Note: Descriptions are shown in the official language in which they were submitted.
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
TITLE OF THE INVENTION
ARYLOXYACETIC ACIDS FOR DIABETES AND LIPID DISORDERS
FIELD OF THE INVENTION
The instant invention is concerned with aryloxyacetic acids and
pharmaceutically acceptable salts and prodrugs thereof which are useful as
therapeutic compounds, particularly in the treatment of Type 2 diabetes
mellitus, often
referred to as non-insulin dependent diabetes (NIDDM), of conditions that are
often
associated with this disease, and of lipid disorders.
BACKGROUND OF THE INVENTION
Diabetes refers to a disease process derived from multiple causative
factors and characterized by elevated levels of plasma glucose or
hyperglycemia in the
fasting state or after administration of glucose during an oral glucose
tolerance test.
Persistent or uncontrolled hyperglycemia is associated with increased and
premature
morbidity and mortality. Often abnormal glucose homeostasis is associated both
directly and indirectly with alterations of the lipid, lipoprotein and
apolipoprotein
metabolism and other metabolic and hemodynamic disease. Therefore patients
with
Type 2 diabetes mellitus are at especially increased risk of macrovascular and
microvascular complications, including coronary heart disease, stroke,
peripheral
vascular disease, hypertension, nephropathy, neuropathy, and retinopathy.
Therefore,
therapeutical control of glucose homeostasis, lipid metabolism and
hypertension are
critically important in the clinical management and treatment of diabetes
mellitus.
There are two generally recognized forms of diabetes. In type 1
diabetes, or insulin-dependent diabetes mellitus (IDDM), patients produce
little or no
insulin, the hormone which regulates glucose utilization. In type 2 diabetes,
or
noninsulin dependent diabetes mellitus (NIDDM), patients often have plasma
insulin
levels that are the same or even elevated compared to nondiabetic subjects;
however,
these patients have developed a resistance to the insulin stimulating effect
on glucose
and lipid metabolism in the main insulin-sensitive tissues, which are muscle,
liver and
adipose tissues, and the plasma insulin levels, while elevated, are
insufficient to
overcome the pronounced insulin resistance.
Insulin resistance is not primarily due to a diminished number of
insulin receptors but to a post-insulin receptor binding defect that is not
yet
understood. This resistance to insulin responsiveness results in insufficient
insulin
-1-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
activation of glucose uptake, oxidation and storage in muscle and inadequate
insulin
repression of lipolysis in adipose tissue and of glucose production and
secretion in the
liver. .
The available treatments for type 2 diabetes, which have not changed
substantially in many years, have recognized limitations. While physical
exercise and
reductions in dietary intake of calories will dramatically improve the
diabetic
condition, compliance with this treatment is very poor because of well-
entrenched
sedentary lifestyles and excess food consumption, especially of foods
containing high
amounts of saturated fat. Increasing the plasma level of insulin by
administration of
sulfonylureas (e.g. tolbutamide and glipizide), which stimulate the pancreatic
(3-cells
to secrete more insulin, and/or by injection of insulin after the response to
sulfonylureas fails, will result in high enough insulin concentrations to
stimulate the
very insulin-resistant tissues. However, dangerously low levels of plasma
glucose can
result from these last two treatments, and increasing insulin resistance due
to the even
higher plasma insulin levels can occur. The biguanides increase insulin
sensitivity
resulting in some correction of hyperglycemia. However, the two biguanides,
phenformin and metformin, can induce lactic acidosis and nausea/diarrhea,
respectively.
The glitazones (i.e. 5-benzylthiazolidine-2,4-diones) are a more
recently described class of compounds with potential for a novel mode of
action in
ameliorating many symptoms of type 2 diabetes. These agents substantially
increase
insulin sensitivity in muscle, liver and adipose tissue in several animal
models of type
2 diabetes resulting in partial or complete correction of the elevated plasma
levels of
glucose without occurrence of hypoglycemia.
Disorders of lipid metabolism or dyslipidemias include various
conditions characterized by abnormal concentrations of one or more lipids
(i.e.
cholesterol and triglycerides), and/or apolipoproteins (i.e., apolipoproteins
A, B, C
and E), and/or lipoproteins (i.e., the macromolecular complexes formed by the
lipid
and the apolipoprotein that allow lipids to circulate in blood, such as LDL,
VLDL and
IDL) . Cholesterol is mostly carried in Low Density Lipoproteins (LDL), and
this
component is commonly known as the "bad" cholesterol because it has been shown
that elevations in LDL-cholesterol correlate closely to the risk of coronary
heart
disease. A smaller component of cholesterol is carried in the High Density
Lipoproteins and is commonly known as the "good" cholesterol. In fact, it is
known
that the primary function of HDL is to accept cholesterol deposited in the
arterial wall
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
and to transport it back to the liver for disposal through the intestine.
Although it is
desirable to lower elevated levels of LDL cholesterol, it is also desirable to
increase
levels of HDL cholesterol. Generally, it has been found that increased levels
of HDL
are associated with lower risk for coronary heart disease (CHD). See, for
example,
Gordon, et al., Am. J. Med., 62, 707-714 (1977); Stampfer, et al., N. England
J.
Med., 325, 373-381 (1991); and Kannel, et al., Ann. Internal Med., 90, 85-91
(1979).
An example of an HDL raising agent is nicotinic acid, a drug with limited
utility
because doses that achieve HILL raising are associated with undesirable
effects, such
as flushing.
Dyslipidemias were originally classified by Fredrickson according to
the combination of alterations mentioned above. The Fredrickson classification
includes 6 phenotypes (i.e., I, IIa, IIb, III, IV and V) with the most common
being the
isolated hypercholesterolemia (or type IIa) which is usually accompained by
elevated
concentrations of total and LDL cholesterol. The initial treatment for
hypercholesterolemia is often to modify the diet to one low in fat and
cholesterol,
coupled with appropriate physical exercise, followed by drug therapy when LDL-
lowering goals are not met by diet and exercise alone
A second common form of dyslipidemia is the mixed or combined
hyperlipidemia or type IIb and III of the Fredrickson classification. This
dyslipidemia
is often prevalent in patients with type 2 diabetes, obesity and the metabolic
syndrome. In this dyslipidemia there are modest elevations of LDL-cholesterol,
accompanied by more pronounced elevations of small dense LDL-cholesterol
particles, VLDL and/or IDL (i.e., triglyceride rich lipoproteins), and total
triglycerides. In addition, concentrations of HDL are often low.
Peroxisome proliferators are a structurally diverse group of compounds
that when administered to rodents elicit dramatic increases in the size and
number of
hepatic and renal peroxisomes, as well as concomitant increases in the
capacity of
peroxisornes to metabolize fatty acids via increased expression of the enzymes
of the
beta-oxidation cycle. Compounds of this group include but are not limited to
the
fibrate class of lipid modulating drugs, herbicides and phthalate
plasticizers.
Peroxisome proliferation is also triggered by dietary or physiological factors
such as a
high-fat diet and cold acclimatization.
Three sub-types of peroxisome proliferator activated receptor (PPAR)
have been discovered and described; they are peroxisome proliferator activated
receptor alpha (PPARa), peroxisome proliferator activated receptor gamma
-3-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
(PPAR~y) and peroxisome proliferator activated receptor delta (PPARB).
Identification
of PPARcc, a member of the nuclear hormone receptor superfamily activated by
peroxisome proliferators, has facilitated analysis of the mechanism by which
peroxisome proliferators exert their pleiotropic effects. PPARa is activated
by a
number of medium and long-chain fatty acids, and it is involved in
stimulating (3-oxidation of fatty acids. PPARoc is also associated with the
activity of
fibrates and fatty acids in rodents and humans. Fibric acid derivatives such
as
clofibrate, fenofibrate, benzafibrate, ciprofibrate, beclofibrate and
etofibrate, as well
as gemfibrozil, each of which are PPARa ligands and/or activators, produce a
substantial reduction in plasma triglycerides as well as some increase in HDL.
The
effects on LDL cholesterol are inconsistent and might depend upon the compound
and/or the dyslipidemic phenotype. For these reasons, this class of compounds
has
been primarily used to treat hypertriglyceridemia (i.e, Fredrickson Type IV
and V)
and/or mixed hyperlipidemia.
The PPAR7 receptor subtypes are involved in activating the program of
adipocyte differentiation and are not involved in stimulating peroxisome
proliferation
in the liver. There are two known protein isoforms of PPAR~ : PPAR~yl and
PPAR~y2
which differ only in that PPAR~y2 contains an additional 28 amino acids
present at the
amino terminus. The DNA sequences for the human isotypes are described in
Elbrecht, et al., BBRC 224;431-437 (1996). In mice, PPAR~y2 is expressed
specifically in fat cells. Tontonoz et al., Cell 79: 1147-1156 (1994) provide
evidence
to show that one physiological role of PPAR~y2 is to induce adipocyte
differentiation.
As with other members of the nuclear hormone receptor superfamily, PPAR~y2
regulates the expression of genes through interaction with other proteins and
binding
to hormone response elements, for example in the 5' flanking regions of
responsive
genes. An example of a PPAR~y2 responsive gene is the tissue-specific
adipocyte P2
gene. Although peroxisome proliferators, including the fibrates and fatty
acids,
activate the transcriptional activity of PPAR's, only prostaglandin J2
derivatives have
been identified as potential natural ligands of the PPAR~y subtype, which also
binds
thiazolidinedione antidiabetic agents with high affinity.
