med. Chem. 1998 (41)
5151-7) discloses a series of
2-(benzimidazol-2-ylthio)-1-(3,4-dihydroxyphenyl)-1- -ethanones as
competitive human glucagon receptor antagonists. WO 99/01423 and WO
00/39088 (Novo Nordisk A/S) disclose different series of alkylidene
hydrazides as glucagon antagonists/inverse agonists. These known
glucagon antagonists differ structurally from the present
compounds.
[0008] Definitions
[0009] The following is a detailed definition of the terms used to
describe the compounds of the invention:
[0010] "Halogen" designates an atom selected from the group
consisting of F, Cl, Br and I.
[0011] The term "C.sub.1-6-alkyl" as used herein represents a
saturated, branched or straight hydrocarbon group having from 1 to
6 carbon atoms. Representative examples include, but are not
limited to, methyl, ethyl, n-propyl, isopropyl, butyl, isobutyl,
sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, tert-pentyl,
n-hexyl, isohexyl and the like.
[0012] The term "C.sub.2-6-alkenyl" as used herein represents a
branched or straight hydrocarbon group having from 2 to 6 carbon
atoms and at least one double bond. Examples of such groups
include, but are not limited to, vinyl, 1-propenyl, 2-propenyl,
iso-propenyl, 1,3-butadienyl, 1-but-enyl, 2-butenyl, 3-butenyl,
2-methyl-1-propenyl, 1-pentenyl, 2-pentenyl, 3-pentenyl,
4-pentenyl, 3-methyl-2-butenyl, 1-hexenyl, 2-hexenyl, 3-hexenyl,
2,4-hexadienyl, 5-hexenyl and the like.
[0013] The term "C.sub.2-6-alkynyl" as used herein represents a
branched or straight hydrocarbon group having from 2 to 6 carbon
atoms and at least one triple bond. Examples of such groups
include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl,
1-butynyl, 2-butynyl, 3-butynyl, 1-pentynyl, 2-pentynyl,
3-pentynyl, 4-pentynyl, 1-hexynyl, 2-hexynyl, 3-hexynyl, 4-hexynyl,
5-hexynyl, 2,4-hexadiynyl and the like.
[0014] The term "C.sub.1-6-alkoxy" as used herein refers to the
radical --O--C.sub.1-6-alkyl, wherein C.sub.1-6-alkyl is as defined
above. Representative examples are methoxy, ethoxy, n-propoxy,
isopropoxy, butoxy, sec-butoxy, tert-butoxy, pentoxy, isopentoxy,
hexoxy, isohexoxy and the like.
[0015] The term "C.sub.3-8-cycloalkyl" as used herein represents a
saturated, carbocyclic group having from 3 to 8 carbon atoms.
Representative examples are cyclopropyl, cyclobutyl, cyclopentyl,
cyclohexyl, cycloheptyl, cyclooctyl and the like.
[0016] The term "C.sub.4-8-cycloalkenyl" as used herein represents
a non-aromatic, carbocyclic group having from 4 to 8 carbon atoms
containing one or two double bonds. Representative examples are
1-cyclopentenyl, 2-cyclopentenyl, 3-cyclopentenyl, 1-cyclohexenyl,
2-cyclohexenyl, 3-cyclohexenyl, 2-cycloheptenyl, 3-cycloheptenyl,
2-cyclooctenyl, 1,4-cyclooctadienyl and the like.
[0017] The term "heterocyclyl" as used herein represents a
non-aromatic 3 to 10 membered ring containing one or more
heteroatoms selected from nitrogen, oxygen and sulfur and
optionally containing one or two double bonds. Representative
examples are pyrrolidinyl, piperidyl, piperazinyl, morpholinyl,
thiomorpholinyl, aziridinyl, tetrahydrofuranyl and the like.
[0018] The term "aryl" as used herein is intended to include
carbocyclic, aromatic ring systems such as 6 membered monocyclic
and 9 to 14 membered bi- and tricyclic, carbocyclic, aromatic ring
systems. Representative examples are phenyl, biphenylyl, naphthyl,
anthracenyl, phenanthrenyl, fluorenyl, indenyl, azulenyl and the
like. Aryl is also intended to include the partially hydrogenated
derivatives of the ring systems enumerated above. Non-limiting
examples of such partially hydrogenated derivatives are
1,2,3,4-tetrahydronaphthyl, 1,4-dihydronaphthyl and the like.
[0019] The term "arylene" as used herein is intended to include
divalent, carbocyclic, aromatic ring systems such as 6 membered
monocyclic and 9 to 14 membered bi- and tricyclic, divalent,
carbocyclic, aromatic ring systems. Representative examples are
phenylene, biphenylylene, naphthylene, anthracenylene,
phenanthrenylene, fluorenylene, indenylene, azulenylene and the
like. Arylene is also intended to include the partially
hydrogenated derivatives of the ring systems enumerated above.
Non-limiting examples of such partially hydrogenated derivatives
are 1,2,3,4-tetrahydronaphthylene, 1,4-dihydronaphthylene and the
like.
[0020] The term "aryloxy" as used herein denotes a group --O-aryl,
wherein aryl is as defined above.
[0021] The term "aroyl" as used herein denotes a group --C(O)-aryl,
wherein aryl is as defined above.
[0022] The term "heteroaryl" as used herein is intended to include
aromatic, heterocyclic ring systems containing one or more
heteroatoms selected from nitrogen, oxygen and sulfur such as 5 to
7 membered monocyclic or 8 to 14 membered bi- and tricyclic
aromatic, heterocyclic ring systems containing one or more
heteroatoms selected from nitrogen, oxygen and sulfur.
Representative examples are furyl, thienyl, pyrrolyl, oxazolyl,
thiazolyl, imidazolyl, isoxazolyl, isothiazolyl, 1,2,3-triazolyl,
1,2,4-triazolyl, pyranyl, pyridyl, pyridazinyl, pyrimidinyl,
pyrazinyl, 1,2,3-triazinyl, 1,2,4-triazinyl, 1,3,5- triazinyl,
1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxa-diazoly
1,3,4-oxadiazolyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl,
1,2,5-thiadiazolyl, 1,3,4-thiadiazolyl, tetrazolyl, thiadiazinyl,
indolyl, isoindolyl, benzofuryl, benzothienyl, indazolyl,
benzimidazolyl, benzthiazolyl, benzisothiazolyl, benzoxazolyl,
benzisoxazolyl, purinyl, quinazolinyl, quinolizinyl, quinolinyl,
isoquinolinyl, quinoxalinyl, naphthyridinyl, pteridinyl,
carbazolyl, azepinyl, diazepinyl, acridinyl and the like.
Heteroaryl is also intended to include the partially hydrogenated
derivatives of the ring systems enumerated above. Non-limiting
examples of such partially hydrogenated derivatives are
2,3-dihydrobenzofuranyl, pyrrolinyl, pyrazolinyl, indolinyl,
oxazolidinyl, oxazolinyl, oxazepinyl and the like.
[0023] "Aryl-C.sub.1-6-alkyl", "heteroaryl-C.sub.1-6-alkyl",
"aryl-C.sub.2-6-alkenyl" etc. mean C.sub.1-6-alkyl or
C.sub.2-6-alkenyl as defined above, substituted by an aryl or
heteroaryl as defined above, for example: 1
[0024] The term "optionally substituted" as used herein means that
the groups in question are either unsubstituted or substituted with
one or more of the substituents specified. When the groups in
question are substituted with more than one substituent the
substituents may be the same or different.
[0025] Certain of the above defined terms may occur more than once
in the structural formulae, and upon such occurrence each term
shall be defined independently of the other.
[0026] Furthermore, when using the terms "independently are" and
"independently selected from" it should be understood that the
groups in question may be the same or different.
DESCRIPTION OF THE INVENTION
[0027] The present invention is based on the unexpected observation
that the compounds of the general formula (I) disclosed below show
a high binding affinity for the glucagon receptor and antagonize
the action of glucagon.
[0028] Accordingly, the invention is concerned with compounds of
the general formula (I): 2
[0029] wherein
[0030] R.sup.2 is hydrogen or C.sub.1-6-alkyl,
[0031] Z is arylene or a divalent radical derived from a 5 or 6
membered heteroaromatic ring containing 1 or 2 heteroatoms selected
from nitrogen, oxygen and sulfur,
[0032] which may optionally be substituted with one or two groups
R.sup.7 and R.sup.8 selected from halogen, --CN, --CF.sub.3,
--OCF.sub.3, --NO.sub.2, --OR.sup.9, --NR.sup.9R.sup.10 and
C.sub.1-6-alkyl,
[0033] wherein R.sup.9 and R.sup.10 independently are hydrogen or
C.sub.1-6-alkyl,
[0034] X is 3
[0035] wherein
[0036] r is 0 or 1,
[0037] q and s independently are 0, 1,2 or 3,
[0038] R.sup.11, R.sup.12, R.sup.13 and R.sup.14 independently are
hydrogen or C.sub.1-6-alkyl,
[0039] D is 4
[0040] wherein
[0041] R.sup.15, R.sup.16, R.sup.17 and R.sup.18 independently
are
[0042] hydrogen, halogen, --CN, --CH.sub.2CN, --CHF.sub.2,
--CF.sub.3, --OCF.sub.3, --OCHF.sub.2, --OCH.sub.2CF.sub.3,
--OCF.sub.2CHF.sub.2, --S(O).sub.2CF.sub.3, --SCF.sub.3,
--NO.sub.2, --OR.sup.21, --NR.sup.21R.sup.22, --SR.sup.21,
--NR.sup.21S(O).sub.2R.sup.22, --S(O).sub.2NR.sup.21R.sup.22,
--S(O)NR.sup.21R.sup.22, --S(O)R.sup.21, --S(O).sub.2R.sup.21,
--C(O)NR.sup.21R.sup.22, --OC(O)NR.sup.21R.sup.22,
--NR.sup.21C(O)R.sup.22, --CH.sub.2C(O)NR.sup.21R.sup.22,
--OCH.sub.2C(O)NR.sup.21R.sup.22, --CH.sub.2OR.sup.21,
--CH.sub.2NR.sup.21R.sup.22, --OC(O)R.sup.21, --C(O)R.sup.21 or
--C(O)OR.sup.21,
[0043] C.sub.1-6-alkyl, C.sub.2-6-alkenyl or C.sub.2-6-alkynyl,
[0044] which may optionally be substituted with one or more
substituents selected from halogen, --CN, --CF.sub.3, --OCF.sub.3,
--NO.sub.2, --OR.sup.21, --NR.sup.21R.sup.22 and
C.sub.1-6-alkyl,
[0045] C.sub.3-8-cycloalkyl, C.sub.4-8-cycloalkenyl, heterocyclyl,
C.sub.3-8-cycloalkyl-C.sub.1-6-alkyl,
C.sub.3-8-cycloalkyl-C.sub.1-6-alko- xy, C.sub.3-8-cycloalkyloxy,
C.sub.3-8-cycloalkyl-C.sub.1-6-alkylthio, C.sub.3-8-cycloalkylthio,
C.sub.3-8-cycloalkyl-C.sub.2-6-alkenyl,
C.sub.3-8-cycloalkyl-C.sub.2-6-alkynyl,
