Title of Invention

NOVEL 1,2,4-THIADIAZOLE DERIVATIVES AS MELANOCORTIN RECEPTOR MODULATORS

Abstract The present invention is directed to novel 1,2,4- thiadiazole derivatives of the formula (II) wherein R1, R2- and R4, are ring containing radicals, useful as agonists or antagonists of the melanocortin receptor. More particularly, the compounds of the present invention are useful for the treatment of metabolic CNS and dermatological disorders.
Full Text NOVEL 1,2,4-THIADIAZOLE DERIVATIVES AS MELANOCORTIN RECEPTOR
MODULATORS
CROSS REFERENCE TO RELATED APPLICATION
This application claims the benefit of U. S. Provisional Application
60/337,927, filed on November 08, 2001, which is incorporated by reference
herejn in Its entirety.
FIELD OF THE INVENTION
The present invention provides novel 1,2,4-thiadiazole derivatives useful
for the treatment of a disorder mediated by a melanocortin receptor. More
particularly, the compounds of the present invention are useful for the treatment
of metabolic, CNS and dermatologic disorders such as obesity, impaired oral
glucose tolerance, elevated blood glucose levels, type II diabetes, Syndrome
X, diabetic retinopathy, acute neurodegenerative disorders, chronic
neurodegenerative disorders, plexopathies, male erectile dysfunction, dry eyes,
acne, dry skin, aged skin, seborrheic dermatitis, rosacea, excessive ear wax,
meibomian gland disorder, pseudofolliculitis, yeast infections, dandruff,
hiradenitis suppurativa, ocular rosacea and eccrine gland disorder.
BACKGROUND OF THE INVENTION
Melanocortins are neuropeptides that arise from proopiomelanocortin
(POMC), which is most prevalently expressed in the arcuate nucleus of the
hypothalamus, pituitary lobes, and the nucleus tractus solarius of the
brainstem. [Gantz, I., et al., Molecular Cloning, Expression, and Gene
Localization of a Fourth Melanocortin Receptor, J. Biolog. Chem., 1993, 268,
15174-15179.] These peptides include ACTH, a-MSH, B-MSH, Y1.3-MSH, and
synthetic analogue NDP-aMSH (Wikberg, J E S, Melanocortin receptors: new
opportunities in drug discovery, Exp. Opin. Ther. Patents, 2000,11(1), 61-76).
These peptides bind to five types of melanocortin receptors (MC1-MC5),
which are G-protein coupled receptors that all positively modulate adenylate
cyclase. The MC4 and MC5 receptors are widely distributed in the brain and

spinal cord, whereas the MC3 receptor is located mainly in the hypothalamus.
[Gantz, I., et al., supra.] The MC4 receptor is selectively activated by cxMSH
and can induce neurite outgrowth in Neuro 2A cells. (Adan RAH, et al.,
Molecular Brian Research, 1996,36, pp 37-44; Mountjoy, K.G., Mortud, MX,
Low, M.J., Simerly, R.B. and Cone, R.D., Mol. Endocrinol., 1994, 8, pp 1298-
1308). ACHT is a less potent activator of the MC4 receptor than aMSH.
(Adan, R.A.H., Cone, R.D., Burbach, J.P.H. and Gispen, W.H., mol.
Pharmacol., 1994, 46, pp 1182-1190). The MC5 receptor is activated, in order
of degree, by NDP ˜ a-MSH > ACHT (1-24) > a MSH ACHT (1-39) = ß MSH >>
?MSH (The Melanocortin Receptors, Cone, R.D., Editor, Human Press Inc.,
Totowa. N. J., 2000, Chen, W., pp449-472)
In whole animals, studies in the rat sciatic nerve crush model have
demonstrated that a-MSH increases neurite outgrowth and, as the most potent
of the ACTH derived peptides, it significantly promotes nerve terminal
branching, endplate area, and perimeter. [Bijlsma, W. A., et al., The Enhanced
Recovery of Sensorimotor Function in Rats is Related to the Melantropic
Moiety of ACTH/MSH Neuropeptides, Eur. J. Pharmacol, 1983, 92, 231-236;
Van der Neut. R., et al., Stimulation by Melanocortins of Neurite Outgrowth
from Spinal and Sensory Neurons In Vitro, Peptides, 1992,13,1109-1115; Van
Der Zee, C. E. E. M., et al., a-MSH and Org 2766 in Peripheral Nerve
Regeneration: Different Route of Delivery, Eur. J. Pharmacol., 1988,147, 351-
357; Strand, F. L., et al., Melanocortins as Factors in Somatic Neuromuscular
Growth and Regrowth, Pharmac. Ther., 1994, 62, 1-27], Furthermore,
recovery of motor function after nerve injury is shortened by application of o-
MSH and other melanocortins. [Strand, F. L, et al., supra]
Mice in which the MC4 receptor is rendered inactive by gene targeting
become obese, suggesting that the MC4 receptor is involved in feeding.
[Huszar, D., et al., Targeted Disruption of the Melanocortins Receptor Results
in Mice, Cell, 1997, 88,131-141] This is substantiated by a report that various
MC4 peptide agonists inhibit feeding behavior in agouti mice. [Fan, W., et al.,

Role of Melanocortingenic Neurons in Feeding and the Agouti Obesity
Syndrome, Nature, 1997, 385, 165-168]. a-MSH induces grooming behavior in
rats, but the significance of this is not clear and may not be mediated via the
MC4 receptor. [Adan, R. A. H., et al., Differential Effects of Melanocortin
Peptides on Neural Melanocortin Receptors, Molecular Pharmacology, 1994,
46,1182-1190).
The melanocortins aMSH and ACTH are also known for their ability to
stimulate pigmentation and adrenal glucocorticoid secretion, respectively. The
role of melanocortins, particularly aMSH, in the regulation of sebaceous gland
activity (an exocrine gland with holocrine type of secretion) was shown
originally in rats. More particularly, the studies showed that removal of the
intermediate lobe of the pituitary (which produces the POMC peptides) resulted
in decreased sebaceous lipid production, with complete restoration to normal
levels after replacement therapy with aMSH (Thody, A.J. and Shuster, Nature,
237, 346-347,1972). In a study of rats following total hypophysectomy,
treatment with aMSH resulted in an increase of sebum production, although full
restoration of sebum production was achieved only after treatment with a
combination of aMSH and testosterone (Thody, A.J., Shuster, S.. J. Endocr.
64,503-610,1975; EbUng, F.J., Ebling, E., Randall. V. and Skinner, J., J.
Endocr. 66, 407-412,1975). Knock-out mice where the MC5 receptor was
deleted were observed to display a severe defect in water repulsion and
thermo-regulation, due to decreased production of sebaceous lipids (Chen, W.
Kelly, M.A., Opitz-Araya, X., Thomas, R.E., Low, M.J., and Cone, R., Cell, 91,
788-798,1997).
The MC5 receptor is known to be expressed in human sebaceous
glands, and may be involved in the regulation of human sebaceous lipid
synthesis. Human MC5-R has been cloned and characterized (Chhajlani, V.,
Muceniece, R., Wikberg, JES., Biochem. Biophys. Res. Commun. 195, 866-
873,1993). Moreover, presence of MC5-R m RNA in human sebaceous
glands has been shown by RT-PCR and the protein was detected by

immunohistochemistry and Western blot analysis (Thiboutot, D., Sivarajah,
Gililand, K., Cong, Z. and Clawson, G., J. Invest. Dermatol. 115(4). 614-619,
2000).
Human sebum differs in its composition from other mammals. The main
lipids in human sebum are triglycerides, wax esters and squalene (Greene,
R.S., Downing, D.T., Poci, P.E., Strauss, J.S., J/0 54, 240-247,1970).
Squalene, for instance is not found in many mammals with the exception of
otter and beaver. Triglyceride, which is a major component of human sebum is
poorly represented in other species and in many (e.g. chimpanzee) appears to
be totally absent (Thody, A.J., Shuster, S., Physiolog. Rev. 69, 383-415,1989).
Moreover melanocortins can have different effects on cells from different
species. For example both aMSH (EC50=3.7nM) and ACTH (EC50=16.4nM)
are potent lipolytic agents for rabbit adipocytes, whereas in the rat only ACTH
(EC50=1.34nM) has potent lipolytic activity (Ramachadran, J., Lee, V., 428,
339-346,1987; Richter, W. O., Schwandt, P., Neuropeptides 9, 59-74,1987).
Despite lipolytic activity in rodents and rabbits, ACTH has very little effect on
lipolysis in isolated human and non-human primate adipocytes, even at
concentrations as high as 1|iM (Ng, T.B. Comparative Biochem. 97,441-446,
1990). Thus defining the role of melanocortins and their receptors in animal
sebaceous model systems is not necessarily predictive of their role in a human
sebaceous lipid regulation.
Recently, Basu et.al., in WIPO publication W099/55679 disclosed
isoquinoline derivatives, small molecule non-peptide compounds, which
showed low micromolar affinities for the MC1 and MC4 receptors, reduction of
dermal inflammation induced by arachidonic acids, and reductions of body
weight and food intake.
Nargund et. al., in WIPO publication WO99/64002 disclosed
spiropiperidine derivatives as melanocortin receptor agonists, useful for the
treatment of diseases and disorders such as obesity, diabetes and sexual
dysfunction.