The human nuclear receptor gene PPARB (hPPARB) has been cloned
from a human osteosarcoma cell cDNA library and is fully described in A.
Schmidt et
al., Molecular Efzdocr-i~zology, 6 :1634-1641 (1992). It should be noted that
PPARB is
-4-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
also referred to in the literature as PPAR(3 and as NUCl, and each of these
names
refers to the same receptor; in Schmidt et al. the receptor is referred to as
NUC1.
In W096/01430, a human PPAR subtype, hNUCIB, is disclosed. The
amino acid sequence of hNUCIB differs from human PPARB (referred to therein as
hNUCl) by one amino acid, i.e., alanine at position 292. Based on in vivo
experiments described therein, the authors suggest that hNUCIB protein
represses
hPPARcc and thyroid hormone receptor protein activity.
It has been disclosed in W097/28149 that agonists of PPARB are
useful in raising HDL plasma levels. W097/27857, 97/28115, 97/28137 and
97/27847 disclose compounds that are useful as antidiabetic, antiobesity, anti-
atherosclerosis and antihyperlipidemic agents, and which may exert their
effect
through activation of PPARs.
It is generally believed that glitazones exert their effects by binding to
the peroxisome proliferator activated receptor (PPAR) family of receptors,
controlling
certain transcription elements having to do with the biological entities
listed above.
See Hulin et al., Current Pharm. Design (1996) 2, 85-102.
A number of glitazones that are PPAR agonists have been approved for
use in the treatment of diabetes. These include troglitazone, rosiglitazone
and
pioglitazone, all of which are primarily or exclusively PPAR~y agonists. Many
of the
newer PPAR agonists that are currently under development or are in clinical
trials
have dual PPARoc and'y activity. These are expected to improve both insulin
sensitivity and the lipid profile in patients having NIDDM.
Although glitazones are beneficial in the treatment of NIDDM, there
have been some serious adverse events associated with the use of the
compounds.
The most serious of these has been liver toxicity, which has resulted in a
number of
deaths. The most serious problems have occurred using troglitazone. Because of
the
problems that have occurred with the glitazones, researchers in a number of
laboratories have been investigating classes of PPAR agonists that are not
glitazones
and do not contain 1,3-thiazolidinedione moieties.
Compounds that are not glitazones but are agonists of PPAR sub-types
are expected to be useful in the treatment of diabetes and associated
conditions.
PPARa agonists should improve the lipid profile and alleviate dyslipidemias by
reducing elevated LDL levels and elevated triglyceride levels and/or
increasing HDL
-5-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
levels. PPAR~y agonists should improve insulin sensitivity, reducing the need
for
insulin injections in patients with NIDDM. The role of PPARB is less well
defined.
SUMMARY OF THE INVENTION
The class of compounds described herein is a new class of PPAR
agonists that do not contain a 1,3-thiazolidinedione moiety and therefore are
not
glitazones. The class of compounds includes compounds that are primarily
PPARoc
agonists and compounds that are mixed PPARaI~ agonists. These compounds are
useful in the treatment, control andlor prevention of diabetes, hyperglycemia,
mixed
or diabetic dyslipidemia, and other lipid disorders (including isolated
hypercholesterolemia as manifested by elevations in LDL-C andlor non-HDL-C
and/or hyperapoBliproteinemia, hypertriglyceridemia and/or increase in
triglyceride-
rich-lipoproteins, or low HDL cholesterol concentrations), atherosclerosis,
obesity,
vascular restenosis, inflammatory conditions, neoplastic conditions, and other
PPARa
and/or'y mediated diseases, disorders and conditions.
The present invention provides compounds having the structure of
Formula I, including pharmaceutically acceptable salts and prodrugs of these
compounds:
R5
-X
Y
Rs ~ Ra.
Z
R1~
2 'C02H
R
In the compounds of Formula I:
-6-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
R1 and R2 are each independently selected from the group consisting
of H, F, C1_5 alkyl, C2_5 alkenyl, and C2_5 alkynyl, wherein said alkyl,
alkenyl, and
alkynyl may be linear or branched and are optionally substituted with 1-3
halogen
atoms; or optionally R1 and R2 together form a C3_6 cycloalkyl;
R3 and R4 are each independently selected from the group consisting
of C1-C5 alkyl, C2_5 alkenyl, C2_5 alkynyl, and chlorine, provided that R3 and
R4 are
not both chlorine, wherein said allcyl, alkenyl, and alkynyl groups may be
linear or
branched and are optionally substituted with 1-5 fluorine atoms;
X is N or CR;
Y is O, S, or NR;
ZisOorS;
Each R group is selected from the group consisting of H, C1_5 alleyl,
C2_5 alkenyl, and C2_5 alkynyl, wherein said alkyl, aIkenyl, and alkynyl may
be linear
or branched and are optionally substituted with 1-5 fluorine atoms and/or one -
OC1_
3 alkyl, said -OC1_3 alkyl being optionally substituted with 1-7 fluorine
atoms; and
R5 is selected from the group consisting of H, C1_6 alleyl, C2_g
alkenyl, C2_6 allcynyl, C(_10 Aryl, -0C1-6 alkyl, -OC2_6 alkenyl, -OC2_6
allcynyl, -
OC(_10 Aryl, C3_6 Cycloalkyl, 5-6-rnembered Heterocyclyl, 5-6-membered
Heteroaryl, -OC3_6 Cycloallcyl, -O 5-6-membered Heterocyclyl, -O 5-6 membered
Heteroaryl, and a C1_q. alkyl group which comprises at a position interrupting
the
chain or at the end of the chain a group selected from C(_10 Aryl, C3-6
Cycloallcyl,
5-6-membered Heterocyclyl, and 5-6-membered Heteroaryl, wherein each of said
alkyl, alkenyl, alkynyl, -Oallcyl, -Oallcenyl; and -Oallcynyl is linear or
branched and is
optionally substituted with 1-5 fluorine atoms and/or one -OCH3 or -OCF3
group,
and each of said Aryl, Cycloalkyl, Heteroaryl, Heterocyclyl, -OAryl, -
OCycloalkyl, -
OHeteroaryl, and -OHeterocyclyl groups is optionally substituted with 1-7
halogen
atoms andlor one -OCH3 or -OCF3 group.
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
These compounds are effective in lowering glucose, lipids, and insulin
in diabetic animals. The compounds are expected to be efficacious in the
treatment ,
control and/or prevention of non-insulin dependent diabetes mellitus (NIDDM)
in
humans and in the treatment, control, andlor prevention of conditions
associated with
NIDDM, including hyperlipidemia, dyslipidemia, obesity, hypercholesterolemia,
hypertrigyceridemia, atherosclerosis, vascular restenosis, inflammatory
conditions,
neoplastic conditions, and other PPARa andlor'y mediated diseases, disorders
and
conditions.
DETAILED DESCRIPTION OF THE INVENTION
The invention has numerous embodiments. Several subsets of
compounds having different heterocyclic rings are included, as follows:
Compounds of Formula I, in which X is N and Y is O;
Compounds of Formula I, in which X is N and Y is S;
Compounds of Formula I, in which X is N and Y is NR;
Compounds of Formula I, in which X is CR and Y is O;
Compounds of Formula I, in which X is CR and Y is S; and
Compounds of Formula I, in which X is CR and Y is NR.
In further subsets of the compounds of Formula I described above, in
which the compounds of Formula I have different heterocycles fused to the
aromatic
ring (i.e. different values of X and Y), R5 is selected from the group
consisting of H,
C1_5 alkyl, C2_5 allcenyl, C2_5 alkynyl, OC1_5 alkyl, OC2_5 allcenyl, OC2_5
allcynyl,
and phenyl; in these compounds, the alkyl, alkenyl, alkynyl, -Oalkyl, -
Oalkenyl, and
-Oallcynyl are optionally substituted with 1-5 fluorine atoms, and phenyl is
optionally
substituted with 1-5 halogens. In a preferred subset, R5 is selected from the
group
consisting of H, C1_5 alkyl, C2_5 alkenyl, C2_5 alkynyl, OC1_5 alkyl, OC2_5
allcenyl,
_g_
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
and OC2_5 alkynyl, where alkyl, alkenyl, allcynyl, -Oalkyl, -Oalkenyl, and -
Oalkynyl
are optionally substituted with 1-5 fluorine atoms.
In preferred compounds, R1 and R2 are each H or C1_3 alkyl, and the
number of carbon atoms in R1 and R2 together is 0-5.
In preferred embodiments, R3 and R4 are each independently C1-5
alkyl. In additional preferred embodiments, one of R3 and R4 is C2-5 alkyl,
and the
other of R3 and R4 is C1_5 allcyl. In another preferred embodiment, both R3
and R4
are C2_5 alkyl. In other preferred compounds, one of R3 and R4 is Cl or C1_5
alkyl,
and the other of R3 and R4 is C2_5 allcyl. In general, the alkyl groups are
linear or
branched. In the most preferred compounds R3 and R4 are linear when they are
alkyl.
In preferred compounds, R5 is selected from C1-5 alkyl and -OC1-5
allcyl, where the alkyl and -Oalkyl are optionally substituted with 1-5
fluorine atoms.