C.sub.4-8-cycloalkenyl-C.sub.1-6-- alkyl,
C.sub.4-8-cycloalkenyl-C.sub.2-6-alkenyl,
C.sub.4-8-cycloalkenyl-C.- sub.2-6-alkynyl,
heterocyclyl-.sub.1-6-alkyl, heterocyclyl-C.sub.2-6-alken- yl,
heterocyclyl-C.sub.2-6-alkynyl, aryl, aryloxy, aryloxycarbonyl,
aroyl, aryl-C.sub.1-6-alkoxy, aryl-C.sub.1-6-alkyl,
aryl-C.sub.2-6-alkenyl, aryl-C.sub.2-6-alkynyl, heteroaryl,
heteroaryl-C.sub.1-6-alkyl, heteroaryl-C.sub.2-6-alkenyl or
heteroaryl-C.sub.2-6-alkynyl,
[0046] of which the cyclic moieties optionally may be substituted
with one or more substituents selected from halogen, --CN,
--CF.sub.3, --OCF.sub.3, --NO.sub.2, --OR.sup.21,
--NR.sup.21R.sup.22 and C.sub.1-6-alkyl,
[0047] wherein R.sup.21 and R.sup.22 independently are hydrogen,
C.sub.1-6-alkyl or aryl,
[0048] or R.sup.21 and R.sup.22 when attached to the same nitrogen
atom together with the said nitrogen atom may form a 3 to 8
membered heterocyclic ring optionally containing one or two further
heteroatoms selected from nitrogen, oxygen and sulfur, and
optionally containing one or two double bonds,
[0049] or two of the groups R.sup.15 to R.sup.18 when placed in
adjacent positions together may form a bridge
--(CR.sup.23R.sup.24).sub.a--O--(CR.-
sup.25R.sup.26).sub.c--O--,
[0050] wherein
[0051] a is 0, 1 or 2,
[0052] c is 1 or2,
[0053] R.sup.23, R.sup.24, R.sup.25 and R.sup.26 independently are
hydrogen, C.sub.1-6-alkyl or fluorine,
[0054] R.sup.19 and R.sup.20 independently are hydrogen,
C.sub.1-6-alkyl, C.sub.3-8-cycloalkyl or
C.sub.3-8-cycloalkyl-C.sub.1-6-alkyl,
[0055] E is 5
[0056] wherein
[0057] R.sup.27 and R.sup.28 independently are
[0058] hydrogen, halogen, --CN, --CF.sub.3, --OCF.sub.3,
--OR.sup.32, --NR.sup.32R.sup.33, C.sub.1-6-alkyl,
C.sub.3-8-cycloalkyl, C.sub.4-8-cycloalkenyl or aryl,
[0059] wherein the cyclic moieties optionally may be substituted
with one or more substituents selected from halogen, --CN,
--CF.sub.3, --OCF.sub.3, --NO.sub.2, --OR.sup.32,
--NR.sup.32R.sup.33 and C.sub.1-6-alkyl,
[0060] wherein
[0061] R.sup.32 and R.sup.33 independently are hydrogen or
C.sub.1-6-alkyl, or
[0062] R.sup.32 and R.sup.33 when attached to the same nitrogen
atom together with the said nitrogen atom may form a 3 to 8
membered heterocyclic ring optionally containing one or two further
heteroatoms selected from nitrogen, oxygen and sulfur, and
optionally containing one or two double bonds,
[0063] R.sup.29, R.sup.30 and R.sup.31 independently are
[0064] hydrogen, halogen, --CHF.sub.2, --CF.sub.3, --OCF.sub.3,
--OCHF.sub.2, --OCH.sub.2CF.sub.3, --OCF.sub.2CHF.sub.2,
--SCF.sub.3, --OR.sup.34, --NR.sup.34R.sup.35, --SR.sup.34,
--S(O)R.sup.34, --S(O).sub.2R.sup.34, --C(O)NR.sup.34R.sup.35,
--OC(O)NR.sup.34R.sup.35, --NR.sup.34C(O)R.sup.35,
--OCH.sub.2C(O)NR.sup.34R.sup.35, --C(O)R.sup.34 or
--C(O)OR.sup.34,
[0065] C.sub.1-6-alkyl, C.sub.2-4-alkenyl or C.sub.2-4-alkynyl,
[0066] which may optionally be substituted with one or more
substituents selected from halogen, --CN, --CF.sub.3, --OCF.sub.3,
--NO.sub.2, --OR.sup.34, --NR.sup.34R.sup.35 and
C.sub.1-6-alkyl,
[0067] C.sub.3-8-cycloalkyl, C.sub.4-8-cycloalkenyl, heterocyclyl,
C.sub.3-4-cycloalkyl-C.sub.1l-alkyl,
C.sub.3-8-cycloalkyl-C.sub.2-6-alken- yl,
C.sub.3-8-cycloalkyl-C.sub.2-6-alkynyl,
C.sub.4-8-cycloalkenyl-C.sub.1- -6-alkyl,
C.sub.4-8-cycloalkenyl-C.sub.2-6-alkenyl,
C.sub.4-8-cycloalkenyl-C.sub.2-6-alkynyl,
heterocyclyl-C.sub.1-6-alkyl, heterocyclyl-C.sub.2-4-alkenyl,
heterocyclyl-C.sub.2-6-alkynyl, aryl, aryloxy, aroyl,
aryl-C.sub.1-6-alkoxy, aryl-C.sub.1-6-alkyl,
aryl-C.sub.2-6-alkenyl, aryl-C.sub.2-6-alkynyl, heteroaryl,
heteroaryl-C.sub.1-6-alkyl, heteroaryl-C.sub.2-6-alkenyl or
heteroaryl-C.sub.2-6-alkynyl,
[0068] of which the cyclic moieties optionally may be substituted
with one or more substituents selected from halogen, --CN,
--CF.sub.3, --OCF.sub.3, --NO.sub.2, --OR.sup.34,
--NR.sup.34R.sup.35 and C.sub.1-6-alkyl,
[0069] wherein R.sup.34 and R.sup.35 independently are hydrogen,
C.sub.1-6-alkyl or aryl,
[0070] or R.sup.34 and R.sup.35 when attached to the same nitrogen
atom together with the said nitrogen atom may form a 3 to 8
membered heterocyclic ring optionally containing one or two further
heteroatoms selected from nitrogen, oxygen and sulfur, and
optionally containing one or two double bonds,
[0071] or two of the groups R.sup.29, R.sup.30 and R.sup.31 when
attached to the same ring carbon atom or different ring carbon
atoms together may form a radical
--O--(CH.sub.2).sub.t--CR.sup.36R.sup.37--(CH.sub.2).sub.l- --O--,
--(CH.sub.2).sub.t--CR.sup.36R.sup.37--(CH.sub.2).sub.l-- or
--S--(CH.sub.2).sub.t--CR.sup.36R.sup.37--(CH.sub.2).sub.l--S--,
[0072] wherein
[0073] t and l independently are 0, 1,2, 3, 4 or 5,
[0074] R.sup.36 and R.sup.37 independently are hydrogen or
C.sub.1-6-alkyl, as well as any optical or geometric isomer or
tautomeric form thereof including mixtures of these or a
pharmaceutically acceptable salt thereof.
[0075] In one embodiment R.sup.2 is hydrogen.
[0076] In another embodiment Z is 6
[0077] wherein R.sup.7 and R.sup.8 are as defined for formula
(I).
[0078] In still another embodiment Z is 7
[0079] In further embodiment X is 8
[0080] wherein q is 0 or 1, r is 0 or 1, s is 0, 1 or 2, and
R.sup.12 and R.sup.13 independently are hydrogen or
C.sub.1-6-alkyl.
[0081] In still a further embodiment X is --C(O)NH--,
--C(O)NHCH.sub.2--, --C(O)NHCH(CH.sub.3)--,
--C)O)NHCH.sub.2CH.sub.2--, --C(O)CH.sub.2--, --C(O)CH.dbd.CH--,
--(CH.sub.2).sub.s--, --C(O)--, --C(O)O--or --NHC(O)--, wherein s
is 0 or 1.
[0082] further embodiment X is --C(O)NH, --C(O)NHCH.sub.2--,
--C(O)NHCH(CH.sub.3)--, --C(O)NHCH.sub.2CH.sub.2--,
--C(O)CH.sub.2--, --CH.sub.2--, --C(O)--or --NHC(O)--, such as
--C(O)NH--.
[0083] In another embodiment D is 9
[0084] wherein R.sup.15, R.sup.16, R.sup.17, R.sup.18, R.sup.19 and
R.sup.20 are as defined for formula (I).
[0085] In still another embodiment D is 10
[0086] wherein R.sup.15, R.sup.16 and R.sup.17 are as defined for
formula (I).
[0087] In an embodiment thereof R.sup.15, R.sup.16 and R.sup.17
independently are hydrogen, halogen, --CN, --NO.sub.2, --CF.sub.3,
--OCF.sub.3, --SCF.sub.3, C.sub.1-6-alkyl, C.sub.1-6-alkoxy,
--S--C.sub.1-6-alkyl, --C(O)OR.sup.21, --C(O)R.sup.21,
--CH.sub.2OR.sup.21, --C(O)NR.sup.21R.sup.22, --S(O).sub.2R.sup.21,
--S(O).sub.2CF.sub.3, --S(O).sub.2NR.sup.21R.sup.22,
C.sub.3-8-cycloalkyl or aryl, or two of the groups R.sup.15,
R.sup.16 and R.sup.17 when placed in adjacent positions together
form a bridge --(CR.sup.23R.sup.24).sub.a--
-O--(CR.sup.25R.sup.26).sub.c--O--, wherein R.sup.21 and R.sup.22
independently are hydrogen or C.sub.1-6-alkyl, and a, c, R.sup.23,
R.sup.24 R.sup.25 and R.sup.26 are as defined for formula (I).
[0088] In another embodiment thereof R.sup.15, R.sup.16 and
R.sup.17 independently are hydrogen, halogen, --CN, --CF.sub.3,
--OCF.sub.3 or C.sub.1-6-alkoxy, such as hydrogen, halogen,
--CF.sub.3 or --OCF.sub.3.
[0089] In a further embodiment E is 11
[0090] wherein R.sup.27, R.sup.28, R.sup.29, R.sup.30 and R.sup.31
are as defined for formula (I).
[0091] In still a further embodiment E is 12
[0092] wherein R.sup.27 and R.sup.28 are as defined for formula
(I).
[0093] In an embodiment thereof R.sup.27 and R.sup.28 are
independently
[0094] hydrogen, C.sub.1-6-alkyl,
[0095] C.sub.3-8-cycloalkyl, C.sub.4-8-cycloalkenyl or phenyl,
which may optionally be substituted as defined for formula (I).
[0096] In another embodiment thereof R.sup.27 is hydrogen and
R.sup.28 is
[0097] C.sub.1-6-alkyl,
[0098] C.sub.4-8-cycloalkenyl or C.sub.3-8-cycloalkyl, which may
optionally be substituted as defined for formula (I).
[0099] In another embodiment E is 13
[0100] wherein R.sup.29, R.sup.30 and R.sup.31 are as defined for
formula (I).
[0101] In still another embodiment E is 14
[0102] wherein R.sup.29, R.sup.30 and R.sup.31 are as defined for
formula (I).
[0103] In an embodiment thereof R.sup.29, R.sup.30 and R.sup.31 are
independently
[0104] hydrogen, --CHF.sub.2, --CF.sub.3, --OCF.sub.3,
--OCHF.sub.2, --OCH.sub.2CF.sub.3, --OCF.sub.2CHF.sub.2,
--SCF.sub.3, --OR.sup.34, --NR.sup.34R.sup.35, --SR.sup.34,
--S(O)R.sup.34, --S(O).sub.2R.sub.34, --C(O)NR.sub.34R.sup.35,
--OC(O)NR.sub.34R.sup.35, --NR.sup.34C(O)R.sup.35,
--OCH.sub.2C(O)NR.sup.34R.sup.35, --C(O)R.sup.34 or
--C(O)OR.sup.34,
[0105] C.sub.1-6-alkyl, C.sub.2-6-alkenyl or C.sub.2-6-alkynyl,
[0106] which may optionally be substituted with one or more
substituents selected from halogen, --CN, --CF.sub.3, --OCF.sub.3,
--NO.sub.2, --OR.sup.34, --NR.sup.34R.sup.35 and
C.sub.1-6-alkyl,
[0107] C.sub.3-8-cycloalkyl or C.sub.4-8-cycloalkenyl,
[0108] which may optionally be substituted with one or more
substituents selected from halogen, --CN, --CF.sub.3, --OCF.sub.3,
--NO.sub.2, --OR.sup.34, --NR.sup.34R.sup.35 and
C.sub.1-6-alkyl,
[0109] wherein R.sup.34 and R.sup.35 independently are hydrogen,
C.sub.1-6-alkyl or aryl,
[0110] or R.sup.34 and R.sup.35 when attached to the same nitrogen
atom together with the said nitrogen atom may form a 3 to 8
membered heterocyclic ring optionally containing one or two further
heteroatoms selected from nitrogen, oxygen and sulfur, and
optionally containing one or two double bonds.