Thus there exist a need for small molecule modulators of the
melanocortin receptor, more particularly the melanocortin-3, melanocortin-4
and / or the melanocortin-5 receptors.
SUMMARY OF THE INVENTION!
The present invention is directed to compounds of the general formula (II)
wherein
R1 is selected from the group consisting of heteroaryl, heteroaryt-alkyl,
heterocycloalkyl, heterocycloalkyl-alkyf, cycloalkyl, cycloalkyl-alkyl, substituted
aryl and substituted aralkyl; (R1 is not unsubstituted aryl or unsubstituted aralkyl);
wherein the heteroaryl, heteroaryl-alkyl, heterocycloalkyl, heterocycloalkyl-
alkyl, cycloalkyl or cycloalkyt-alky) group is optionally substituted with one or
more substituents independently selected from halogen, hydroxy, alkyl, alkoxy
halogenated alkyl, halogenated alkoxy, amino, alkylamino or di(alkyl)amino;
wherein the aryl or aralkyl group is substituted with one or more
substituents independently selected from halogen, hydroxy, alkyl, alkoxy,
halogenated alkyl, halogenated alkoxy, amino, alkylamino or di(alkyl)amino;
R2 is selected from the group consisting of heteroaryl, heteroaryl-alkyl,
heterocycloalkyl, heterocycloalkyl-alkyl, cycloalkyl, cycloalkyl-alkyl, substituted
aryl and substituted aralkyl; (R2 is not unsubstituted aryl or unsubstitutedd
aralkyl);
wherein the heteroaryl, heteroaryl-alkyl, heterocycloalkyl, heterocycloalkyl-
alkyl. cycloalkyl or cycloalkyl-alkyl group is optionally substituted with one or
more substituents independently selected from halogen, hydroxy, alkyl, alkoxy,
halogenated alkyl, halogenated alkoxy, amino, alkylamino or di(alkyl)amino;

wherein the aryl or aralkyl group is substituted with one or more
substituents independently selected from halogen, hydroxy, alkyl, alkoxy;
halogenated alkyl, halogenated alkoxy, amino, alkylamino or di(alkyl)amino;
R4 is selected from the group consisting of aryl, aralkyl, heteroaryl,
heterocycloalkyl, and cycloalkyl-alkyl; wherein the aryl, aralkyl, heteroaryl,
heterocycloalkyl or cycloalkyl-alkyl group is optionally substituted with one or
more substituents independently selected from halogen, hydroxy, alkyl, alkoxy,
halogenated alkyl, halogenated alkoxy, amino, alkylamino or di(alkyl)amrno;
provided that when R1 is aryl substituted with one halogen and R4 is aryl
substituted with one halogen, then R2 is not morpholinyl;
and pharmaceutically acceptable salts thereof.
The present invention is further directed to compounds of the general
formula (II)
wherein
R1 is selected from the group consisting of heteroaryl, heteroaryl-alkyl,
heterocycloalkyl, heterocycloalkyl-alkyl, cycloalkyl, cycloalkyl-alkyl, substituted
aryl and substituted aralkyl; (R1 is not unsubstituted aryl or unsubstituted aralkyl);
wherein the heteroaryl, heteroaryl-alkyl, heterocycloalkyl, heterocycloalkyl-
alkyl, cycloalkyl or cycloalkyl-alkyl group is optionally substituted with one or
more substituents independently selected from halogen, hydroxy, alkyl, alkoxy;
halogenated alkyl, halogenated alkoxy, amino, alkylamino or di(alkyl)amino;
wherein the aryl or aralkyl group is substituted with one or more
substituents independently selected from halogen, hydroxy, alkyl, alkoxy;
halogenated alkyl, halogenated alkoxy, amino, alkylamino or di(alkyl)amino;
R2 is selected from the group consisting of aryl, aralkyl, heteroaryl,
heteroaryl-alkyl, heterocycloalkyl, heterocycloalkyl-alkyl, cycloalkyl, cycloalkyl-
alkyl, substituted aryl and substituted aralkyl;

wherein the aryl, aralkyl, heteroaryl, heteroaryl-alkyl, heterocycloalkyl,
heterocycloalkyl-alkyl, cycloalkyl or cycloalkyl-alkyl group is optionally substituted
with one or more substituents independently selected from halogen, hydroxy,
alkyl, alkoxy; halogenated alkyl, halogenated alkoxy, amino, alkylamino or
di(alkyi)amino;
R4 is selected from the group consisting of heteroaryl, heterocycloalkyl,
cycloalkyl-alkyl, substituted aryl and substituted aralkyl; (R4 is not unsubstituted
aryl or unsubsttuted aralkyl);
wherein the heteroaryl, heterocycloalkyl or cycloalkyl-alkyl group is
optionally substituted with one or more substituents independently selected from
halogen, hydroxy, alkyl, alkoxy; halogenated alkyl, halogenated alkoxy, amino,
alkylamino or di(alkyl)amino;
wherein the aryl or aralkyl group is substituted with one or more
substituents independently selected from halogen, hydroxy, alkyl, alkoxy;
halogenated alkyl, halogenated alkoxy, amino, alkylamino or di(alkyl)amino;
provided that when R1 is aryl substituted with one halogen and R4 is aryl
substituted with one halogen, then R2 is not morpholinyl;
provided further that when R1 is methylphenyl and R4 is methoxyphenyl,
then R2 is selected from the group consisting of aralkyl, heteroaryl, heteroaryl-
alkyl, heterocycloalkyl, heterocycloalkyl-alkyl, cycloalkyl, cycloalkyl-alkyl and
substituted aryl;
wherein the aralkyl, heteroaryl, heterocycloalkyl or cycloalkyl group is
optionally substituted with one or more substituents independently selected from
halogen, hydroxy, alkyl, alkoxy; halogenated alkyl, halogenated alkoxy, amino,
alkylamino or di(alkyl)amino;
wherein the aryl group is substituted with one or more substituents
independently selected from halogen, hydroxy, alkyl, alkoxy; halogenated alkyl,
haloc mated alkoxy, amino, alkylamino or di(alkyl)amino;
and pharmaceutical^ acceptable salts thereof.
The present invention is further directed to a method for treating disorders
mediated by a melanocortin receptor comprising administering to a subject in
need thereof a therapeutically effective amount of a compound of formula (II)

wherein
R1 is selected from the group consisting of alkyl, aryl, aralkyl, heteroaryl,
heteroaryl-alkyl, heterocycloalkyl, heterocycloalkyl-alkyl, cycloalkyl and
cycloalkyl-alkyl; wherein the aryl, aralkyl, heteroaryl, heteroaryl-alkyl,
heterocycloalkyl, heterocycloalkyl-alkyl, cycloalkyl or cycloalkyl-alkyl group is
optionally substituted with one or more substituents independently selected from
halogen, hydroxy, alkyl, alkoxy; haiogenated alkyl, haiogenated alkoxy, amino,
alkylamino or di(alkyl)amino;
R2 is selected from the group consisting of alkyl, aryl, aralkyl, heteroaryl,
heterocycloalkyl and cycloalkyl-alkyl; wherein the aryl, aralkyl, heteroaryl,
heteroaryl-alkyl, heterocycloalkyl, heterocycloalkyl-alkyl, cycioalkyt or cycloalkyl-
alkyl group is optionally substituted with one or more substituents independently
selected from halogen, hydroxy, alkyl, alkoxy; haiogenated alkyl, haiogenated
alkoxy, amino, alkylamino or di(alkyl)amino;
R4 is selected from the group consisting of hydrogen, alkyl, aryl, aralkyl,
heteroaryl, heterocycloalkyl, and cycloalkyl-alkyl; wherein the aryl, aralkyl,
heteroaryl, heterocycloalkyl or cycloalkyl-alkyl group is optionally substituted
with one or more substituents independently selected from halogen, hydroxy,
alkyl, alkoxy; haiogenated alkyl, haiogenated alkoxy, amino, alkylamino or
di(alkyl)amino;
and pharmaceutical^ acceptable salts thereof.
Illustrative of the invention is a pharmaceutical composition comprising a
pharmaceutically acceptable carrier and any of the compounds described
above. An illustration of the invention is a pharmaceutical composition made
by mixing any of the compounds described above and a pharmaceutically
acceptable carrier. Illustrating the invention is a process for making a