In preferred embodiments, Z is O.
In preferred embodiments, X is N and Y is O, so that the compounds
are benzisoxazoles.
In highly preferred embodiments of the groups of compounds above,
R5 is C1_3 alkyl, -OC1_3 alkyl, CF3, C2F5, -OCF3 or -OC2F5~ and R3 and R4 are
each n-propyl.
Specific examples of compounds of this invention are provided as
Examples 1-29.
The invention further includes pharmaceutical compositions
comprising any of the compounds described above and a pharmaceutically
acceptable
carrier.
-9-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
The compounds as defined above are useful in the following methods
of treating, controlling, and preventing diseases, as well as other diseases
not listed
below:
(1) a method for treating, controlling or preventing diabetes
mellitus, and particularly non-insulin dependent diabetes mellitus, in a
mammalian
patient in need of such treatment which comprises administering to the patient
a
therapeutically effective amount of a compound of Formula I;
(2) a method for treating, controlling, or preventing hyperglycemia
in a mammalian patient in need of such treatment which comprises administering
to
the patient a therapeutically effective amount of a compound of Formula I;
(3) a method for treating, controlling, or preventing lipid disorders,
hyperlipidemia, or low HDL in a mammalian patient in need of such treatment
which
comprises administering to the patient a therapeutically effective amount of a
compound of Formula I;
(4) a method for treating, controlling, or preventing obesity in a
mammalian patient in need of such treatment which comprises administering to
the
patient a therapeutically effective amount of a compound of Formula I;
(5) a method for treating, controlling, or preventing
hypercholesterolemia in a mammalian patient in need of such treatment which
comprises administering to the patient a therapeutically effective amount of a
compound of Formula I;
(6) a method for treating, controlling, or preventing
hypertriglyceridemia in a mammalian patient in need of such treatment which
comprises administering to the patient a therapeutically effective amount of a
compound of Formula I;
(7) a method for treating, controlling, or preventing dyslipidemia,
including low HDL cholesterol, in a mammalian patient in need of such
treatment
which comprises administering to the patient a therapeutically effective
amount of a
compound of Formula I;
(8) a method for treating, controlling, or preventing atherosclerosis
in a mammalian patient in need of such treatment which comprises administering
to
the patient a therapeutically effective amount of a compound of Formula I. It
is
understood that the sequellae of atherosclerosis (angina, claudication, heart
attack,
stroke, etc.) are thereby treated.
-10-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
Definitions
"Ac" is acetyl, which is CH3C(O)-.
"Alkyl", as well as other groups having the prefix "ally", such as alkoxy
or alkanoyl, means carbon chains which may be linear or branched or
combinations
thereof, unless the carbon chain is defined otherwise. Examples of alkyl
groups
include methyl, ethyl, propyl, isopropyl, butyl, sec- and tert-butyl, pentyl,
hexyl,
heptyl, octyl, nonyl, and the lilce.
"Alkenyl" means carbon chains which contain at least one carbon-
carbon double bond, and which may be linear or branched or combinations
thereof.
Examples of alkenyl include vinyl, allyl, isopropenyl, pentenyl, hexenyl,
heptenyl, 1-
propenyl, 2-butenyl, 2-methyl-2-butenyl, and the like.
"Alkynyl" means carbon chains which contain at least one carbon-
carbon triple bond, and which may be linear or branched or combinations
thereof.
Examples of alkynyl include ethynyl, propargyl, 3-methyl-1-pentynyl, 2-
heptynyl and
the like.
"Cycloalkyl" means mono- or bicyclic saturated carbocyclic rings, each
having from 3 to 10 carbon atoms. The team also includes a rnonocyclic ring
fused to
an aryl group in which the point of attachment is on the non-aromatic portion.
Examples of cycloalkyl include cyclopropyl, cyclopentyl, cyclohexyl,
cycloheptyl, and
the like.
"Aryl" (and "arylene") means mono- or bicyclic aromatic rings
containing only carbon ring atoms. The term also includes an aryl group fused
to a
monocyclic cycloalkyl or monocyclic heterocyclic group in which the points) of
attachment is on the aromatic portion. The preferred aryl is phenyl.
"Heterocycle"
and "heterocyclic" means a fully or partially saturated monocyclic, bicyclic
or tricyclic
ring containing at least one heteroatom selected from N, S and O, each of said
rings
having from 3 to 10 atoms. Examples of aryl include phenyl, naphthyl, indanyl,
indenyl, and tetrahydronaphthyl. Examples of aryl fused to heterocyclic groups
include 2,3-dihydrobenzofuranyl, benzopyranyl, 1,4-benzodioxanyl,and the like.
Examples of heterocycles include tetrahydrofuran, piperazine, and morpholine.
"Heteroaryl" (and heteroarylene) means a mono-, bi- or tricyclic
aromatic ring containing at least one ring heteroatom selected from N, O and S
(including SO and S02), with each ring containing 5 to 6 atoms. Examples of
heteroaryl include pyrrolyl, isoxazolyl, isothiazolyl, pyrazolyl, pyridyl,
oxazolyl,
oxadiazolyl, thiadiazolyl, thiazolyl, imidazolyl, triazolyl, tetrazolyl,
furanyl, triazinyl,
-11-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
thienyl, pyrimidyl, pyridazinyl, pyrazinyl, benzisoxazolyl, benzoxazolyl,
benzothiazolyl, benzimidazolyl, benzofuranyl, benzothiophenyl (including S-
oxide
and dioxide), furo(2,3-b)pyridyl, quinolyl, indolyl, isoquinolyl, dibenzofuran
and the
like.
"Halogen" includes fluorine, chlorine, bromine and iodine.
The term "composition," as in pharmaceutical composition, is
intended to encompass a product comprising the active ingredient(s), and the
inert
ingredients) that make up the carrier, as well as any product which results,
directly or
indirectly, from combination, complexation or aggregation of any two or more
of the
ingredients, or from dissociation of one or more of the ingredients, or from
other types
of reactions or interactions of one or more of the ingredients. Accordingly,
the
pharmaceutical compositions of the present invention encompass any composition
made by admixing a compound of the present invention and a pharmaceutically
acceptable carrier.
Optical Isomers - Diastereomers - Geometric Isomers - Tautomers
Compounds of Formula I may contain one or more asymmetric centers
and can thus occur as racemates and racemic mixtures, single enantiomers,
diastereomeric mixtures and individual diastereomers. The present invention is
meant
to comprehend all such isomeric forms of the compounds of Formula I.
Some of the compounds described herein contain olefinic double
bonds, and unless specified otherwise, are meant to include both E and Z
geometric
isomers.
Some of the compounds described herein may exist with different
points of attachment of hydrogen, referred to as tautomers. Such an example
may be a
ketone and its enol form, known as lceto-enol tautomers. The individual
tautomers as
well as mixtures thereof are encompassed with compounds of Formula I.
Compounds of the Formula I may be separated into diastereoisomeric
pairs of enantiomers by, for example, fractional crystallization from a
suitable solvent,
for example methanol or ethyl acetate or a mixture thereof. The pair of
enantiomers
thus obtained may be separated into individual stereoisomers by conventional
means,
for example by the use of an optically active acid or base as a resolving
agent.
Alternatively, any enantiomer of a compound of the general Formula I
or Ia may be obtained by stereospecific synthesis using optically pure
starting
materials or reagents of known configuration.
-12-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
S alts
The term "pharmaceutically acceptable salts" refers to salts prepared
from pharmaceutically acceptable non-toxic bases or acids including inorganic
or
organic bases and inorganic.or organic acids. Salts derived from inorganic
bases
include aluminum, ammonium, calcium, copper, ferric, ferrous, lithium,
magnesium,
manganic salts, manganous, potassium, sodium, zinc, and the like. Particularly
preferred are the ammonium, calcium, magnesium, potassium, and sodium salts.
Salts
in the solid form may exist in more than one crystal structure, and may also
be in the
form of hydrates. Salts derived from pharmaceutically acceptable organic non-
toxic
bases include salts of primary, secondary, and tertiary amines, substituted
amines
including naturally occurring substituted amines, cyclic amines, and basic ion
exchange resins, such as arginine, betaine, caffeine, choline, N,N'-
dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-
dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethyl-morpholine, N-
ethylpiperidine, glucamine, glucosamine, histidine, hydrabamine,
isopropylamine,
lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins,
procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine,
tromethamine, and the like.
When the compound of the present invention is basic, salts may be
prepared from pharmaceutically acceptable non-toxic acids, including inorganic
and
organic acids. Such acids include acetic, benzenesulfonic, benzoic,
camphorsulfonic,
citric, ethanesulfonic, fumaric, gluconic, glutamic, hydrobromic,
hydrochloric,
isethionic, lactic, malefic, malic, mandelic, methanesulfonic, mucic, nitric,
pamoic,
pantothenic, phosphoric, succinic, sulfuric, tartaric, p-toluenesulfonic acid,
and the
like. Particularly preferred are citric, hydrobromic, hydrochloric, malefic,
phosphoric,
sulfuric, and tartaric acids.
It will be understood that, as used herein, references to the compounds
of Formula I are meant to also include the pharmaceutically acceptable salts.
