U.S. patent application number 10/541493 was filed with the patent office on 2006-03-09 for kynurenine 3-hydroxylase inhibitors for the treatment of diabetes by increasing the number of islets of langerhans cells.
This patent application is currently assigned to MERCK PATENT GMBH. Invention is credited to Annick Arbellot De Vacqueur, Valerie Autier, Catherine Kargar, Micheline Kergoat, Dominique Marais, Gerard Moinet.
Application Number | 20060052456 10/541493 |
Document ID | / |
Family ID | 32598906 |
Filed Date | 2006-03-09 |
United States Patent
Application |
20060052456 |
Kind Code |
A1 |
Autier; Valerie ; et
al. |
March 9, 2006 |
Kynurenine 3-hydroxylase inhibitors for the treatment of diabetes
by increasing the number of islets of langerhans cells
Abstract
The present invention relates to the use of at least one
compound with inhibitory activity on kynurenine 3-hydroxylase, for
the preparation of a medica ment for increasing the number of
islets of Langerhans cells, which is intended especially for the
prevention and/or treatment of diabetes.
Inventors: |
Autier; Valerie; (Gif sur
Yvette, FR) ; Arbellot De Vacqueur; Annick; (Fontenay
les Briis, FR) ; Kergoat; Micheline; (Bures sur
Yvette, FR) ; Moinet; Gerard; (Orsay, FR) ;
Marais; Dominique; (Meulan, FR) ; Kargar;
Catherine; (Versailles, FR) |
Correspondence
Address: |
MILLEN, WHITE, ZELANO & BRANIGAN, P.C.
2200 CLARENDON BLVD.
SUITE 1400
ARLINGTON
VA
22201
US
|
Assignee: |
MERCK PATENT GMBH
FANKLINE STRASSE 250
DARMSTADT GERMAY
DE
64293
|
Family ID: |
32598906 |
Appl. No.: |
10/541493 |
Filed: |
December 18, 2003 |
PCT Filed: |
December 18, 2003 |
PCT NO: |
PCT/EP03/14538 |
371 Date: |
July 7, 2005 |
Current U.S.
Class: |
514/570 ;
514/571 |
Current CPC
Class: |
A61P 3/08 20180101; A61K
31/22 20130101; A61K 31/235 20130101; A61P 43/00 20180101; A61P
3/06 20180101; A61P 3/04 20180101; A61K 31/194 20130101; A61K
31/192 20130101; A61P 9/12 20180101; A61K 31/22 20130101; A61K
31/19 20130101; A61K 31/194 20130101; A61P 37/06 20180101; A61K
45/06 20130101; A61K 31/06 20130101; A61K 31/235 20130101; A61K
31/216 20130101; A61K 31/55 20130101; A61K 31/06 20130101; A61K
31/19 20130101; A61K 31/192 20130101; A61K 31/55 20130101; A61K
2300/00 20130101; A61K 2300/00 20130101; A61P 3/10 20180101; A61K
31/216 20130101; A61K 31/426 20130101; A61K 31/00 20130101; A61K
31/426 20130101; A61K 2300/00 20130101; A61K 2300/00 20130101; A61K
2300/00 20130101; A61K 2300/00 20130101; A61K 2300/00 20130101;
A61K 2300/00 20130101; A61K 2300/00 20130101 |
Class at
Publication: |
514/570 ;
514/571 |
International
Class: |
A61K 31/192 20060101
A61K031/192; A61K 31/19 20060101 A61K031/19 |
Foreign Application Data
Date |
Code |
Application Number |
Jan 7, 2003 |
FR |
03-00-107 |
Dec 4, 2003 |
FR |
03-14-0263 |
Claims
1. Use of a kynurenine 3-hydroxylase inhibitor for the manufacture
of a medicament for increasing the number of islets of Langerhans
cells.
2. Use of a kynurenine 3-hydroxylase inhibitor according to claim
1, in the context of the treatment and/or prevention of diabetes,
its complications and/or its related pathologies.
3. Use of a kynurenine 3-hydroxylase inhibitor for the manufacture
of a medicament for the treatment of prediabetes.
4. Use according to claim 3, for which the said prediabetes is an
insulin-dependent prediabetes.
5. Use according to claim 3, for which the said prediabetes is a
non-insulin-dependent prediabetes.
6. Use of a kynurenine 3-hydroxylase inhibitor for the manufacture
of a medicament for the the treatment and/or prevention of
insulin-dependent diabetes.
7. Use of a kynurenine 3-hydroxylase inhibitor for the manufacture
of a medicament for the prevention of non-insulin-dependent
diabetes.
8. Use of a kynurenine 3-hydroxylase inhibitor for the manufacture
of a medicament for the treatment of early non-insulin-dependent
diabetes.
9. Use according to claim 3, for which the said treatment or
prevention is by increasing the number of islets of Langerhans
cells.
10. Use of a kynurenine 3-hydroxylase inhibitor in combination with
one or more immunosuppressants, for the manufacture of a medicament
for the prevention and/or treatment of insulin-dependent
diabetes.
11. Use according to claim 1, which is suitable for the said
treatment and/or the said prevention in the case of a patient with
an impairment in the number of islets of Langerhans cells.
12. Use according to claim 11, for which the said patient shows a
decrease in the number of islets of Langerhans cells of at least
40%.
13. Use according to claim 11, for which the said patient shows a
decrease in the number of islets of Langerhans cells of between 50%
and 90%.
14. Use according to claim 1, which is suitable for the said
treatment and/or the said prevention in the case of a patient with
glucose intolerance.
15. Use according to claim 14, for which the said patient presents
a fasting glycaemia of between 1.10 g/l and 1.26 g/l and a
glycaemia after meals of between 1.40 g/l and 2 g/l after
meals.
16. Use according to claim 1, which is suitable for the said
treatment and/or the said prevention in the case of a patient with
one or more anti-islets of Langerhans cells immunological
markers.
17. Use according to claim 16, for which the said marker(s)
indicate(s) the existence of an autoimmune response of the body
directed against the antigenic markers of the body's islets of
Langerhans cells.
18. Use according to claim 16, for which the said marker(s) is
(are) chosen from the anti-islet (ICA), anti-glutamic acid
decarboxylase (GAD), anti-tyrosine phosphatase (IA-2) and
anti-(pro)insulin (AIA) auto-antibodies, or the
anti-carboxypeptidase H, anti-64 kD and anti-heat shock protein
antibodies.
19. Use according to claim 1, which is suitable for the said
treatment and/or the said prevention in the case of a patient with
insulin resistance.
20. Use according to claim 19, for which the said patient responds
partially or not at all to insulin secreted by the beta cells or
injected.
21. Use according to claim 1, for which the said patient presents a
level of glycated haemoglobin of higher than 7%.
22. Use according to claim 1, for which the said patient has islets
of Langerhans cells showing an anomaly of insulin secretion in
response to glucose.
23. Use according to claim 1, for which the said patient presents a
suppression of the early peak of insulin secretion.
24. Use according to claim 1, for which the said patient shows
related hyperglycaemia and obesity.
25. Use according to claim 24, for which the said patient suffers
from paediatric obesity.
26. Use according to claim 1, which is suitable for the said
treatment and/or the said prevention in the case of a patient
presenting any diabetic risk factor.
27. Use according to claim 25, for which the said risk factor is
chosen from familial history, gestational diabetes, excess weight,
obesity, insufficient physical exercise, high blood pressure, a
high level of triglycerides, hyperlipidaemia and inflammation.
28. Use according to claim 1, comprising the in vitro treatment of
isolated islets of Langerhans cells with the said kynurenine
3-hydroxylase inhibitor.
29. Process for increasing the number or the insulin-secreting
capacity of islets of Langerhans cells, comprising the in vitro
application of a kynurenine 3-hydroxylase inhibitor to the said
cells.
30. Pharmaceutical composition comprising a kynurenine
3-hydroxylase inhibitor in combination with one or more
immunosuppressants.
31-32. (canceled)
33. Use according to claim 1, for which the said kynurenine
3-hydroxylase inhibitor is a compound of the general formula (I) or
(II): ##STR79## in which: W represents a divalent radical chosen
from the following radicals: ##STR80## R.sup.1 represents a radical
chosen from linear or branched alkyl containing from 1 to 14 carbon
atoms and optionally substituted, alkenyl, alkynyl, cycloalkyl,
cycloalkenyl, a heterocyclic radical, an aryl radical and a
heteroaryl radical; R.sup.2 is chosen from hydrogen, a halogen
atom, hydroxyl, thiol, carboxyl, alkyl, alkenyl, alkynyl, alkoxy,
alkylthio, alkylcarbonyl, alkoxycarbonyl, aryl, heteroaryl,
cycloalkyl and a heterocyclic radical; R.sup.3 is chosen from
hydrogen, a halogen atom, hydroxyl, thiol, alkyl, alkenyl, alkynyl,
alkoxy, alkylthio, aryl, heteroaryl, cycloalkyl and a heterocyclic
radical; R.sup.2 and R.sup.3 together also possibly forming a group
.dbd.CR.sup.16R.sup.17; or alternatively together forming, with the
carbon atom that bears them, a cycloalkyl radical or a heterocyclic
radical; R.sup.4 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, heteroaryloxy, --N(R.sup.12R.sup.12'),
--N(R.sup.12)OR.sup.13, linear or branched alkyl containing from 1
to 14 carbon atoms and optionally substituted, alkenyl, alkynyl,
cycloalkyl, cycloalkenyl, aryl, heteroaryl and a heterocyclic
radical; R.sup.5, R.sup.6, R.sup.7 and R.sup.8, which may be
identical or different, are chosen, independently of each other,
from hydrogen, a halogen atom, and a nitro, cyano, hydroxyl,
trifluoromethyl, alkyl, alkoxy, cycloalkyl or aryl radical; the
radicals R.sup.5 and R.sup.6, on the one hand, or R.sup.6 and
R.sup.7, on the other hand, may also form, together with the carbon
atoms to which they are attached, a benzene ring optionally
substituted by one or more groups, which may be identical or
different, chosen from a halogen atom, a trifluoromethyl, cyano or
nitro radical, an alkyl radical and an alkoxy radical; R.sup.9
represents hydrogen or an alkyl radical; R.sup.10 is chosen from an
alkyl, an aryl and a heteroaryl radical; R.sup.12 and R.sup.12',
which may be identical or different, are chosen, independently of
each other, from hydrogen and an alkyl, alkenyl, alkynyl,
alkylcarbonyl, aryl or heteroaryl radical; or alternatively
R.sup.12 and R.sup.12' may form, together with the nitrogen atom to
which they are attached, a monocyclic or bicyclic heterocyclic
group containing a total of 5 to 10 atoms, among which 1, 2, 3 or 4
are chosen, independently of each other, from nitrogen, oxygen and
sulfur, the said heterocyclic radical also optionally comprising 1,
2, 3 or 4 double bonds and optionally being substituted by one or
more chemical groups, which may be identical or different, chosen
from hydroxyl, halogen atom, alkyl, alkenyl, alkynyl, alkoxy,
alkylthio, aryl, heteroaryl, heterocyclic radical and
trifluoromethyl; R.sup.13 is chosen from hydrogen and an alkyl,
alkenyl, alkynyl, aryl, heteroaryl, --N(R.sup.12R.sup.12') or
--N(R.sup.12)OR.sup.13 radical; R.sup.14 is chosen from hydrogen, a
halogen atom, hydroxyl, thiol, carboxyl, alkyl, alkenyl, alkynyl,
alkoxy, alkylthio, alkylcarbonyl, alkoxycarbonyl, aryl, arylalkyl,
heteroaryl, cycloalkyl and a heterocyclic radical; R.sup.14 may
also form a bond with R.sup.2, thus forming a double bond between
the carbon atoms respectively bearing the substituents R.sup.14 and
R.sup.2; or alternatively R.sup.14 forms, with R.sup.2 and with the
carbon atoms that bear them, a ring containing a total of 3, 4, 5,
6 or 7 carbon atoms, among which 1, 2 or 3 may be replaced with an
atom chosen from nitrogen, oxygen and sulfur, the said ring
possibly comprising one or more unsaturations in the form of (a)
double bond(s), and being optionally substituted by one or more
radicals, which may be identical or different, chosen from oxo,
alkoxy, alkoxycarbonyl and alkylcarbonyloxy; R.sup.15 is chosen
from hydrogen, a halogen atom, hydroxyl, thiol, carboxyl, alkyl,
alkenyl, alkynyl, alkylcarbonyl, alkoxycarbonyl, alkoxy,
alkenyloxy, alkynyloxy, aryloxy, cycloalkyloxy, heteroaryloxy,
heterocyclyloxy, alkylthio, alkenylthio, alkynylthio, arylthio,
cycloalkylthio, heteroarylthio, heterocyclylthio, aryl, heteroaryl,
cycloalkyl and a heterocyclic radical; R.sup.14 and R.sup.15 also
possibly forming, together with the carbon atom that bears them, a
cycloalkyl radical or a heterocyclic radical; R.sup.16 and
R.sup.17, which may be identical or different, are chosen,
independently of each other, from hydrogen, a halogen atom, an
alkyl, aryl, heteroaryl or cycloalkyl radical and a heterocyclic
radical; or alternatively R.sup.16 and R.sup.17 form, together with
the carbon atom that bears them, a cycloalkyl radical or a
heterocyclic radical; and R.sup.11 is chosen from hydrogen and an
alkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, cycloalkyl or
cycloalkylalkyl radical, and any protecting group for an amine
function; and also the possible geometrical and/or optical isomers
thereof, and possible tautomeric forms thereof; the solvates and
hydrates of these compounds; and also the possible salts thereof
with a pharmaceutically acceptable acid or base, or alternatively
the pharmaceutically acceptable prodrugs of these compounds.
34-52. (canceled)
53. Process for manufacturing a medicament for the the treatment
and/or prevention of diabetes, its complications and/or its related
pathologies, by increasing the number of islets of Langerhans
cells, in which at least one compound of the formula (I) or (II) as
defined in one of claim 1 is subjected to an in vitro test of
inhibition of kynurenine 3-hydroxylase, and the molecules
responding positively to the said tests are then conditioned in the
form of a pharmaceutical composition, optionally with addition of a
pharmaceutically acceptable filler or vehicle.
54. Process for screening candidate compounds for activity in the
prevention or treatment of diabetes, its complications and/or its
related pathologies, by increasing the number of islets of
Langerhans cells by inhibiting kynurenine 3-hydroxylase, the said
candidates not corresponding to formula (I) or (II) as defined in
one of claim 33, in which process the candidate compounds are
subjected to an in vitro test of inhibition of kynurenine
3-hydroxylase, and the candidate that has responded positively to
this test is selected.
Description
[0001] The present invention relates to compounds with inhibitory
activity on kynurenine 3-hydroxylase and in particular to their use
as products for pharmaceutical use by increasing the number of
islets of Langerhans cells in the case of patients in need thereof,
especially for the prevention and treatment of diabetes and related
its complications and/or related pathologies (obesity,
hypertension, etc.).