pharmaceutical composition comprising mixing any of the compounds
described above and a pharmaceutically acceptable carrier.
Exemplifylng the invention are methods of treating disorders mediated
by the melanocortin receptor in a subject in need thereof comprising
administering to the subject a therapeutically effective amount of any of the
compounds or pharmaceutical compositions described above.
An embodiment of the present invention is the use of any of the
compounds described herein for the treatment of a disorder selected from the
group consisting of metabolic disorders, CNS disorders and dermatological
disorders.
An example of the invention is a method for treating a disorder selected
from the group consisting of obesity, impaired oral glucose tolerance, elevated
blood glucose levels, type II diabetes, Syndrome X, diabetic retinopathy, spinal
cord injury, nerve injury, acute neurodegenerative disorders, chronic
neurodegenerative disorders, plexopathies, male erectile dysfunction, dry eyes,
acne, dry skin, aged skin, seborrheic dermatitis, rosacea, excessive ear wax,
meibomian gland disorder, pseudofolliculitis, yeast infections, dandruff,
hidradenitis suppurativa, ocular rosacea and eccrine gland disorder in a subject
in need thereof comprising administering to the subject a therapeutically
effective amount of any of the compounds or pharmaceutical compositions
described above.
Another example of the invention is the use of any of the compounds
described herein in the preparation of a medicament for treating: (a) obesity,
(b) impaired oral glucose tolerance, (c) elevated blood glicose levels, (d) type
II diabetes, (e) Syndrome X, (0 diabetic retinopathy, (g) an acute
neurodegenerative disorder, (h) a chronic neurodegenerative disorder, (i) a
plexopathy, (j) male erectile dysfunction, (k) dry eyes, (I) acne, (m) dry skin, (n)
aged skin, (o) seborrheic dermatitis, (p) rosacea, (q) excessive ear wax, (r)
meibomian gland disorder, (s) pseudofolliculitis, (t) yeast infections, (u)

dandruff, (v) hidradenitis suppurativa, (w) ocular rosacea or (x) eccrine gland
disorder, in a subject in need thereof.
DETAIL ED DESCRIPTION OF THE INVENTION
The present invention is directed to novel substituted 1,2,4-thiadlazole
derivatives useful for the treatment of disorders mediated by a melanocortin
receptor. More particularly, the present invention is directed to compounds of
formula (II)
wherein R1, R2 and R4 are as herein defined, useful as melanocortin
receptor agonists or antagonists.
The present invention is further directed to a method of treating a
disorder mediated by a melanocortin receptor, preferably a disorder which is
susceptible to treatment by agonism or antagonism of a melanocortin receptor.
Preferably the melanocortin receptor is selected from the group consisting of
the melanocortin-3, melanocortin-4 and melanocortin-5 receptor, more
preferably the melanocortin receptor is melanocortin-4 or melanocortin-5.
In an embodiment of the present invention, R1 is selected from the group
consisting of aryl, aralkyl and heteroaryl; wherein the aryt, aralkyl or heteroaryl
group is optionally substituted with one or more substituents independently
selected from halogen, hydroxy, alkyl. alkoxy; trihalomethyl, trihalomethoxy,
amino, alkylamino or di(alkyl)amino. Preferably, R1 is selected from the group
consisting of aryl; wherein aryl group is optionally substituted with one or more
substituents independently selected from halogen, alkyl and alkoxy. More
preferably, R1 is selected from the group consisting of phenyl, 2-chlorophenyl, 4-
chlorophenyl, 2-methylphenyl, 4-methylphenyl, 2-methoxyphenyl and 4-
methoxyphenyl. Most preferably R1 is 2-methoxyphenyl.

In another embodiment of the present invention R1 is selected from the
group consisting of substituted aryl and substituted aralkyl, wherein the aryl or
aralkyl is substituted with one to two substituents independently selected from
halogen, hydroxy, alkyl, alkoxy, trihalomethyl, trihalomethoxy, amino, alkylamino
or di(alkyl)amino. Preferably, R1 is selected from the group consisting of
substituted aryl; wherein the aryl group is substituted with a substituent selected
from halogen, alkyl or alkoxy. More preferably, R1 is 2-methoxyphenyl.
In yet another embodiment of the present invention, R1 is selected from
the group consisting of substituted aryl and substituted aralkyl; wherein the aryl
or aralkyl group is substituted with one or more substituents independently
selected from halogen, hydroxy, alkyl, alkoxy; halogenated alkyl, halogenated
alkoxy, amino, alkylamino or di(alkyl)amino.
in an embodiment of the present invention, R2 is selected from the group
consisting of aryl, aralkyl and heteroaryl; wherein the aryl, aralkyl or heteroaryl
group is optionally substituted with one or more substituents independently
selected from halogen, hydroxy, alkyl, alkoxy; trihalomethyl, trihalomethoxy,
amino, alkylamino or di(alkyl)amino. Preferably, R2 is selected from the group
consisting of aryl; wherein the aryl group is optionally substituted with one or
more substituents independently selected from alkyl and alkoxy. More
preferably, R2 is selected from the group consisting of phenyl, 4-methylphenyl,
2-methoxyphenyl and 4-methoxyphenyl. Most preferably, R2 is selected from
the group consisting of phenyl and 2-methoxyphenyl.
In another embodiment of the present invention R2 is selected from the
group consisting of substituted aryl and substituted aralkyl, wherein the aryl or
aralkyl is substituted with one to two substituents independently selected form
halogen, hydroxy, alkyl, alkoxy, trihalomethyl, trihalomethoxy, amino, alkylamino
or di(alkyl)amino. Preferably, R2 is selected from the group consisting of
substituted aryl; wherein the aryl group is substituted with a substituent selected
from alkyl or alkoxy. More preferably, R2 is 2-methoxyphenyl.

In yet another embodiment of the present invention, R2 is selected from
the group consisting of aryl and aralkyl; wherein the aryl or aralkyt group is
optionally substituted with one or more substituents independently selected from
halogen, hydroxy, alkyl, alkoxy; halogenated alkyt, halogenated alkoxy, amino,
alkylamino or di(alkyl)amino.
In an embodiment of the present invention, R4 is selected from the
group consisting of aryl, aralkyl and heteroaryl; wherein the aryl, aralkyl or
heteroaryl group is optionally substituted with one or more substituents
independently selected from halogen, hydroxy, alkyl, alkoxy; trihalomethyl,
trihalomethoxy, amino, alkylamino or di(alkyl)amino. Preferably, R4 is selected
from the group consisting of aryl, aralkyl, and heteroaryl; wherein the aryl or
aralkyl group is optionally substituted with one or more substituents
independently selected from halogen, alkyl and alkoxy. More preferably, R4 is
selected from the group consisting of phenyl, 2-chlorophenyl, 4-chlorophenyl,
4-bromophenyl, 2-methylphenyl, 4-methylphenyl, 2-methoxyphenyl, 4-
methoxyphenyl, benzyl, 2-chlorobenzyl, 4-chlorobenzyl, 2-methylbenzyl, 4-
methylbenzyl, 2-methoxybenzyl, 4-methoxybenzyl, 2,6-difluorophenyl, 3,5-
difluorophenyl, 2-chloro-6-methylphenyl and 3-pridyl. Most preferably, R4 is
selected from the group consisting of phenyl, 2-methylphenyl, 4-methylphenyl,
2-methoxyphenyl and 4-methoxyphenyl.
In another embodiment of the present invention R4 is selected from the
group consisting of aryl, aralkyl or heteroaryl; wherein the aryl, aralkyl or
heteroaryl group is optionally substituted with one or more substituents
independently selected from halogen, hydroxy, alkyl, alkoxy, trihalomethyl,
trihalomethoxy, amino, alkylamino, di(alkyl)amino. . 'referably, R4 is selected
from the group consisting of aryl and substituted aryl; wherein the substituents
on the aryl are one to two independently selected halogen, alkyl or alkoxy. More
preferably, R4 is selected from the group consisting of phenyl, 4-methoxyphenyl,
2,6-difluorophenyl, 2-chloro-6-methylphenyl and 3,5-difluorophenyl.