[0111] In another embodiment thereof R.sup.29, R.sup.30 and
R.sup.31 are independently
[0112] hydrogen, C.sub.1-6-alkoxy, --CF.sub.3, --OCF.sub.3 or
--NR.sup.34R.sup.35, wherein R.sup.34 and R.sup.35 are as defined
for formula (I), or
[0113] C.sub.1-6-alkyl, C.sub.3-8-cycloalkyl or
C.sub.4-8-cycloalkenyl, which are optionally substituted as defined
for formula (I).
[0114] In still another embodiment thereof R.sup.29, R.sup.30 and
R.sup.31 are independently
[0115] hydrogen or
[0116] C.sub.1-6-alkyl, C.sub.3-8-cycloalkyl or
C.sub.4-8-cycloalkenyl, which are optionally substituted as defined
for formula (I).
[0117] In a further embodiment thereof R.sup.29, R.sup.30 and
R.sup.31 are independently hydrogen, C.sub.1-6-alkyl,
C.sub.3-8-cycloalkyl or C.sub.4-8-cycloalkenyl, wherein
C.sub.3-8-cycloalkyl or C.sub.4-8-cycloalkenyl are optionally
substituted with C.sub.1-6-alkyl.
[0118] In still a further embodiment thereof R.sup.29 and R.sup.31
are both hydrogen and R.sup.30 is C.sub.1-6-alkyl,
C.sub.3-8-cycloalkyl or C.sub.4-8-cycloalkenyl, wherein
C.sub.3-8-cycloalkyl or C.sub.4-8-cycloalkenyl are optionally
substituted with C.sub.1-6-alkyl.
[0119] In yet a further embodiment thereof R.sup.29 and R.sup.31
are both hydrogen and R.sup.30 is C.sub.1-6-alkyl.
[0120] In still another embodiment thereof R.sup.29 and R.sup.31
are both hydrogen and R.sup.30 is C.sub.4-8-cycloalkenyl which is
optionally substituted with C.sub.1-6-alkyl.
[0121] In another aspect, the invention is concerned with compounds
of the general formula (I'): 15
[0122] wherein
[0123] R.sup.2 is hydrogen or C.sub.1-6-alkyl,
[0124] Z is arylene or a divalent radical derived from a 5 or 6
membered heteroaromatic ring containing 1 or 2 heteroatoms selected
from nitrogen, oxygen and sulfur,
[0125] which may optionally be substituted with one or two groups
R.sup.7 and R.sup.8 selected from halogen, --CN, --CF.sub.3,
--OCF.sub.3, --NO.sub.2, --OR.sup.9, --NR.sup.9R.sup.10 and
C.sub.1-6-alkyl,
[0126] wherein R.sup.9 and R.sup.10 independently are hydrogen or
C.sub.1-6-alkyl,
[0127] X is 16
[0128] wherein
[0129] r is 0 or 1,
[0130] q and s independently are 0, 1, 2 or 3,
[0131] R.sup.11, R.sup.12, R.sup.13, and R.sup.14 independently are
hyrogen or C.sub.1-6-alkyl,
[0132] D is 17
[0133] wherein
[0134] R.sup.15, R.sup.16, R.sup.17 and R.sup.18 independently
are
[0135] hydrogen, halogen, --CN, --CH.sub.2CN, --CHF.sub.2,
--CF.sub.3, --OCF.sub.3, --OCHF.sub.2, --OCH.sub.2CF.sub.3,
--OCF.sub.2CHF.sub.2, --S(O).sub.2CF.sub.3, --SCF.sub.3,
--NO.sub.2, --OR.sup.21, --NR.sup.21R.sup.22, --SR.sup.21,
--NR.sup.21S(O).sub.2R.sup.22, --S(O).sub.2NR.sup.21R.sup.22,
--S(O)NR.sup.21R.sup.22, --S(O)R.sup.21, --S(O).sub.2R.sup.21,
--C(O)NR.sup.21R.sup.22, --OC(O)NR.sup.21R.sup.22,
--NR.sup.21C(O)R.sup.22, --CH.sub.2C(O)NR.sup.21R.sup.22,
--OCH.sub.2C(O)NR.sup.21R.sup.22, --CH.sub.2OR.sup.21,
--CH.sub.2NR.sup.21R.sup.22, --OC(O)R.sup.21, --C(O)R.sup.21 or
--C(O)OR.sup.21,
[0136] C.sub.1-6-alkyl, C.sub.2-6-alkenyl or C.sub.2-6-alkynyl,
[0137] which may optionally be substituted with one or more
substituents selected from halogen, --CN, --CF.sub.3, --OCF.sub.3,
--NO.sub.2, --OR.sup.21, --NR.sup.21R.sup.22 and
C.sub.1-6-alkyl,
[0138] C.sub.3-8-cycloalkyl, C.sub.4-8-cycloalkenyl, heterocyclyl,
C.sub.3-8-cycloalkyl-C.sub.1-6-alkyl,
C.sub.3-8-cycloalkyl-C.sub.1-6-alko- xy, C.sub.3-8-cycloalkyloxy,
C.sub.3-8-cycloalkyl-C.sub.1-6-alkylthio, C.sub.3-8-cycloalkylthio,
C.sub.3-8-cycloalkyl-C.sub.2-6-alkenyl,
C.sub.3-8-cycloalkyl-C.sub.2-6-alkynyl,
C.sub.4-8-cycloalkenyl-C.sub.1-6-- alkyl,
C.sub.4-8-cycloalkenyl-C.sub.2-6-alkenyl,
C.sub.4-8-cycloalkenyl-C.- sub.2-6-alkynyl,
heterocyclyl-C.sub.1-6-alkyl, heterocyclyl-C.sub.2-6-alke- nyl,
heterocyclyl-C.sub.2-6-alkynyl, aryl, aryloxy, aryloxycarbonyl,
aroyl, aryl-C.sub.1-6-alkoxy, aryl-C.sub.1-6-alkyl,
aryl-C.sub.2-6-alkenyl, aryl-C.sub.2-6-alkynyl, heteroaryl,
heteroaryl-C.sub.1-6-alkyl, heteroaryI-C.sub.2-6-alkenyl or
heteroaryl-C.sub.2-6-alkynyl,
[0139] of which the cyclic moieties optionally may be substituted
with one or more substituents selected from halogen, --CN,
--CF.sub.3, --OCF.sub.3, --NO.sub.2, --OR.sup.21,
--NR.sup.21R.sup.22 and C.sub.1-6-alkyl,
[0140] wherein R.sup.21 and R.sup.22 independently are hydrogen,
C.sub.1-6-alkyl or aryl,
[0141] or R.sup.21 and R.sup.22 when attached to the same nitrogen
atom together with the said nitrogen atom may form a 3 to 8
membered heterocyclic ring optionally containing one or two further
heteroatoms selected from nitrogen, oxygen and sulfur, and
optionally containing one or two double bonds,
[0142] or two of the groups R.sup.15 to R.sup.18 when placed in
adjacent positions together may form a bridge
--(CR.sup.23R.sup.24).sub.a--O--(CR.-
sup.25R.sup.26).sub.c--O--,
[0143] wherein
[0144] a is 0, 1 or 2,
[0145] c is 1 or 2,
[0146] R.sup.23, R.sup.24, R.sup.25 and R.sup.26 independently are
hydrogen, C.sub.1-6-alkyl or fluorine,
[0147] R.sup.19 and R.sup.20 independently are hydrogen,
C.sub.1-6-alkyl, C.sub.3-8-cycloalkyl or
C.sub.3-8-cycloalkyl-C.sub.1-6-alkyl,
[0148] E is 18
[0149] wherein
[0150] R.sup.27 and R.sup.28 independently are
[0151] hydrogen, halogen, --CN, --CF.sub.3, --OCF.sub.3,
--OR.sup.32, --NR.sup.32R.sup.33, C.sub.1-6-alkyl,
C.sub.3-8-cycloalkyl, C.sub.4-8-cycloalkenyl or aryl,
[0152] wherein the aryl group optionally may be substituted with
one or more substituents selected from halogen, --CN, --CF.sub.3,
--OCF.sub.3, --NO.sub.2, --OR.sup.32, --NR.sup.32R.sup.33 and
C.sub.1-6-alkyl,
[0153] wherein
[0154] R.sup.32 and R.sup.33 independently are hydrogen or
C.sub.1-6-alkyl, or
[0155] R.sup.32 and R.sup.33 when attached to the same nitrogen
atom together with the said nitrogen atom may form a 3 to 8
membered heterocyclic ring optionally containing one or two further
heteroatoms selected from nitrogen, oxygen and sulfur, and
optionally containing one or two double bonds,
[0156] R.sup.29, R.sup.30 and R.sup.31 independently are
[0157] hydrogen, halogen, --CHF.sub.2, --CF.sub.3, --OCF.sub.3,
--OCHF.sub.2, --OCH.sub.2CF.sub.3, --OCF.sub.2CHF.sub.2,
--SCF.sub.3, --OR.sup.34, --NR.sup.34R.sup.35, --SR.sup.34,
--S(O)R.sup.34, --S(O).sub.2R.sup.34,
C(O)NR.sup.34R.sup.35--OC(O)NR.sup.34R.sup.35,
--NR.sup.34C(O)R.sup.35, --OCH.sub.2C(O)NR.sup.34R.sup.35,
--C(O)R.sup.34 or --C(O)OR.sup.34,
[0158] C.sub.1-6-alkyl, C.sub.2-6-alkenyl or C.sub.2-6-alkynyl,
[0159] which may optionally be substituted with one or more
substituents selected from halogen, --CN, --CF.sub.3, --OCF.sub.3,
--NO.sub.2, --OR.sup.34, --NR.sup.34R.sup.35 and
C.sub.1-6-alkyl,
[0160] C.sub.3-8-cycloalkyl, C.sub.4-8-cycloalkenyl, heterocyclyl,
C.sub.3-8-cycloalkyl-C.sub.1-6-alkyl,
C.sub.3-8-cycloalkyl-C.sub.2-6-alke- nyl,
C.sub.3-8-cycloalkyl-C.sub.2-6-alkynyl,
C.sub.4-8-cycloalkenyl-C.sub.- 1-6-alkyl,
C.sub.4-8-cycloalkenyl-C.sub.2-6-alkenyl,
C.sub.4-8-cycloalkenyl-C.sub.2-6-alkynyl,
heterocyclyl-C.sub.1-6-alkyl, heterocyclyl-C.sub.2-6-alkenyl,
heterocyclyl-C.sub.2-6-alkynyl, aryl, aryloxy, aroyl,
aryl-C.sub.1-6-alkoxy, aryl-C.sub.1-6-alkyl,
aryl-C.sub.2-6-alkenyl, aryl-C.sub.2-6-alkynyl, heteroaryl,
heteroaryl-C.sub.1-6-alkyl, heteroaryl-C.sub.2-6-alkenyl or
heteroaryl-C.sub.2-6-alkynyl,
[0161] of which the cyclic moieties optionally may be substituted
with one or more substituents selected from halogen, --CN,
--CF.sub.3, --OCF.sub.3, --NO.sub.2, --OR.sup.34,
--NR.sup.34R.sup.35 and C.sub.1-6-alkyl,
[0162] wherein R.sup.34 and R.sup.35 independently are hydrogen,
C.sub.1-6-alkyl or aryl,
[0163] or R.sup.34 and R.sup.35 when attached to the same nitrogen
atom together with the said nitrogen atom may form a 3 to 8
membered heterocyclic ring optionally containing one or two further
heteroatoms selected from nitrogen, oxygen and sulfur, and
optionally containing one or two double bonds,
[0164] or two of the groups R.sup.29, R.sup.30 and R.sup.31 when
attached to the same ring carbon atom or different ring carbon
atoms together may form a radical
--O--(CH.sub.2).sub.t--CR.sup.36R.sup.37--(CH.sub.2).sub.l- --O--,
--(CH.sub.2).sub.t--CR.sup.36R.sup.37--(CH.sub.2).sub.l-- or
--S--(CH.sub.2).sub.t--CR.sup.36R.sup.37--(CH.sub.2).sub.l--S--,
[0165] wherein
[0166] t and l independently are 0, 1, 2, 3, 4 or 5,
[0167] R.sup.36 and R.sup.37 independently are hydrogen or
C.sub.1-6-alkyl, as well as any optical or geometric isomer or
tautomeric form thereof including mixtures of these or a
pharmaceutically acceptable salt thereof.