Metabolites - Prodru~s
Metabolites of the compounds of this invention that are therapeutically
active also are within the scope of the claimed parent compound. Prodrugs,
which are
compounds that are converted to the claimed compounds as they are being
administered to a patient or after they have been administered to a patient,
are also
-13-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
within the scope of the claimed active compound. A non-limiting example of a
prodrug of the carboxylic acids of this invention would be an ester of the
carboxylic
acid group, for example a C1 to C~ ester, which may be linear or branched, or
an ester
which has functionality that makes it more easily hydrolyzed after
administration to a
patient.
Prodrugs of this class of compounds may be described as compounds
having the Formula Ia:
R5
-X
Y
Rs ~ R4
Z
R1~
2 'C(O)RE
R
la
wherein R6 is a group that is easily removed under physiological
conditions during or after administration to a mammalian patient to yield a
compound
having Formula I, or the carboxylate anion thereof (in solution), or a
pharmaceutically acceptable salt thereof , where R1, R2, R3, R4, R5, X, Y, Z,
and R
are as defined above for compounds having Formula I.
Examples of prodrugs of Formula Ia include compounds in which R6
is selected from the group consisting of -ORS, -OCH20R~, -OCH(CH3)OR~, -
OCH20C(O)R~, -OCH(CH3)OC(O)R~~ -OCH20C(O)OR~, -OCH(CH3)OC(O)OR~~
-NR$R8~ and -O~$Rg, where each R~ is independently selected from C1_6 alkyl
optionally substituted with one or two groups selected from -C02H, -CONH2 , -
NH2, -OH, -OAc, NHAc, and phenyl; and wherein each R$ is independently
selected from H and R~. Compounds having Formula Ia, where R( has the chemical
structure described above, are described as prodrugs. However, regardless of
whether
they are active as prodrugs, yielding compounds or salts of Formula I, or
whether they
have a different means of exhibiting pharmaceutical activity, the compounds of
Formula Ia are included in this invention. Such compounds are claimed herein,
regardless of the mechanism leading to their activity.
-14-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
The description of utility, pharmaceutical compositions, combination
therapies, administration, dosage, and the like are all described in terms of
compounds
of Formula I. These descriptions of utility, etc. also apply to compounds of
Formula
Ia.
Utilities
Compounds of the present invention are potent agonists of varioius
peroxisome proliferator activator receptor subtypes, particularly PPARcc
and/or
PPAR~y. Compounds of the present invention may be selective agonists of one
receptor subtype, e.g. PPARy or PPARa agonists, or they may be agonists of
more
than one receptor subtypes, e.g. dual PPARocJ~y agonists. Compounds of the
present
invention are useful in treating, controlling or preventing diseases,
disorders or
conditions, wherein the treatment is mediated by the activation of an
individual PPAR
subtype (cc or y), or a combination of PPAR subtypes (e.g. ally). Thus one
aspect of
the present invention provides a method for the treatment, control or
prevention of
such diseases, disorders, or conditions in a mammal which comprises
administering to
such mammal a therapeutically effective amount of a compound of Formula I. The
diseases, disorders or conditions for which compounds of the present invention
are
useful in treating, controlling or preventing include, but are not limited to,
(1) diabetes
mellitus, and especially non-insulin dependent diabetes mellitus (NIDDM), (2)
hyperglycemia, (3) low glucose tolerance, (4) insulin resistance, (5) obesity,
(6)
lipid disorders, (7) dyslipidemia, (8) hyperlipidemia, (9)
hypertriglyceridemia, (10)
hypercholesterolemia, (11) low HDL levels, (12) high LDL levels, (13)
atherosclerosis and its sequelae, (14) vascular restenosis, (15) irritable
bowel
syndrome, (16) inflamatory bowel disease, including Crohn's disease and
ulcerative
colitis, (17) other inflammatory conditions, (18) pancreatitis, (19) abdominal
obesity, (20) neurodegenerative disease, (21) retinopathy, (22) neoplastic
conditions,
(23) adipose cell tumors, (24) adipose cell carcinomas, such as liposarcoma,
(25)
prostate cancer and other cancers, including gastric, breast, bladder and
colon cancers,
(26) angiogenesis, (27) Alzheimer's disease, (28) psoriasis, (29)high blood
pressure, (30) Syndrome X, (31) ovarian hyperandrogenism (polycystic ovarian
syndrome), and other disorders where insulin resistance is a component.
Another aspect of the invention provides a method for the treatment,
control, or prevention of hypercholesterolemia, atherosclerosis, low HDL
levels, high
LDL levels, hyperlipidemia, hypertriglyceridemia, and/or dyslipidemia, which
-15-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
comprises administering to a mammal in need of such treatment a
therapeutically
effective amount of an agonist of PPARa and/or PPAR~y or a PPARoc%y dual
agonist.
The PPAR agonist may be used alone or advantageously may be administered with
a
cholesterol biosynthesis inhibitor, particularly an HMG-CoA reductase
inhibitor such
as lovastatin, simvastatin, pravastatin, fluvastatin, atorvastatin,
rivastatin, itavastatin,
or ZD-4522. The PPAR agonist may also be used advantageously in combination
with other lipid lowering drugs such as cholesterol absorption inhibitors (for
example
stanol esters, sterol glycosides such as tiqueside, and azetidinones such as
ezetimibe),
ACAT inhibitors (such as avasimibe), and with niacin, bile acid sequestrants,
microsomal triglyceride transport inhibitors, and bile acid reuptake
inhibitors. These
combination treatments may also be effective for the treatment, control or
prevention
of one or more related conditions selected from the group consisting of
hypercholesterolemia, atherosclerosis, hyperlipidemia, hypertriglyceridemia,
dyslipidemia, high LDL, and low HDL.
Another aspect of the invention provides a method of treating
inflammatory conditions, including inflammatory. bowel disease, Crohn's
disease, and
ulcerative colitis by administering an effective amount of a PPAR agonist,
which may
be a PPARoc agonist, a PPAR~ agonist, or a PPARor,/~y dual agonist. Additional
inflammatory diseases that may be treated with the instant invention include
gout,
rheumatoid arthritis, osteoarthritis, multiple sclerosis, asthma, ARDS,
psoriasis,
vasculitis, ischemialreperfusion injury, frostbite, and related diseases.
Administration and Dose Ranges
Any suitable route of administration may be employed for providing a
mammal, especially a human, with an effective dose of a compound of the
present
invention. For example, oral, rectal, topical, parenteral, ocular, pulmonary,
nasal, and
the like may be employed. Dosage forms include tablets, troches, dispersions,
suspensions, solutions, capsules, creams, ointments, aerosols, and the like.
Preferably
compounds of Formula I are administered orally.
The effective dosage of active ingredient employed may vary
depending on the particular compound employed, the mode of administration, the
condition being treated and the severity of the condition being treated. Such
dosage
may be ascertained readily by a person skilled in the art.
When treating or preventing diabetes mellitus and/or hyperglycemia or
hypertriglyceridemia or other diseases for which compounds of Formula I are
indicated, generally satisfactory results are obtained when the compounds of
the
-16-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
present invention are administered at a daily dosage of from about 0.1
milligram to
about 100 milligram per kilogram of animal body weight, preferably given as a
single
daily dose or in divided doses two to six times a day, or in sustained release
form. For
most large mammals, the total daily dosage is from about 1.0 milligrams to
about
1000 milligrams, preferably from about 1 milligrams to about 50 milligrams. In
the
case of a 70 kg adult human, the total daily dose will generally be from about
7
milligrams to about 350 milligrams. This dosage regimen may be adjusted to
provide
the optimal therapeutic response.
Pharmaceutical Compositions
Another aspect of the present invention provides pharmaceutical
compositions which comprise a compound of Formula I and a pharmaceutically
acceptable carrier. The pharmaceutical compositions of the present invention
comprise a compound of Formula I or a pharmaceutically acceptable salt or
prodrug
thereof as an active ingredient, as well as a pharmaceutically acceptable
carrier and
optionally other therapeutic ingredients. The term "pharmaceutically
acceptable salts"
refers to salts prepared from pharmaceutically acceptable non-toxic bases or
acids
including inorganic bases or acids and organic bases or acids.
The compositions include compositions suitable for oral, rectal,
topical, parenteral (including subcutaneous, intramuscular, and intravenous),
ocular
(ophthalmic), pulmonary (nasal or buccal inhalation), or nasal administration,
although the most suitable route in any given case will depend on the nature
and
severity of the conditions being treated and on the nature of the active
ingredient.
They may be conveniently presented in unit dosage form and prepared by any of
the
methods well-known in the art of pharmacy.
In practical use, the compounds of Formula I can be combined as the
active ingredient in intimate admixture with a pharmaceutical carrier
according to
conventional pharmaceutical compounding techniques. The carrier may take a
wide
variety of forms depending on the form of preparation desired for
administration, e.g.,
oral or parenteral (including intravenous). In preparing the compositions for
oral
dosage form, any of the usual pharmaceutical media may be employed, such as,
for
example, water, glycols, oils, alcohols, flavoring agents, preservatives,
coloring agents
and the like in the case of oral liquid preparations, such as, for example,
suspensions,
elixirs and solutions; or carriers such as starches, sugars, microcrystalline
cellulose,
diluents, granulating agents, lubricants, binders, disintegrating agents and
the like in
-17-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
the case of oral solid preparations such as, for example, powders, hard and
soft
capsules and tablets, with the solid oral preparations being preferred over
the liquid
preparations.