In yet another embodiment of the present invention, R4 is selected from
the group consisting of substituted aryf and substituted aralkyl; wherein the aryl
or aralkyl group is substituted with one or more substituents independently
selected from halogen, hydroxy, alkyl, alkoxy; halogenated alkyl, halogenated
alkoxy, amino, alkylamino or di(alkyl)amino.
As used herein, unless otherwise note, the term "disorders mediated by
a melanocortin receptor" include, but are not limited to, obesity, impaired oral
glucose tolerance, elevated blood glucose levels, type II diabetes, Syndrome
X, diabetic retinopathy, acute neurodegenerative disorders, chronic
neurodegenerative disorders, plexopathies, male erectile dysfunction, dry eyes,
acne, dry skin, aged skin, seborrheic dermatitis, rosacea, excessive ear wax,
meibomian gland disorder, pseudofolliculitis, yeast infections, dandruff,
hidradenitis suppurativa, ocular rosacea and eccrine gland disorder.
As used herein, unless otherwise noted, the term "metabolic disorders"
include, but are not limited to, obesity, impaired oral glucose tolerance,
elevated blood glucose levels, type II diabetes and Syndrome X.
As used herein, unless otherwise noted, the term "CNS disorders"
include, but are not limited to, diabetic retinopathy, acute neurodegenerative
disorders, chronic neurodegenerative disorders and plexopathies.
As used herein, unless otherwise noted, the term "dermatological
disorders" include, but are not limited to, dry eyes, acne, dry skin, aged skin,
seborrheic dermatitis, rosacea, excessive ear wax, meibomian gland disorder,
pseudofolliculitis, yeast infections, dandruff, hidradenitis suppurativa, ocular
rosacea and eccrine gland disorder.
As used herein, acute neurodegenerative disorders include various
types of acute neurodegenerative disorders associated with neuronal cell death
or compromise including cerebrovascular insufficiency, focal or diffuse brain
trauma, diffuse brain damage, and spinal cord injury, that is, cerebral ischemia

or infarction including embolic occlusion and thrombotic occlusion, reperfusion
following acute ischemia, perinatal hypoxic-ischemic injury, cardiac arrest, as
well as intracranial hemorrhage of any type (including, but not limited to,
epidural, subdural, subarachnoid and intracerebral), and intracranial and
intravertebral lesions (including, but not limited to, contusion, penetration,
shear, compression and laceration), and whiplash shaken infant syndrome.
As used herein, chronic neurodegenerative disorders included within the
methods of the present invention include Alzheimer's disease. Pick's disease,
diffuse Lewy body disease, progressive supranuclear palsy (Steel-Richardson
syndrome), multisystem degeneration (Shy-Drager syndrome), chronic epileptic
conditions associated with neurodegeneration, motor neuron diseases
including amyotrophic lateral sclerosis, degenerative ataxias, cortical basal
degeneration, ALS-Parkinson's-Dementia complex of Guam, subacute
sclerosing panencephalitis, Huntington's disease, Parkinson's disease,
synucieinopathies (including multiple system atrophy), primary progressive
aphasia, striatonigral degeneration, Machado-Joseph disease / spinocerebellar
ataxia type 3 and olivopontocerebellar degenerations, Gilles De La Tourette's
disease, bulbar and pseudobulbar palsy, spinal and spinobulbar muscular
atrophy (Kennedy's disease), primary lateral sclerosis, familial spastic
paraplegia, Werdnig-Hoffmann disease, Kugelberg-Welander disease, Tay-
Sach's disease, Sandhoff disease, familial spastic disease, Wohlfart-
Kugelberg-Welander disease, spastic paraparesis, progressive multifocal
leukoencephalopathy, familial dysautonomia (Riley-Day syndrome), and prion
diseases (including, but not limited to Creutzfeldt-Jakob, Gerstmann-
Straussler-Scheinker disease, Kuru and fatal familial insomnia).
As used herein, plexopathies include plexus palsies, multifocal
neuropathies, sensory neuropathies, motor neuropathies, sensory-motor
neuropathies, infections neuropathies, autonomic neuropathies, sensory-
autonomic neuropathies, demyelinating neuropathies (including, but not limited
to Guillain-Barre syndrome and chronic inflammatory demyelinating
polyradiculoneuropathy), other inflammatory and immune neuropathies,
14
neuropathies induced by drugs, neuropathies induced by pharmacological
treatments, neuropathies induced by toxins, traumatic neuropathies (including,
but not limited to compression, crush, laceration and segmentation
neuropathies), metabolic neuropathies, endocrine and paraneoplastic
neuropathies, and other neuropathies such as Charcot-Marie-Tooth disease
(type 1a, 1b, 2,4a,1-X linked), Friedreich's ataxia, metachromatic
leukodystrophy, Refsum's disease, adrenomyeloneuropathy, Ataxia-
telangiectasia, Dejerine-Sottas neuropathy (types A and B), Lambert-Eaton
syndrome, and disorders of the cranial nerves.
As used herein, unless otherwise noted, the term "halogen" shall include
iodine, bromine, chlorine and fluorine.
As used herein, the term "alkyl" whether used alone or as part of a
substituent group, include straight and branched chains comprising one to
eight carbon atoms. For example, alkyl radicals include methyl, ethyl, propyl,
isopropyl, butyl, isobutyl, sec-butyl, t-butyl, pentyl and the like. Unless
otherwise noted, "lower" when used with alkyl means a carbon chain
composition of one to four carbon atoms.
The term "alkenyT, whether used alone or as part of a substituent group,
shall include straight and branched alkene chains comprising two to eight
carbon atoms. Suitable examples include vinyl, 1-propenyl, 2-propenyl, 1-
butenyl. 2-butenyl, 1-pentenyl. 2-pentenyJ, 1-isobut-2-enyl, and the like.
Similarly, the term "alkynyl", whether used alone or as part of a substituent
group, shall include straight and branched alkyne chains comprising two to
eight carbon atoms. Suitable examples include 2-propynyl, 2-butynyl, 1-
butynyl, 1-pentynyl, and the like.
As used herein, unless otherwise noted, "alkoxy" shall denote an oxygen
ether radical of the above described straight or branched chain alkyl groups. For
example, methoxy, ethoxy, n-propoxy, sec-butoxy, t-butoxy, n-hexyloxy and the
like.

As used herein, unless otherwise noted, the term "cycloalkyl shall
denote saturated monocyclic ring structures comprising three to eight ring
carbons, preferably 5 to 7 carbons. Suitable examples include cyclopropyl,
cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl and cyclooctyl.
As used herein, the term "aryl" indicates aromatic carbocyclic ring
structures such as phenyl, naphthyl, and the like.
As used herein, unless otherwise noted, "aralkyl" shall mean any lower
alkyl group substituted with an aryl group. For example, benzyl, phenylethyl,
phenylpropyl, naphthylmethyl, and the like.
As used herein, unless otherwise noted, "heteroaryl shall denote any five
or six membered monocyclic aromatic ring structure containing at least one
heteroatom selected from the group consisting of O, N and S, optionally
containing one to three additional heteroatoms independently selected from the
group consisting of O, N and S; or a nine or ten membered bicyclic aromatic ring
structure containing at least one heteroatom selected from the group consisting
of 0, N and S, optionally containing one to four additional heteroatoms
independently selected from the group consisting of O, N and S. The heteroaryl
group may be attached at any carbon atom of the ring such that the result is a
stable structure.
Examples of suitable heteroaryl groups include, but are not limited to,
pyrrolyl, furyl, thienyl, oxazolyl, imidazolyt, purazolyl, isoxazolyl, isothiazolyl,
triazolyl, thiadiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, pyranyl, furazanyl,
indolizinyl, indolyl, isoindolinyl, indazolyl, isoxazolyl, benzofuryl, benzothienyl,
benzimidazolyl, benzthiazolyl, purinyl, quinolizinyl, quinolinyl, isoquinolinyl,
isothiazolyl, cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, naphthyridinyl,
pteridinyl, and the like. Preferred heteroaryl groups include pyridyl, thienyl and
imidazolyl.