[0168] In specific embodiments thereof, R.sup.2, Z, E, X and D are
as described in the embodiments above.
[0169] The compounds of the present invention may have one or more
asymmetric centres and it is intended that any optical isomers, as
separated, pure or partially purified optical isomers or racemic
mixtures thereof are included within the scope of the
invention.
[0170] Furthermore, when a double bond or a fully or partially
saturated ring system is present in the molecule geometric isomers
may be formed. It is intended that any geometric isomers, as
separated, pure or partially purified geometric isomers or mixtures
thereof are included within the scope of the invention. Likewise,
molecules having a bond with restricted rotation may form geometric
isomers. These are also intended to be included within the scope of
the present invention.
[0171] Furthermore, some of the compounds of the present invention
may exist in different tautomeric forms and it is intended that any
tautomeric forms that the compounds are able to form are included
within the scope of the present invention.
[0172] The present invention also encompasses pharmaceutically
acceptable salts of the present compounds. Such salts include
pharmaceutically acceptable acid addition salts, pharmaceutically
acceptable metal salts, ammonium and alkylated ammonium salts. Acid
addition salts include salts of inorganic acids as well as organic
acids. Representative examples of suitable inorganic acids include
hydrochloric, hydrobromic, hydroiodic, phosphoric, sulfuric, nitric
acids and the like. Representative examples of suitable organic
acids include formic, acetic, trichloroacetic, trifluoroacetic,
propionic, benzoic, cinnamic, citric, fumaric, glycolic, lactic,
maleic, malic, malonic, mandelic, oxalic, picric, pyruvic,
salicylic, succinic, methane-sulfonic, ethanesulfonic, tartaric,
ascorbic, pamoic, bismethylene salicylic, ethanedisulfonic,
gluconic, citraconic, aspartic, stearic, palmitic, EDTA, glycolic,
p-aminobenzoic, glutamic, benzenesulfonic, p-toluenesulfonic acids
and the like. Further examples of pharmaceutically acceptable
inorganic or organic acid addition salts include the
pharmaceutically acceptable salts listed in J. Pharm. Sci. 1977,
66, 2, which is incorporated herein by reference. Examples of metal
salts include lithium, sodium, potassium, magnesium salts and the
like.
[0173] Examples of ammonium and alkylated ammonium salts include
ammonium, methyl-, dimethyl-, trimethyl-, ethyl-, hydroxyethyl-,
diethyl-, butyl-, tetramethylammonium salts and the like.
[0174] Also intended as pharmaceutically acceptable acid addition
salts are the hydrates which the present compounds are able to
form.
[0175] Furthermore, the pharmaceutically acceptable salts comprise
basic amino acid salts such as lysine, arginine and ornithine.
[0176] The acid addition salts may be obtained as the direct
products of compound synthesis. In the alternative, the free base
may be dissolved in a suitable solvent containing the appropriate
acid, and the salt isolated by evaporating the solvent or otherwise
separating the salt and solvent.
[0177] The compounds of the present invention may form solvates
with standard low molecular weight solvents using methods well
known to the person skilled in the art. Such solvates are also
contemplated as being within the scope of the present
invention.
[0178] The invention also encompasses prodrugs of the present
compounds, which on administration undergo chemical conversion by
metabolic processes before becoming pharmacologically active
substances. In general, such prodrugs will be functional
derivatives of the compounds of the general formula (I), which are
readily convertible in vivo into the required compound of the
formula (I). Conventional procedures for the selection and
preparation of suitable prodrug derivatives are described, for
example, in "Design of Prodrugs", ed. H. Bundgaard, Elsevier,
1985.
[0179] The invention also encompasses active metabolites of the
present compounds. The compounds according to the present invention
act to antagonize the action of glucagon and are accordingly useful
for the treatment and/or prevention of disorders and diseases in
which such an antagonism is beneficial.
[0180] Accordingly, the present compounds may be applicable for the
treatment and/or prevention of hyperglycemia, IGT (impaired glucose
tolerance), insulin resistance syndromes, syndrome X,
[0181] Type 1 diabetes, Type 2 diabetes, hyperlipidemia,
dyslipidemia, hypertriglyceridemia, hyperlipoproteinemia,
hypercholesterolemia, arteriosclerosis including atherosclerosis,
glucagonomas, acute pancreatitis, cardiovascular diseases,
hypertension, cardiac hypertrophy, gastrointestinal disorders,
obesity, diabetes as a consequence of obesity, diabetic
dyslipidemia, etc.
[0182] Furthermore, they may be applicable as diagnostic agents for
identifying patients having a defect in the glucagon receptor, as a
therapy to increase gastric acid secretions and to reverse
intestinal hypomobility due to glucagon administration.
[0183] They may also be useful as tool or reference molecules in
labelled form in binding assays to identify new glucagon
antagonists.
[0184] Accordingly, in a further aspect the invention relates to a
compound according to the invention for use as a medicament.
[0185] The invention also relates to pharmaceutical compositions
comprising, as an active ingredient, at least one compound
according to the invention together with one or more
pharmaceutically acceptable carriers or excipients.
[0186] The pharmaceutical composition is preferably in unit dosage
form, comprising from about 0.05 mg to about 1000 mg, preferably
from about 0.1 mg to about 500 mg and especially preferred from
about 0.5 mg to about 200 mg of the compound according to the
invention.
[0187] Furthermore, the invention relates to the use of a compound
according to the invention for the preparation of a pharmaceutical
composition for the treatment and/or prevention of a disorder or
disease, wherein a glucagon antagonistic action is beneficial.
[0188] The invention also relates to a method for the treatment
and/or prevention of disorders or diseases, wherein a glucagon
antagonistic action is beneficial the method comprising
administering to a subject in need thereof an effective amount of a
compound according to the invention.
[0189] In a preferred embodiment of the invention the present
compounds are used for the preparation of a medicament for the
treatment and/or prevention of any glucagon-mediated conditions and
diseases.
[0190] In a preferred embodiment of the invention the present
compounds are used for the preparation of a medicament for the
treatment and/or prevention of hyperglycemia.
[0191] In yet a preferred embodiment of the invention the present
compounds are used for the preparation of a medicament for lowering
blood glucose in a mammal. The present compounds are effective in
lowering the blood glucose, both in the fasting and the
postprandial stage.
[0192] In another preferred embodiment of the invention the present
compounds are used for the preparation of a pharmaceutical
composition for the treatment and/or prevention of IGT.
[0193] In still another preferred embodiment of the invention the
present compounds are used for the preparation of a pharmaceutical
composition for the treatment and/or prevention of Type 2
diabetes.
[0194] In yet another preferred embodiment of the invention the
present compounds are used for the preparation of a pharmaceutical
composition for the delaying or prevention of the progression from
IGT to Type 2 diabetes.
[0195] In yet another preferred embodiment of the invention the
present compounds are used for the preparation of a pharmaceutical
composition for the delaying or prevention of the progression from
non-insulin requiring Type 2 diabetes to insulin requiring Type 2
diabetes.
[0196] In a further preferred embodiment of the invention the
present compounds are used for the preparation of a pharmaceutical
composition for the treatment and/or prevention of Type 1 diabetes.
Such treatment and/or prevention is normally accompanied by insulin
therapy.
[0197] In a further preferred embodiment of the invention the
present compounds are used for the preparation of a pharmaceutical
composition for the treatment and/or prevention of obesity.
[0198] In yet a further preferred embodiment of the invention the
present compounds are used for the preparation of a pharmaceutical
composition for the treatment and/or prevention of disorders of the
lipid metabolism.
[0199] In still a further preferred embodiment of the invention the
present compounds are used for the preparation of a pharmaceutical
composition for the treatment and/or prevention of an appetite
regulation or energy expenditure disorder.
[0200] In a further aspect of the invention, treatment of a patient
with the present compounds is combined with diet and/or
exercise.
[0201] In still a further aspect of the invention the present
compounds are administered in combination with one or more further
active substances in any suitable ratios. Such further active
substances may eg be selected from antiobesity agents,
antidiabetics, antihypertensive agents, agents for the treatment of
complications resulting from or associated with diabetes and agents
for the treatment of complications and disorders resulting from or
associated with obesity.
[0202] Thus, in a further aspect of the invention the present
compounds may be administered in combination with one or more
antiobesity agents or appetite regulating agents.
[0203] Such agents may be selected from the group consisting of
CART (cocaine amphetamine regulated transcript) agonists, NPY
(neuropeptide Y) antagonists, MC4 (melanocortin 4) agonists, MC3
(melanocortin 3) agonists, orexin antagonists, TNF (tumor necrosis
factor) agonists, CRF (corticotropin releasing factor) agonists,
CRF BP (corticotropin releasing factor binding protein)
antagonists, urocortin agonists, P3 adrenergic agonists such as
CL-316243, AJ-9677, GW-0604, LY362884, LY377267 or AZ-40140, MSH
(melanocyte-stimulating hormone) agonists, MCH
(melanocyte-concentrating hormone) antagonists, CCK
(cholecystokinin) agonists, serotonin re-uptake inhibitors such as
fluoxetine, seroxat or citalopram, serotonin and noradrenaline
re-uptake inhibitors, mixed serotonin and noradrenergic compounds,
5HT (serotonin) agonists, bombesin agonists, galanin antagonists,
growth hormone, growth factors such as prolactin or placental
lactogen, growth hormone releasing compounds, TRH (thyreotropin
releasing hormone) agonists, UCP 2 or 3 (uncoupling protein 2 or 3)
modulators, leptin agonists, DA agonists (bromocriptin, doprexin),
lipase/amylase inhibitors, PPAR (peroxisome proliferator-activated
receptor) modulators, RXR (retinoid X receptor) modulators, TR,
agonists, AGRP (Agouti related protein) inhibitors, H3 histamine
antagonists, opioid antagonists (such as naltrexone), exendin-4,
GLP-1 and ciliary neurotrophic factor.
[0204] In one embodiment of the invention the antiobesity agent is
leptin.
[0205] In another embodiment the antiobesity agent is
dexamphetamine or amphetamine.
[0206] In another embodiment the antiobesity agent is fenfluramine
or dexfenfluramine.
[0207] In still another embodiment the antiobesity agent is
sibutramine.