[0002] Diabetes mellitus represents a very heterogeneous group of
diseases all having a certain number of characteristics in common:
elevation of glycaemia and increased long-term risk of developing
cardiovascular complications.
[0003] In 1985, according to the criteria of the WHO, two major
types of diabetes are distinguished: insulin-dependent diabetes
(IDD), which involves the manifestation of immunological phenomena,
and non-insulin-dependent diabetes (NIDD), which were previously
known as type-1 and type-2 diabetes, respectively (World Health
Organization, 1985). The diabetes is said to be insulin-dependent
if its symptoms (thirst, polyuria, coma, etc.) are associated with
hyperglycaemia and ketosis: the administration of insulin is then
vital from the early stages of the disease. In the majority of
other cases, even if persistence of the hyperglycaemia secondarily
necessitates the administration of insulin, the diabetes is
considered as non-insulin-dependent and is treated in general using
oral antidiabetic agents. Non-insulin-dependent diabetes currently
affects 110 million people worldwide. This number shows no sign of
decreasing, since it is forecast that 216 million people will be
affected by 2010.
[0004] Maintaining a sugar balance requires strict coordination
between the organs (brain, liver, pancreas, muscles and adipose
tissue mainly) involved in energy metabolism.
[0005] In non-insulin-dependent diabetes, the liver and the
pancreas are the main participants. Specifically, it has been
clearly demonstrated that excessive production of glucose by the
liver is responsible for fasted hyperglycaemia in diabetics
(Consoli et al., Diabetes, Vol. 38 (1989), 550-557). Similarly,
impairment in pancreatic function (number of islets of Langerhans
cells, secretion of insulin and glucagon in response to glucose)
contributes to the development of postprandial hyperglycaemia
(Polonsky et al., N. Engl. J. Med., 318 (1988), 1231-39).
[0006] Insulin-dependent diabetes is an autoimmune disease that
destroys the beta cells of the pancreas. This disease involves
genetic factors (genes of the HLA (human leukocyte antigen) system
and of insulin itself) and also environmental factors of
nutritional and/or viral origin.
[0007] In addition to the hyperglycaemia symptoms and the
complications resulting therefrom, the two types of diabetes have
in common a defect of pancreatic origin.
[0008] The pancreas is a mixed organ comprising exocrine tissue,
the role of which is the synthesis and secretion of the enzymes
required for digestion, and an endocrine tissue composed of several
types of cells, the role of which is to synthesis and secrete the
hormones involved in maintaining carbohydrate homeostasis. The
endocrine cells are grouped together in the exocrine pancreas in
the form of small structures of complex cellular organisation known
as islets of Langerhans. These islets are composed of four major
cell types:
[0009] beta cells, which secrete insulin
[0010] alpha cells, which secrete glucagon
[0011] delta cells, which secrete somatostatin
[0012] PP cells, which secrete pancreatic polypeptide.
[0013] The amount of circulating insulin is controlled by rapid
changes in the amount of hormone released by individual beta cells
as a function of the variations in plasmatic glucose. However, a
longer-term regulation also exists, which makes it possible to
adapt the production of insulin by means of changes in the total
mass of beta cells. The pancreas is capable of adapting its mass of
beta cells when the demand for insulin increases. The increase in
this demand is observed in physiological and physiopathological
situations in which there is a reduction in the biological efficacy
of insulin (insulin resistance). Besides an anomaly of secretion of
pancreatic hormones (glucagon and insulin), an insufficiency in the
number of islets of Langerhans cells and more particularly of beta
cells may also contribute towards the secretory deficit and thus
towards the establishment of hyperglycaemia in the case of type I
and 11 diabetics (Kloppel G. et al. Surv. Synth. Path. Res (1985),
4:110125). Several studies performed on animal models of diabetes
show that the genetic terrain is an important parameter in the
growth of beta cells (Andersson A., Diabetologia (1983); 25:
269-272; Swenne I. Diabetes, (1984), 32:14-19).
[0014] In the course of diabetes, three stages are distinguished in
the evolution:
[0015] not requiring insulin
[0016] requiring insulin
[0017] insulin required for survival.
[0018] Separation of the description of the types of diabetes and
of their evolutive stages shows the importance of avoiding the
assimilation of insulin-dependent diabetes and diabetes treated
with insulin. However, for non-insulin-dependent diabetes, an early
stage and a late stage are conventionally distinguished, relating
to the duration and seriousness of the diabetic condition.
[0019] The main treatment for type I diabetes consists of the
subcutaneous injection of insulin. The clinical manifestation of
diabetes is always preceded by a longer or shorter asymptomatic
period known as prediabetes, during which organs can, however,
become affected long before the diabetes is diagnosed.
[0020] In 2002, the American Diabetes Association suggested a new
definition of prediabetes, namely a condition characterised by
blood glucose concentrations that are higher normal, but lower than
those corresponding to the predefined criteria of diabetes. A
normal glycaemic equilibrium is characterised by a fasting
glycaemia of less than 1.10 g/l and a glycaemia after meals of less
than 1.40 g/l. If the fasting glycaemia is 1.26 g/l or greater
and/or increases to more than 2 g/l after meals, diabetes is
diagnosed.
[0021] More specifically, the prediabetic condition corresponding
to type I diabetes may be defined by the presence of immunological
markers, such as those described by Buysschaert et al, Louvain Med.
119, S251-S258, 2000, especially including the anti-islet (ICA),
anti-glutamic acid decarboxylase (GAD), anti-tyrosine phosphatase
(IA-2) and anti-(pro)insulin (AIA) auto-antibodies, or the
anti-carboxypeptidase H, anti-64 kD and anti-heat shock protein
antibodies.
[0022] The type II prediabetic condition is characterised mainly by
a disappearance of the early peak of insulin secretion, the
consequence of which is glucose intolerance (also known as IGT, for
"impaired glucose tolerance") or impaired fasting glycaemia (also
known as IFG, for "impaired fasting glucose").
[0023] No medicament exists for effectively preventing diabetes. It
is thus desirable to provide novel routes for the prevention and/or
treatment of prediabetes or diabetes. The main treatment of type I
diabetes consists of the subcutaneous injection of insulin. For
type II diabetes, it is legitimate to propose a medicinal treatment
when the level of glycated haemoglobin (A1c) remains higher than 7%
after 3 to 6 months of the only hygieno-dietetic measurements. It
is necessary to do this if the A1c remains higher than 8% (Nathan,
N.E.J.M., (2002), 17: 1342-1349). Type II diabetes is generally
treated using oral active medicaments. Although many oral
antidiabetic agents are nowadays available, none of them makes it
possible to achieve a normalisation of the glycaemic control
parameters. The diabetic complications associated with
hyperglycaemia inevitably appear. The main weakness of these
medicaments is that they address only one defect at a time, either
insulin resistance (thiazolidinediones or biguanides) or insulin
secretion (sulfonylureas, glinides, etc.). Furthermore, no
medicament capable of increasing the number and functionality of
the islets of Langerhans cells is available at the present time.
Finally, some of them have non-negligible adverse effects.
Sulfonylureas in particular present a major risk of hypoglycaemia,
which demands that the dosage of these medicaments be scrupulously
defined and complied with from patient to patient. Simultaneous
correction of the two defects mentioned above without risks of
associated hypoglycaemia would constitute a fundamental
breakthrough in the treatment of type II diabetes and its
complications. The prevention of the associated cardiovascular
risk, which represents one of the major complications, would also
be of important benefit to diabetic patients.
[0024] With the pancreatic and hepatic function as the central
focus in diabetic pathology in the present invention, the Inventors
focused on a metabolic pathway, namely the metabolism of
tryptophan. Tryptophan is an amino acid whose involvement in
controlling carbohydrate metabolism has previously been reported
(Tsiolakis D. and V. Marks, Horm. Metabol. Res., 16 (1964),
226-229). Its complex metabolisation via kynurenine leads to the
production of NAD+. Some of the intermediate metabolites have also
been described as possibly being involved in glycaemia control
(Connick J. and Stone, Medical hypothesis, 18 (1985), 371-376) and
in particular in the mechanisms for controlling the production of
glucose by the liver ("Effect of tryptophan and its metabolites on
GNG in mammalian tissues", Pogson et al., 1975) and/or in insulin
secretion and synthesis (Noto Y. and Okamoto, Acta Diabet. Lat., 15
(1978), 273-282; Rogers and Evangelista, Proc. Soc. Exp., 178
(1985), 275-278). Among the active metabolites of this pathway are
tryptophan itself, kynurenine and kynurenic acid. The concentration
of these metabolites is controlled by three enzymes: kynurenine
3-hydroxylase, kynureninase and kynurenine aminotransferase.
Kynurenine aminotransferase has also been suspected of being
involved in the hypertension physiopathology of SHR rats
(Spontaneously Hypertensive Rat; Kwok et al., JBC, 35779-35782,
September 2002) which are otherwise insulin-resistant. Despite
that, the joint action of these metabolites on glucose production
by the liver and on insulin secretion in response to glucose has
not been demonstrated in the prior art. In particular, it has not
been demonstrated that some of these metabolites can restore a
physiological response to glucose, the secretion of the pancreatic
hormones (insulin and glucagon), in animals rendered diabetic by
injection of streptozotocin, which would thus make it possible to
correct the insulin secretion defect without giving rise to any
risk of hypoglycaemia.
[0025] It is well described in the prior art that certain
metabolites of the kynurenine pathway, such as quinolinic acid and
kynurenic acid, act as neurotoxic agents and neuroprotective
agents, respectively, on the nervous system. These effects are
linked to their capacity to modulate glutamate receptors and/or
nicotinic receptors (Schwarcz R. and Pellicciardi R., JPET 303
(2002), 1-10; Stone and Darlington, Nature Reviews, 1 (2002),
609-620). The presence of glutamate receptors in the pancreas is
described in the prior art, as is their involvement in pancreatic
hormone secretion (Weaver C. et al., J. Biol. Chem., 271 (1996),
12977-12984), but it has not been demonstrated that these glutamate
receptors are controlled by the kynurenine metabolites in this
organ.
[0026] The research conducted with the aim of meeting the
objectives of the present invention has made it possible to
demonstrate, surprisingly, that the modulation of tryptophan
metabolism in the kynurenine pathway via the pancreatic inhibition
of kynurenine 3-hydroxylase allows an increase in the number of
islets of Langerhans cells and thus plays an important role
especially in the prevention and treatment of diabetic diseases,
its complications and/or its related pathologies (obesity,
hypertension, etc.).
[0027] One of the objectives of the present invention consequently
consists in providing novel therapeutic means which have curative
and/or preventive activity for the prevention of diabetes, its
complications and/or its related pathologies, by increasing the
number of islets of Langerhans cells, and which are free of the
risk of hypoglycaemia.
[0028] The present invention also proposes, as another objective, a
process for the treatment of diabetes that makes it possible to
avoid the side effects and especially hypoglycaemia, the said
process using therapeutic means whose mechanism of action for this
type of pathology is not described or suggested in the prior
art.
[0029] Certain compounds are known (see patents U.S. Pat. No.
6,048,896 and U.S. Pat. No. 6,323,240), which have inhibitory
activity on the kynurenine 3-hydroxylase and which are useful in
the treatment of neurodegenerative diseases, including diseases of
the central nervous system, sclerosis and glaucoma-related
retinopathy. Such compounds were already known as having analgesic
and anti-inflammatory properties.
[0030] The research conducted with the aim of meeting the
objectives of the present invention has made it possible to
demonstrate, surprisingly, that the inhibition of kynurenine
3-hydroxylase plays an important role in the prevention and
treatment of diabetic diseases, in particular non-insulin-dependent
diabetes, its complications and/or its related pathologies.
[0031] It has thus been discovered that compounds with inhibitory
activity on kynurenine 3-hydroxylase increase the number of islets
of Langerhans cells and are especially useful for the prevention
and treatment of diabetes, its complications and/or its related
pathologies.
[0032] The present inventors have now discovered, entirely
unexpectedly, that kynurenine 3-hydroxylase inhibitors show
activity towards pancreatic beta cells.
[0033] Specifically, according to the present invention, the
kynurenine 3-hydroxylase inhibitors increase the number of islets
of Langerhans cells and in particular the beta cells.
[0034] The use of kynurenine 3-hydroxylase inhibitors should thus
make it possible to compensate for the reduction in the number of
pancreatic islets of Langerhans cells in the course of the diabetic
condition, in addition to their effect on the function of these
cells.
[0035] According to the invention, kynurenine 3-hydroxylase
inhibitors thus make it possible to prevent diabetes and its
effects.
[0036] According to the invention, kynurenine 3-hydroxylase
inhibitors thus make it possible to specifically target the
treatment of hyperglycaemia as a function of the type of diabetes,
its degree of progress and/or the population concerned.
[0037] Also, the use of kynurenine 3-hydroxylase inhibitors makes
it possible to act selectively on the increase in the number of
islets of Langerhans cells. This therefore makes it possible to
selectively target patients with an anomaly of insulin secretion of
the islets of Langerhans cells in response to glucose and/or an
impairment in their number.
[0038] Specifically, the use of kynurenine 3-hydroxylase inhibitors
makes it possible to treat and/or prevent insulin-dependent
diabetes, by increasing the mass of insulin-secreting islets of
Langerhans cells.
[0039] More particularly, kynurenine 3-hydroxylase inhibitors make
it possible to prevent insulin-dependent diabetes by increasing the
number of insulin-secreting islets of Langerhans cells before the
disease has been declared, more particularly during
prediabetes.
[0040] Also, the use of kynurenine 3-hydroxylase inhibitors makes
it possible to treat and/or prevent early non-insulin-dependent
diabetes, by increasing the number of functional cells. This is
particularly advantageous insofar as this use makes it possible to
avoid increasing the number of non-functional beta cells and
reducing the mass of beta cells, respectively, above or below the
normal value, which thus makes it possible to advantageously avoid
the appearance of diabetes, its symptoms and/or its
complications.
[0041] Also, the use of kynurenine 3-hydroxylase inhibitors makes
it possible to treat and/or prevent non-insulin-dependent diabetes
at an advanced stage, known as a late stage, by replacing the
non-functional beta cells with functional beta cells.
[0042] Also, the use of kynurenine 3-hydroxylase inhibitors makes
it possible to treat and/or prevent late non-insulin-dependent
diabetes by regenerating the number of beta cells, following the
failure and/or a reduction in the number of the beta cells.
[0043] According to the invention, the kynurenine 3-hydroxylase
inhibitors may be administered orally, in monotherapy, to prevent
and/or treat non-insulin-dependent diabetes.
[0044] According to the invention, the kynurenine 3-hydroxylase
inhibitors can be used in vitro for the treatment of pancreatic
stem cells; the said treated cells may be transplanted into a
patient to prevent and/or treat non-insulin-dependent diabetes.
[0045] According to the invention, the kynurenine 3-hydroxylase
inhibitors can be used in vitro for the treatment of pancreatic
stem cells; the said treated cells may be transplanted into a
patient to prevent and/or treat insulin-dependent diabetes.