As used herein, the term "heterocycloalkyl" shall denote any five to seven
membered monocyclic, saturated, partially unsaturated or partially aromatic ring
structure containing at least one heteroatom selected from the group consisting
of O, N and S, optionally containing one to three additional heteroatoms
independently selected from the group consisting of O, N and S; or a nine to ten
membered saturated, partially unsaturated or partially aromatic bicyclic ring
system containing at least one heteroatom selected from the group consisting of
0, N and S, optionally containing one to four additional heteroatoms
independently selected from the group consisting of O, N and S. The
heterocycloalkyl group may be attached at any carbon atom of the ring such that
the result is a stable structure.
Examples of suitable heterocycloalkyl groups include, but are not limited
to, pyrrolinyl, pyrrolidinyl, dioxalanyl, imidazolinyl, imidazolidinyl, pyrazolinyl,
pyrazolidinyl, piperidinyl, dioxanyl, morpholinyl, dithianyl, thiomorpholinyl,
piperazinyl, trithianyl, indolinyl, chromenyl, 3,4-methylenedioxyphenyl, 2,3-
dihydrobenzofuryl, and the like.
As used herein, the notation "*" shall denote the presence of a
stereogenic center.
Where the compounds according to this invention have at least one
chiral center, they may accordingly exist as enantiomers. Where the
compounds possess two or more chiral centers, they may additionally exist as
diastereomers. It is to be understood that all such isomers and mixtures
thereof are encompassed within the scope of the present invention.
Furthermore, some of the crystalline forms for the compounds may exist as
polymorphs and as such are intended to be included in the present invention.
In addition, scnie of the compounds may form solvates with water (i.e.,
hydrates) or common organic solvents, and such solvates are also intended to
be encompassed within the scope of this invention.
When a particular group is "substituted" (e.g., cycloalkyl, aryl, aralkyl,
heteroaryl, heterocycloalkyl), that group may have one or more substituents,

preferably from one to five substituents, more preferably from one to three
substituents, most preferably from one to two substituents, independently
selected from the list of substituents.
It is intended that the definition of any substituent or variable at a
particular location in a molecule be independent of its definitions elsewhere in
that molecule, it is understood that substituents and substitution patterns on
the compounds of this invention can be selected by one of ordinary skill in the
art to provide compounds that are chemically stable and that can be readily
synthesized by techniques know in the art as well as those methods set forth
herein.
Under standard nomenclature used throughout this disclosure, the
terminal portion of the designated side chain is described first, followed by the
adjacent functionality toward the point of attachment. Thus, for example, a
MphenylCi-C6alkylaminocarbonylCi-C6alkyl" substituent refers to a group of the
formula

The term "subject" as used herein, refers to an animal, preferably a
mammal, most preferably a human, who is or has been the object of treatment,
observation or experiment.
As used herein, the term "composition" is intended to encompass a
product comprising the specified ingredients in the specified amounts, as well
as any product which results, directly or indirectly, from combinations of the
specified ingredients in the specified amounts.
The term "therapeutically effective amount" as used herein, means that
amount of active compound or pharmaceutical agent that elicits the biological or

medicinal response in a tissue system, animal or human that is being sought by
a researcher, veterinarian, medical doctor or other clinician, which includes
alleviation of the symptoms of the disease or disorder being treated.
For use in medicine, the salts of the compounds of this invention refer to
non-toxic "pharmaceutically acceptable salts". Cther salts may, however, be
useful in the preparation of compounds according to this invention or of their
pharmaceutically acceptable salts. Suitable pharmaceutically acceptable salts
of the compounds of this invention include acid addition salts which may, for
example, be formed by mixing a solution of the compound with a solution of a
pharmaceutically acceptable acid such as hydrochloric acid, sulfuric acid,
fumaric acid, maleic acid, succinic acid, acetic acid, benzoic acid, citric acid,
tartaric acid, carbonic acid or phosphoric acid.
The present invention includes within its scope prodrugs of the
compounds of this invention. In general, such prodrugs will be functional
derivatives of the compounds which are readily convertible in vivo into the
required compound. Thus, in the methods of treatment of the present
invention, the term "administering" shall encompass the treatment of the
various disorders described with the compound specifically disclosed or with a
compound which may not be specifically disclosed, but which converts to the
specified compound in vivo after administration to the patient. Conventional
procedures for the selection and preparation of suitable prodrug derivatives are
described, for example, in "Design of Prodrugs", ed. H. Bundgaard, Elsevier,
1985.
Abbreviations used in the instant specification, particularly the Schemes
and Examples, are as follows:
BHT = 2,6-bis-(t-butyl)-4-methyl-phenol
BSA = Bovine Serum Albumin
cAMP or = Cyclic-adenosine monophosphate
cyclic AMP

DCE = 1,2-dichloroethane
DEAD = Diethyl azodicarboxylate
DM = Differentiation Medium
DMF = Dimethyl formamide
DMEM = Dulbeccos Minimal Essential Medium
DMSO Dimethylsulfoxide
DPBS = Dulbeccos phosphate buffered saline
EDTA = Ethylene Diamine Tetraacetic Acid
FBS = Fetal bovine serum
GDP = Guanosine Diphosphate
GTP = Guanosine Triphosphate
GM = Growth Medium
HBSS = Hank's Buffered salt Solution
HEPES = 4-(2-Hydroxyethyl)-1-piperizine ethane
sulfonic acid
HS = Human Serum
IgG = Immunoglobulin G
% Inh = Percent Inhibition
MEM = Minimum Essential Medium
NBS = N-bromosuccinimide
NCS = N-chlorosuccinimide
NDP aMSH = [Nle4, D-Phe7]aMSH, an analog of aMSH
PBS = Phosphate Buffered saline
PEG = Polyethylene Glycol
PNC = Penicillin
rt or RT = Room Temperature
SPA = Scintillation Proximity Assay
bTM = Streptomycin
TLC = Thin layer chromatography
TM = Transition Medium
TMS = Trimethylsilyl
Compounds of formula (I) wherein R3 is hydrogen may be prepared
according to the process outlined in Scheme 1.

More particularly, a suitably substituted cyano compound of formula (III),
a known compound or compound prepared by known methods, is reacted with
a suitably substituted primary amine of formula (IV), a known compound or
compound prepared by known methods, in the presence of a base such as
NaNH2. NaH, NaN(TMS)2, and the like, preferably NaNH2, at an elevated
temperature, preferably at about reflux, to yleld the corresponding compound
of formula (V).
The compound of formula (V) is reacted with a suitably substituted
thiocyanate of formula (VI), a known compound or compound prepared by
known methods, in the presence of DCE, at an elevated temperature,
preferably about 45°C, to yleld the corresponding compound of formula (VII).
The compound of formula (VII) is subjected to ring closure/oxidation, in
the presence of Br2, at room temperature, to yleld the corresponding
compound of formula (la).
Compounds of formula (II) may be prepared from suitably substituted
compounds of formula (I) wherein R3 is hydrogen according to the process
outlined in Scheme 2.


More particularly, a suitably substituted compound of formula (la) is
treated with a base such as NaHC03, Na2CO3, NaOH, and the like, preferably
NaHCO3, at room temperature, to yleld the corresponding compound of
formula (II).
Compounds of formula (II) may also be prepared from suitably
substituted compounds of formula (VII) according to the process outlined in
Scheme 3.

More particularly, a suitably substituted compound of formula (VII) is
reacted with an oxidizing agent such as NBS, NCS, DEAD, and the like,
preferably NBS, at room temperature, to yleld the corresponding compound of
formula (II). Preferably, the compound of formula (II) is extracted from a basic
aqueous solution such as NaHCOa, Na2CO3, NaOH, and the like.
Compounds of formula (I) wherein R3 is alkyl may be prepared from a
suitably substituted compound of formula (II) according to the process outlined
in Scheme 4.

Accordingly, a suitably substituted compound of formula (II) is reacted
with a suitably substituted compound of formula (VIII), a known compound or
compound prepared by known methods, wherein X is trifluorornethylsulfonate,
Br-, and I-, at room temperature, to yleld the corresponding compound of
formula (I).
Compounds of formula (I) wherein R3 is hydrogen may be prepared from
a suitably substituted compound of formula (II) according to the process
outlined in Scheme 5.

Accordingly, a suitably substituted compound of formula (II) is reacted
with a pharmaceutically acceptable acid such as HCI, HBr, HNO3, and the like,
preferably HCI, at room temperature, to yleld the corresponding compound of
formula (la), wherein X" is CI-, Br-, NO3-. and the like.
Where the processes for the preparation of the compounds according to
trie invention give rise to mixture of stereoisomers, these isomers may be
separated by conventional techniques such as preparative chromatography.
The compounds may be prepared in racemic form, or individual enantiomers
may be prepared either by enantiospecific synthesis or by resolution. The
compounds may, for example, be resolved into their components enantiomers
by standard techniques, such as the formation of diastereomeric pairs by salt

formation with an optically active acid, such as (-)-di-p-toluoyl-d-tartaric acid
and/or (+)-di-p-toluoyl-l-tartaric acid followed by fractional crystallization and
regeneration of the free base. The compounds may also be resolved by
formation of diastereomeric esters or amides, followed by chromatographic
separation and removal of the chiral auxiliary. Alternatively, the compounds
may be resolved using a chiral HPLC column.
During any of the processes for preparation of the compounds of the
present invention, it may be necessary and/or desirable to protect sensitive or
reactive groups on any of the molecules concerned. This may be achieved by
means of conventional protecting groups, such as those described in
Protective Groups in Oroanic Chemistry, ed. J.F.W. McOmie, Plenum Press,
1973; and T.W. Greene & P.G.M. Wuts, Protective Groups in Oroanic
Synthesis. John Wliey & Sons, 1991. The protecting groups may be removed
at a convenient subsequent stage using methods known from the art.
The utility of the compounds of the present invention to treat disorders
mediated by a melanocortin receptor can be determined according to the
procedures described in Examples 4-11 herein. The present invention
therefore provides a method of treating such disorders, which comprises
administering any of the compounds as defined herein in a quantity effective to
treat the disorder (i.e. in a therapeutically effective amount). The compound
may be administered to a patient afflicted with such a disorder by any
conventional route of administration, including, but not limited to, intravenous,
oral, subcutaneous, intramuscular, intradermal, parenteral and transdermal.
The present invention also provides pharmaceutical compositions
comprising one or more copounds of this invention in association with a
pharmaceutically acceptable carrier.
To prepare the pharmaceutical compositions of this invention, one or
more compounds of formula (I) and/or (II) or a salt thereof (the active
ingredient), is intimately admixed with a pharmaceutical carrier according to