[0208] In a further embodiment the antiobesity agent is
orlistat.
[0209] In another embodiment the antiobesity agent is mazindol or
phentermine.
[0210] In still another embodiment the antiobesity agent is
phendimetrazine, diethylpropion, fluoxetine, bupropion, topiramate
or ecopipam.
[0211] Suitable antidiabetic agents include insulin, insulin
analogues and derivatives such as those disclosed in EP 792 290
(Novo Nordisk ANS), eg N.sup..epsilon.B29-tetradecanoyl des (B30)
human insulin, EP 214 826 and EP 705 275 (Novo Nordisk A/S), eg
Asp.sup.B28 human insulin, U.S. Pat. No. 5,504,188 (Eli Lilly), eg
LyS.sup.B28 Pro.sup.B29 human insulin, EP 368 187 (Aventis), eg
Lantus.RTM., which are all incorporated herein by reference, GLP-1
and GLP-1 derivatives such as those disclosed in WO 98/08871 (Novo
Nordisk A/S), which is incorporated herein by reference, as well as
orally active hypoglycemic agents.
[0212] The orally active hypoglycemic agents preferably comprise
imidazolines, sulfonylureas, biguanides, meglitinides,
oxadiazolidinediones, thiazolidinediones, insulin sensitizers,
.alpha.-glucosidase inhibitors, agents acting on the ATP-dependent
potassium channel of the .beta.-cells eg potassium channel openers
such as those disclosed in WO 97/26265, WO 99/03861 and WO 00/37474
(Novo Nordisk A/S) which are incorporated herein by reference, or
mitiglinide, or a potassium channel blocker, such as BTS-67582,
nateglinide, glucagon antagonists such as those disclosed in WO
99/01423 and WO 00/39088 (Novo Nordisk A/S and Agouron
Pharmaceuticals, Inc.), which are incorporated herein by reference,
GLP-1 agonists such as those disclosed in WO 00/42026 (Novo Nordisk
A/S and Agouron Pharmaceuticals, Inc.), which are incorporated
herein by reference, insulin secretagogues, such as glimepride,
DPP-IV (dipeptidyl peptidase-IV) inhibitors, PTPase (protein
tyrosine phosphatase) inhibitors, inhibitors of hepatic enzymes
involved in stimulation of gluconeogenesis and/or glycogenolysis,
glucose uptake modulators, GSK-3 (glycogen synthase kinase-3)
inhibitors, compounds modifying the lipid metabolism such as
antilipidemic agents, compounds lowering food intake, PPAR
(peroxisome proliferator-activated receptor) and RXR (retinoid X
receptor) agonists, such as ALRT-268, LG-1268 or LG-1069.
[0213] In one embodiment of the invention the present compounds are
administered in combination with insulin,
N.sup..epsilon.B29-tetradecanoy- l des (B30) human insulin,
Asp.sup.B28 human insulin, Lys.sup.B28pro.sup.B29 human insulin or
Lantus, or a mix-preparation comprising one or more of these.
[0214] In a further embodiment of the invention the present
compounds are administered in combination with a sulfonylurea eg
tolbutamide, chlorpropamide, tolazamide, glibenclamide, glipizide,
glimepiride, glicazide or glyburide.
[0215] In another embodiment of the invention the present compounds
are administered in combination with a biguanide eg mefformin.
[0216] In yet another embodiment of the invention the present
compounds are administered in combination with a meglitinide eg
repaglinide or nateglinide.
[0217] In still another embodiment of the invention the present
compounds are administered in combination with a thiazolidinedione
insulin sensitizer eg troglitazone, ciglitazone, pioglitazone,
rosiglitazone, isaglitazone, darglitazone, englitazone,
CS-011/CI-1037 or T 174 or the compounds disclosed in WO 97/41097,
WO 97/41119, WO 97/41120, WO 00/41121 and WO 98/45292 (Dr. Reddy's
Research Foundation), which are incorporated herein by
reference.
[0218] In still another embodiment of the invention the present
compounds may be administered in combination with an insulin
sensitizer eg such as GI 262570, YM-440, MCC-555, JTT-501,
AR-H039242, KRP-297, GW-409544, CRE-16336, AR-H049020, LY510929,
MBX-102, CLX-0940, GW-501516 or the compounds disclosed in WO
99/19313, WO 00/50414, WO 00/63191, WO 00/63192, WO 00/63193 (Dr.
Reddy's Research Foundation) and WO 00/23425, WO 00/23415, WO
00/23451, WO 00/23445, WO 00/23417, WO 00/23416, WO 00/63153, WO
00/63196, WO 00/63209, WO 00/63190 and WO 00/63189 (Novo Nordisk
A/S), which are incorporated herein by reference.
[0219] In a further embodiment of the invention the present
compounds are administered in combination with an
.alpha.-glucosidase inhibitor eg voglibose, emiglitate, miglitol or
acarbose. In another embodiment of the invention the present
compounds are administered in combination with an agent acting on
the ATP-dependent potassium channel of the .beta.-cells eg
tolbutamide, glibenclamide, glipizide, glicazide, BTS-67582 or
repaglinide.
[0220] In yet another embodiment of the invention the present
compounds may be administered in combination with nateglinide.
[0221] In still another embodiment of the invention the present
compounds are administered in combination with an antilipidemic
agent eg cholestyramine, colestipol, clofibrate, gemfibrozil,
lovastatin, pravastatin, simvastatin, probucol or
dextrothyroxine.
[0222] In another aspect of the invention, the present compounds
are administered in combination with more than one of the
above-mentioned compounds eg in combination with mefformin and a
sulfonylurea such as glyburide; a sulfonylurea and acarbose;
nateglinide and mefformin; acarbose and mefformin; a sulfonylurea,
mefformin and troglitazone; insulin and a sulfonylurea; insulin and
mefformin; insulin, metfonmin and a sulfonylurea; insulin and
troglitazone; insulin and lovastatin; etc.
[0223] Furthermore, the present compounds may be administered in
combination with one or more antihypertensive agents. Examples of
antihypertensive agents are .beta.-blockers such as alprenolol,
atenolol, timolol, pindolol, propranolol and metoprolol, ACE
(angiotensin converting enzyme) inhibitors such as benazepril,
captopril, enalapril, fosinopril, lisinopril, quinapril and
ramipril, calcium channel blockers such as nifedipine, felodipine,
nicardipine, isradipine, nimodipine, diltiazem and verapamil, and
.alpha.-blockers such as doxazosin, urapidil, prazosin and
terazosin. Further reference can be made to Remington: The Science
and Practice of Pharmacy, 19.sup.th Edition, Gennaro, Ed., Mack
Publishing Co., Easton, Pa., 1995.
[0224] It should be understood that any suitable combination of the
compounds according to the invention with diet and/or exercise, one
or more of the above-mentioned compounds and optionally one or more
other active substances are considered to be within the scope of
the present invention.
[0225] Pharmaceutical Compositions
[0226] The compounds of the invention may be administered alone or
in combination with pharmaceutically acceptable carriers or
excipients, in either single or multiple doses. The pharmaceu-tical
compositions according to the invention may be formulated with
pharmaceutically acceptable carriers or diluents as well as any
other known adjuvants and excipients in accordance with
conventional techniques such as those disclosed in Remington: The
Science and Practice of Pharmacy, 19th Edition, Gennaro, Ed., Mack
Publishing Co., Easton, Pa., 1995.
[0227] The pharmaceutical compositions may be specifically
formulated for administration by any suitable route such as the
oral, rectal, nasal, pulmonary, topical (including buccal and
sublingual), transdermal, intracisternal, intraperitoneal, vaginal
and parenteral (including subcutaneous, intramuscular, intrathecal,
intravenous and intradermal) route, the oral route being preferred.
It will be appreciated that the preferred route will depend on the
general condition and age of the subject to be treated, the nature
of the condition to be treated and the active ingredient
chosen.
[0228] Pharmaceutical compositions for oral administration include
solid dosage forms such as capsules, tablets, dragees, pills,
lozenges, powders and granules. Where appropriate, they can be
prepared with coatings such as enteric coatings or they can be
formulated so as to provide controlled release of the active
ingredient such as sustained or prolonged release according to
methods well known in the art.
[0229] Liquid dosage forms for oral administration include
solutions, emulsions, suspensions, syrups and elixirs.
[0230] Pharmaceutical compositions for parenteral administration
include sterile aqueous and non-aqueous injectable solutions,
dispersions, suspensions or emulsions as well as sterile powders to
be reconstituted in sterile injectable solutions or dispersions
prior to use. Depot injectable formulations are also contemplated
as being within the scope of the present invention.
[0231] Other suitable administration forms include suppositories,
sprays, ointments, cremes, gels, inhalants, dermal patches,
implants etc. A typical oral dosage is in the range of from about
0.001 to about 100 mg/kg body weight per day, preferably from about
0.01 to about 50 mg/kg body weight per day, and more preferred from
about 0.05 to about 10 mg/kg body weight per day administered in
one or more dosages such as 1 to 3 dosages. The exact dosage will
depend upon the frequency and mode of administration, the sex, age,
weight and general condition of the subject treated, the nature and
severity of the condition treated and any concomitant diseases to
be treated and other factors evident to those skilled in the
art.
[0232] The formulations may conveniently be presented in unit
dosage form by methods known to those skilled in the art. A typical
unit dosage form for oral administration one or more times per day
such as 1 to 3 times per day may contain from 0.05 to about 1000
mg, preferably from about 0.1 to about 500 mg, and more preferred
from about 0.5 mg to about 200 mg.
[0233] For parenteral routes such as intravenous, intrathecal,
intramuscular and similar administration, typically doses are in
the order of about half the dose employed for oral
administration.
[0234] The compounds of this invention are generally utilized as
the free substance or as a pharmaceutically acceptable salt
thereof. One example is an acid addition salt of a compound having
the utility of a free base. When a compound of the formula (I)
contains a free base such salts are prepared in a conventional
manner by treating a solution or suspension of a free base of the
formula (I) with a chemical equivalent of a pharmaceutically
acceptable acid. Representative examples are mentioned above.
Physiologically acceptable salts of a compound with a hydroxy group
include the anion of said compound in combination with a suitable
cation such as sodium or ammonium ion.
[0235] For parenteral administration, solutions of the novel
compounds of the formula (I) in sterile aqueous solution, aqueous
propylene glycol or sesame or peanut oil may be employed. Such
aqueous solutions should be suitably buffered if necessary and the
liquid diluent first rendered isotonic with sufficient saline or
glucose. The aqueous solutions are particularly suitable for
intravenous, intramuscular, subcutaneous and intraperitoneal
administration. The sterile aqueous media employed are all readily
available by standard techniques known to those skilled in the
art.
[0236] Suitable pharmaceutical carriers include inert solid
diluents or fillers, sterile aqueous solution and various organic
solvents. Examples of solid carriers are lactose, terra alba,
sucrose, cyclodextrin, talc, gelatine, agar, pectin, acacia,
magnesium stearate, stearic acid and lower alkyl ethers of
cellulose. Examples of liquid carriers are syrup, peanut oil, olive
oil, phospholipids, fatty acids, fatty acid amines, polyoxyethylene
and water. Similarly, the carrier or diluent may include any
sustained release material known in the art, such as glyceryl
mono-stearate or glyceryl distearate, alone or mixed with a wax.
The pharmaceutical compositions formed by combining the novel
compounds of the formula (I) and the pharmaceutically acceptable
carriers are then readily administered in a variety of dosage forms
suitable for the disclosed routes of administration. The
formulations may conveniently be presented in unit dosage form by
methods known in the art of pharmacy.
[0237] Formulations of the present invention suitable for oral
administration may be presented as discrete units such as capsules
or tablets, each containing a predetermined amount of the active
ingredient, and which may include a suitable excipient.