[0046] According to the invention, the kynurenine 3-hydroxylase
inhibitors may be administered in combination with one or more
agents for reducing the body's immune response, to prevent and/or
treat insulin-dependent diabetes.
[0047] According to a first subject, the present invention thus
relates to the use of a kynurenine 3-hydroxylase inhibitor for the
manufacture of a medicament for increasing the number of islets of
Langerhans cells.
[0048] According to a second subject, the present invention relates
to the use of a kynurenine 3-hydroxylase inhibitor for the
manufacture of a medicament for the the treatment and/or prevention
of insulin-dependent diabetes.
[0049] According to another subject, the present invention relates
to the use of a kynurenine 3-hydroxylase inhibitor for the
manufacture of a medicament for the prevention and/or treatment of
insulin-dependent prediabetes.
[0050] According to another subject, the present invention relates
to the use of a kynurenine 3-hydroxylase inhibitor for the
manufacture of a medicament for the prevention of
non-insulin-dependent diabetes.
[0051] According to another subject, the present invention relates
to the use of a kynurenine 3-hydroxylase inhibitor for the
manufacture of a medicament for the treatment of early
non-insulin-dependent diabetes.
[0052] According to another subject, the present invention relates
to the use of a kynurenine 3-hydroxylase inhibitor for the
manufacture of a medicament for the treatment of late
non-insulin-dependent diabetes.
[0053] According to another subject, the present invention relates
to pharmaceutical compositions comprising a kynurenine
3-hydroxylase inhibitor in combination with one or more
immunosuppressants.
[0054] According to another subject, the present invention also
relates to the use of a kynurenine 3-hydroxylase inhibitor in
combination with one or more immunosuppressants, for the
manufacture of a medicament for the prevention and/or treatment of
insulin-dependent diabetes.
[0055] According to a preferred aspect, the present invention
relates to any of the uses mentioned above in the case of a patient
with an impairment in the number of islets of Langerhans cells.
Preferably, the impairment in the number of islets of Langerhans
cells represents a decrease of at least 40% in the number of cells,
more preferably a decrease of between 50% and 90%, and even more
preferably between 60% and 85%.
[0056] According to a preferred aspect, the present invention
relates to any of the uses mentioned above in the case of a patient
presenting anti-islets of Langerhans cells immunological
markers.
[0057] According to a preferred aspect, the present invention
relates to any of the uses mentioned above in the case of a patient
presenting any diabetic risk factor.
[0058] According to a preferred aspect, the present invention
relates to any of the uses mentioned above in the case of a patient
with insulin resistance.
[0059] According to a preferred aspect, the present invention
relates to any of the uses mentioned above in the case of a patient
presenting markers, such as glycated haemoglobin at concentrations
higher than 7%.
[0060] According to a preferred aspect, the present invention
relates to any of the uses mentioned above in the case of a patient
whose islets of Langerhans cells show an anomaly of insulin
secretion in response to glucose.
[0061] According to a preferred aspect, the present invention
relates to any of the uses mentioned above in the case of a patient
with related hyperglycaemia and obesity.
[0062] According to another aspect, the present invention relates
to any of the uses mentioned above, comprising the in vitro
treatment of isolated islets of Langerhans cells with a kynurenine
3-hydroxylase inhibitor.
[0063] According to another subject, the present invention also
relates to the method for the in vitro treatment of isolated islets
of Langerhans cells with a kynurenine 3-hydroxylase inhibitor.
[0064] The culturing and transplantation of the said islets of
Langerhans cells may especially be performed by application or
adaptation of the methods described by Docherty et al., Current
Opinion in Pharmacology.2001, 1 :641-650.
[0065] According to the present invention, the term "prediabetes"
means a condition characterised by one or more of the following
factors: the presence of anti-islets of Langerhans cells
immunological markers, an impairment in the number of islets of
Langerhans cells, suppression of the early peak of insulin
secretion, glucose intolerance, an impairment tin fasting glycaemia
and/or any diabetic risk factor.
[0066] According to the invention, the expression "impairment in
fasting glycaemia and/or glucose intolerance" means a fasting
glycaemia of between 1.10 g/l and 1.26 g/l and a glycaemia after
meals of between 1.40 g/l and 2 g/l after meals.
[0067] According to the invention, the expression "anti-islets of
Langerhans cells immunological markers" means any immunological
marker indicating the existence of an autoimmune response of the
body directed against the antigenic markers of the body's islets of
Langerhans cells. These markers include auto-antibodies, such as
those described by Buysschaert et al., Louvain Med. 119, S251-S258,
2000. These antibodies are chosen from the anti-islet (ICA),
anti-glutamic acid decarboxylase (GAD), anti-tyrosine phosphatase
(IA-2) and anti-(pro)insulin (AIA) auto-antibodies, or the
anti-carboxypeptidase H, anti-64 kD and anti-heat shock protein
antibodies.
[0068] According to the invention, the expression "impairment in
the number of islets of Langerhans cells" means a decrease of at
least 40% in the number of cells. Preferably, the impairment in the
number of islets of Langerhans cells represents a decrease of at
least 40% in the number of cells, more preferably a decrease of
between 50% and 90% and even more preferably between 60% and
85%.
[0069] According to the invention, the expression "anomaly of
insulin secretion in response to glucose" means any impairment in
the normal capacity of the islets of Langerhans cells to secrete
insulin in response to glucose.
[0070] According to the invention, the expression "diabetic risk
factor" means any complaint directly or indirectly associated with
the appearance of diabetes. These especially comprise familial
history, gestational diabetes, excess weight, obesity, insufficient
physical exercise, high blood pressure, a high level of
triglycerides, inflammation, hyperlipidaemia, etc.
[0071] According to the invention, the term "immunosuppressant"
means any physical agent (for example x-rays) chemical agent (for
example azathioprine or mercaptopurine) or biological agent (for
example anti-lymphocyte serum) for reducing or inhibiting the
stimulation of an immune response of the body with an antigen.
[0072] According to the invention, the term "islets of Langerhans
cells" means the alpha, beta, delta and PP cells mentioned above;
more preferably, the islets of Langerhans cells represent the beta
cells.
[0073] It has especially been discovered that the compounds
corresponding to the general formula (I) or to the general formula
(II) described hereinbelow generally have inhibitory activity on
kynurenine 3-hydroxylase. Among the compounds described in formulae
(I) and (II), some families of compounds are known to have activity
that is useful in the treatment of diabetes, and especially the
families of compounds corresponding to patent application
WO-A-98/07681 and the families corresponding to patent application
EP-A-0 885 869. The compounds with substantial activity on
kynurenine 3-hydroxylase are especially preferred. The term
"substantial activity" means any inhibitory activity on the enzyme
by the in vitro test process defined below, thus making it possible
to obtain an effective therapeutic action on the enzyme. In
particular, an enzymatic activity of less than or equal to 70%,
advantageously less than or equal to 50% and even more preferably
less than or equal to 30% relative to the control, is
preferred.
[0074] It has thus been discovered that, within these families of
compounds, it is possible to use compounds that are characterised
by inhibitory activity on kynurenine 3-hydroxylase to obtain an
improved treatment or improved medicaments, or for a different
purpose, to increase the mass of beta cells and especially to
prevent or treat diabetes, and also the complications of this
diabetes, via a novel route that offers unexpected advantages. They
also make it possible to improve the prevention and treatment of
diabetes, especially of non-insulin-dependent diabetes, by
administration of a therapeutically effective amount to patients in
need of inhibition of kynurenine 3-hydroxylase.
[0075] In particular, the compounds of family Ih are found to be
noteworthy kynurenine 3-hydroxylase inhibitors and agents for
increasing the mass of beta cells, especially antidiabetic
agents.
[0076] Confirmation of the existence of inhibitory activity on
kynurenine 3-hydroxylase may be made by any known means and
especially, in a particularly simple manner, by subjecting the
compound to an in vitro test that will be defined hereinbelow.
[0077] More specifically, the compounds with inhibitory activity on
kynurenine 3-hydroxylase belong to the general formula (I) or to
the general formula (II) below: ##STR1## in which:
[0078] W represents a divalent radical chosen from the following
radicals: ##STR2##
[0079] R.sup.1 represents a radical chosen from linear or branched
alkyl containing from 1 to 14 carbon atoms and optionally
substituted, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, a
heterocyclic radical, an aryl radical and a heteroaryl radical;
[0080] R.sup.2 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, carboxyl, alkyl, alkenyl, alkynyl, alkoxy, alkylthio,
alkylcarbonyl, alkoxycarbonyl, aryl, heteroaryl, cycloalkyl and a
heterocyclic radical;
[0081] R.sup.3 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, alkyl, alkenyl, alkynyl, alkoxy, alkylthio, aryl,
heteroaryl, cycloalkyl and a heterocyclic radical;
[0082] R.sup.2 and R.sup.3 together also possibly forming a group
.dbd.CR.sup.16R.sup.17; or alternatively together forming, with the
carbon atom that bears them, a cycloalkyl radical or a heterocyclic
radical;
[0083] R.sup.4 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, heteroaryloxy, --N(R.sup.12R.sup.12'),
--N(R.sup.12)OR.sup.13, linear or branched alkyl containing from 1
to 14 carbon atoms and optionally substituted, alkenyl, alkynyl,
cycloalkyl, cycloalkenyl, aryl, heteroaryl and a heterocyclic
radical;
[0084] R.sup.5, R.sup.6, R.sup.7 and R.sup.8, which may be
identical or different, are chosen, independently of each other,
from hydrogen, a halogen atom, and a nitro, cyano, hydroxyl,
trifluoromethyl, alkyl, alkoxy, cycloalkyl or aryl radical;
[0085] the radicals R.sup.5 and R.sup.6, on the one hand, or
R.sup.6 and R.sup.7, on the other hand, may also form, together
with the carbon atoms to which they are attached, a benzene ring
optionally substituted by one or more groups, which may be
identical or different, chosen from a halogen atom, a
trifluoromethyl, cyano or nitro radical, an alkyl radical and an
alkoxy radical;
[0086] R.sup.9 represents hydrogen organ alkyl radical;
[0087] R.sup.10 is chosen from an alkyl, an aryl and a heteroaryl
radical;
[0088] R.sup.12 and R.sup.12', which may be identical or different,
are chosen, independently of each other, from hydrogen and an
alkyl, alkenyl, alkynyl, alkylcarbonyl, aryl or heteroaryl radical;
or alternatively R.sup.12 and R.sup.12' may form, together with the
nitrogen atom to which they are attached, a monocyclic or bicyclic
heterocyclic group containing a total of 5 to 10 atoms, among which
1, 2, 3 or 4 are chosen, independently of each other, from
nitrogen, oxygen and sulfur, the said heterocyclic radical also
optionally comprising 1, 2, 3 or 4 double bonds and optionally
being substituted by one or more chemical groups, which may be
identical or different, chosen from hydroxyl, halogen atom, alkyl,
alkenyl, alkynyl, alkoxy, alkylthio, aryl, heteroaryl, heterocyclic
radical and trifluoromethyl;
[0089] R.sup.13 is chosen from hydrogen and an alkyl, alkenyl,
alkynyl, aryl, heteroaryl, --N(R.sup.12R.sup.12') or
--N(R.sup.12)OR.sup.13 radical;
[0090] R.sup.14 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, carboxyl, alkyl, alkenyl, alkynyl, alkoxy, alkylthio,
alkylcarbonyl, alkoxycarbonyl, aryl, arylalkyl, heteroaryl,
cycloalkyl and a heterocyclic radical; R.sup.14 may also form a
bond with R.sup.2, thus forming a double bond between the carbon
atoms respectively bearing the substituents R.sup.14 and R.sup.2;
or alternatively R.sup.14 forms, with R.sup.2 and with the carbon
atoms that bear them, a ring containing a total of 3, 4, 5, 6 or 7
carbon atoms, among which 1, 2 or 3 may be replaced with an atom
chosen from nitrogen, oxygen and sulfur, the said ring possibly
comprising one or more unsaturations in the form of (a) double
bond(s), and being optionally substituted by one or more radicals,
which may be identical or different, chosen from oxo, alkoxy,
alkoxycarbonyl and alkylcarbonyloxy;
[0091] R.sup.15 is chosen from hydrogen a halogen atom, hydroxyl,
thiol, carboxyl, alkyl, alkenyl, alkynyl, alkylcarbonyl,
alkoxycarbonyl, alkoxy, alkenyloxy, alkynyloxy, aryloxy,
cycloalkyloxy, heteroaryloxy, heterocyclyloxy, alkylthio,
alkenylthio, alkynylthio, arylthio, cycloalkylthio, heteroarylthio,
heterocyclylthio, aryl, heteroaryl, cycloalkyl and a heterocyclic
radical;
[0092] R.sup.14 and R.sup.15 also possibly forming, together with
the carbon atom that bears them, a cycloalkyl radical or a
heterocyclic radical;
[0093] R.sup.16 and R.sup.17, which may be identical or different,
are chosen, independently of each other, from hydrogen, a halogen
atom, an alkyl, aryl, heteroaryl or cycloalkyl radical and a
heterocyclic radical; or alternatively
[0094] R.sup.16 and R.sup.17 form, together with the carbon atom
that bears them, a cycloalkyl radical or a heterocyclic radical;
and
[0095] R.sup.11 is chosen from hydrogen and an alkyl, aryl,
arylalkyl, heteroaryl, heteroarylalkyl, cycloalkyl or
cycloalkylalkyl radical, and any protecting group for is an amine
function;
[0096] and also the possible geometrical and/or optical isomers
thereof, and possible tautomeric forms thereof;
[0097] the solvates and hydrates of these compounds;
[0098] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0099] The following definitions specify the natures of the various
groups and radicals defined above. Unless otherwise indicated,
these definitions apply for all the terms of the present invention
thus explained.
[0100] The term "halogen atom" denotes a fluorine, chlorine,
bromine or iodine atom.
[0101] The term "alkyl" denotes a linear or branched alkyl radical
containing from 1 to 6 carbon atoms, optionally substituted by one
or more chemical groups chosen from hydroxyl, carboxyl, cyano,
nitro, --N(R.sup.12R.sup.12'), --N(R.sup.12)OR.sup.13, aryl,
heteroaryl, cycloalkyl, heterocyclic radical, alkyl, alkenyl,
alkynyl, alkoxy, alkylcarbonyl, alkoxycarbonyl, halogen atom,
trifluoromethyl, thiol, --SR.sup.13', --S(O)R.sup.13' and
--S(O.sub.2)R.sup.13', with R.sup.13' having the same definition as
R.sup.13, with the exception of hydrogen. The possible substituents
on the alkyl radical containing from 1 to 14 carbon atoms may be
identical to those described above.