conventional pharmaceutical compounding techniques, which carrier may take
a wide variety of forms depending of the form of preparation desired for
administration, e.g., oral or parenteral such as intramuscular. In preparing the
compositions in oral dosage form, any of the usual pharmaceutical media may
be employed. Thus, for liquid oral preparations, such as for example,
suspensions, elixirs and solutions, suitable carriers and additives include water,
glycols, oils, alcohols, flavoring agents, preservatives, coloring agents and the
like; for solid oral preparations such as, for example, powders, capsules,
caplets, gelcaps and tablets, suitable carriers and additives include starches,
sugars, diluents, granulating agents, lubricants, binders, disintegrating agents
and the like. Because of their ease in 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
sugar coated or enteric coated by standard techniques. For parenterals, the
carrier will usually comprise sterile water, though other ingredients, for
example, for purposes such as aiding solubility or for preservation, may be
included. Injectable suspensions may also be prepared, in which case
appropriate liquid carriers, suspending agents and the like may be employed.
Topical formulations included within the present invention, include but
are not limited to creams, lotions, multiple emulsions, microemulsions,
liposomal creams or gels, gels, solutions, suspensions, ointments, foaming
aerosols, hard or soft gelatin capsules, masks, sticks, roll-ons, powders, spray
forms, and the like. The topical formulations may contain, in addition to the
active ingredient(s), one or more non-active components including, but are not
limited to chelating agents, buffering agents, colorants, preservatives,
fragrances, emulsifiers, surfactants, opacifylng agents, emollients, solvents,
sunscreens, viscosity modifylng agents, antioxidants moisturizers,
permeations enhancers, film formers, and the like.
Topical formulations for acne treatment included within the present
invention may also contain one or more of the following components, including
comedolytic/keratolytic agents, antimicrobial agents and steroidal or non-

steroidal anti-inflammatory agents. (Comedolytic agents refer to any
compound capable of rupturing a comedo. Keratolytic agents refer to any
compound capable of breaking apart keratinocyes resulting in exfoliation of the
epidermis.) Suitable comedolytic/keratofytic agents include, but are not limited
to retinoids, salicylic acid, giycoiic acid, cetyl betaine, and the like. Suitable
antimicrobial agents include, but are not limited to benzoyl peroxide,
erythromycin, tetracycline, clindamycin, azelaic acid, and the like. Topical
formulations typically contain 0.01-1% active ingredient.
The pharmaceutical compositions herein will contain, per dosage unit,
e.g., tablet, capsule, powder, injection, teaspoonful and the like, an amount of
the active ingredient necessary to deliver an effective dose as described
above. The non-topical pharmaceutical compositions herein will contain, per
unit dosage unit, e.g., tablet, capsule, powder, injection, suppository,
teaspoonful and the like, of from about 0.03 mg to 100 mg/kg (preferred 0.1-30
mg/kg) and may be given at a dosage of from about 0.1-300 mg/kg/day
(preferred 1-50 mg/kg/day). The dosages, however, may be varied depending
upon the requirement of the patients, the severity of the condition being treated
and the compound being employed. The use of either daily administration or
post-periodic dosing may be employed.
Preferably these compositions are in unit dosage forms from such as
tablets, pills, capsules, powders, granules, sterile parenteral solutions or
suspensions, metered aerosol or liquid sprays, drops, ampoules, autoinjector
devices or suppositories; for oral parenteral, intranasal, sublingual or rectal
administration, or for administration by inhalation or insufflation. Alternatively,
the composition may be presented in a form suitable for once-weekly or once-
monthly administration; for example, an insoluble salt of the active compound
such as the decanoate salt, may be adapted to provide a depot preparation for
intramuscular injection. For preparing solid compositions such as tablets, the
principal active ingredient is mixed with a pharmaceutical carrier, e.g.
conventional tableting ingredients such as com starch, lactose, sucrose,
sorbitol, talc, stearic acid, magnesium stearate, dicalcium phosphate or gums,

and other pharmaceutical diluents, e.g. water, to form a solid preformulation
composition containing a homogeneous mixture of a compound of the present
invention, or a pharmaceutically acceptable salt thereof. When referring to
these preformulation compositions as homogeneous, it is meant that the active
ingredient is dispersed evenly throughout the composition so that the
composition may be readily subdivided into equally effective dosage forms
such as tablets, pills and capsules. This solid preformulation composition is
then subdivided into unit dosage forms of the type described above containing
from 0.1 to about 500 mg of the active ingredient of the present invention. The
tablets or pills of the novel composition can be coated or otherwise
compounded to provide a dosage form affording the advantage of prolonged
action. For example, the tablet or pill can comprise an inner dosage and an
outer dosage component, the latter being in the form of an envelope over the
former. The two components can be separated by an enteric layer which
serves to resist disintegration in the stomach and permits the inner component
to pass intact into the duodenum or to be delayed in release. A variety of
material can be used for such enteric layers or coatings, such materials
including a number of polymeric acids with such materials as shellac, cetyl
alcohol and cellulose acetate.
The liquid forms in which the novel compositions of the present
invention may be incorporated for administration orally or by injection include,
aqueous solutions, suitably flavoured syrups, aqueous or oil suspensions, and
flavoured emulsions with edible oils such as cottonseed oil, sesame oil,
coconut oil or peanut oil, as well as elixirs and similar pharmaceutical vehicles.
Suitable dispersing or suspending agents for aqueous suspensions, include
synthetic and natural gums such as tragacanth, acacia, alginate, dextran,
sodium carboxymeth/lcellulose, methylcellulose, polyvinyl-pyrrolidone or
gelatin.
The method of treating disorders mediated by a melanocortin receptor
described in the present invention may also be carried out using a
pharmaceutical composition comprising any of the compounds as defined herein

and a pharmaceutical^ acceptable carrier. The non-topical pharmaceutical
composition may contain between about 0.01 mg and 100 mg, preferably about
5 to 50 mg, of the compound, and may be constituted into any form suitable for
the mode of administration selected, earners include necessary and inert
pharmaceutical excipients, including, but not limited to, binders, suspending
agents, lubricants, flavorants, sweeteners, preservatives, dyes, and coatings.
Compositions suitable for oral administration include solid forms, such as pills,
tablets, caplets, capsules (each including immediate release, timed release and
sustained release formulations), granules, and powders, and liquid forms, such
as solutions, syrups, elixers, emulsions, and suspensions. Forms useful for
parenteral administration include sterile solutions, emulsions and suspensions.
Advantageously, compounds of the present invention may be
administered in a single daily dose, or the total daily dosage may be
administered in divided doses of two, three or four times daily. Furthermore,
compounds for the present invention can be administered in intranasal form via
topical use of suitable intranasal vehicles, or via transdermal skin patches well
known to those of ordinary skill in that art. To be administered in the form of a
transdermal delivery system, the dosage administration will, of course, be
continuous rather than intermittent throughout the dosage regimen.
For instance, for oral administration in the form of a tablet or capsule, the
active drug component can be combined with an oral, non-toxic pharmaceutically
acceptable inert carrier such as ethanol, glycerol, water and the like. Moreover,
when desired or necessary, suitable binders; lubricants, disintegrating agents
and coloring agents can also be incorporated into the mixture. Suitable binders
include, without limitation, starch, gelatin, natural sugars such as glucose or
beta-lactose, com sweeteners, natural and syndetic gums such as acacia,
tragacanth or sodium oleate, sodium stearate, magnesium stearate, sodium
benzoate, sodium acetate, sodium chloride and the like. Disintegrators include,
without limitation, starch, methyl cellulose, agar, bentonfte, xanthan gum and the
like.