Because of their ease of administration, tablets and capsules represent
the most advantageous oral dosage unit form in which case solid pharmaceutical
carriers are obviously employed. If desired, tablets may be coated by standard
aqueous or nonaqueous techniques. Such compositions and preparations should
contain at least 0.1 percent of active compound. The percentage of active
compound
in these compositions may, of course, be varied and may conveniently be
between
about 2 percent to about 60 percent of the weight of the unit. The amount of
active
compound in such therapeutically useful compositions is such that an effective
dosage
will be obtained. The active compounds can also be administered intranasally
as, for
example, liquid drops or spray.
The tablets, pills, capsules, and the like may also contain a binder such
as gum tragacanth, acacia, corn starch or gelatin; excipients such as
dicalcium
phosphate; a disintegrating agent such as corn starch, potato starch, alginic
acid; a
lubricant such as magnesium stearate; and a sweetening agent such as sucrose,
lactose
or saccharin. When a dosage unit form is a capsule, it may contain, in
addition to
materials of the above type, a liquid carrier such as a fatty oil.
Various other materials may be present as coatings or to modify the
physical form of the dosage unit. For instance, tablets may be coated with
shellac,
sugar or both. A syrup or elixir may contain, in addition to the active
ingredient,
sucrose as a sweetening agent, methyl and propylparabens as preservatives, a
dye and
a flavoring such as cherry or orange flavor.
Compounds of formula I may also be administered parenterally.
Solutions or suspensions of these active compounds can be prepared in water
suitably
mixed with a surfactant such as hydroxy-propylcellulose. Dispersions can also
be
prepared in glycerol, liquid polyethylene glycols and mixtures thereof in
oils. Under
ordinary conditions of storage and use, these preparations contain a
preservative to
prevent the growth of microorganisms.
The pharmaceutical forms suitable for injectable use include sterile
aqueous solutions or dispersions and sterile powders for the extemporaneous
preparation of sterile injectable solutions or dispersions. In all cases, the
form must
be sterile and must be fluid to the extent that easy syringability exists. It
must be
stable under the conditions of manufacture and storage and must be preserved
against
-18-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
the contaminating action of microorganisms such as bacteria and fungi. The
carrier
can be a solvent or dispersion medium containing, for example, water, ethanol,
polyol
(e.g. glycerol, propylene glycol and liquid polyethylene glycol), suitable
mixtures
thereof, and vegetable oils.
Combination Therapy
Compounds of Formula I may be used in combination with other drugs
that may also be useful in the treatment, prevention, suppression or
amelioration of
the diseases or conditions for which compounds of Formula I are useful. Such
other
drugs may be administered, by a route and in an amount commonly used therefor,
contemporaneously or sequentially with a compound of Formula I. When a
compound of Formula I is used contemporaneously with one or more other drugs,
a
pharmaceutical composition in unit dosage form containing such other drugs and
the
compound of Formula I is preferred. However, the combination therapy also
includes
therapies in which the compound of Formula I and one or more other drugs are
administered on different overlapping schedules. It is also contemplated that
when
used in combination with one or more other active ingredients, the compound of
the
present invention and the other active ingredients may be used in Iower doses
than
when each is used singly. Accordingly, the pharmaceutical compositions of the
present invention include those that contain one or more other active
ingredients, in
addition to a compound of Formula I.
Examples of other active ingredients that may be administered in
combination with a compound of Formula I, and either administered separately
or in
the same pharmaceutical composition, include, but are not limited to:
(a) insulin sensitizers including (i) PPAR~y agonists such as the
glitazones (e.g. troglitazone, pioglitazone, englitazone, MCC-555,
rosiglitazone, and
the like), and compounds disclosed in W097/27857, 97/28115, 97!28137 and
97/27847; (ii) biguanides such as metformin and phenformin; (iii) protein
tyrosine
phosphatase-1B (PTP-1B) inhibitors, and (iv) dipeptidyl peptidase IV (DP-IV)
inhibitors;
(b) insulin or insulin mimetics;
(c) sulfonylureas such as tolbutamide and glipizide, or related
materials;
(d) a-glucosidase inhibitors (such as acarbose);
-19-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
(e) cholesterol lowering agents such as (i) HMG-CoA reductase
inhibitors (lovastatin, simvastatin, pravastatin, fluvastatin, atorvastatin,
rivastatin,
itavastatin, ZD-4522 and other statins), (ii) sequestrants (cholestyramine,
colestipol,
and diallcylaminoalkyl derivatives of a cross-linked dextran), (iii) nicotinyl
alcohol,
nicotinic acid or a salt thereof, (iv) PPARa agonists such as fenofibric acid
derivatives (gemfibrozil, clofibrate, fenofibrate and benzafibrate), (v)
PPARaJy dual agonists, such as KRP-297, (vi) inhibitors of cholesterol
absorption,
such as for example beta-sitosterol, (vii) acyl CoA:cholesterol
acyltransferase
inhibitors, such as for example avasimibe, and (viii) anti-oxidants, such as
probucol;
(f) PPARB agonists such as those disclosed in W097/28149;
(g) antiobesity compounds such as fenfluramine, dexfenfluramine,
phentiramine, sulbitramine, orlistat, neuropeptide Y5 inhibitors, and (33
adrenergic
receptor agomsts;
(h) an ileal bile acid transporter inhibitor; and
(i) agents intended for use in inflammatory conditions such as aspirin,
non-steroidal anti-inflammatory drugs, glucocorticoids, azulfidine, and cyclo
oxygenase 2 selective inhibitors.
The above combinations include combinations of a compound of the
present invention not only with one other active compound, but also with two
or
more other active compounds. Non-limiting examples include combinations of
compounds having Formula I with two or more active compounds selected from
biguanides, sulfonylureas, HMG-CoA reductase inhibitors, other PPAR agonists,
PTP-1B inhibitors, DP-IV inhibitors, and anti-obesity compounds.
BIOLOGICAL ASSAYS
A) PPAR Binding Assays
For preparation of recombinant human PPAR~y, PPARB, and
PPARa: Human PPAR~yz, human PPARB and human PPARa were expressed as gst-
fusion proteins in E. coli. The full length human cDNA for PPAR~y2 was
subcloned
into the pGEX-2T expression vector (Pharmacia). The full length human cDNAs
for
PPAR~ and PPARoc were subcloned into the pGEX-KT expression vector
(Pharmacia). E. coli containing the respective plasmids were propagated,
induced, and
harvested by centrifugation. The resuspended pellet was broken in a French
press and
-20-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
debris was removed by centrifugation at 12,000 X g. Recombinant human PPAR
receptors were purified by affinity chromatography on glutathione sepharose.
After
application to the column, and one wash, receptor was eluted with glutathione.
Glycerol (10%) was added to stabilize the receptor and aliquots were stored at
-80°C.
For binding to PPAR~y, an aliquot of receptor was incubated in TEGM (10 mM
Tris,
pH 7.2, 1 mM EDTA, 10% glycerol, 7 ~,L1100 mL 13-mercaptoethanol, 10 mM
Na molybdate, 1 mM dithiothreitol, 5 ~.g/mL aprotinin, 2 ~,g/mL leupeptin, 2
~,g/mL benzamidine and 0.5 mM PMSF) containing 0.1% non-fat dry milk and
nM [3H2] AD5075, (21 Ci/mmole), ~ test compound as described in
10 Berger et al (Novel peroxisome proliferator-activated receptor (PPAR~y) and
PPARB ligands produce distinct biological effects. J. Biol. Chem. (1999),
274: 6718-6725. Assays were incubated for ~16 hr at 4°C in a final
volume of
150 ~,L. Unbound ligand was removed by incubation with 100 ~,L
dextran/gelatin-coated charcoal, on ice, for ~10 min. After centrifugation at
3000 rpm for 10 min at 4°C, 50 p,L of the supernatant fraction was
counted in
a Topcount.
For binding to PPARB, an aliquot of receptor was incubated in TEGM
(10 mM Tris, pH 7.2, 1 mM EDTA, 10% glycerol, 7 ,uL/100 mL f3-mercaptoethanol,
10 mM Na molybdate, 1 mM dithiothreitol, 5 ~.g/mL aprotinin, 2 ~,g/mL
leupeptin, 2
~,g/mL benzamide and 0.5 mM PMSF) containing 0.1% non-fat dry mills and 2.5 nM
[3H2]-Comp'd A, (17 Ci/mrnole), ~ test compound as described in Berger et al
(Novel peroxisome proliferator-activated receptory (PPAR~y) and PPARB ligands
produce distinct biological effects.1999 J Biol Chem 274: 6718-6725). (Comp'd
A is
3-chloro-4-(3-(7-propyl-3-trifluoromethyl-6-benz-[4,5]-
isoxazoloxy)propylthio)phenylacetic acid, Ex. 20 in WO 97/28137). Assays were
incubated for ~16 hr at 4°C in a final volume of 150 pL. Unbound ligand
was
removed by incubation with 100 ~,L dextran/gelatin-coated charcoal, on ice,
for ~10
min. After centrifugation at 3000 rpm for 10 min at 4°C, 50 ~,L of the
supernatant
fraction was counted in a Topcount.