[0102] The term "alkenyl" denotes an alkenyl radical containing one
or more double bonds; the said radical, which is linear or
branched, and which contains from 2 to 6 carbon atoms, is
optionally substituted by one or more chemical groups chosen from
hydroxyl, carboxyl, cyano, nitro, --N(R.sup.12R.sup.12'),
--N(R.sup.12)OR.sup.13, aryl, heteroaryl, cycloalkyl, heterocyclic
radical, alkyl, alkenyl, alkynyl, alkoxy, alkylcarbonyl,
alkoxycarbonyl, halogen atom, trifluoromethyl, thiol, --SR.sup.13',
--S(O)R.sup.13' and --S(O.sub.2)R.sup.13', with R.sup.13' having
the same definition as R.sup.13, with the exception of
hydrogen.
[0103] The term "alkynyl" denotes an alkynyl radical containing one
or more triple bonds; the said radical, which is linear or
branched, and which contains from 2 to 6 carbon atoms, is
optionally substituted by one or more chemical groups chosen from
hydroxyl, carboxyl, cyano, nitro, --N(R.sup.12R.sup.12'),
--N(R.sup.12)OR.sup.13, aryl, heteroaryl, cycloalkyl, heterocyclic
radical, alkyl, alkenyl, alkynyl, alkoxy, alkylcarbonyl,
alkoxycarbonyl, halogen atom, trifluoromethyl, thiol, --SR.sup.13',
--S(O)R.sup.13' and --S(O.sub.2)R.sup.13', with R.sup.13' having
the same definition as R.sup.13, with the exception of
hydrogen.
[0104] The term "alkoxy" should be understood as being
"alkyl"+"oxy", in which the term "alkyl" is as defined above. The
substituents of the alkoxy radical are identical to those defined
for the alkyl radical.
[0105] The term "cycloalkyl" denotes a bridged or non-bridged
monocyclic, bicyclic or tricyclic cycloalkyl radical containing
from 3 to 13 carbon atoms, optionally substituted by one or more
chemical groups, which may be identical or different, chosen from
hydroxyl, carboxyl, cyano, nitro, --N(R.sup.12R.sup.12'),
--N(R.sup.12)OR.sup.13, aryl, heteroaryl, cycloalkyl, heterocyclic
radical, alkyl, alkenyl, alkynyl, alkoxy, alkylcarbonyl,
alkoxycarbonyl, halogen atom, trifluoromethyl, thiol, --SR.sup.13',
--S(O)R.sup.13' and --S(O.sub.2)R.sup.13', with R.sup.13' having
the same definition as R.sup.13, with the exception of
hydrogen.
[0106] The term "cycloalkenyl" denotes a cycloalkyl radical as
defined above comprising at least one double bond.
[0107] The term "heterocyclic radical" or "heterocyclyl" denotes a
monocyclic or bicyclic radical containing a total of 5 to 10 atoms,
among which 1, 2, 3 or 4 are chosen, independently of each other,
from nitrogen, oxygen and sulfur, the said heterocyclic radical
also optionally comprising 1, 2, 3 or 4 double bonds and being
optionally substituted by one or more chemical groups, which may be
identical or different, chosen from hydroxyl, carboxyl, cyano,
nitro, --N(R.sup.12R.sup.12'), --N(R.sup.12)OR.sup.13, aryl,
heteroaryl, cycloalkyl, heterocyclic radical, alkyl, alkenyl,
alkynyl, alkoxy, alkylcarbonyl, alkoxycarbonyl, halogen atom,
trifluoromethyl, thiol, --SR.sup.13', --S(O)R.sup.13' and
--S(O.sub.2)R.sup.13', with R.sup.13' having the same definition as
R.sup.3, with the exception of hydrogen.
[0108] The term "aryl" denotes a monocyclic, bicyclic or tricyclic
aryl radical containing from 6 to 14 carbon atoms, optionally
substituted by one or more chemical groups, which may be identical
or different, chosen from hydroxyl, carboxyl, cyano, nitro,
--N(R.sup.12R.sup.12') --N(R.sup.12)OR.sup.13; aryl, heteroaryl,
cycloalkyl; heterocyclic: radical, alkyl, alkenyl, alkynyl, alkoxy,
alkylcarbonyl, alkoxycarbonyl, halogen atom, trifluoromethyl,
thiol, --SR.sup.13', --S(O)R.sup.13' and --S(O.sub.2)R.sup.13',
with R.sup.13' having the same definition as R.sup.13, with the
exception of hydrogen.
[0109] The term "heteroaryl" denotes a monocyclic or bicyclic
heteroaryl radical containing a total of 5 to 10 atoms, among which
1, 2, 3 or 4 are chosen, independently of each other, from
nitrogen, oxygen and sulfur, the said heteroaryl radical being
optionally substituted by one or more chemical groups, which may be
identical or different, chosen from hydroxyl, carboxyl, cyano,
nitro, --N(R.sup.12R.sup.12'), --N(R.sup.12)OR.sup.13, aryl,
heteroaryl, cycloalkyl, heterocyclic radical, alkyl, alkenyl,
alkynyl, alkoxy, alkylcarbonyl, alkoxycarbonyl, halogen atom,
trifluoromethyl, thiol, --SR.sup.13', --S(O)R.sup.13' and
--S(O.sub.2)R.sup.13', with R.sup.13' having the same definition as
R.sup.13, with the exception of hydrogen.
[0110] A preferred aryl radical is the phenyl radical or the
1-naphthyl, 2-naphthyl or fluorenyl radical.
[0111] Among the alkyl and alkoxy radicals substituted by an aryl
radical, the benzyl, benzyloxy, phenethyl, phenylethoxy,
naphthylmethyl and naphthylmethoxy radicals are particularly
preferred.
[0112] Among the cycloalkyl radicals that are preferred are
cyclopropyl, cyclopentyl, cyclohexyl, the adamantyl radical and
radicals derived from tetralin and from decalin.
[0113] The terms "heteroaryl radical" and "heterocyclic radical"
preferably mean a pyridyl, furyl, thienyl, 1-quinolyl, 2-quinolyl,
tetrahydrofuryl, tetrahydropyranyl, pyrrolyl, imidazolyl,
pyrazolyl, thiazolyl, isothiazolyl, oxazolyl, isoxazolyl,
morpholino, piperazinyl, piperidyl, pyranyl, thiopyranyl, indanyl,
benzothienyl or benzofuryl radical.
[0114] For the compounds of the formulae (I) and (II) presented
above, the term "geometrical isomer" means a cis/trans or E/Z
isomerism. More particularly, for the compounds of the formula (I)
and when R.sup.14 forms a bond with R.sup.2, thus forming a double
bond between the carbon atoms respectively bearing the substituents
R.sup.14 and R.sup.2, this double bond may be of E or Z
configuration. These geometrical isomers, which may or may not be
pure, alone or as a mixture, form an integral part of the compounds
of the formula (I).
[0115] The term "optical isomer" includes all the forms of isomers,
alone or as mixtures, arising from the presence of one or more axes
and/or centres of symmetry in the molecule, and resulting in the
rotation of a beam of polarised light. The term "optical isomer"
more particularly includes the enantiomers and diastereoisomers, in
pure form or as a mixture.
[0116] In particular, for the compounds of the formula (I), and
when the substituents R.sup.2 and R.sup.3, on the one hand, and/or
the substituents R.sup.16 and R.sup.17, on the other hand, are
different, the carbon atoms bearing these pairs of substituents are
asymmetric, and thus lead to enantiomers and/or diastereoisomers.
These optical isomers, which may or may not be pure, alone or as a
mixture, form an integral part of the compounds of the formula
(I).
[0117] Among the acids capable of forming pharmaceutically
acceptable salts with the compounds of the formula (I) or of the
formula (II) above, non-limiting examples that may be mentioned
include hydrochloric acid, phosphoric acid, sulfuric acid, tartaric
acid, citric acid, maleic acid, acetic acid, fumaric acid,
alkylsulfonic acid and camphoric acid.
[0118] Among the bases capable of forming pharmaceutically
acceptable salts with the compounds of the formula (I) or of the
formula (II) above, non-limiting examples that may be mentioned
include sodium hydroxide, potassium hydroxide, diethylamine,
triethylamine, ethanolamine, diethanolamine, arginine and
lysine.
[0119] The compounds of the formulae (I) and (II) above also
comprise the prodrugs of these compounds.
[0120] The term "prodrugs" means compounds which, once administered
to the patient, are chemically and/or biologically transformed by
the living body, into compounds of the formula (I) or (II).
[0121] Examples of prodrugs of compounds of the formula (I) above
are those for which R.sup.4 represents a radical --OP, in which P
is a leaving group, for example a sugar residue, such as sucrose,
which can thus lead to compounds in which R.sup.4 represents --OH.
Such prodrugs are included in the field of the present
invention.
[0122] A large number of compounds of the formulae (I) and (II)
defined above are known, especially by the patent publications and
patent applications U.S. Pat. No. 6,048,896, U.S. Pat. No.
6,323,240, EP 0 885 869 and U.S. Pat No. 5,877,193. These
publications provide the processes for the preparation of these
various compounds, to which processes a person skilled in the art
may refer, or may adapt, to synthesise all the compounds of the
formulae (I) and (II).
[0123] According to one variant of the present invention, the
compounds of the formula (I) that are preferred are those having
the following characteristics, taken separately or in
combination:
[0124] W represents a divalent radical chosen from the following
radicals: ##STR3##
[0125] R.sup.1 represents a radical chosen from linear or branched
alkyl containing from 1 to 14 carbon atoms and optionally
substituted, alkenyl, cycloalkyl, cycloalkenyl, a heterocyclic
radical, an aryl radical and a heteroaryl radical;
[0126] R.sup.2 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, carboxyl, alkyl, alkenyl, alkoxy, alkylthio, alkylcarbonyl,
alkoxycarbonyl and aryl;
[0127] R.sup.3 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, alkyl, alkenyl, alkoxy, alkylthio and aryl;
[0128] R.sup.2 and R.sup.3 together also possibly forming a group
.dbd.CR.sup.16R.sup.17;
[0129] R.sup.4 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, heteroaryloxy, --N(R.sup.12R.sup.12'),
--N(R.sup.12)OR.sup.13, linear or branched alkyl containing from 1
to 14 carbon atoms and optionally substituted, cycloalkyl,
cycloalkenyl, aryl, heteroaryl and a heterocyclic radical;
[0130] R.sup.12 and R.sup.12', which may be identical or different,
are chosen, is independently of each other, from hydrogen and an
alkyl, alkenyl, alkynyl, alkylcarbonyl, aryl or heteroaryl
radical;
[0131] R.sup.13 is chosen from hydrogen and an alkyl, alkenyl,
alkynyl, aryl, --N(R.sup.12R.sup.12') or --N(R.sup.12)OR.sup.13
radical;
[0132] R.sup.14 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, carboxyl, alkyl, alkenyl, alkoxy, alkylthio, alkylcarbonyl,
alkoxycarbonyl, aryl and arylalkyl; R.sup.14 may also form a bond
with R.sup.2, thus forming a double bond between the carbon atoms
respectively bearing the substituents R.sup.14 and R.sup.2; or
alternatively R.sup.14 forms, with R.sup.2 and with the carbon
atoms that bear them, a ring containing a total of 3, 4, 5 or 6
carbon atoms, among which 1, 2 or 3 may be replaced with an atom
chosen from nitrogen and oxygen, the said ring possibly comprising
one or more unsaturations in the form of (a) double bond(s), and
being optionally substituted by one or more radicals, which may be
identical or different, chosen from oxo, alkoxy, alkoxycarbonyl and
alkylcarbonyloxy;
[0133] R.sup.15 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, carboxyl, alkyl, alkenyl, alkylcarbonyl, alkoxycarbonyl,
alkoxy, alkylthio and aryl;
[0134] R.sup.16 is chosen from hydrogen and an alkyl or aryl
radical;
[0135] R.sup.17 represents a hydrogen atom; and
[0136] R.sup.11 is chosen from hydrogen and any protecting group
for an amine function;
[0137] and also the possible geometrical and/or optical isomers
thereof, and possible tautomeric forms thereof;
[0138] the solvates and hydrates of these compounds;
[0139] and the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0140] According to another variant of the present invention, this
invention relates to the use of compounds of the, formula (Ia) that
have inhibitory activity on kynurenine 3-hydroxylase, for the
preparation of a medicament for the prevention and/or treatment of
diabetes. These compounds of the formula (Ia) have the general
structure (I) as defined above, in which:
[0141] W represents a divalent radical chosen from the following
radicals: ##STR4##
[0142] R.sup.1 represents a radical chosen from linear or branched
alkyl containing from 1 to 14 carbon atoms and optionally
substituted, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, a
heterocyclic radical, an aryl radical and a heteroaryl radical;
[0143] R.sup.2 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, carboxyl, alkyl, alkenyl, alkynyl, alkoxy, alkylthio,
alkylcarbonyl, alkoxycarbonyl, aryl, heteroaryl, cycloalkyl and a
heterocyclic radical;
[0144] R.sup.3 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, alkyl, alkenyl, alkynyl, alkoxy, alkylthio, aryl,
heteroaryl, cycloalkyl and a heterocyclic radical;
[0145] R.sup.2 and R.sup.3 together also possibly forming a group
.dbd.CR.sup.16R.sup.17, or alternatively forming, together with the
carbon atom that bears them, a cycloalkyl radical or a heterocyclic
radical;
[0146] R.sup.4 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, heteroaryloxy, --N(R.sup.12R.sup.12'),
--N(R.sup.12)OR.sup.13, linear or branched alkyl containing from 1
to 14 carbon atoms and optionally substituted, alkenyl, alkynyl,
cycloalkyl, cycloalkenyl, aryl, heteroaryl and a heterocyclic
radical;
[0147] R.sup.12 and R.sup.12', which may be identical or different,
are chosen, independently of each other, from hydrogen and an
alkyl, alkenyl, alkynyl, alkylcarbonyl, aryl or heteroaryl radical;
or alternatively R.sup.12 and R.sup.12' may form, together with the
nitrogen atom to which they are attached, a monocyclic or bicyclic
heterocyclic group containing a total of 5 to 10 atoms, among which
1, 2, 3 or 4 are chosen, independently of each other, from
nitrogen, oxygen and sulfur, the said heterocyclic radical also
optionally comprising 1, 2, 3 or 4 double bonds and optionally
being substituted by one or more chemical groups, which may be
identical or different, chosen from hydroxyl, halogen atom, alkyl,
alkenyl, alkynyl, alkoxy, alkylthio, aryl, heteroaryl, heterocyclic
radical and trifluoromethyl;
[0148] R.sup.13 is chosen from hydrogen and an alkyl, alkenyl,
alkynyl, aryl, heteroaryl, --N(R.sup.12R.sup.12') or
--N(R.sup.12)OR.sup.13 radical;
[0149] R.sup.14 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, carboxyl, alkyl, alkenyl, alkynyl, alkoxy, alkylthio,
alkylcarbonyl, alkoxycarbonyl, aryl, arylalkyl, heteroaryl,
cycloalkyl and a heterocyclic radical; R.sup.14 may also form a