The liquid forms in suitably flavored suspending or dispersing agents such
as the synthetic and natural gums, for example, tragacanth, acacia, methyl-
cellulose and the like. For parenteral administration, sterile suspensions and
solutions are desired. Isotonic preparations which generally contain suitable
preservatives are employed when intravenous administration is desired.
The compound of the present invention can also be administered in the
form of liposome delivery systems, such as small unilamellar vesicles, large
unilamellar vesicles, and multilamellar vesicles. Liposomes can be formed from
a variety of phospholipids, such as cholesterol, stearylamine or
phophatidylcholines.
Compounds of the present invention may also be delivered by the use of
monoclonal antibodies as individual carriers to which the compound molecules
are coupled. The compounds of the present invention may also be coupled with
soluble polymers as targetable drug carriers. Such polymers can include
polyvinylpyrrolidone, pyran copolymer, polyhydroxypropylmethacrylamidephenol,
polyhydroxy-ethylaspartamidephenol, or polyethyl eneoxidepolylysine substituted
with palmitoyl residue. Furthermore, the compounds of the present invention
may be coupled to a class of biodegradable polymers useful in achieving
controlled release of a drug, for example, polylactic acid, polyepsilon
caprolactone, polyhydroxy butyeric acid, polyorthoesters, polyacetals,
polydihydropyrans, polycyanoacrylates and cross-linked or amphipathic block
copolymers of hydrogels.
Compounds of this invention may be administered in any of the foregoing
compositions and according to dosage regimens established in the art whenever
treatment of disorders mediated by a melanocortin receptor is required.
The daily dosage of the products may be varied over a wide range from
0.01 to 1,000 mg per adult human per day. For oral administration, the
compositions are preferably provided in the form of tablets containing, 0.01,0.05,
0.1, 0.5,1.0,2.5, 5.0,10.0,15.0, 25.0, 50.0,100,150, 200, 250 and 500

milligrams of the active ingredient for the symptomatic adjustment of the dosage
to the patient to be treated. An effective amount of the drug is ordinarily supplied
at a dosage level of from about 0.01 mg/kg to about 100 mg/kg of body weight
per day. Preferably, the range is from about 0.03 to about 10 mg/kg of body
weight per day. The compounds may be administered on a regimen of 1 to 4
times per day.
Optimal dosages to be administered may be readily determined by those
skilled in the art, and will vary with the particular compound used, the mode of
administration, the strength of the preparation, the mode of administration, and
the advancement of the disease condition. In addition, factors associated with
the particular patient being treated, including patient age, weight, diet and time of
administration, will result in the need to adjust dosages.
The following Examples are set forth to aid in the understanding of the
invention, and are not intended, and should not be construed to limit in any way
the invention set forth in the claims which follow thereafter.

STEP A:
A mixture of o-anisidine (15.0 g, 121.8 mmol) and sodium amide (50 wt.
% suspension in toluene) (11.40 g, 146.2 mmol) in anhydrous toluene (200ml)
was stirred for 1 hour at room temperature. To the mixture was added

benzonitrile (9.86 ml, 96.6 mmol) and heated under reflux for 16 hours. The
reaction mixture was cooled and 1 .ON HCI (150 ml) was added to quench the
reaction. Activated carbon was added and the reaction mixture was filtered
through a Celite pad. The pH of the mixture was adjusted to about 14 by
addition of 1.0 N NaOH (200 ml). The aqueous layer was extracted with
chloroform (3x150 ml). The combined organic layer was dried over anhydrous
MgSO4, and evaporated. The resulting solid was washed with hexane and
dried over the vacuum to yleld the product as a pale white solid.
1H NMR (300MHz. CDCI3) ™ 3.83 (s, 3H), 4.79 (s, 2H), 6.96(rn, 3H),
7.04 (m, 1H), 7.43 (m, 3H), 7.93 (d, 2H)
MS (APCI, MH+) 227
STEP B:
A mixture of N-arylbenzamidine (3.0 g, 13.27mmol), prepared as in Step
A, and 4-tolylisothiocyanate (2.18 g, 14.60) in anhydrous chloroform (30 ml)
was heated under reflux for 16 hours. The reaction mixture was cooled and
the solvent was evaporated. The resulting residue was purified by flash
column chromatography with a mobile phase of 25% hexane in
dichloromethane. The combined fractions were evaporated; and the resulting
solid was dried over vacuum to yleld the product as a yellow solid.
1H NMR (300MHz. CDCI3) ™ 2.36 (s. 3H). 3.66 (s, 3H), 6.76 (m. 1H),
6.97 (t. 1H), 7.17-7.39 (m. 7H). 7.47 (d, 2H). 7.60 (d. 2H), 8.20 (s. 1H), 14.18
(s, 1H)
MS (ES. MH+) 376.29
STEP C:
To the solution of thiourea (2.90 g, 7.73 mmol), prepared as in STEP B,
in anhydrous chloroform (15 ml) was slowly added bromine (438 µl, 8.51
mmol). After stirring for 16 hours, the solvent was evaporated. The resulting
solid was washed with anhydrous ethyl ether. The crude product was re-
crystallized from 20% water in ethanol to yleld the product as a yellow solid.
1H NMR (300MHz, CDCI3) ™ 2.39 (s, 3H), 3.66 (s, 3H), 6.96 (d, 1H),
7.06 (t, 1H), 7.20 - 8.07 (m, 7H), 7.61 (d, 2H), 7.80 (d, 2H), 12.40 (s. 1H)

STEP A:
A mixture of o-anisidine (13.5 g, 110.0 mmol) and sodium amide (50 wt.
% suspension in toluene) (9.40 g, 120.0 mmol) in anhydrous toluene (200ml)
was stirred for 1 hour at room temperature. To the mixture was added 2-
methoxybenzonitrile (16 ml, 131.0 mmol) and the reaction mixture was then
heated under reflux for 16 hours. The reaction mixture was cooled and 1.0N
HCI (150 ml) was added to quench the reaction. Activated carbon was added
and the reaction mixture was filtered through a Celite pad. The pH of the
mixture was adjusted to about 14 by addition of 1.0 N NaOH (200 ml). The
aqueous layer was extracted with chloroform (3x150 ml). The combined
organic layer was dried over anhydrous MgSO4, and evaporated. The resulting
solid was washed with hexane and dried over the vacuum to yleld the product
as a pale white solid.
MS (APCI, MH+) 257
STEP B:
A mixture of N-arylbenzamidine (15.5 g, 60.6 mmol), prepared as In
STEP A and phenylisothiocyanate (8.70 mL, 72.7 mmol) in anhydrous
chloroform (30 ml) was heated at 45°C for 16 hours. The reaction mixture was
cooled and the solvent was evaporated. The resulting residue was purified by