Furthermore, the orally available formulations may be in the form
of a powder or granules, a solution or suspension in an aqueous or
non-aqueous liquid, or an oil-in-water or water-in-oil liquid
emulsion.
[0238] If a solid carrier is used for oral administration, the
preparation may be tabletted, placed in a hard gelatine capsule in
powder or pellet form or it can be in the form of a troche or
lozenge. The amount of solid carrier will vary widely but will
usually be from about 25 mg to about 1 g. If a liquid carrier is
used, the preparation may be in the form of a syrup, emulsion, soft
gelatine capsule or sterile injectable liquid such as an aqueous or
non-aqueous liquid suspension or solution.
[0239] A typical tablet that may be prepared by conventional
tabletting techniques may contain:
2 Core: Active compound (as free compound or salt thereof) 5.0 mg
Lactosum Ph. Eur. 67.8 mg Cellulose, microcryst. (Avicel) 31.4 mg
Amberlite .RTM. IRP88* 1.0 mg Magnesii stearas Ph. Eur. q.s.
Coating: Hydroxypropyl methylcellulose approx. 9 mg Mywacett 9-40
T** approx. 0.9 mg *Polacrillin potassium NF, tablet disintegrant,
Rohm and Haas. **Acylated monoglyceride used as plasticizer for
film coating.
[0240] If desired, the pharmaceutical composition of the invention
may comprise the compound of the formula (I) in combination with
further pharmacologically active substances such as those described
in the foregoing.
EXAMPLES
[0241] The compounds according to the invention may be prepared
according to the general procedures outlined below. All starting
materials are known or may easily be prepared from known starting
materials.
[0242] All temperatures are set forth in degrees Celsius and unless
otherwise indicated, all parts and percentages are by weight when
referring to yields and all parts are by volume when referring to
solvents and eluents.
[0243] The following terms are intended to have the following
meanings:
[0244] DCM dichloromethane
[0245] DMF: N,N-dimethylformamide
[0246] DMSO: dimethyl sulphoxide
[0247] M.p.: melting point
[0248] TFA: trifluoroacetic acid
[0249] THF: tetrahydrofuran
[0250] EDAC: 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide
hydrochloride
[0251] HOBt: 1-hydroxybenzotriazole
[0252] HOAt: 3-hydroxy-3H-[1,2,3]triazolo[4,5-b]pyridine
[0253] EGTA: ethylene glycol bis(O-aminoethyl ether)
N,N,N'N'-tetracetic acid
[0254] IBMX: isobutylmethylxanthine
[0255] General Procedure (A) for Synthesis of Compounds of the
General Formula (Ia) According to the Invention: 19
[0256] wherein D and E are as defined for formula (I) and R is
C.sub.1-6-alkyl.
[0257] Intermediates to be Used in Step A:
[0258] 4-Cyclohex-1-enylphenylamine is described in the literature:
v Brown et al., Justus Liebigs, Ann. Chem., 1929 (472), 1-89.
[0259] 4-(4-tert-Butylcyclohex-1-enyl)Dhenylamine was prepared by
similar method. p 4-Cyclohexylphenylamine is commercially available
(e.g. from Lancaster or Avocado).
[0260] Bicyclohexyl-4-ylamine is described in the literature: H.
Booth et al., J. Chem. Soc. (B), 1971, 1047-1050.
[0261] Intermediates to be Used in step D:
3-Amino-2,2-difluoropropionic acid methyl ester hydrochloride
[0262] 20
Step 1: 2,2-Difluorosuccinic acid 1-methyl ester
[0263] 21
[0264] Commercially available 2,2-difluorosuccinic acid (8.0 g,
51.9 mmol) was dissolved in THF and cooled on ice before addition
of 1,3-dicyclohexylcarbodiimide (11.8 g, 57.2 mmol). The mixture
was stirred at room temperature for 3 hours, cooled and filtered.
The filtrate was concentrated in vacuo. The residue was stirred
with methanol (50 ml) for 3 hours at room temperature and then
solvents were removed in vacuo to afford the crude ester as an oil
(8.4 g). The oil was purified by column chromatography on silica
gel column and elution with a 0-100% ethyl acetate/toluene gradient
to afford 2,2-difluorosuccinic acid 1-methyl ester as a colorless
oil (6.8 g).
[0265] .sup.1H NMR (DMSO-d.sub.6): .delta.3.37 (t, 2H), 3.85 (s,
3H), 13.22 (br s, 1H).
Step 2: 3-Benzyloxycarbonylamino-2,2-difluoropropionic acid methyl
ester
[0266] 22
[0267] 2,2-Difluorosuccinic acid 1-methyl ester (6.7 g, 39.9 mmol)
and thionyl chloride were heated at reflux for 1 hour. The mixture
was co-evaporated with dry toluene (3.times.) to leave an oil. The
residue was dissolve in dry toluene (100 ml) and heated to
70.degree. C. before adding trimethylsilyl azide (6.3 ml, 47.5
mmmol) over 30 min. The mixture was stirred for 16 hours at
80.degree. C. and evaporated to an oil. The residue was dissolved
in dry toluene, charged with dry benzyl alcohol (5.0 ml, 50.2 mmol)
and stirring was continued for 16 hours at 80.degree. C. The
mixture was concentrated in vacuo and the residue was partitioned
between ethyl acetate (100 ml) and water (100 ml). The organic
layer was washed twice with water and dried over magnesium
sulphate, filtered and concentrated in vacuo. The residue was
subjected to flash column chromatography using DCM as eluent (200
ml) removing highly polar impurities before a second column
chromatography eluting with 10% ethyl acetate/toluene. Evaporation
of proper fractions in vacuo afforded 3.3 g (23%) of
3-benzyloxycarbonylamino-2,2-difluoropropionic acid methyl ester as
an oil.
[0268] .sup.1H NMR (DMSO-d.sub.6): .delta.3.62-3.77 (m, 2H), 3.78
(s, 3H), 5.05 (s, 2H), 7.35 (s, 5H), 7.90 (t, 1H).
Step 3: 3-Benzyloxycarbonylamino-2,2-difluoropropionic acid
[0269] 3-Benzyloxycarbonylamino-2,2-difluoropropionic acid methyl
ester (3.3 g, 12.1 mmol), THF (50 ml), methanol (50 ml) and 1N
sodium hydroxide solution (50ml) were mixed and stirred for 16
hours at room temperature. After adjusting the pH to acidic
reaction and removal of organic solvent by evaporation, the aqueous
residue was extracted twice with ethyl acetate. The combined
organic layers were washed twice with 5% brine, dried over
magnesium sulphate and filtered through a short silica column
before concentration in vacuo to afford 2.6 g (83%) of
3-benzyloxycarbonylamino-2,2-difluoropropionic acid as an oil.
[0270] .sup.1H NMR (DMSO-d.sub.6): .delta.3.67 (dt, 2H), 5.05 (s,
2H), 7.35 (s, 5H), 7.83 (t, 1H).
Step 4: 3-Amino-2,2-difluoropropionic acid
[0271] 23
[0272] 3-Benzyloxycarbonylamino-2,2-difluoropropionic acid (2.6 g,
10.0 mmol) was dissolved in abs. ethanol (100 ml) and Pd--C, 10%
(100 mg) was added under a nitrogen atmosphere. The mixture is
hydrogenated for 3 hours at 40 psi, filtered on micro filter,
washed with abs. ethanol and a small amount of water. The
colourless solution was concentrated to approx. 10 ml and acetone
(75 ml) was added dropwise. The precipitate was filtered off and
dried to afford 0.8 g (64%) of 3-amino-2,2-difluoropropi- onic acid
as white crystals. M.p. 125-127.degree. C.
[0273] .sup.1H NMR (DMSO-d.sub.6): .delta.3.28 (t, 2H), 8.42 (bs,
2H).
Step 5: 3-Amino-2,2-difluoropropionic acid methyl ester
hydrochloride
[0274] 24
[0275] Thionyl chloride (1.2 ml, 16.5 mmol) was added dropwise to
ice cold dry methanol (20 ml) over 10 min.
3-Amino-2,2-difluoropropionic acid (0.7 g, 5.6 mmol) was added and
stirring was continued in the cold for additional 15 min. The
cooling source was removed and stirring was continued at room
temperature for 16 hours. The mixture was co-evaporated with
toluene (3.times.) to give a white suspension, which upon
filtration and drying at 40.degree. C. for 16 hours gave 0.96 g
(98%) of the title compound.
[0276] .sup.1H NMR (DMSO-d.sub.6): .delta.3.65 (t, 2H), 3.89 (s,
3H), 9.07 (br s, 3H).
[0277] .sup.13C NMR (DMSO-d.sub.6): .delta.39.8 (CH.sub.2), 54.6
(CH.sub.3), 112.8 (CF.sub.2), 162 (C.dbd.O).
[0278] Using the general procedure (A) the following compounds have
been prepared. The procedure is further illustrated by the
following example.
Example 1
3-{4-[1-(4-Cyclohex-1-enylphenyl)-3-(3,5-dichlorophenyl)ureidomethyl]benzo-
ylamino}-2,2-difluoropropionic acid)
[0279] 25
[0280] Step A:
[0281] To a solution of E-NH.sub.2 (eg 4-cyclohexenylaniline,
prepared as described above) (0.023 mol) and methyl
4-formylbenzoate (3.77 g, 0.023 mol) in DCM (50 ml) and methanol
(15 ml) is added a catalytic amount of acetic acid. After stirring
the solution for 3 hours, Na(OAc).sub.3BH (24 g, 0.115 mol) is
added. The reaction is allowed to stir at room temperature for 16
hours. The reaction mixture is diluted with ethyl acetate and
washed with aqueous sodium bicarbonate (3.times.), brine
(2.times.), dried over magnesium sulphate, filtered, and
concentrated to give a solid. The crude product is purified by
column chromatography eluting with mixtures of ethyl acetate and
heptane to give 4-[(4-cyclohex-1-enylphenylamino)methyl]benzoic
acid methyl ester (A1) (5 g, 0.015 mol).
[0282] .sup.1H NMR (DMSO-d.sub.6): .delta.1.56 (m, 2H), 1.67 (m,
2H), 2.11 (m, 2H), 2.25 (m, 2H), 3.81 (s, 3H), 4.34 (d, 2H), 5.89
(t, 1H), 6.34 (t, 1H), 6.49 (d, 2H), 7.10 (d, 2H), 7.47 (d, 2H),
7.90 (d, 2H).
[0283] Step B:
[0284] The above 4-[(4-cyclohex-1-enylphenylamino)methyl]benzoic
acid methyl ester (5 g, 0.015 mol) is dissolved in anhydrous DCM
and diisopropylethylamine ( 5.8 g, 0.045 mol) is added. To this
solution is added an isocyanate (D-N.dbd.C.dbd.O) (0.018 mol), eg.
3,5-dichlorophenyl isocyanate. After stirring the reaction mixture
for 3 hours, the solution is diluted with ethyl acetate and washed
with 1N hydrochloric acid (2.times.), water, brine, dried over
magnesium sulphate, filtered, and concentrated in vacuo. The
residue is purified by column chromatography eluting with mixtures
of ethyl acetate and heptane to give (A2).
[0285] Step C:
[0286] To a solution of (A2) (2 mmol) in THF (30 ml) and methanol
(10 ml) is added an excess of 2 M lithium hydroxide (10 ml). After
stirring the reaction mixture for 3 hours, the solution is
concentrated. The residue is taken up in ethyl acetate and washed
with 1 N hydrochloric acid (2.times.), water (2.times.), brine, and
dried over magnesium sulphate. Evaporation of the solvent affords
(A3).