bond with R.sup.2, thus forming a double bond between the carbon
atoms respectively bearing the substituents R.sup.14 and R.sup.2;
or alternatively R.sup.14 forms, with R.sup.2 and with the carbon
atoms that bear them, a ring containing a total of 3, 4, 5, 6 or 7
carbon atoms, among which 1, 2 or 3 may be replaced with an atom
chosen from nitrogen, oxygen and sulfur, the said ring possibly
comprising one or more unsaturations in the form of (a) double
bond(s), and being optionally substituted by one or more radicals,
which may be identical or different, chosen from oxo, alkoxy,
alkoxycarbonyl and alkylcarbonyloxy;
[0150] R.sup.15 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, carboxyl, alkyl, alkenyl, alkynyl, alkylcarbonyl,
alkoxycarbonyl, alkoxy, alkenyloxy, alkynyloxy, aryloxy,
cycloalkyloxy, heteroaryloxy, heterocyclyloxy, alkylthio,
alkenylthio, alkynylthio, arylthio, cycloalkylthio, heteroarylthio,
heterocyclylthio, aryl, heteroaryl, cycloalkyl and a heterocyclic
radical;
[0151] R.sup.14 and R.sup.15 also possibly forming, together with
the carbon atom that bears them, a cycloalkyl radical or a
heterocyclic radical;
[0152] R.sup.16 and R.sup.17, which may be identical or different,
are chosen, independently of each other, from hydrogen, a halogen
atom, an alkyl, aryl, heteroaryl or cycloalkyl radical and a
heterocyclic radical; or alternatively R.sup.16 and R.sup.17 form,
together with the carbon atom that bears them, a cycloalkyl radical
or a heterocyclic radical; and
[0153] R.sup.11 is chosen from hydrogen and an alkyl, aryl,
arylalkyl, heteroaryl, heteroarylalkyl, cycloalkyl or
cycloalkylalkyl radical, and any protecting group for an amine
function;
[0154] with the restriction that when R.sup.3, R.sup.2 and R.sup.14
each represent hydrogen, then R.sup.15 is other than an alkyl
radical, optionally substituted by aryl, heteroaryl, cycloalkyl and
a heterocyclic radical;
[0155] and also the possible geometrical and/or optical isomers
thereof, and possible tautomeric forms thereof;
[0156] the solvates and hydrates of these compounds;
[0157] and the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0158] Among the compounds (Ia) defined above, the compounds that
will also be preferred are those of the family (Ib) belonging to
formula (I) in which:
[0159] W represents a divalent radical chosen from the radicals:
##STR5##
[0160] R.sup.1 represents a phenyl radical, optionally substituted
by 1, 2 or 3 groups chosen from cyano, nitro, phenyl, benzyl,
alkyl, alkenyl containing from 2 to 4 carbon atoms, alkynyl
containing from 2 to 4 carbon atoms, alkoxy, thiol --SR.sup.13',
--S(O)R.sup.13' and --S(O.sub.2)R.sup.13', and a halogen atom;
[0161] R.sup.2 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, optionally substituted alkyl, in particular benzyl, alkenyl
containing from 2 to 4 carbon atoms, alkoxy, alkylthio and
phenyl;
[0162] R.sup.3 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, optionally substituted alkyl, in particular benzyl, alkenyl
containing from 2 to 4 carbon atoms, alkoxy, alkylthio and
phenyl;
[0163] R.sup.2 and R.sup.3 together also possibly forming a group
.dbd.CR.sup.16R.sup.17;
[0164] R.sup.4 is chosen from hydroxyl, optionally substituted
alkoxy, in particular benzyloxy, alkenyloxy containing from 2 to 4
carbon atoms, alkynyloxy containing from 2 to 4 carbon atoms,
phenoxy, --N(R.sup.12R.sup.12') and --N(R.sup.12)OR.sup.13;
[0165] R.sup.12 and R.sup.12', which may be identical or different,
are chosen, independently of each other, from hydrogen, an
optionally substituted alkyl radical, in particular benzyl, alkenyl
containing from 2 to 4 carbon atoms, alkynyl containing from 2 to 4
carbon atoms, and phenyl;
[0166] R.sup.13 is chosen from hydrogen, an optionally substituted
alkyl radical, in particular benzyl, alkenyl containing from 2 to 4
carbon atoms, alkynyl containing from 2 to 4 carbon atoms, and
phenyl;
[0167] R13' is chosen from an optionally substituted alkyl radical,
in particular benzyl, alkenyl containing from 2 to 4 carbon atoms,
alkynyl containing from 2 to 4 carbon atoms, phenyl and
--N(R.sup.12R.sup.12');
[0168] R.sup.14 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, optionally substituted alkyl, in particular benzyl, alkenyl
containing from 2 to 4 carbon atoms, alkoxy, alkylthio and phenyl;
R.sup.14 may also form a bond with R.sup.2, thus forming a double
bond between the carbon atoms respectively bearing the substituents
R.sup.14 and R.sup.2;
[0169] R.sup.15 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, optionally substituted alkyl, in particular benzyl, alkenyl
containing from 2 to 4 carbon atoms, alkoxy, alkylthio and
phenyl;
[0170] R.sup.16 is chosen from hydrogen, a halogen atom, hydroxyl,
thiol, optionally substituted alkyl, in particular benzyl, alkenyl
containing from 2 to 4 carbon atoms, alkoxy, alkylthio and phenyl;
and
[0171] R.sup.17 represents a hydrogen atom;
[0172] with the restriction that when R.sup.3, R.sup.2 and R.sup.14
each represent hydrogen, then R.sup.15 is other than an alkyl
radical, optionally substituted by aryl, heteroaryl, cycloalkyl and
a heterocyclic radical;
[0173] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0174] the solvates and hydrates of these compounds;
[0175] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0176] In the vast majority, the compounds (Ib) defined above show
entirely advantageous inhibitory activity on kynurenine
3-hydroxylase. As a result, these compounds are most particularly
preferred and simple to use for any of the abovementioned uses
according to the invention.
[0177] According to another variant of the invention, this
invention relates to the use of compounds of the family (Ic) as
kynurenine 3-hydroxylase inhibitors in any of the abovementioned
uses according to the invention. These compounds of family (Ic)
have the general structure (I) as defined above, in which:
[0178] W represents the divalent radical: ##STR6##
[0179] R.sup.1 represents a radical chosen from linear or branched
alkyl containing from 1 to 14 carbon atoms and optionally
substituted, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, a
heterocyclic radical, an aryl radical and a heteroaryl radical;
[0180] R.sup.2 represents hydrogen;
[0181] R.sup.3 represents hydrogen;
[0182] R.sup.4 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, heteroaryloxy, --N(R.sup.12R.sup.12') and
--N(R.sup.12)OR.sup.13;
[0183] R.sup.12 and R.sup.12', which may be identical or different,
are chosen, independently of each other, from hydrogen and an
alkyl, alkenyl, alkynyl, alkylcarbonyl, aryl or heteroaryl radical;
or alternatively R.sup.12 and R.sup.12' may form, together with the
nitrogen atom to which they are attached, a monocyclic or bicyclic
heterocyclic group containing a total of 5 to 10 atoms, among which
1, 2, 3 or 4 are chosen, independently of each other, from
nitrogen, oxygen and sulfur, the said heterocyclic radical also
optionally comprising 1, 2, 3 or 4 double bonds and optionally
being substituted by one or more chemical groups, which may be
identical or different, chosen from hydroxyl, halogen atom, alkyl,
alkenyl, alkynyl, alkoxy, alkylthio, aryl, heteroaryl, heterocyclic
radical and trifluoromethyl;
[0184] R.sup.13 is chosen from hydrogen and an alkyl, alkenyl,
alkynyl, aryl, heteroaryl, --N(R.sup.12R.sup.12') or
--N(R.sup.12)OR.sup.13 radical;
[0185] R.sup.14 represents hydrogen;
[0186] R.sup.15 represents hydrogen;
[0187] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0188] the solvates and hydrates of these compounds;
[0189] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0190] According to another variant, the invention relates to the
use of compounds of the family (Id) as kynurenine 3-hydroxylase
inhibitors in any of the above-mentioned uses according to the
invention, the said compounds (Id) having the general structure (I)
as defined above, in which:
[0191] W represents the divalent radical: ##STR7##
[0192] R.sup.1 represents a radical chosen from linear or branched
alkyl containing from 1 to 14 carbon atoms and optionally
substituted, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, a
heterocyclic radical, an aryl radical and a heteroaryl radical;
[0193] R.sup.2 represents hydrogen;
[0194] R.sup.3 represents hydrogen;
[0195] R.sup.4 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, heteroaryloxy, --N(R.sup.12R.sup.12') and
--N(R.sup.12)OR.sup.13;
[0196] R.sup.12 and R.sup.12', which may be identical or different,
are chosen, independently of each other, from hydrogen and an
alkyl, alkenyl, alkynyl, alkylcarbonyl, aryl or heteroaryl radical;
or alternatively R.sup.12 and R.sup.12' may form, together with the
nitrogen atom to which they are attached, a monocyclic or bicyclic
heterocyclic group containing a total of 5 to 10 atoms, among which
1, 2, 3 or 4 are chosen, independently of each other, from
nitrogen, oxygen and sulfur, the said heterocyclic radical also
optionally comprising 1, 2, 3 or 4 double bonds and optionally
being substituted by one or more chemical groups, which may be
identical or different, chosen from hydroxyl, halogen atom, alkyl,
alkenyl, alkynyl, alkoxy, alkylthio, aryl, heteroaryl, heterocyclic
radical and trifluoromethyl;
[0197] R.sup.13 is chosen from hydrogen and an alkyl, alkenyl,
alkynyl, aryl, heteroaryl, --N(R.sup.12R.sup.12') or
--N(R.sup.12)OR.sup.13 radical;
[0198] R.sup.14 represents hydrogen; and
[0199] R.sup.15 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, cycloalkyloxy, heteroaryloxy and
heterocyclyloxy;
[0200] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0201] the solvates and hydrates of these compounds;
[0202] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0203] Another preferred group of compounds consists of the
compounds of family (Ie) as kynurenine 3-hydroxylase inhibitors in
any of the abovementioned uses according to the invention, the said
compounds (Ie) belonging to the general formula (I) as defined
above, in which:
[0204] W represents the divalent radical: ##STR8##
[0205] R.sup.1 represents a radical chosen from linear or branched
alkyl containing from 1 to 14 carbon atoms and optionally
substituted, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, a
heterocyclic radical, an aryl radical and a heteroaryl radical;
[0206] R.sup.2 and R.sup.14 together form a bond, thus forming a
double bond between the carbon atoms respectively bearing R.sup.2
and R.sup.14;
[0207] R.sup.3 represents hydrogen;
[0208] R.sup.4 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, heteroaryloxy, --N(R.sup.12R.sup.12') and
--N(R.sup.12)OR.sup.13;
[0209] R.sup.12 and R.sup.12', which may be identical or different,
are chosen, independently of each other, from hydrogen and an
alkyl, alkenyl, alkynyl, alkylcarbonyl, aryl or heteroaryl radical;
or alternatively R.sup.12 and R.sup.12' may form, together with the
nitrogen atom to which they are attached, a monocyclic or bicyclic
heterocyclic group containing a total of 5 to 10 atoms, among which
1, 2, 3 or 4 are chosen, independently of each other, from
nitrogen, oxygen and sulfur, the said heterocyclic radical also
optionally comprising 1, 2, 3 or 4 double bonds and optionally
being substituted by one or more chemical groups, which may be
identical or different, chosen from hydroxyl, halogen atom, alkyl,
alkenyl, alkynyl, alkoxy, alkylthio, aryl, heteroaryl, heterocyclic
radical and trifluoromethyl;
[0210] R.sup.13 is chosen from hydrogen and an alkyl, alkenyl,
alkynyl, aryl, heteroaryl, --N(R.sup.12R.sup.12') or
--N(R.sup.12)OR.sup.13 radical; and
[0211] R.sup.15 represents hydrogen;
[0212] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0213] the solvates and hydrates of these compounds;
[0214] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0215] According to another variant of the present invention, this
invention relates to the use of compounds of family (If) as
kynurenine 3-hydroxylase inhibitors in any of the abovementioned
uses according to the invention, the said compounds (If) belonging
to the general formula (I) as defined above, in which:
[0216] W represents the divalent radical: ##STR9##
[0217] R.sup.1 represents a radical chosen from linear or branched
alkyl containing from 1 to 14 carbon atoms and optionally
substituted, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, a
heterocyclic radical, an aryl radical and a heteroaryl radical;
[0218] R.sup.2 and R.sup.14 together form a bond, thus forming a
double bond between the carbon atoms respectively bearing R.sup.2
and R.sup.14;
[0219] R.sup.3 represents hydrogen;
[0220] R.sup.4 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, heteroaryloxy, --N(R.sup.2R.sup.2') and
--N(R.sup.2)OR.sup.13;
[0221] R.sup.12 and R.sup.12', which may be identical or different,
are chosen, independently of each other, from hydrogen and an
alkyl, alkenyl, alkynyl, alkylcarbonyl, aryl or heteroaryl radical;
or alternatively R.sup.12 and R.sup.12' may form, together with the
nitrogen atom to which they are attached, a monocyclic or bicyclic
heterocyclic group containing a total of 5 to 10 atoms, among which
1, 2, 3 or 4 are chosen, independently of each other, from
nitrogen, oxygen and sulfur, the said heterocyclic radical also
optionally comprising 1, 2, 3 or 4 double bonds and optionally
being substituted by one or more chemical groups, which may be
identical or different, chosen from hydroxyl, halogen atom, alkyl,
alkenyl, alkynyl, alkoxy, alkylthio, aryl, heteroaryl, heterocyclic
radical and trifluoromethyl;
[0222] R.sup.13 is chosen from hydrogen and an alkyl, alkenyl,
alkynyl, aryl, heteroaryl, --N(R.sup.12R.sup.12') or
--N(R.sup.12)OR.sup.13 radical; and
[0223] R.sup.15 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, cycloalkyloxy, heteroaryloxy and
heterocyclyloxy;
[0224] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0225] the solvates and hydrates of these compounds;
[0226] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compound.