-21-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
For binding to PPARoc, an aliquot of receptor was incubated in TEGM
(10 mM Tris, pH 7.2, 1 mM EDTA, 10% glycerol, 7 ~,L/100 mL 13-mercaptoethanol,
mM Na molybdate, 1 mM dithiothreitol, 5 ~,glmL aprotinin, 2 p,g/mL leupeptin,
2
~,g/mL benzamide and 0.5 mM PMSF) containing 0.1 % non-fat dry milk and 5.0 nM
5 [3H2]-Comp'd B, (34 Ci/mmole), ~ test compound. (Comp'd B is (3-(4-(3-phenyl-
7-
propyl-6-benz-[4,5]-isoxazoloxy)butyloxy))phenylacetic acid, Ex.62 in WO
97/28137). Assays were incubated for ~16 hr at 4°C in a final volume of
150 ~,L.
Unbound ligand was removed by incubation with 100 p,L dextran/gelatin-coated
charcoal, on ice, for ~10 min. After centrifugation at 3000 rpm for 10 min at
4°C,
10 50 ~.L of the supernatant fraction was counted in a Topcount.
B). Gal-4 hPPAR Transactivation Assays
The chimeric receptor expression constructs, pcDNA3-hPPAR~/GALA~,
pcDNA3-hPPARB/GAI~I-, pcDNA3-hPPARaJGAI~I. were prepared by inserting the
yeast GAL4 transcription factor DBD adjacent to the ligand binding domains
(LBDs)
of hPPAR~y, hPPARB, hPPARa, respectively. The reporter construct, pUAS(5X)-tIc-
luc was generated by inserting 5 copies of the GAL4 response element upstream
of the
herpes virus minimal thymidine kinase promoter and the Iuciferase reporter
gene.
pCMV-lacZ contains the galactosidase Z gene under the regulation of the
cytomegalovirus promoter. COS-1 cells were seeded at 12 X 103 cells/well in 96
well
cell culture plates in high glucose Dulbecco's modified Eagle medium (DMEM)
containing 10% charcoal stripped fetal calf serum (Gemini Bio-Products,
Calabasas,
CA), nonessential amino acids, 100 units/ml Penicillin G and 100 mg/ml
Streptomycin sulfate at 37 °C in a humidified atmosphere of 10% C02.
After 24 h,
transfections were performed with Lipofectamine (GIBCO BRL, Gaithersburg, MD)
according to the instructions of the manufacturer. Briefly, transfection mixes
for each
well contained 0.48 ~ul of Lipofectamine, 0.00075 ~,g of pcDNA3-PPAR/GAL4
expression vector, 0.045 ~.g of pUAS(5X)-tk-luc reporter vector and 0.0002 ~.g
of
pCMV-lacZ as an internal control for transactivation efficiency. Cells were
incubated
-22-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
in the transfection mixture for 5 h at 37° C in an atmosphere of 10%
C02. The cells
were then incubated for ~48 h in fresh high glucose DMEM containing 5%
charcoal
stripped fetal calf serum, nonessential amino acids, 100 units/ml Penicillin G
and 100
mg/ml Streptomycin sulfate ~ increasing concentrations of test compound. Since
the
compounds were solubilized in DMSO, control cells were incubated with
equivalent
concentrations of DMSO; final DMSO concentrations wexe < 0.1%, a concentration
which was shown not to effect transactivation activity. Cell lysates were
produced
using Reporter Lysis Buffer (Promega, Madison, WI) according to the
manufacturer's
instructions. Luciferase activity in cell extracts was determined using
Luciferase
Assay Buffer (Promega, Madison, WI) in~an ML3000 luminometer (Dynatech
Laboratories, Chantilly, VA). (3-galactosidase activity was determined using
(3-D-
galactopyranoside (Calbiochem, San Diego, CA).
C. Ifi Vivo Studies
Male db/db mice (10-11 week old C57B1/KFJ, Jackson Labs, Bar
Harbor, ME) were housed 5/cage and allowed ad lib. access to ground Purina
rodent
chow and water. The animals, and their food, were weighed every 2 days and
were
dosed daily by gavage with vehicle (0.5% carboxymethylcellulose) ~ test
compound
at the indicated dose. Drug suspensions were prepared daily. Plasma glucose,
and
triglyceride concentrations were determined from blood obtained by tail bleeds
at 3-5
day intervals during the study period. Glucose, and triglyceride,
determinations were
performed on a Boehringer Mannheim Hitachi 911 automatic analyzer (Boehringer
Mannheim, Indianapolis, IN) using heparinized plasma diluted 1:6 (v/v) with
normal
saline. Lean animals were age-matched heterozygous mice maintained in the same
manner.
EXAMPLES
-23-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
The following Examples are provided only to illustrate the invention,
including methods of making the compounds of the invention, and are not to be
construed as limiting the invention in any manner.
INTERMEDIATE 1
Step 1. Preparation of 1,3-diall.Yloxybenzene:
OH ~Br
K2C03, DMF
OH
To a solution of resorcinol (1,200 g, 10.9 mmol) in DMF (10.89 L) was added
K2C03 (4,463 g). Allyl bromide (3,811 mL) was added slowly (keeping the
temperature below 30 °C). The reaction mixture was stirred at ambient
temperature
overnight, poured into water (94 L), extracted with Et20 (3 x 20L). Combined
organic
layers were washed with water (3 x 15 L) and brine (10 L), dried over MgS04,
filtered through Na2S04, and evaporated in vacuo. The residue was pumped dry
on
high vacuum to give the crude product as a red/yellow oil, which was used in
the next
step without further purification. 1H NMR (CDC13, 400 MHz) 8 4.5 (d, 4H), 5.27
(m,
2H), 5.42 (m, 2H), 6.05 (m, 2H), 6.5 (m, 3H), 7.16 (m, 1H).
Step 2. Preparation of 2,4-diallylresorcinol:
O 187 °C I ~ OH
CI ~ /~ ~ u\
O ~ / OH
CI
-24-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
The crude starting material (2,278 g) was dissolved in 1,2-dichlorobenzene
(11.4 L) in a 22 L 4-neck flask equipped with a mechanical stirrer, a
thermocouple, a
distillation condenser and a nitrogen inlet. A portion of the solvent (1.7 L)
was
distilled off at 187 °C before the distillation condenser was switched
to a reflux
condenser. The reaction was refluxed at 187 °C overnight. Ice-water (4
L) was
added, followed by NaOH (320 g). The mixture was poured into hexanes (12 L)
and
layers were separated. The organic layer was extracted with aqueous NaOH (2 N,
2 x
4 L). The combined aqueous layers were acidified with concentrated HCl (ice
was
added to maintain the temperature below 30 °C), then extracted with
Et20 (3 x 4 L).
The combined organic layers were dried over MgS04, filtered through Na2S04,
and
evaporated in vacuo. Purification by chromatography (12 kg silica gel paclced
in
hexanes, eluted with 10% EtOAc/hexanes) to give the desired product. 1H NMR
(CDC13, 400 MHz) 8 3.31 (m, 2H), 3.48 (m, 2H), 5.2 (m, 4H), 6.0 (m, 2H), 6.40
(d,
1H), 6.85 (d, 1H).
Step 3. Preparation of 2,4-dipropylresorcinol:
OH H2, 10% PdIC I ~ OH
EtOAc /
OH OH
Starting material (1237.2 g, 6.5 mmol) was split into two runs. To each
solution of the bis-allyl starting material (618.6 g) in ethyl acetate (2,780
mL) was
added 10% Pd/C (46 g). Hydrogenation was carried out at rt under 40 psi
hydrogen
atmosphere for 1.5 h. The reaction was filtered through super cell, and the
solvent was
evaporated in vacuo. The combined crude products from the two runs were
slurried in
hexanes, filtered, and dried to give the product. 1H NMR (CDC13, 400 MHz) S
1.00
(m, overlapping signals, 6H), 1.6 (m, overlapping signals, 4H), 2.48 (t, 2H),
2.60 (t,
2H), 4.56 (s, 1H), 4.70 (s, 1H), 6.31 (d, 1H), 6.80 (d, 1H).
Step 4. Preparation of 2,4-dihydrox -~~3,5dipropyl-1',l',l'-
trifluoroacetonephenone:
-25-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
F3C O
OH
(CF3C0)20 I ~ OH
AICI3, CH2CI2 /
OH
OH
To a solution of the starting material (785.7 g, 4.04 mmol) in CH2C12 (20,600
mL) was added AlCl3 (1,763 g). Trifluoroacetic anhydride (814 mL) was added
slowly at 0 °C. The reaction mixture was stirred overnight at rt,
poured into ice-water,
extracted with CH2Cl2 (3 x). The combined organic layers were washed with sat.
NaHC03 and brine, dried over MgS04, filtered through Na2S04, and evaporated in
vacuo. Purification by chromatography (10 kg silica gel, packed in hexanes,
eluted
with 5% EtOAclhexanes) to give the desired product as a yellow solid. 1H NMR
(CDCl3, 400 MHz) & 1.00 (m, overlapping signals, 6H), 1.60 (m, overlapping
signals,
4H), 2.55 (t, J = 7.4 Hz, 2H), 2.66 (t, J = 7.4 Hz, 2H), 5.65 (s, 1H), 7.45
(s, 1H); MS
(ESI) 291 (M +1).