[0287] Step D:
[0288] To a solution of (A3) (0.81 mmol) in DMF (4 ml) are added
3-[(dimethyliminium)(dimethylamino)methyl]-1,2,3-benzotriazol-1-ium-1-ola-
te hexafluorophosphate (0.37 g, 0.90 mmol), diisopropylethylamine
(0.30 g, 2.4 mmol), and 3-amino-2,2-difluoropropionic acid methyl
ester hydrochloride (2.4 mmol). After stirring the solution for 16
hours, the reaction is diluted with ethyl acetate and washed with 1
N hydrochloric acid (3.times.), brine (3.times.), dried over
magnesium sulphate, filtered, and concentrated. The residue is
purified by column chromatography and eluted with mixtures of ethyl
acetate and heptane to afford (A4).
[0289] Step E:
[0290] (A4) is dissolved in THF (6 ml) and methanol (3 ml). A
solution of 2 M lithium hydroxide (3 ml) is then added and the
reaction is stirred at room temperature for 30 min. The solvents
are evaporated under reduced pressure. The residue is taken up in
ethyl acetate and washed with 1 N hydrochloric acid (2.times.),
brine (2.times.), dried over magnesium sulphate, filtered, and
concentrated in vacuo to afford the title compound of the general
formula (1a).
[0291] .sup.1H NMR (DMSO-d.sub.6): .delta.1.60 (m, 2H), 1.70 (m,
2H), 2.18 (m, 3H), 2.33 (m, 2H), 3.93 (m, 2H), 6.18 (m, 1H),
7.12-7.22 (m, 3H), 7.37 (d, 2H), 7.41 (d, 2H), 7.62 (d, 2H), 7.78
(s, 2H), 8.54 (s, 1H), 8.85 (t,1H).
Example 2
3-{4-[1-[4-(4-tert-Butylcyclohex-1-enyl)phenyl]-3-(3,5-dichlorophenyl)urei-
domethyl]benzoyl-amino}-2,2-difluororroiionic acid
[0292] 26
[0293] .sup.1H-NMR (DMSO-d.sub.6): .delta.0.88 (s, 9H), 1.23 (m,
2H), 1.92 (bm, 2H), 2.13-2.43 (m, 3H), 3.93 (m, 2H), 4.94 (s, 2H),
6.18 (m, 1H), 7.13 (m, 1H), 7.17 (d, 2H), 7.32 (d, 2H), 7.39 (d,
2H), 7.62 (s, 2H), 7.74 (d, 2H), 8.47 (t, 1H), 8.53 (s, 1H), 8.83
(t, 1H); M.p. 125-127.degree. C.
[0294] General Procedure (B) for the Synthesis of Compounds of the
General Formula (1a) According to the Invention: 27
[0295] wherein D and E are as defined for formula (I) and R is
C.sub.1-6-alkyl.
Preparation of methyl
2,2-difluoro-3-[(4-formylbenzoyl)amino]proiionate as starting
material:
[0296] To a solution of the 4-formylbenzoic acid in a suitable
solvent such as DCM, DMF or THF is added diisopropylethylamine (3
eq) and
3-[(dimethyliminium)(dimethylamino)methyl]-1,2,3-benzotriazol-1-ium-1-ola-
te hexafluorophosphate (1.1 eq). The reaction is allowed to stir
for 30 min before ethyl or methyl 3-amino-2,2-difluoropropionate
hydrochloride (1.1 eq) is added.
[0297] The solution is stirred at room temperature for 4 hours. The
solvents are evaporated under reduced pressure. The residue is
taken up in ethyl acetate and 1N hydrochloric acid. The organic
layer is separated and washed with water (2.times.), aqueous sodium
hydrogen carbonate (3.times.), brine (2.times.), dried over
magnesium sulphate and concentrated to give the desired
product.
[0298] Step A:
[0299] The aldehyde (0.011 mmol) in DCM is dispensed into the wells
of a deepwell plate containing the desired amines (E-NH.sub.2) in
DCM. To this solution is added sodium triacetoxyborohydride (1.5
eq) followed by a catalytic amount of acetic acid. The reaction is
left to proceed for 15 hours.
[0300] Step B:
[0301] To the resulting amines from step A is added the desired
isocyanate (D-N.dbd.C.dbd.O) (0.011 mmol) in DCM. The reaction
mixtures are agitated for three hours and the solvents are removed
under reduced pressure to give the desired ureas.
[0302] Step C:
[0303] The residue obtained in step B is dissolved in DMF and
aqueous 2 M lithium hydroxide (10 eq.) is added into each reaction
well. The samples are shaken overnight and filtered. Aqueous 1 N
hydrochloric acid is then added to give the desired carboxylic
acids.
[0304] General Procedure (C) for Solid Phase Synthesis of Compounds
of the General Formula (1a) According to the Invention: 28
[0305] wherein D and E are as defined for formula (I), R is
C.sub.1-6-alkyl and Resin denotes a polystyrene resin with a linker
such as the Wang linker: 29
[0306] wherein PS denotes polystyrene.
[0307] Step A:
[0308] The reaction is known (Wang S. J., J. Am. Chem. Soc. 95,
1328, 1973) and is generally performed by stirring polystyrene
resin loaded with a linker such as the Wang linker with a 4-10
molar excess of Fmoc-protected amino acid activated with a 2-5
molar excess of diisopropylcarbodiimide or dicyclohexylcarbodiimide
in the presence of a catalyst such as N,N-4-dimethylaminopyridine.
The esterification is carried out in a solvent such as THF,
dioxane, toluene, DCM, DMF, N-methylpyrrolidinone or a mixture of
two or more of these. The reactions are performed between 0.degree.
C. to 80.degree. C., preferably between 20.degree. C. to 40.degree.
C. When the esterification is complete excess of reagents is
removed by filtration. The resin is successively washed with the
solvent used in the reaction, followed by washings with methanol.
The resin bound product can be further dried and analyzed.
[0309] Step B:
[0310] The Fmoc protecting group is removed using a solution of 20%
piperidine in DMF which is added to the resin and vortexed for 0.5
hours. After draining the resin is washed with DMF containing HOBt
(50 mg/ml) and DMF. The acylation (The combinatorial index, Ed.
Bunin, B. A. 1998, Academic Press, p. 78) is performed by adding an
excess of acid in a solvent such as DMF, N-methylpyrrolidinone,
THF, DCM, 1,2-dichloroethane, acetonitrile, DMSO or a mixture of
two or more of these, optionally in the presence of a base such as
N-methylmorpholine, triethylamine, diisopropylethylamine,
dicyclohexylmethylamine or another tertiary amine, followed by a
coupling reagent such as dicyclohexylcarbodiimide,
diisopropylcarbodiimide, 1,1'-carbonyldiimidazole,
2-(1H-9-azabenzotriazole-1-yl)-1,1,3,3-tetramethyluronium
hexafluorophosphate or bromo-tris-pyrrolidinophosphonium
hexafluorophosphate in a solvent such as DMF,
N-methylpyrrolidinone, THF, DCM, 1,2-dichloroethane, acetonitrile,
DMSO or a mixture of two or more of these, optionally in the
presence of a side reaction inhibitor such as
3-hydroxy-4-oxo-3,4-dihydro-1,2,3-benzotriazine, HOBt or
1-hydroxy-7-azabenzotriazole. The reaction is performed between
20.degree. C. and 40.degree. C., preferably at 25.degree. C. Excess
reagents are filtered off and the resin is washed several times
with the solvent used during the reaction.
[0311] Step C:
[0312] The reaction is generally known (The combinatorial index,
Ed. Bunin, B. A. 1998, Academic Press, p. 133) and is generally
performed by stirring resin bound aldehyde or ketone with an excess
of amine at low pH (by addition of an acid, such as acetic acid or
formic acid) in a solvent such as THF, DMF, N-methylpyrrolidinone,
methanol, ethanol, DMSO, DCM, 1,2-dichloroethane, trimethyl
orthoformate, triethyl orthoformate, or a mixture of two or more of
these. As reducing agent sodium cyanoborohydride may be used. The
reaction is performed between 20.degree. C. and 120.degree. C.,
preferably at 25.degree. C.
[0313] Step D:
[0314] The reaction is generally known (Organic synthesis on solid
phase. Dorwald, F. Z. 2000, Wiley VCH, p. 331) and is generally
performed by stirring resin bound amine with an excess of
isocyanate in a solvent such as THF, DMF, N-methylpyrrolidinone,
DCM, 1,2-dichloroethane, toluene or a mixture of two or more of
these. The reaction is performed between 20.degree. C. and
80.degree. C., preferably at 25.degree. C.
[0315] Step E:
[0316] The reaction is known (The combinatorial index, Ed. Bunin B.
A., 1998, Academic press, p.21) and is generally performed by
stirring resin bound intermediate obtained in step D with a 50-95%
solution of TFA. The final cleavage is carried out in a solvent
such as THF, DCM, 1,2-dichloroethane, 1,3-dichloropropane, toluene
or a mixture of two or more of these. The reaction is performed
between 0.degree. C. and 80.degree. C., preferably between
20.degree. C. and 40.degree. C. When the reaction is complete the
product is removed by filtration. The resin is successively washed
with DCM. The product and washings are collected. The solvent is
removed and the product is dried in vacuo.
[0317] Optionally, the resin can be a 2-chlorotrityl resin. In that
case, step A is a nucleophilic reaction of Fmoc-protected beta
alanine with 2-chlorotritylchloride resin in the presence of a
base, such as triethylamine or N,N-diisopropyl-N-ethylamine. All
other steps are identical to those described above with the
exception of step E, cleavage from the resin. This can be done with
only 5% TFA in DCM.
[0318] More specifically, preparation of the compounds of the
invention according to the general procedure (C) may be prepared as
follows:
Step A: Resin Bound Fmoc .beta.-alanine (C1)
[0319] 150 .mu.mol Fmoc .alpha.,.alpha.-difluoro-.beta.-alanine is
dissolved in 500 .mu.l of a mixture of DMF and
diisopropylethylamine (430:70) and added to 50 mg polystyrene resin
functionalised with a Wang linker. 200 .mu.mol PyBrOP dissolved in
DMF (500 .mu.l) is added. After shaking the suspension for,4 hours
at 25.degree. C., the resin is isolated by filtration and washed
with 3.times.1 ml DMF.
Step B: Resin Bound 3-(4-formylbenzoylamino)propionic acid (C2)
[0320] To the above resin bound Fmoc
.alpha.,.alpha.-difluoro-.beta.-alani- ne (C1) is added 1000 .mu.l
of a 20% solution of piperidine in DMF. Upon shaking for 30 min,
the resin is drained and washed with 1 ml DMF containing HOBt (50
mg/ml) and DMF (2.times.1 ml). Then 200 .mu.mol 4-formylbenzoic
acid (30 mg) and diisopropylethylamine (70 .mu.l) are dissolved in
DMF (430 .mu.l) and added to the resin followed by 200 .mu.mol
PyBrOP dissolved in DMF (500 .mu.l). The mixture is shaken for 4
hours at 25.degree. C. followed by filtration and washing of the
resin with DMF (3.times.1 ml) and trimethylorthoformate (1.times.1
ml).
Step C: (C3)
[0321] The above resin bound 3-(4-formylbenzoylamino)propionic acid
(C2) (50 mg) is treated with a solution of E-NH.sub.2 (500 .mu.mol)
in a mixture of DMF (500 .mu.l) and trimethylorthoformate (500
.mu.l). Glacial acetic acid (100 .mu.l) is added and the mixture is
shaked for 1 hour at 25.degree. C. Sodium cyanoborohydride (750
.mu.mol) suspended in a mixture of DMF and trimethylorthoformate
(1:1, 1 ml) is added and the mixture is vortexed at 25.degree. C.
for 16 hours followed by filtration and washing with a mixture of
DMF and water (4:1, 2.times.1 ml) followed by 3.times.1 ml DMF and
2.times.1 ml DCM to afford (C3).