[0227] Among the compounds of the general formula (I), and
according to another variant of the invention, the compounds are
chosen from the family of compounds (Ig) consisting of: [0228]
4-(4'-methylcyclohexyl)-4-oxobutanoic acid; [0229]
2-hydroxy-4-(3',4'-difluorophenyl)-4-oxobutanoic acid; [0230]
2-methoxy-4-(3',4'-difluorophenyl)-4-oxobutanoic acid; [0231]
2-hydroxy-3-methyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid;
[0232] 2-hydroxy-3-phenyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic
acid; [0233]
2-hydroxy-3-benzyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid;
[0234] 2-methyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0235]
2-methyl-4-(3',4'-difluorophenyl)-4-oxobutanoic acid; [0236]
2-chloro-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0237]
2-chloro-4-(3',4'-difluorophenyl)-4-oxobutanoic acid; [0238]
2-fluoro-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0239]
2-fluoro-4-(3',4'-difluorophenyl)-4-oxobutanoic acid; [0240]
2-thiomethyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0241]
2-methylidene-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0242]
2-phenyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0243]
2-benzyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0244]
3-methyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0245]
3-phenyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0246]
3-benzyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0247] methyl
(R,S)-2-hydroxy-4-(3',4'-dichlorophenyl)-4-oxobutanoate; [0248]
methyl (R,S)-2-benzyl-4-(3',4'-dichlorophenyl)-4-oxobutanoate;
[0249] 4-(3'-fluorophenyl)-4-oxo-2-butenoic acid; [0250]
4-(3'-chlorophenyl)-4-oxo-2-butenoic acid; [0251]
4-(3'-nitrophenyl)-4-oxo-2-butenoic acid; [0252]
4-(3'-fluoro-4'-methoxyphenyl)-4-oxo-2-butenoic acid; [0253]
2-methyl-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid; [0254]
3-methyl-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid; [0255]
3-phenyl-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid; [0256]
3-benzyl-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid; [0257]
2,3-dimethyl-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid; [0258]
2-hydroxy-4-(3'-chlorophenyl)-4-oxo-2-butenoic acid; [0259]
2-hydroxy-4-(3'-fluorophenyl)-4-oxo-2-butenoic acid; [0260]
2-hydroxy-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid; [0261]
2-hydroxy-4-(3',4'-difluorophenyl)-4-oxo-2-butenoic acid; and
[0262] 2-hydroxy-4-(3'-chloro-4'-methoxyphenyl)-4-oxo-2-butenoic
acid;
[0263] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0264] the solvates and hydrates of these compounds;
[0265] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0266] According to another variant of the invention, a family of
compounds (Ih) having the abovementioned general structure (I) is
defined, for which:
[0267] W represents the divalent radical: ##STR10##
[0268] R.sup.1, R.sup.2, R.sup.3, R.sup.12, R.sup.12'; R.sup.13 and
R.sup.14 areas defined above; and
[0269] R.sup.15 is chosen from a thiol, alkylthio, alkenylthio,
alkynylthio, arylthio, cycloalkylthio, heteroarylthio or
heterocyclylthio radical;
[0270] with the restriction that when R.sup.2, R.sup.3 and R.sup.14
each represent hydrogen, then R.sup.15 cannot represent a thiol or
alkylthio radical;
[0271] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0272] the solvates and hydrates of these compounds;
[0273] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0274] The compounds of family (Ih) form a particularly preferred
aspect of the present invention. The compounds of family (Ih) have
entirely noteworthy hypoglycaemiant properties and, in this
respect, are useful as kynurenine 3-hydroxylase inhibitors in any
of the abovementioned uses according to the invention.
[0275] In addition, the compounds of family (Ih) show inhibitory
activity on kynurenine 3-hydroxylase that may be linked to the
observed effect on the increase in the mass of beta cells,
especially in the case of diabetes.
[0276] A preferred subfamily of the compounds of the family (Ih)
consists of the compounds of the family (Ii) belonging to the
general formula (I) in which:
[0277] W represents the divalent radical: ##STR11##
[0278] R.sup.1 represents an aryl radical;
[0279] R.sup.2 represent hydrogen, or forms a bond with
R.sup.14;
[0280] R.sup.3 represents hydrogen;
[0281] R.sup.4 is chosen from hydroxyl, alkoxy, alkenyloxy,
alkynyloxy, aryloxy, heteroaryloxy, --N(R.sup.12R.sup.12') and
--N(R.sup.12)OR.sup.13;
[0282] R.sup.12 and R.sup.12', which may be identical or different,
are chosen, independently of each other, from hydrogen and an
alkyl, alkenyl, alkynyl, alkylcarbonyl, aryl or heteroaryl radical;
or alternatively R.sup.12 and R.sup.12' may form, together with the
nitrogen atom to which they are attached, a monocyclic or bicyclic
heterocyclic group containing a total of 5 to 10 atoms, among which
1, 2, 3 or 4 are chosen, independently of each other, from
nitrogen, oxygen and sulfur, the said heterocyclic radical also
optionally comprising 1, 2, 3 or 4 double bonds and optionally
being substituted by one or more chemical groups, which may be
identical or different, chosen from hydroxyl, halogen atom, alkyl,
alkenyl, alkynyl, alkoxy, alkylthio, aryl, heteroaryl, heterocyclic
radical and trifluoromethyl;
[0283] R.sup.13 is chosen from hydrogen and an alkyl, alkenyl,
alkynyl, aryl, heteroaryl, --N(R.sup.12R.sup.12') or
--N(R.sup.12)OR.sup.13 radical;
[0284] R.sup.14 represents hydrogen, or forms a bond with R.sup.2;
and
[0285] R.sup.15 represents an arylthio radical;
[0286] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0287] the solvates and hydrates of these compounds;
[0288] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0289] Among the compounds of family (Ii) that are also preferred
are the compounds of family (Ij) corresponding to the general
formula (I), in which:
[0290] W represents the divalent radical: ##STR12##
[0291] R.sup.1 represents a phenyl radical;
[0292] R.sup.2 represents hydrogen;
[0293] R.sup.3 represents hydrogen;
[0294] R.sup.4 is chosen from hydroxyl and an alkoxy radical;
[0295] R.sup.14 represents hydrogen; and
[0296] R.sup.15 represents a phenylthio radical;
[0297] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0298] the solvates and hydrates of these compounds;
[0299] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0300] By way of illustration, examples of compounds of family (Ih)
are:
[0301] compound Ih-1: [0302]
2-(2'-naphthylthio)-4-phenyl-4-oxobutanoic acid;
[0303] compound Ih-2: [0304] 2-phenylthio-4-phenyl-4-oxobutanoic
acid;
[0305] compound Ih-3: [0306]
2-(4'-fluorophenylthio)-4-phenyl-4-oxobutanoic acid;
[0307] compound Ih-4: [0308]
2-(4'-chlorophenylthio)-4-phenyl-4-oxobutanoic acid;
[0309] compound Ih-5: [0310]
2-(4'-methylphenylthio)-4-phenyl-4-oxobutanoic acid;
[0311] compound Ih-6: [0312]
2-(4'-methoxyphenylthio)-4-phenyl-4-oxobutanoic acid;
[0313] compound Ih-7: [0314]
2-cyclohexylthio-4-phenyl-4-oxobutanoic acid;
[0315] compound Ih-8: [0316] 2-benzylthio-4-phenyl-4-oxobutanoic
acid;
[0317] compound Ih-9: [0318] ethyl
2-phenylthio-4-phenyl-4-oxobutanoate;
[0319] compound Ih-10: [0320] ethyl
2-(4'-fluorophenylthio)-4-phenyl-4-oxobutanoate;
[0321] compound Ih-11: [0322] ethyl
2-(4'-chlorophenylthio)-4-phenyl-4-oxobutanoate;
[0323] compound Ih-12: [0324] ethyl
2-(4'-methylphenylthio)-4-phenyl-4-oxobutanoate;
[0325] compound Ih-13: [0326] ethyl
2-(4'-methoxyphenylthio)-4-phenyl-4-oxobutanoate;
[0327] compound Ih-14: [0328] ethyl
2-(2'-naphthylthio)-4-phenyl-4-oxobutanoate;
[0329] compound Ih-15: [0330] ethyl
2-cyclohexylthio-4-phenyl-4-oxobutanoate;
[0331] compound Ih-16: [0332] ethyl
2-benzylthio-4-phenyl-4-oxobutanoate;
[0333] compound Ih-17: [0334]
2-phenylthio-4-(4'-methoxyphenyl)-4-oxobutanoic acid;
[0335] compound Ih-18: [0336]
2-(4'-fluorophenylthio)-4-(4'-methoxyphenyl)-4-oxobutanoic
acid;
[0337] compound Ih-19: [0338]
2-(4'-chlorophenylthio)-4-(4'-methoxyphenyl)-4-oxobutanoic
acid;
[0339] compound Ih-20: [0340]
2-(4'-methylphenylthio)-4-(4'-methoxyphenyl)-4-oxobutanoic
acid;
[0341] compound Ih-21: [0342]
2-(4'-methoxyphenylthio)-4-(4'-methoxyphenyl)-4-oxobutanoic
acid;
[0343] compound Ih-22: [0344]
2-(2'-naphthylthio)-4-(4'-methoxyphenyl)-4-oxobutanoic acid;
[0345] compound Ih-23: [0346]
2-cyclohexylthio-4-(4'-methoxyphenyl)-4-oxobutanoic acid;
[0347] compound Ih-24: [0348]
2-benzylthio-4-(4'-methoxyphenyl)-4-oxobutanoic acid;
[0349] compound Ih-25: [0350]
2-phenylthio-4-(4'-chlorophenyl)-4-oxobutanoic acid;
[0351] compound Ih-26: [0352]
2-(4'-fluorophenylthio)-4-(4'-chlorophenyl)-4-oxobutanoic acid;
[0353] compound Ih-27: [0354]
2-(4'-chlorophenyl)-4-(4'-chlorophenyl)-4-oxobutanoic acid;
[0355] compound Ih-28: [0356]
2-(4'-methylphenylthio)-4-(4'-chlorophenyl)-4-oxobutanoic acid;
[0357] compound Ih-29: [0358]
2-(4'-methoxyphenylthio)-4-(4'-chlorophenyl)-4-oxobutanoic
acid;
[0359] compound Ih-30: [0360]
2-(2'-naphthylthio)-4-(4'-chlorophenyl)-4-oxobutanoic acid;
[0361] and 2-carboxymethylthio-4-phenyl-4-oxobutanoic acid (f);
[0362] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0363] the solvates and hydrates of these compounds;
[0364] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the
pharmaceutically,acceptable prodrugs of these compounds.
[0365] It has been discovered, unexpectedly, that the compounds of
the formula (I) according to the variants described above show
particularly advantageous activity when R.sup.1 is aryl or
heteroaryl; these groups are thus most particularly preferred.
[0366] According to one particular aspect of the invention, among
the different variants of the formula (I) above that are preferred
are the compounds for which, when R.sup.2.dbd.R.sup.3.dbd.H, W is
other than --CH(CH.sub.2--X)-- in which X=alkyl, aryl, cycloalkyl,
pyridyl, pyrimidyl, pyrrolyl, furyl, thienyl, tetrahydrofuryl,
tetrahydropyranyl, piperidyl or pyrrolidinyl, which are optionally
substituted.
[0367] According to another particular aspect of the invention, the
compounds of the formula (I) are different from: [0368] racemic
2-benzyl-4-(4-methoxyphenyl)-4-oxobutanoic acid and the R and S
isomers thereof; [0369] racemic
2-benzyl-4-(4-fluorophenyl)-4-oxobutanoic acid and the R and S
isomers thereof; [0370]
2-cyclohexylmethyl-4-(4-methoxyphenyl)-4-oxobutanoic acid; [0371]
2-benzyl-4-phenyl-4-oxobutanoic acid; [0372]
2-(.beta.-naphthylmethyl)-4-phenyl-4-oxobutanoic acid; [0373]
2-benzyl-4-(.beta.-naphthyl)-4-oxobutanoic acid; [0374]
2-[(4-chlorophenyl)methyl]-4-(4-methoxyphenyl)-4-oxobutanoic acid;
[0375] 2-benzyl-4-(4-methylphenyl)-4-oxobutanoic acid; [0376]
4-(4-fluorophenyl)-2-[(4-methoxyphenyl)methyl]-4-oxobutanoic acid;
[0377] 2-benzyl-4-(3,4-methylenedioxyphenyl)-4-oxobutanoic acid;
[0378] 2-benzyl-4-cyclohexyl-4-oxobutanoic acid; [0379]
4-phenyl-2-[(tetrahydrofur-2-yl)methyl]-4-oxobutanoic acid.
[0380] Among the compounds of the formula (II) defined above that
are preferred are the compounds of the family (IIa) corresponding
to the general formula (II) in which:
[0381] R.sup.5, R.sup.6, R.sup.7 and R.sup.8 are as defined
above;
[0382] R.sup.9 represents hydrogen; and
[0383] R.sup.10 is chosen from a phenyl radical, optionally
substituted in position 3 and/or 4 with an alkyl or alkoxy radical,
preferably methyl or methoxy, and a naphthyl radical;
[0384] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0385] the solvates and hydrates of these compounds;
[0386] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0387] Another family (IIb) of compounds of the formula (II) is
represented by the compounds of the general formula (II) in
which:
[0388] R.sup.5, R.sup.6, R.sup.7 and R.sup.8, which may be
identical or different, are chosen, independently of each other,
from hydrogen, a halogen atom, a nitro radical and a
trifluoromethyl radical;
[0389] the radicals R.sup.6 and R.sup.7 also possibly forming,
together with the carbon atoms to which they are attached, a
benzene ring, optionally substituted by one or more groups, which
may be identical or different, chosen from a halogen atom and a
trifluoromethyl, nitro or alkoxy radical; and
[0390] R.sup.9 and R.sup.10 are as defined above;
[0391] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0392] the solvates and hydrates of these compounds;
[0393] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0394] According to one preferred variant of the invention, the
compounds of the formula (II) are chosen from the list consisting
of: [0395]
4-methoxy-N-(4-naphthalen-2-ylthiazol-2-yl)benzenesulfonamide;
[0396] 4-amino-N-[4-(3-nitrophenyl)thiazol-2-yl]benzenesulfonamide;
[0397]
4-methyl-N-[4-(3-nitrophenyl)thiazol-2-yl]benzenesulfonamide;
[0398]
3,4-dimethoxy-N-[4-(3-nitrophenyl)thiazol-2-yl]benzenesulfonamide;
[0399]
4-methoxy-N-[4-(3-nitrophenyl)thiazol-2-yl]benzenesulfonamide;
[0400] 2-naphthalenesulfonic
acid[4-(3-nitrophenyl)thiazol-2-yl]benzenesulfonamide; [0401]
N-[4-(2-fluoro-5-trifluoromethylphenyl)thiazol-2-yl]-4-methylbenzenesulfo-
namide; [0402]
N-[4-(3-fluoro-5-trifluoromethylphenyl)thiazol-2-yl]-4-methylbenzenesulfo-
namide; [0403]
4-methyl-N-[4-(4-nitrophenyl)thiazol-2-yl]benzenesulfonamide;
[0404]
4-amino-N-[4-(2-fluoro-5-trifluoromethylphenyl)thiazol-2-yl]benzenesulfon-
amide; and [0405]
3,4-dimethoxy-N-[4-(2-fluoro-5-trifluoromethylphenyl)thiazol-2-yl]benzene-
sulfenamide;
[0406] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0407] the solvates and hydrates of these compounds;
[0408] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0409] Among the variants of the formulae (I) and (II) described
above, the compounds that are preferred according to the invention
are those with substantial inhibitory activity on kynurenine
3-hydroxylase as defined above.
[0410] The compounds of the formulae (I) and (II) defined above are
useful as kynurenine 3-hydroxylase inhibitors for any of the uses
according to the invention defined above.