Step 5. Preparation of 2,4-dih day-3,5-dipropyl-1',1',l'-
trifluoroacetonephenone
oxime:
NH20H.HC1
NaOAc, MeOH
To a mixture of NaOAc (2,677 g, 32.5 mol) and hydroxyamine hydrochloride
(2,000 g, 28.8 mol) in methanol (1 L) was added a solution of the starting
phenol
(1,139.8 g, 3.93 mol) in methanol (26 L). The yellow suspension was refluxed
for 18
h. TLC showed significant amount of starting material remained. Additional
hydroxyamine hydrochloride (1,000 g), NaOAc (1,338 g) and methanol (4 L) were
added. The reaction mixture was refluxed overnight. TLC indicated the complete
consumption of the starting material. The reaction was poured into ice-water
(32 L),
extracted with EtOAc (2 x 16 L). The combined organic layers were washed with
brine, dried, and evaporated in vacuo. Chromatography (10 lcg silica gel, 15%
-26-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
EtOAc/hexanes) gave the desired product as yellow solid. 1H NMR (CDCl3, 400
MHz) 8 0.98 (m, overlapping signals, 6H), 1.60 (m, overlapping signals, 4H),
2.50 (t,
2H), 2.68 (t, 2H), 5.00 (s, broad, 1H), 5.80 (s, broad, 1H), 6.92 (s, 1H).
Step 6. Preparation of 5,7-dipropyl-6-h day-3-trifluoromethyl-1,2-
benzisoxazole:
F3C NOH F3C .-N
1 ) Ac20 p
Ohi 2) pyridine, Et3N
/ /
OH OH
A solution of the starting oxime (600 g) in Ac20 (3 L) was stirred at rt
overnight. The solvent was removed in vacuo. The residue was coevaporated with
toluene (4 x ) to give the crude 2,4-dihydroxy-3,5-dipropyl-1',1',1'-
trifluoroacetonephenone O-acetyl oxime. This crude product was dissolved in
pyridine (6 L) and Et3N (684 mL). The reaction was refluxed (112 °C)
for 3 h, and
allowed to cool overnight. The solvent was evaporated in vacuo. The residue
was
coevaporated with toluene (2 x), then partitioned between EtOAc and 1 N HCl.
The
organic layer was washed with 1 N HCl and brine, dried, filtered, and
concentrated in
vacuo to give a black oil. Purification by flash chromatography (10 kg silica
gel, 5%
EtOAc/hexanes) gave the desired product. 1H NMR (CDCl3, 400 MHz) 8 1.00 (m,
overlapping signals, 6H), 1.70 (m, overlapping signals, 4H), 2.68 (t, 2H),
2.90 (t, 2H),
5.21 (s, 1H), 7.33 (s, 1H); MS (ESI) 288.3 (M +1).
INTERMEDIATE 2
-27-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
Similarly prepared as Intermediate 1 using 6-propylresorcinol. 1H
NMR (CDC13, 400 MHz) 8 1.03 (t, 3H), 1.71 (m, 2H), 2.72 (t, 2H), 5.5 (s,
broad,
1H), 7.06 (s, 1H), 7.51 (s, 1H).
INTERMEDIATE 3
To a solution of Intermediate 2 (0.22 g, 0.89 mmol) and sulfuryl chloride
(0.096 mL, 1.2 mmol) in CH2C12 (5 mL)was adde Et20 (0.5 mL). The reaction was
stirred at room temperature overnight, partitioned between water and Et20. The
organic layer was washed with saturated NaHC03 aqueous solution and brine,
dried
over MgS04, filtered and concentrated in vacuo to give 7-chloro-6-hydroxy-5-
propyl-
3-trifluoromethyl-1,2-benzisoxazole. 1H NMR (CDC13, 400 MHz) 8 1.01 (t, 3H),
1.71 (m, 2H), 2.77 (t, 2H), 6.15 (s, 1H), 7.44 (s, 1H).
EXAMPLE 1
F3C .N
O
O
home
/ ~(O
Preparation of methyl 2-[(5,7-dipropyl-3-trifluoromethyl-1,2-benzisoxazol-6-
yl)oxyl-
2-methylpropionate:
-28-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
Cs2C03, DMF
Br
~C02Me
To a solution of starting phenol (20 g, 69.7 mmol) in DMF (200 mL) were
added methyl a-bromoisobutyrate (126.7 g, 0.7 mol) and cesium carbonate (228g,
0.7
mol) and the mixture was stirred at 60 °C for seven days. Reaction was
worl~ed up by
partitioning between ether and water. The aqueous phase was extracted with
ether and
the organic phase was washed with water, then brine, dried over Mg2SOq.,
filtered,
and evaporated in vacuo. Purification by chromatography gave the desired
product.1H
NMR (CDC13): 8 1.00 (m, 6H), 1.53 (s, 6H), 1.69 (m, 2H), 1.76 (m, 2H), 2.62
(t, 2H),
2.85 (m, 2H), 3.89 (s, 3H), 7.39 (s, 1H).
EXAMPLE 2
Preparation of 2-f (5,7-dipropyl-3-trifluoromethyl-1,2-benzisoxazol-6-yl)oxyl-
2-
meth, r~opionic acid:
FsC _N F3C -N
O O
NaOH
O O
home ~OH
~O( / ~O
To a solution of the starting methyl ester (8 g, 20.7 mmol) in methanol (50
mL) was added aqueous sodium hydroxide (1.0 N, 60 mL). Enough THF (100 mL)
-29-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
was added to bring the mixture back to a clear solution. The mixture was
heated at 80
°C for 2 h. TLC showed that reaction was complete. The reaction was
partitioned
between 1N HCl and ether. The organic phase was washed with water and brine,
dried over magnesium sulfate, filtered, and evaporated in vacuo. The solid
residue
was recrystallized from pentane to afford the desired product as colorless
crystals.1H
NMR (CDCl3): 8 1.01 (m, 6H), 1.59 (s, 6H), 1.70 (m, 2H), 1.78 (m, 2H), 2.68
(t, 2H),
2.92 (m, 2H), 7.43 (s, 1H).
EXAMPLE 3
Similarly prepared as Example 2 using Intermediate 3. 1H NMR (CDC13, 400
MHz) S 1.03 (t, 3H), 1.68 (s, 6H), 1.72 (m, 2H), 2.75 (t, ZH), 7.52 (s, 1H).
EXAMPLE 4
F3C ._N
v
O
O
~O Et
~(O
Preparation of ethyl 2-f (5,7-dipropyl-3-trifluoromethyl-1,2-benzisoxazol-6-
oxylbut r
-30-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
Cs2C03, DMF
Br
~C02Et
To a solution of Intermediate 1 (150 mg, 0.52 mmol) and ethyl 2-
bromobutyrate (152 mg, 0.78 mmol) in DMF (5 mL) was added cesium carbonate
(254 mg, 0.78 mmol). The mixture was stirred at room temperature for 6 h, then
partitioned between ether and water. The organic phase was washed with water
and
brine, dried over magnesium sulfate, filtered, and concentrated in vacuo.
Purification
by flash chromatography gave the desired product. 1H NMR(CDCl3): 81.01 (m,
6H),
1.08 (t, 3H), 1.24 (t, 3H), 1.69 (m, 2H), 1.78 (m, 2H), 2.04 (m, 2H), 2.72 (m,
1H),
2.82 (m, 1H), 2.97 (m, 2H), 4.19 (m, 2H), 4.54 (m, 1H), 7.41 (s, 1H).
EXAMPLE 5
F3C _.N
O
O
OH
O
Step 1. Preparation of ethyl 2-f (5,7-dipropyl-3-trifluoromethyl-1,2-
benzisoxazol-6-
ox.1-~ylvalerate:
-31-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
F3C _N F3C .-N
O O
LDA, HMPA
O
O
~OEt OEt
~O O
To a solution of the title compound of Example 5 (80 mg) in THF (5 mL) at -
78 °C was added LDA (1.5 M, 0.27 mL) followed by HMPA (0.069 mL). After
5 min
at -78 °C, iodopropane (98 ~.L) was introduced. The mixture was stirred
at -78 °C for
1 h, then allowed to slowly warm to room temperature over 2 h. Aqueous worlcup
and
purification by chromatography gave the desired product as a colorless oil. 1H
NMR(CDC13): 8 0.87 (t, 3H), 0.95 (t, 3H), 1.01 (m, overlapping signals, 6H),
1.25 (m,
1), 1.50 - 1.80 (m, overlapping signals, 5H), 1.82 - 2.10 (m, overlapping
signals, 4H),
2.68 (m, 2H), 2.91 (m, 2H), 4.18 (q, 2H), 7.38 (s, 1H).
Step 2. Preparation of 2-f (5,7-dipropyl-3-trifluoromethyl-1,2-benzisoxazol-6-
, l
2-ethylvaleric acid:
t-BuOK
To a solution of the ethyl ester (30 mg) in DMSO (5 mL) at room temperature
was added tBuOK (0.5 g). The reaction was stirred at room temperature
overnight,
partitioned between 1.0 N aqueous HCl and EtOAc. The organic phase was washed
with water and brine, dried over magnesium sulfate, filtered, and concentrated
in
vacuo. Purification by preparative HPLC gave the desired product. 1H NMR
(CDC13)
-32-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
8 0.87 (t, 3H), 0.95 (t, 3H), 1.01 (m, 6H), 1.25 (m, 1H), 1.56 - 1.77 (m, 5H),
1.82 -
2.10 (m, 4H), 2.69 (m, 2H), 2.91 (m, 2H), 7.42 (s, 1H).