[0322] Step D: (C4)
[0323] 200 .mu.mol isocyanate (D-N.dbd.C.dbd.O) dissolved in 500
.mu.l DCM is added to (C3) (50 mg). Shaking the mixture for 16
hours at 25.degree. C. followed by filtration and washing of the
resin with 4.times.1 ml DMF, 2.times.1 ml water, 3.times.1 ml THF
and 5.times.1 ml DCM afford (C4).
[0324] Step E:
[0325] (C4) (50 mg) is treated with 1 ml 50% TFA in DCM for 1 hour
at 25.degree. C. The product is filtered off and the resin is
washed with 1 ml DCM. The combined extracts are concentrated in
vacuo. The residue is purified by chromatography and/or
crystallisation to afford the compounds of the general formula (Ia)
according to the invention.
[0326] The following preferred compounds are within the scope of
the present invention and may be prepared according to the general
procedures disclosed above.
303132333435363738394041424344454647
[0327] Pharmacological Methods
[0328] In the following section binding assays as well as
functional assays useful for evaluating the efficiency of the
compounds of the invention are described.
[0329] Binding of compounds to the glucagon receptor may be
determined in a competition binding assay using the cloned human
glucagon receptor.
[0330] Antagonism may be determined as the ability of the compounds
to inhibit the amount of cAMP formed in the presence of 5 nM
glucagon.
[0331] Glucagon Binding Assay (I)
[0332] Receptor binding is assayed using cloned human receptor (Lok
et al., Gene 140, 203-209 (1994)). The receptor inserted in the
pLJ6' expression vector using EcoRI/SSt1 restriction sites (Lok et
al.) is expressed in a baby hamster kidney cell line (A3 BHK
570-25). Clones are selected in the presence of 0.5 mg/ml G-418 and
are shown to be stable for more than 40 passages. The Kd is shown
to be 0.1 nM.
[0333] Plasma membranes are prepared by growing cells to
confluence, detaching them from the surface and resuspending the
cells in cold buffer (10 mM tris/HCl, pH 7.4 containing 30 mM NaCl,
1 mM dithiothreitol, 5 mg/l leupeptin (Sigma), 5 mg/l pepstatin
(Sigma), 100 mg/l bacitracin (Sigma) and 15 mg/l recombinant
aprotinin (Novo Nordisk A/S)), homogenization by two 10-s bursts
using a Polytron PT 10-35 homogenizer (Kinematica), and
centrifugation upon a layer of 41 w/v % sucrose at 95.000.times.g
for 75 min. The white band located between the two layers is
diluted in buffer and centrifuged at 40.000.times.g for 45 min. The
precipitate containing the plasma membranes is suspended in buffer
and stored at -80.degree. C. until use.
[0334] Glucagon is iodinated according to the chloramine T method
(Hunter and Greenwood, Nature 194, 495 (1962)) and purified using
anion exchange chromatography (Jorgensen et al., Hormone and Metab.
Res. 4, 223-224 (1972). The specific activity is 460 .mu.Ci/.mu.g
on the day of iodination. Tracer is stored at -18.degree. C. in
aliquots and are used immediately after thawing.
[0335] Binding assays are carried out in triplicate in filter
microtiter plates (MADV N65, Millipore). The buffer used in this
assay is 50 mM HEPES, 5 mM EGTA, 5 mM MgCl.sub.2, 0.005% tween 20,
pH 7.4. Glucagon is dissolved in 0.05 M HCl, added an equal amount
(w/w) of human serum albim and freeze-dried. On the day of use, it
is dissolved in water and diluted in buffer to the desired
concentrations.
[0336] Test compounds are dissolved and diluted in DMSO. 140 .mu.l
buffer, 25 .mu.l glucagon or buffer, and 10 .mu.l DMSO or test
compound are added to each well. Tracer (50.000 cpm) is diluted in
buffer and 25 .mu.l are added to each well. 1-4 .mu.g freshly
thawed plasma membrane protein diluted in buffer is then added in
aliquots of 25 .mu.l to each well. Plates are incubated at
30.degree. C. for 2 hours. Non-specific binding is determined with
10.sup.-6 M of glucagon. Bound tracer and unbound tracer are then
separated by vacuum filtration (Millipore vacuum manifold). The
plates are washed with 2.times.100 .mu.l buffer/ well. The plates
are air dried for a couple of hours, whereupon the filters are
separated from the plates using a Millipore Puncher. The filters
are counted in a gamma counter.
[0337] Functional Assay (I)
[0338] The functional assay is carried out in 96 well microtiter
plates (tissue culture plates, Nunc). The resulting buffer
concentrations in the assay are 50 mM tris/HCl, 1 mM EGTA, 1.5 mM
magnesium sulphate, 1.7 mM ATP, 20 .mu.pM GTP, 2 mM IBMX, 0.02%
tween-20 and 0.1% human serum albim. pH is 7.4. Glucagon and
proposed antagonist are added in aliquots of 35 .mu.l diluted in 50
mM tris/HCl, 1 mM EGTA, 1.85 mM magnesium sulphate, 0.0222%
tween-20 and 0.111% human serum albim, pH 7.4.20 .mu.l of 50 mM
tris/HCl, 1 mM EGTA, 1.5 mM magnesium sulphate, 11.8 mM ATP, 0.14
mM GTP, 14 mM IBMX and 0.1% human serum albim, pH 7.4 is added. GTP
is dissolved immediately before the assay.
[0339] 50 .mu.l containing 5 .mu.g of plasma membrane protein is
added in a tris/HCl, EGTA, magnesium sulphate, human serum albumin
buffer (the actual concentrations are dependent upon the
concentration of protein in the stored plasma membranes).
[0340] The total assay volume is 140 .mu.l. The plates are
incubated for 2 hours at 37.degree. C. with continuous shaking.
Reaction is terminated by addition of 25 .mu.l 0.5 N HCl. cAMP is
measured by the use of a scintillation proximity kit
(Amersham).
[0341] Glucagon Binding Assay (II)
[0342] BHK (baby hamster kidney cell line) cells are transfected
with the human glucagon receptor and a membrane preparation of the
cells is prepared. Wheat Germ Agglutinin derivatized SPA beads
containing a scintillant (WGA beads) (Amersham) bound the
membranes. .sup.125I-glucagon bound to human glucagon receptor in
the membranes and excited the scintillant in the WGA beads to light
emission. Glucagon or samples binding to the receptor competed with
.sup.125I-glucagon.
[0343] All steps in the membrane preparation are kept on ice or
performed at 4.degree. C. BHK cells are harvested and centrifuged.
The pellet is resuspended in homogenisation buffer (25 mM HEPES,
pH=7.4, 2.5 mM CaCl.sub.2, 1.0 mM MgCl.sub.2, 250 mg/l bacitracin,
0.1 mM Pefabloc), homogenised 2.times.10 sec using Polytron 10-35
homogenizer (Kinematica) and added the same amount of
homogenisation buffer as used for resuspension. After
centrifugation (15 min at 2000.times.g) the supernatant is
transferred to cold centrifuge tubes and centrifuged for 45 min at
40.000.times.g. The pellet is resuspended in homogenisation buffer,
homogenised 2.times.10 sec (Polytron) and additional homogenisation
buffer is added. The suspension is centrifuged for 45 min at
40.000.times.g and the pellet is resuspended in resuspension buffer
(25 mM HEPES, pH=7.4, 2.5 mM CaCl.sub.2, 1.0 mM MgCl.sub.2) and
homogenised 2.times.10 sec (Polytron). The protein concentration is
normally around 1.75 mg/ml. Stabilisation buffer (25 mM HEPES,
pH=7.4, 2.5 mM CaCl.sub.2, 1.0 mM MgCl.sub.2, 1% bovine serum
albumin, 500 mg/l bacitracin, 2.5 M sucrose) is added and the
membrane preparation is stored at -80.degree. C.
[0344] The glucagon binding assay is carried out in opti plates
(Polystyrene Microplates, Packard). 50 .mu.l assay buffer (25 mM
HEPES, pH=7.5, 2.5 mM CaCl.sub.2, 1.0 mM MgCl.sub.2, 0.003%
Tween-20, 0.005% bacitracin, 0.05% sodium azide) and 5 .mu.l
glucagon or test compound (in DMSO) are added to each well. 50
.mu.l tracer (.sup.125I-porcine glucagon, 50.000 cpm) and 50 .mu.l
membranes (7.5 .mu.g) containing the human glucagon receptor are
then added to the wells. Finally 50 .mu.l WGA beads containing 1 mg
beads are transferred to the well. The plates are incubated for 4
hours on a shaker and then settled for 8-48 hours. The opti plates
are counted in a Topcounter. Non-specific binding is determined
with 500 nM of glucagon.
[0345] The compounds according to the examples showed IC.sub.50
values below 1000 nM when tested in the glucagon binding assay
(II).
[0346] GIP Binding Assay
[0347] BHK (baby hamster kidney cell line) cells are transfected
with the human GIP receptor and a membrane preparation of the cells
is prepared. Wheat Germ Agglutinin derivatized SPA beads containing
a scintillant (WGA beads) (Amersham) bound the membranes.
.sup.125I-GIP bound to human GIP receptor in the membranes and
excited the scintillant in the WGA beads to light emission. GIP or
samples binding to the receptor competed with .sup.125I-GIP.
[0348] All steps in the membrane preparation are kept on ice or
performed at 4.degree. C. BHK cells are harvested and centrifuged.
The pellet is resuspended in homogenisation buffer (25 mM HEPES,
pH=7.4, 2.5 mM CaCl.sub.2, 1.0 mM MgCl.sub.2, 250 mg/l bacitracin,
0.1 mM Pefabloc), homogenised 2.times.10 sec using Polytron 10-35
homogenizer (Kinematica) and added the same amount of
homogenisation buffer as used for resuspension. After
centrifugation (15 min at 2000.times.g) the supernatant is
transferred to cold centrifuge tubes and centrifuged for 45 min at
40.000.times.g. The pellet is resuspended in homogenisation buffer,
homogenised 2.times.10 sec (Polytron) and additional homogenisation
buffer is added. The suspension is centrifuged for 45 min at
40.000.times.g and the pellet is resuspended in resuspension buffer
(25 mM HEPES, pH=7.4, 2.5 mM CaCl.sub.2, 1.0 mM MgCl.sub.2) and
homogenised 2.times.10 sec (Polytron). The protein concentration is
normally around 1.75 mg/ml. Stabilisation buffer (25 mM HEPES,
pH=7.4, 2.5 mM CaCl.sub.2, 1.0 mM MgCl.sub.2, 1% bovine serum
albumin, 500 mg/l bacitracin, 2.5 M sucrose) is added and the
membrane preparation is stored at -80.degree. C.
[0349] The GIP binding assay is carried out in opti plates
(Polystyrene Microplates, Packard). 50 .mu.l assay buffer (25 mM
HEPES, pH=7.5, 2.5 mM CaCl.sub.2, 1.0 mM MgCl.sub.2, 0.003%
Tween-20, 0.005% bacitracin, 0.05% sodium azide) and 5 .mu.l GIP or
test compound (in DMSO) are added to each well. 50 .mu.l tracer
(.sup.125I-porcine GIP, 50.000 cpm) and 50 .mu.l membranes (20
.mu.g) containing the human GIP receptor are then added to the
wells. Finally 50 .mu.l WGA beads containing 1 mg beads are
transferred to the well. The plates are incubated for 3.5 hours on
a shaker and then settled for 8-48 hours. The opt plates are
counted in a Topcounter. Non-specific binding is determined with
500 nM of GIP.
[0350] The compounds according to the examples show a higher
affinity for the glucagon receptor compared to the GIP
receptor.
* * * * *