[0411] The pharmaceutical uses or compositions according to the
invention thus comprise as active principle a pharmacologically
effective amount of at least one kynurenine 3-hydroxylase
inhibitor, preferably a compound of the formula (I) or of the
formula (II), alone or in combination with one or more fillers,
vehicles, colorants or sweeteners, i.e. any suitable and
pharmaceutically acceptable non-toxic, inert excipient usually used
in the production of pharmaceutical compositions.
[0412] The said compositions are administered to patients in need
thereof, i.e. to individuals whose condition might be prevented or
improved by increasing the number of islets of Langerhans
cells.
[0413] According to the invention, the kynurenine 3-hydroxylase
inhibitors may be useful in combination with an active agent
usually used in the treatment of diabetes, as a main active
principle or as an adjuvant and/or potentiator of the said
agent.
[0414] The pharmaceutical compositions thus obtained will be in
various forms, the most advantageous being gel capsules,
suppositories, injectable or drinkable solutions, patches, plain,
sugar-coated, film-coated or sublingual tablets, sachets, packets,
lozenges, creams, ointments, dermal gels, aerosols, etc.
[0415] The working dose may be adapted according to the nature and
severity of the pathology to be treated, the administration route
and also the patient's age and weight. In general, the unit dose
will range between 5 mg and 2000 mg per day, in one or more dosage
intakes, advantageously between 10 mg and 1000 mg, for example
between 50 mg and 800 mg.
[0416] It has been discovered, surprisingly, that the kynurenine
3-hydroxylase inhibitors have the twofold activity of controlling
the secretion of both glucagon and insulin. Specifically, in the
absence of glucose, the secretion of glucagon is stimulated whereas
that of insulin is not. In the presence of glucose, the secretion
of insulin is potentiated whereas the secretion of glucagon remains
normally inhibited.
[0417] Such a dual activity affords a considerable improvement over
the processes for the treatment of diabetes currently used.
Specifically, the risks of hypoglycaemia are very greatly reduced,
or even virtually nonexistent, even when the prescribed doses
and/or number of administrations are exceeded or have been poorly
controlled.
[0418] The abovementioned uses according to the invention thus make
it possible to minimise or eliminate the risk of hypoglycaemia.
[0419] Among the compounds of the formula (I) that have inhibitory
activity on kynurenine 3-hydroxylase, non-limiting examples that
may be mentioned include: [0420]
4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0421]
4-(3',4'-difluorophenyl)-4-oxobutanoic acid; [0422] methyl
4-(3',4'-dichlorophenyl)-4-oxobutanoate; [0423]
(R,S)-2-hydroxy-4-(3'-chlorophenyl)-4-oxobutanoic acid; [0424]
(R,S)-2-hydroxy-4-(3'-fluorophenyl)-4-oxobutanoic acid; [0425]
(R,S)-2-hydroxy-4-(3'-nitrophenyl)-4-oxobutanoic acid; [0426]
(R,S)-2-hydroxy-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0427]
(S)-2-hydroxy-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0428]
(R)-2-hydroxy-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0429]
methyl (R,S)-2-hydroxy-4-(3',4'-dichlorophenyl)-4-oxobutanoate;
[0430] (R,S)-2-hydroxy-4-(3',4'-difluorophenyl)-4-oxobutanoic acid;
[0431] (R,S)-2-methoxy-4-(3',4'-difluorophenyl)-4-oxobutanoic acid;
[0432] (R,S)-2-methoxy-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid;
[0433] (R,S)-2-methyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid;
[0434] (R,S)-3-methyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid;
[0435] 2-hydroxy-3-benzyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic
acid; [0436] (R,S)-2-methyl-4-(3',4'-difluorophenyl)-4-oxobutanoic
acid; [0437] (R,S)-2-chloro-4-(3',4'-dichlorophenyl)-4-oxobutanoic
acid; [0438]
(R,S)-2-methylidene-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid;
[0439] (R,S)-3-phenyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid;
[0440] methyl
(R,S)-2-benzyl-4-(3',4'-dichlorophenyl)-4-oxobutanoate; [0441]
(R,S)-2-phenyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0442]
(R,S)-2-benzyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid; [0443]
(E)-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid; [0444]
(E)-4-(3',4'-difluorophenyl)-4-oxo-2-butenoic acid; [0445]
(E)-4-(3'-fluorophenyl)-4-oxo-2-butenoic acid; [0446]
(E)-4-(3'-chlorophenyl)-4-oxo-2-butenoic acid; [0447]
(E)-4-(3'-nitrophenyl)-4-oxo-2-butenoic acid; [0448]
(E)-2-methyl-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid; [0449]
3-methyl-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid; [0450]
3-benzyl-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid; [0451]
(E)-2-hydroxy-4-(3'-chlorophenyl)-4-oxo-2-butenoic acid; [0452]
(E)-2-hydroxy-4-(3'-fluorophenyl)-4-oxo-2-butenoic acid; [0453]
(E)-2-hydroxy-4-(4'-chlorophenyl)-4-oxo-2-butenoic acid; [0454]
(E)-2-hydroxy-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoic acid;
[0455] (E)-2-hydroxy-4-(3',4'-difluorophenyl)-4-oxo-2-butenoic
acid; [0456] methyl
(E)-2-hydroxy-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoate; and
[0457] ethyl
(E)-2-hydroxy-4-(3',4'-dichlorophenyl)-4-oxo-2-butenoate;
[0458] and also the possible geometrical and/or optical isomers
thereof, and the possible tautomeric forms thereof;
[0459] the solvates and hydrates of these compounds;
[0460] and also the possible salts thereof with a pharmaceutically
acceptable acid or base, or alternatively the pharmaceutically
acceptable prodrugs of these compounds.
[0461] The invention also relates to a process for increasing the
number of islets of Langerhans cells, comprising the
administration, to a patient requiring it, of a dose of one or more
compounds that inhibit kynurenine 3-hydroxylase of the formula (I)
or of the formula (II) defined above, such that it produces a
substantial inhibition of kynurenine 3-hydroxylase in the
patient.
[0462] In particular, the process defined above allows the
prevention or treatment of diabetes and/or its complications,
especially in the case of patients presenting the characteristics
of the diabetes pathology, without this pathology yet having been
declared. The criteria for diagnosing this pathology are defined,
for example, in Diabetes Care, vol. 25, suppl. 1, January 2002.
[0463] Among the complications that may be mentioned especially are
arterial hypertension, diabetes-related inflammatory processes,
diabetic nephropathy, macroangiopathy and microangiopathy,
peripheral diabetic neuropathy and retinopathy of diabetic
origin.
[0464] As mentioned previously, the compounds of the formulae (I)
and (II) defined above have been found to be useful in the
prevention and/or treatment of diabetes and its complications, by
increasing the number of islets of Langerhans cells, according to a
mode of action that is hitherto unknown in this therapeutic
field.
[0465] The invention also relates to a process for manufacturing
medicaments for increasing the number of islets of Langerhans
cells, especially for the the treatment and/or prevention of
diabetes and its complications, by inhibiting kynurenine
3-hydroxylase, in which at least one compound of the formula (I) or
(II) is subjected to an in vitro test of inhibition of kynurenine
3-hydroxylase, and the molecules responding positively to the said
tests are then conditioned in the form of a pharmaceutical
composition, optionally with addition of a pharmaceutically
acceptable filler or vehicle.
[0466] Finally, the invention also relates to a process for
screening candidate compounds for activity in increasing the number
of islets of Langerhans cells, especially for the the treatment
and/or prevention of diabetes or its complications, by inhibiting
kynurenine 3-hydroxylase, the said candidates not corresponding to
formula (I) or (II), in which process the candidate compounds are
subjected to an in vitro test of inhibition of kynurenine
3-hydroxylase, and the candidate that has responded positively to
this test is selected.
[0467] Among the candidates that will be preferred are the
compounds already known as having antidiabetic activity.
[0468] The examples that follow illustrate, without placing any
limitation of any kind on the invention, some of the subjects of
the invention, in particular the preparation processes and the
activities of some of the compounds described above in antidiabetic
activity tests and tests of inhibition of kynurenine
3-hydroxylase.
PREPARATION EXAMPLE
Preparation of 2-(2'-naphthylthio)-4-phenyl-4-oxobutanoic acid
(Compound Ih-1)
[0469] 7.04 g (0.04 mol) of commercial 3-benzoylacrylic acid are
dissolved in 90 mL of methylene chloride. 2-Naphthalenethiol (0.04
mol; 1 equivalent) is then added. The reaction medium is left for
20 hours at 20.degree. C. and then concentrated under vacuum. The
crude solid product isolated is then triturated from isopropyl
ether, filtered off by suction and recrystallised from isopropyl
ether.
[0470] Isolated weight: 5.55 g; yield=41%; melting
point=146-149.degree. C. (capillary melting point).
[0471] Proton NMR (200 MHz, solvent: deuterated DMSO): 3.74 ppm,
multiplet, 2H; 4.43 ppm, broad singlet, 1H; 7.9 ppm, multiplet, 12H
arom.; 12.9 ppm, COOH).
[0472] Infrared spectrometry (cm.sup.-1): 1702.8; 1680.7; 1595.0;
1435.2; 1326.6; 1217.6.
[0473] TLC Analysis:
[0474] silica, eluent: methylcyclohexane, ethyl acetate, acetic
acid (50/45/5): Rf: 0.53.
[0475] The compounds of the family (Ih) as defined above were
prepared according to a similar process.
Preparation of ethyl
2-(4-methoxyphenylthio)-4-phenyl-4-oxobutanoate (Compound
Ih-13)
[0476] 0.408 g of commercial ethyl benzoylacrylate (0.002 mol) is
dissolved in 6 ml of methylene chloride in a round-bottomed flask
under argon. 0.280 9 (1 equivalent) of 4-methoxythiophenol is then
added.
[0477] The reaction medium is left at 20.degree. C. for 72 hours
and then concentrated under vacuum.
[0478] The crude oil isolated is then purified on a column of
silica (eluent: 90/10 cyclohexane/ethyl acetate).
[0479] Isolated weight: 0.390 g; yield=56.6%; oil.
[0480] Proton NMR (200 MHz, Solvent: Deuterated Chloroform):
[0481] 1.06 ppm, triplet, 3H; 3.41 ppm, multiplet, 2H; 3.66 ppm,
singlet, 3H; 4.01 ppm, multiplet, 3H; 6.72 ppm, doublet, 2H arom.;
7.32 ppm, multiplet, 5H arom.; 7.78 ppm, doublet, 2H arom.
[0482] Infrared spectrometry (cm.sup.-1): 1730.6; 1685.1; 1493.9;
1448.8; 1287.6; 1248.21; 1213.6.
[0483] The ethyl ester compounds of family Ih as defined above were
prepared according to a similar process.
[0484] The compounds of family Ih are collated in Tables I 1-4
below. The purities were determined by HPLC/MS. TABLE-US-00001
TABLE I-1 Compounds Ih ##STR13## Purity Yield m.p. (.degree. C.);
R-SH Number Mass (%) (%) (solvent*) ##STR14## 1h 366.41 99 81.1
146-149 (isopropyl ether) ##STR15## 2h 286.35 99 67.6 132-135
(ethanol 85) ##STR16## 3h 304.34 99 68.4 114 116 (isopropyl ether)
##STR17## 4h 320.8 99 72.5 140-142 (ethanol 85) ##STR18## 5h 300.38
99 66 132-134 (ethanol 95) ##STR19## 6h 316.38 99 77.2 116-118
(ethanol 50) ##STR20## 7h 292.4 99 6.8 117 (ethanol 50) ##STR21##
8h 300.38 99 52.7 143-146 (ethanol 95) *recrystallisation
solvent
[0485] TABLE-US-00002 TABLE I-2 Compounds Ih ##STR22## Purity Yield
m.p. (.degree. C.); R-SH Number Mass (%) (%) (solvent*) ##STR23##
9h 341.41 99 33 oil ##STR24## 10h 332.4 97.4 24 oil ##STR25## 11h
348.85 95.2 19.8 oil ##STR26## 12h 328.43 94.4 24.2 oil ##STR27##
13h 344.43 95.7 56.6 oil ##STR28## 14h 364.47 94 9.6 oil ##STR29##
15h 320.42 99 75.8 oil ##STR30## 16h 328.43 99 41.2 oil
[0486] TABLE-US-00003 TABLE I-3 Compounds Ih ##STR31## Purity Yield
m.p. (.degree. C.); R-SH Number Mass (%) (%) (solvent*) ##STR32##
17h 316.38 99 70.4 121-125 (ethanol 50) ##STR33## 18h 334.37 98.2
51.3 108-110 (ethanol 50) ##STR34## 19h 350.82 99 68 120-121
(ethanol 50) ##STR35## 20h 330.41 99 23.2 137-141 (ethanol 70)
##STR36## 21h 346.4 99 68 137-140 (ethanol 70) ##STR37## 22h 366.44
99 87.4 169-169 (ethanol 50) ##STR38## 23h 322.43 96.7 30.4 120-122
(ethanol 50) ##STR39## 24h 330.41 91.8 72.5 105-109 (ethanol
50)
[0487] TABLE-US-00004 TABLE I-4 Compounds Ih ##STR40## Purity Yield
m.p. (.degree. C.); R-SH Number Mass (%) (%) (solvent*) ##STR41##
25h 320.8 98.5 78.5 166-169 (ethanol 85) ##STR42## 26h 338.79 98.6
81.2 140-141 (ethanol 85) ##STR43## 27h 355.24 97.8 82.8 154-156
(ethanol 85) ##STR44## 28h 334.82 99 62.9 151-153 (ethanol 85)
##STR45## 29h 350.82 99 54.2 117-119 (ethanol 70) ##STR46## 30h
370.86 96.6 82.7 141-145 (isopropyl ether)
[0488] Study of the Inhibitory Activity on Kynurenine 3-Hydroxylase
in Rat Liver
[0489] Experimental Protocol
[0490] Rat livers are homogenised (1:8 weight/volume) in a buffer
solution comprising: 0.25 M sucrose; 50 mM pH 7.4 Tris; 1 mM EDTA;
and 1 mM DTT.
[0491] The homogenates are centrifuged for 10 minutes at 12000 rpm.
The pellets are resuspended in the buffer solution described above
(1:2 weight/volume).
[0492] The kynurenine 3-hydroxylase inhibition is determined by
incubating 10 .mu.L of the homogenate with NADPH (2 mM), kynurenine
(100 .mu.M) and various concentrations of the test compounds in a
final volume of 100 .mu.L at 37.degree. C. for 5 minutes.
[0493] The compounds are tested at concentrations of between 1
.mu.M and 300 .mu.M.
3,4-Dimethoxy-N-[4-(3-nitrophenyl)thiazol-2-yl]benzenesulfonamide
is a compound from the company Hoffmann-LaRoche (Basle, see J. Med.
Chem., 40 (1997), 4738). 30H-Kynurenine was tested according to the
protocol described by Carpendo et al. (Neuroscience, 61 (1994),
237-244).