EXAMPLE 6
F3C .-N
O
O O
~O~N~
IO H
Preparation of 2-(acetylamino)ethyl 2-f(5,7-dipropyl-3-trifluorometh 1-y 1,2-
benzisoxazol-6-yl)oxyl-2-methylpropionate:
F3C -N F3C .-N
O
O HO~N~ I \ O
H /
O DCC, DMAP O O
~OH ~O~N~
' ~O'( ''
To a solution of the title compound of Example 1 (500 mg, 1.34 mmol)
in methylene chloride (10 mL) were added N-2-hydroxyethylacetamide (166 mg,
1.50
mmol), DCC (1.5 mL, 1N) and DMAP (16 mg, 0.13 mmol). The reaction was stirred
at room temperature overnight. Precipitates were filtered off. Filtrate and
washings
were combined and evaporated in vacuo. Purification by flash chromatography
gave
the desired product. 1H NMR (CDC13) 8 1.00 (m, 6H), 1.54 (s, 6H), 1.59 - 1.81
(m,
4H), 2.02 (s, 3H), 2.62 (t, 2H), 2.87 (m, 2H), 3.65 (m, 2H), 4.34 (t, 3H),
5.79 (s,
broad, 1H), 7.40 (s, 1H).
EXAMPLE 7
-33-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
Preparation of 2-f (5,7-dipropyl-3-trifluoromethyl-1,2-benzisoxazol-6-yl)ox
meths r~opionamide:
NH4CI/AICI3
To a suspension of NH4Cl (86 mg) in toluene (10 mL) at room
temperature was added A1C13 (2.0 M, 0.8 mL) dropwise. The reaction was stirred
for
3 h. The resultant clear solution was transferred to a solution of methyl 2-
[(5,7-
dipropyl-3-trifluorornethyl-1,2-benzisoxazol-6-yl)oxy]-2-methylpropionate (200
mg)
in toluene (5 mL). The reaction was stirred at 80 °C overnight, cooled
to room
temperature, and then partitioned between EtOAc and 1.0 N aqueous HCl
solution.
The organic phase was washed with water and brine, dried over magnesium
sulfate,
filtered, and concentrated in vacuo. Purification by chromatography gave the
desired
product. 1H NMR (CDC13) 8 1.00 (m, 6H), 1.51 (s, 6H), 1.65 (m, ZH), 1.75 (m,
2H),
2.70 (t, 2H), 2.94 (m, 2H), 5.72 (s, broad, 1H), 6.87 (s, broad, 1H), 7.43 (s,
1H).
EXAMPLE 8
-34-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
Step 1. Preparation of methyl f (5,7-dipropyl-3-trifluoromethyl-1,2-
benzisoxazol-6-
yl)oxylacetate:
F3C .-N
\ O Cs2C03, DMF
w gr~C02Me
OH
To a solution of starting phenol (0.5 g) in DMF (10 mL) were added methyl
bromoacetate (0.4) and cesium carbonate (0.85) and the mixture was stirred at
rt for 4
h. Reaction was worked up by partitioning between EtOAc and water. The organic
phase was washed with water, then brine, dried over Mg2S04, filtered, and
evaporated in vacuo. Purification by chromatography gave the desired
product.1H
NMR (CDC13): 8 1.02 (m, 6H), 1.72 (m, 2H), 1.80 (m, 2H), 2.74 (t, 2H), 2.97
(m,
2H), 3.89 (s, 3H), 4.51 (s, 2H), 7.45 (s, 1H).
Step 2. Preparation of methyl 1-f(5,7-dipropyl-3-trifluoromethyl-1,2-
benzisoxazol-6-
yl)ox~c clopentanecarbox.l~ate:
F3C -N F3C .-N
O O
LiHMDS, HMPA I \
O iii O
home home
O <~I ~O
-35-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
To a solution of the starting ester (100 mg) in THF (4.0 mL) at - 78
°C
were added LiHMDS (1.0 M, 0.31 mL) and HMPA (0.054 mL). After 15 min at -78
°C, 1,4-diiodobutane (96 mg) was introduced. The reaction was allowed
to warm to rt
over 3 h, then cooled to -78 °C before another 2.1 equiv of LiHMDS and
HMPA
were added. The reaction was allowed to slowly warm to rt and stirred
overnight. The
mixture was then partitioned between EtOAc and water. The organic phase was
washed with water and brine, dried over magnesium sulfate, filtered, and
concentrated in vacuo. Purification by chromatography gave the desired
product. 1H
NMR (CDCl3) 8 0.99 (m, 6H), 1.72 (rn, 8H), 2.08 (m, 2H), 2.32 (m, 2H), 2.63
(t,
2H), 2.84 (t, 2H), 3.81 (s, 3H), 7.39 (s, 1H).
Step 3. Preparation of 1-~(5,7-dipropyl-3-trifluoromethyl-1,2-benzisoxazol-6-
l~ylcyclopentanecarboxylic acid:
NaOH
home
To a solution of the starting methyl ester (30 mg) in methanol (2 mL) was
added
aqueous sodium hydroxide (1.0 N, 2 mL). Enough THF (6 mL) was added to bring
the mixture back to a clear solution. The mixture was heated at 60 °C
for 5 h, then
partitioned between 1N HCl and ether. The organic phase was washed with water
and
brine, dried over magnesium sulfate, filtered, and evaporated in vacuo.
Purification
by chromatography gave the desired product.1H NMR (CDC13) 8 0.99 (m, 6H), 1.72
(m, 8H), 2.10 (m, 2H), 2.35 (m, 2H), 2.62 (t, 2H), 2.84 (t, 2H), 7.41 (s, 1H).
The examples listed in Table I below were prepared using the same or similar
protocols as described for the examples (1 - 8) listed above.
-36-
CA 02400021 2002-08-13
WO 01/60807 PCT/USO1/04636
TABLE I.
R5 _N
O
R3 R4
O
R1~OH
I ~'(2
O
Exam Rl R2 R3 R4 R5 Partial'H-Nmr Data (8, ppm,
1e CDCI3)
9 methylmethylCl ro trifluoromethyl1.67 (s,6H), 7.70 (s,lH)
y1
ethyl H ro ro trifuoromethyl4.60 (m,lH),7.44 (s,lH)
y1 y1
11 ethyl methylro ro trifuoromethyl1.15 (t, 3H), 1.28 (s, 3H),
1 y1 7.43 (s,lH)
12 ropyl methylro ro trifluorometh1.28 (s, 3H),1.47 (m, 1H),
y1 y1 1 7.43 (s, 1H)
13 ro H ro ro trifluoromethyl4.G4 (t, 1H), 7.43 (s, 1H)
y1 y1 y1
14 ro ro ro ro trifluoromethyl0.85 (t, 6H), 1.23 (m, 2H),
y1 y1 y1 y1 7.42 (s, 1H)
ethyl eth ro ro trifluoromethyl1.56-1.86 (m, 4H), 7.41
1 y1 y1 (s,lH)
16 methylH Cl ro ethyl 1.45 (t, 3H), 1.64 (d, 3H),
y1 4.95 ( , 1H), 7.55 (s,
1H)
17 methylmethylCI ro ethyl 1.45 (t, 3H), 2.97 ( , 3H),
1 7.53 (s, 1H)
18 meth H Cl ro trifluoromethyl1.67(d, 3H), 5.03 ( , 1H),
1 y1 7.72 (s, 1H)
19 methylmeth ro ro ethyl 1.45 (t, 3H), 3.00 ( , 2H),
1 y1 y1 7.29 (s, 1H)
methylmethylro ro methyl 2.57 (s, 3H), 7.27 (s, 1H)
y1 y1
21 methylmethylro pro methoxy 4.16 (s, 3H), 7.28 (s, 1H)
y1 y1
22 methylH ro ro trifluoromethyl1.61(d,3H), 4.71( ,1H),
y1 y1 7.46(s,lH)
23 methylH ro ro henyl 1.61 (d, 3H), 4.71 ( , 1H),
1 y1 7.57 (s, 1H)
24 methylmethylro ro phenyl 7.54 (s, 1H), 7.58 (m, 3H),
y1 y1 7.96 (d, 2H)
methylethylro ropylhenyl 1.16 (t, 3H), 1.30 (s, 3H),
y1 7.54 (s, 1H)
26 H H ro ro trifluorometh4.58(s,2H), 7.47(s,lH)
y1 y1 1
27 4- H propylpropyltrifuoromethyl4.62 (m, 1H), 5.03(m, 2H),
penten- 5.80 (m, 1H), 7.43(s,lH)
1-yl
28 cyclobut ro ro trifuoromethyl2.37 (m, 2H), 2.58 (m, 2H),
lidene y1 y1 7.50 (s,lH)
29 cyclohexylidene propylpropyltrifuoromethyl1.20 (m, 1H), 2.30 (d,
broad,
. I 2H), 7.40 (s,1H)
-37-