[0494] Results:
[0495] Each of the experiments is repeated once and the IC.sub.50
values (in .mu.mol/L) are calculated and given in the form of a
mean of these two experiments.
[0496] By way of example,
(R)-2-benzyl-4-(4-fluorophenyl)-4-oxobutanoic acid (compound i) has
an IC.sub.50 value of 1.+-.0.2 .mu.mol/L, whereas
3,4-dimethoxy-N-[4-(3-nitrophenyl)thiazol-2-yl]benzenesulfonamide
(compound k) has an IC.sub.50 value of 10.+-.2.1 .mu.mol/L.
[0497] Results concerning representative examples of family Ih are
given in Table II below, in which is indicated the measurement of
the percentage of remaining kynurenine 3-hydroxylase activity
relative to the control (100%). TABLE-US-00005 ##STR47## Kynurenine
3-hydroxylase inhibition R-SH Ih 10 .mu.M % control ##STR48## Ih-1
23.2 ##STR49## Ih-2 70.4 ##STR50## Ih-3 50.4 ##STR51## Ih-4 34.8
##STR52## Ih-5 45.4 ##STR53## Ih-7 81.3 ##STR54## Ih-8 68.6
##STR55## Kynurenine 3-hydroxylase inhibition R-SH Ih 10 .mu.M %
control ##STR56## Ih-9 80.8 ##STR57## Ih-10 66.7 ##STR58## Ih-11
44.6 ##STR59## Ih-12 63.3 ##STR60## Ih-13 55.2 ##STR61## Ih-14 30.0
##STR62## Ih-15 95.0 ##STR63## Ih-16 84.4
[0498] TABLE-US-00006 ##STR64## Kynurenine 3-hydroxylase inhibition
R-SH Ih 10 .mu.M % control ##STR65## Ih-17 16.0 ##STR66## Ih-18 6.6
##STR67## Ih-19 4.1 ##STR68## Ih-20 13.3 ##STR69## Ih-21 17.4
##STR70## Ih-22 8.5 ##STR71## Ih-23 38.1 ##STR72## Ih-24 18.9
##STR73## Kynurenine 3-hydroxylase inhibition R-SH Ih 10 .mu.M %
control ##STR74## Ih-25 67.6 ##STR75## Ih-26 55.5 ##STR76## Ih-27
34.9 ##STR77## Ih-28 50.5 ##STR78## Ih-30 24.3
[0499] Study of the Antidiabetic Activity in N0STZ Rats
[0500] The antidiabetic activity of the compounds of the formulae
(I) and (II) orally was determined on an experimental model of
non-insulin-dependent diabetes, induced in rats with
steptozotocin.
[0501] The model of non-insulin-dependent diabetes is obtained in
the rats by means of a neonatal injection (on the day of birth) of
steptozotocin.
[0502] The diabetic rats used are eight weeks old. The animals are
housed, from the day of birth to the day of the experiment, in an
animal house at a regulated temperature of 21 to 22.degree. C. and
subjected to a fixed cycle of light (from 7 a.m. to 7 p.m.) and
darkness (from 7 p.m. to 7 a.m.). Their food consisted of a
maintenance diet, and water and food were given "ad libitum", with
the exception of fasting two hours before the tests, during which
period the food is removed (post-absorptive state).
[0503] The rats are treated orally for one (D1) or four (D4) days
with the test product. Two hours after the final administration of
the product and 30 minutes after anaesthetising the animals with
pentobarbital sodium (Nembutal.RTM.), a 300 .mu.L blood sample is
taken from the end of the tail.
[0504] Among the compounds of the formula (I), the compounds of the
family (Ih), especially the compounds of the subfamily (Ii), in
particular compound Ih-1 defined previously
(2-(2'-naphthylthio)-4-phenyl-4-oxobutanoic acid) and compound Ih-3
of the subfamily (Ij)
(2-(4'-fluorophenylthio)-4-phenyl-4-oxobutanoic acid) were
evaluated according to the experimental protocol described
above.
[0505] The results presented below are expressed as a percentage
change in the glycaemia on D1 and D4 (number of days of treatment)
relative to D0 (before the treatment). TABLE-US-00007 Compound D1
(20 mg) D1 (200 mg) D4 (20 mg) D4 (200 mg) Ih-3 -3 7 -19 -12 Ih-1 7
10 -12 -21
[0506] These results show the efficacy of the compounds, especially
of the formula (Ih), in reducing glycaemia in the diabetic
animals.
[0507] This antidiabetic activity is correlated with an inhibitory
effect of this family of molecules on kynurenine 3-hydroxylase.
[0508] Study of the Effect on Glucose Production By the Liver
[0509] Materials and Method:
[0510] The hepatocytes are isolated from the liver of Wistar rats
fasted for 24 hours, according to the method described in Methods
Cell Biol., 13 (1975), 29-83.
[0511] The following two methods were used:
[0512] 1) The hepatocytes are cultured for 16 to 18 hours in DMEM
medium in the presence of AMP cyclase/dexamethasone at respective
concentrations of 5.times.10.sup.-5 M and 5.times.10.sup.-7 M, with
preincubation of the products at the test doses. After washing in
pH 7.4 PBS buffer, the cells are incubated for three hours at
37.degree. C. in a Krebs/AMPc/DEX buffer at the abovementioned
concentrations. 0.1 .mu.M insulin is used as reference substance.
Two identical experiments are performed (Table III-1).
[0513] 2) The hepatocytes are cultured for 16 to 18 hours in RPMI
1640 medium free of glucose but supplemented with 1% glutamine, 100
U/mL penicillin, 100 mg/mL streptomycin and 7.times.10.sup.-5 M
hydrocortisone hemisuccinate.
[0514] After washing in pH 7.4 PBS buffer, the cells are incubated
for two hours at 37.degree. C. in a Krebs buffer free of glucose
and of insulin, containing lactate/pyruvate (10/1 mM) in the
presence or absence of the test compounds. 10 .mu.M MICA
(5-methoxyindole-2-carboxylic acid) is used as reference substance.
Two identical experiments are performed (Table III-2).
[0515] Quantification of the glucose is performed via a
calorimetric method using glucose oxidase (IL test.TM. Glucose,
Monarch 181633-80). The protein assay is performed on the rest of
the incubation medium via the Lowry method (BIO-RAD Dc protein
assay, BIO-RAD 5000116).
[0516] The results are expressed as nmoles of glucose produced per
ng of proteins. The statistical test used is the t test.
[0517] Results:
[0518] It was thus demonstrated that tryptophan and kynurenine are
powerful inhibitors of hepatic glucose production in vitro.
[0519] Effect of kynurenine 3-hydroxylase inhibitors By way of
example, compound Ih-1 (Table III 1-3) and
(R)-2-benzyl-4-(4-fluorophenyl)-4-oxobutanoic acid (compound i) and
(R,S)-2-benzyl-4-(3',4'-dichlorophenyl)-4-oxobutanoic acid
(compound j) (Table IV), two kynurenine 3-hydroxylase inhibitors,
were found to be powerful inhibitors of hepatic glucose production
in vitro, as shown by the following results: TABLE-US-00008 TABLE
III-1 Products tested on primary hepatocytes Hepatic Glucose
Production stimulated by AMPc/DEX Test HGP Proteins Products
Concentration % of control % of control Ih-1 1 .mu.M 103 113 10
.mu.M 83 117 100 .mu.M 15 85
[0520] TABLE-US-00009 TABLE III-2 Products tested on primary
hepatocytes Hepatic Production Glucose Basal Lact/Pyr 2 hours Test
HGP Proteins Products Concentration % of control % of control Ih-1
1 .mu.M 110 94 10 .mu.M 127 101 100 .mu.M 75 96
[0521] TABLE-US-00010 TABLE IV Concentration Hepatic glucose
production inhibition Compound (.mu.M) (mmol/mg of protein) (%)
MICA 10 67** Compound i 0 101 .+-. 6 -- 1 88 .+-. 7 13 10 73 .+-. 4
28** 100 39 .+-. 3 62** Compound j 0 101 .+-. 6 -- 1 71 .+-. 3 30**
10 50 .+-. 3 51** 100 35 .+-. 1 65** Compound k 0 587 .+-. 12 -- 10
605 .+-. 24 0 100 460 .+-. 12 22 Kynurenine 0 101 .+-. 6 -- 1 99
.+-. 5 2 10 97 .+-. 6 4 100 66 .+-. 4 25** 1000 22 .+-. 2 78**
Tryptophan 0 587 .+-. 12 -- 10 518 .+-. 8 12 100 111 .+-. 5
81**
[0522] Study of the Effect on the Secretion of the Pancreatic
Hormones Insulin and Glucagon, in N0STZ Diabetic Rats
[0523] Materials and Method:
[0524] The pancreas is taken from animals rendered diabetic by
injection of streptozotocin on the day of birth (Portha et al.,
Diabetes, 23: 889-895; (1974)) and anaesthetised with pentobarbital
(Nembutal: 45 mg/kg; intraperitoneal route).
[0525] The isolation and perfusion of the pancreas were performed
according to a modification (Assan et al., Nature, 239 (1972),
125-126) of the protocol described by Sussman et al. (Diabetes, 15
(1966), 466-472).
[0526] The effect of the compounds or of the reference substances
is tested for 35 minutes (from t=20 minutes to t=55 minutes) in
Krebs buffer in the absence of glucose, and then for 30 minutes
(from t =55 minutes to t =85 minutes) in the presence of 16.5 mM
glucose.
[0527] The concentration of the hormones, insulin and glucagon,
secreted into the medium is measured via a competitive
radioimmunoassay using the kits: Insulin-CT Cis Bio-International,
Schering and Glucagon-10904-Biochem immuno system,
respectively.
[0528] The results are expressed as the mean .+-.SEM (standard
error of mean) of several experiments. The statistical test used is
the Scheffe test.
[0529] Results:
[0530] Effect of Tryptophan and its Metabolites on the Secretion of
Insulin and Glucagon in Perfused Isolated Pancreases from N0 STZ
Diabetic Rats
[0531] FIG. 1 shows that tryptophan stimulates insulin secretion in
a glucose-dependent manner in a diabetic rat pancreas. Similarly,
FIG. 2 shows that tryptophan stimulates glucagon secretion in a
glucose-dependent manner in a diabetic rat pancreas.
[0532] Kynurenic acid, like tryptophan, stimulates the secretion of
insulin (FIG. 3) and of glucagon (FIG. 4) in a glucose-dependent
manner in a diabetic rat pancreas.
[0533] FIG. 5 and FIG. 6 show the secretion profile for insulin and
glucagon, respectively, stimulated with kynurenine (at 10.sup.-4 M
and 10.sup.-5 M) in a glucose-dependent manner in a diabetic rat
pancreas. This stimulation is similar to that obtained with
tryptophan and kynurenic acid.
[0534] Effect of Kynurenine 3-Hydroxylase Inhibitors on the
Secretion of Insulin and Glucagon in Perfused Isolated Pancreases
from N0 STZ Diabetic Rats
[0535] The kynurenine 3-hydroxylase inhibitors show the same
insulin and glucagon secretion profile as for tryptophan,
kynurenine and kynurenic acid. This observation may be seen in
FIGS. 7 and 8 (stimulation of insulin and of glucagon,
respectively, with compound i) and in FIGS. 9 and 10 (stimulation
of insulin and of glucagon, respectively, with compound k).
[0536] Study of the Activity on Isolated Rat Islets
[0537] Effect of the chemical compounds on insulin secretion as a
function of the glucose concentration, in vitro, in isolated islets
of Langerhans in static incubation:
[0538] The islets of Langerhans obtained by digestion of exocrine
pancreatic tissue with collagenase, and then purified on Ficoll
gradient, are incubated for 90 minutes in the presence of two
concentrations of glucose, (2.8 mM or 8 mM), in the presence or
absence of the chemical compound. The insulin secretion is assayed
by RIA in the incubation medium.
[0539] The potential of the various chemical compounds to stimulate
insulin secretion is estimated by calculating the stimulation
factor*.
[0540] A compound stimulates the secretion of insulin if this
factor is greater than or equal to 130% for a given dose of
insulin. * .times. NB .times. : .times. .times. stimulation .times.
.times. factor = ( G + Product ) * 100 G ##EQU1## where [0541]
G=secretion of insulin (pmol/min. islet) in the presence of glucose
alone [0542] G+Product=secretion of insulin (pmol/min. islet) in
the presence of the same concentration of glucose and of the test
chemical compound.
[0543] FIG. 11 shows the insulin secretion for compounds Ih-18 and
(i) at 10.sup.-5 M at glucose concentrations of 2.8 mM and 8
mM.
[0544] Study of the Effect on the Increase in the Mass of Beta
Cells
[0545] Culturing of Rat Foetal Pancreases
[0546] Experimental Protocol
[0547] Embryonic pancreases are collected on day 12.5 of gestation
from gestating females of the Wistar strain, which have received an
overdose of sodium pentobarbital. The embryos are extracted from
the uterus and placed in phosphate-buffered saline (PBS). The
dorsal pancreatic bud is dissected under stereomicroscopy. The
separation of the mesenchyme, which inhibits the development of the
endocrine pancreas, is performed via an enzymatic reaction with
0.05% concentrated collagenase A in the synthetic culture medium
RPMI 1640.
[0548] The pancreatic epithelia thus isolated are inserted into a
collagen gel, which allows three-dimensional culturing to be
performed. The pancreases are cultured in the RPMI 1640 culture
medium supplemented with 10% foetal calf serum and 5.5 mM glucose
and in the absence (control) or in the presence of the test
compounds. The cultures are maintained at 37.degree. C. in the
presence of 5% CO.sub.2 for seven days. The culture medium is
renewed every day.
[0549] At the end of the seven days of culturing, the pancreases
are isolated from the collagen gels and dissociated into individual
cells by means of a trypsin digestion (0.05% trypsin-EDTA) for
three minutes at 37.degree. C. The enzymatic reaction is quenched
by adding RPMI 1640 medium containing 20% foetal calf serum. The
cells are washed with the same medium and then fixed to glass
slides using a cytocentrifuge for five minutes at 125.times.g. The
cells are then treated with 4% paraformaldehyde, and then incubated
overnight at 4.degree. C. with guinea pig anti-insulin antibody
(1:1 500 dilution). After washing several times with PBS, they are
incubated with FITC-coupled, rabbit anti-guinea pig IgG (dilution
1:100) for 75 minutes at room temperature. The cells are finally
mounted in a medium that protects the fluorescence and that
contains DAPI for labelling the cell nuclei. On each slide, a
minimum of 300 nuclei and the amount of cells expressing insulin
are counted. The calculation of the amount of beta cells represents
the proportion of cells expressing insulin to the total number of
nuclei counted. An experiment is performed with a minimum of four
pancreases per group and each experiment is repeated three
times.
[0550] FIGS. 12, 13, 14 and 15 represent the amount of beta cells
expressing insulin in the cultured rat foetal pancreatic buds over
seven days, with or without test compound. The increase in the
number of beta cells is mainly due to stimulation of the neogenesis
of these cells from the stems cells.
* * * * *