flash column chromatography with a mobile phase of 25% hexane in
dichloromethane. The combined fractions were evaporated and the resulting
solid was dried over vacuum to yleld the product as a yellow solid.
MS (ES,MH+) 391.50
STEP C:
To a solution of thiourea (12.95 g, 33.1 mmol), prpeared as in STEP B,
in anhydrous chloroform (15 ml) was slowly added bromine (1.78 mL, 34.75
mmol). After stirring for 16 hours, the solvent was evaporated. The resulting
solid was washed with anhydrous ethyl ether. The crude product was re-
crystallized from 20% water in ethanol to yleld the product as a yellow solid.
MS (ES, MH*) 390.1
Following the procedures disclosed herein, representative compounds
of formula (I) of the present invention were prepared, as listed in Table 1.
To a solution of the compound prepared as in Step B of Example 2
(0.921 g, 2.36 mmol) in anhydrous chloroform (10 mL) was added N-
chlorosuccinimide (326 mg, 2.71 mmol). The reaction mixture was then stirred
for 16 hours, and then stopped and washed twice with aqueous NaHCO3. The
organic layer was dried over magnesium sulfate and the remaining solvent
removed under vacuum to yleld the title product as a solid.
MS (ES, MH+) 390.1
Following the procedures described herein, representative compounds
of formula (II) of the present invention were prepared, as listed in Table2.
Unless otherwise noted, NMR spectra, were measured on a Bruker
Avance 300 MHz NMR spectrometer. Unless otherwise noted, molecular
weights were measured using a Micromass LC platform electrospray mass
spectrometer, as listed in Table 3.
EXAMPLE 4
Melanocortin MC-4 Receptor Binding Assay
Meianocortin [MC-4]-membrane [purchased from Receptor Biology Inc]
was coupled to wheat germ agglutinin coated polyvinyl toluene-Scintillation
Proximity Assay beads [purchased from Amersham Pharmacia Inc.] for 30 min
at 25°C. Into each well of a 96-well Opti plate [purchased from Packard, CA],
2.5 ug of membrane and 0.25 mg of beads were mixed in a volume of 100 |iL
media. The media was 50mM HEPES, pH 7.4 containing 0.1 % bovine serum
albumin, 2 mM CaCb, 2 mM MgCfe and protease inhibitors. Test compounds
(1.5 nL) at 1mM in 30% DMSO-50mM HEPES, pH 7.4 buffer was added to
separate wells on the plate. Radioactive ligand 125l-NDP-meianocyte
stimulating hormone [NEN, 2000Ci/mmol] was added to each well (48.5 u.L per
well, 40 pM final concentration). The plate was then sealed and let stand for 16
hr at 25°C. NDP-Melanocyte stimulating hormone peptide and a-melanocyte
stimulating hormone peptide [purchased from Palomar Research Inc, 1 nM]
were used as reference inhibitor compounds to define non-specific binding (N).
Total binding (T) was defined using 30% DMSO-50 mM HEPES, pH 7.4
buffer. Bound radioactivity for each well (Y), measured at counts per minute
(cpm) was measured in a TopCount [Packard, CA]. Percent inhibition was
calculated as:
EXAMPLE 5
Cvclic-Adenosine monophosphate fcAMPI Stimulation assay
Human Bowes melanoma cells expressing human melanocortin MC-4
receptor were grown to confluence in a 24-well culture plate. The growth
medium was discarded and to each well was added 0.5 mL of Hank's solution.
Test compounds were added to wells of a 96 well plate. NDP-melanocyte
stimulating hormone peptide (1 jaM) was added to the positive control wells
41
while negative control wells received vehicle of 30% DMSO-50mM HEPES, pH
7.4 buffer. The plate was incubated at 37°C and 5% CO2 for 30 min. The
supernatant was discarded and the cells were washed twice with Hank's
solution. Ethanol (80%, 0.5 mL) was added to each well and the plates were
incubated at 4ºC for 30 min. Cyclic AMP content was measured using the
NEN Flashplate kit [NEN]. A melanocortin receptor agonist is defined as a test
compound which resulted in an increase in cAMP production in this assay.
EXAMPLE 6
G-protein Activation Assay
For each assay, membranes expressing the melanocortin MC-4
receptor (5 ng) were incubated for 5 min at 25°C with 0.5 nM 35S-GTP?S in 100
µL of 25 mM HEPES buffer, pH7.5 containing 100 mM NaCI, protease
inhibitors. 0.5 yM GDP, 5 mM 2-mercaptoethanol, 1 mM MgCl2 together with
test compound, 1 µM of NDP-melahocyte stimulating hormone or a
combination of NDP-melanocyte stimulating hormone and test compound.
Basal 35S-GTP?S binding was defined by 10mM HEPES, pH 7.4 buffer
containing 30% DMSO. The reaction was terminated by addition of 50 µl of
termination buffer containing 25 mM HEPES, pH7.5, 20 mM MgCI2, protease
inhibitors, 100 µM GDP, 100 uM GTP, 5 mM 2-mercaptoethanol with
detergents (0.5 % digitonin, 0.2% sodium deoxycholate, and 0.5% NP-40).
The membranes were solubilized for 30 minutes at 25°C. The 35S-GTP?S
bound Gas protein was immunoprecipitated using anti-Gas (0.5 µg) that linked
to anti-rabbit IgG protein A conjugated SPA. Bound radioactivity was
measured in a Topcount [Packard]. Non-specific 35S-GTP?S binding was
defined by 35S-GTP?S immunoprecipitated by normal rabbit IgG (0.5 µg).
Basal binding (B) = Mean counts/minute (cpm)
immunoprecipitated by anti- Gas.
Non-specific binding (NSB) = Mean cpm immunoprecipitated
by normal rabbit IgG.
Specific basal binding (SB) = B-NSB

Cpm in each well = C
Net cpm in each well (N) = C-NSB
% stimulation = [(N -SB)/SB] x 100%
The procedures described above for the melanocortin MC-4 receptor
were repeated for the melanocortin MC-3 receptor. Following the procedures
described, representative compounds of the present invention were tested for
binding in the MC-4 and / or MC-3 assay, as listed in Table 4.
EXAMPLE 7
Rodent Feeding: Food Intake in Food-Deprived Rats (MC-4)
Male Long-Evans rats (180-200 grams) were housed individually and
maintained on a once-a-day feeding schedule (i.e.10 a.m. until 4 p.m.) for five
days following quarantine to allow the animals to acclimate to feeding on
powdered chow (#5002 PMI Certified Rodent Meal) during the allotted time.
The chow was made available in an open jar, anchored in the cage by a wire,
with a metal follower covering the food to minimize spillage. Water was
available ad-libitum.
Animals were fasted for 18 hours prior to testing. At the end of the
fasting period, animals were administered either a test compound or vehicle.
Vehicle and test compound were administered either orally (5 mL/kg) 60
minutes prior to the experiment, subcutaneously (1 mL/kg) 30 minutes prior to
the experiment, or intraperitoneally (1 mL/kg) 30 minutes prior to the
experiment. Test compounds were administered orally as a suspension in
aqueous 0.5% methylcellulose-0.4% Tween 80, or intraperitoneally as a
solution or suspension in PEG 200; compound concentrations typically ranged
from 1 mg/kg to 100 mg/kg, preferably from 10-30 mg/kg. Food intake was
measured at 2, 4, and 6 hours after administration by weighing the special jar
containing the food before the experiment and at the specified times. Upon
completion of the experiment, all animals were given a one-week washout
period before re-testing.
Following the procedure described above, select compounds of the
present invention were tested to measure the effect on food intake in fasted rats,
as listed in Table 5 and 6.
EXAMPLE 8
Neurite Cell Outgrowth Assay
Cell Culture:
Dissociated hippocampal and cortical cell cultures were established
from embryonic day 18 rat fetuses as described by Mattson, MP., Barger,
S.W., Begley, J, and Mark, R.J., Methods Cell Biol., 1994, 46:087-216. Briefly,
fetuses were removed via cesarean section from pregnant moms (Sprague-
Dawley) and anesthetized with halothane according to the AVMA Panel on
Euthanasia. Pups were decapitated and the brains were removed and placed
in HEPES-buffered Hank's Balanced Salt solution (HBSS; Gibco). The
hippocampi and cortices were dissected out and pooled according to tissue-
type. Tissue was trypsinized for 15 min (1mg/ml trypsin-HBSS; Worthington),
rinsed with fresh HBSS, incubated in trypsin inhibitor (1mg/ml; Sigma) for 5
min, rinsed again with fresh HBSS and then triturated in 1ml fresh HBSS with a
fire-polished glass pipette. Dissociated cells were seeded at 30,000 cells/well
onto poly-D-lysine coated 96-well plates (Collaborative BioScience). Each well
contained 100µl of Eagle's Minimal Essential Media (MEM; Gibco)
supplemented with 26mM NaHCO3 (Sigma), 56mM glucose (Sigma), 15mM
KCI (Sigma), 1mM sodium pyruvate (Sigma), 1.1mM L-glutamine (Sigma), 10%
(v/v) heat-inactivated fetal bovine serum (Hyclone), and 0.001% gentamicin
sulfate (Sigma) (pH 7.4). Cells were allowed to attach for 24h in a humidified
37ºC 5% CO2 incubator before experimental treatment. The culture media was
aspirated and exchanged with fresh media every 3 days.
Assay:
Twenty-four hours after plating, cultures were treated with vehicle
(PBS+0.1% BSA), alpha-melanocyte stimulating hormone (a-MSH) or test
compound (diluted in DPBS). Each treatment condition was run in
quadruplicate. On the third day in culture, the media was aspirated off and
replaced with fresh media and test compound. At one week in culture, the
cells were fixed with 10% phosphate-buffered formalin for 15min, then rinsed
with DPBS (Sigma) and placed in blocking serum for 30 min (horse serum;
1:50 dilution in DPBS; Vector Labs). The cultures were rinsed again with
DPBS and then incubated in primary antibody for 2 hr (microtubule-associated
protein-2 (MAP-2) is a selective marker for dendritic processes; anti-mouse
monoclonal (Chemicon); 1:1000 dilution of MAP-2 in antibody diluent (Zymed)).
Negative control wells were incubated in antibody diluent alone. Background
signal was determined by blank wells (cell-free) incubated with or without
antibody. Cultures were rinsed again with DPBS and then placed in
fluorescein for 1 hr (FITC; anti-mouse IgG; rat adsorbed; 1:50 dilution in DPBS;
Vector Labs). Cultures were rinsed a final time with DPBS and the plates were
then read on a Cytofluor 4000 fluorescence plate reader. Neurite outgrowth
was expressed as percent change from control (vehicle).
Selected compounds of the instant invention were tested in the above
assay with results as listed in Table 7. The data are expressed as percent
change over the vehicle response. All compounds were screened at 50nM. The
abbreviation NA indicates no change / not active; the abbreviation ND indicates a
compound not tested / response not determined.
The data above show that cultures treated with select compounds of the
present invention resulted in a significant increase in neurite outgrowth as
measured by MAP2-FITC immunofluorescence. A comparison between the test
compounds and a-MSH indicates that many of the test compounds were
superior to a-MSH in promoting neurite outgrowth at the concentration tested. In
addition, several of the test compounds displayed selective effects on neurite
outgrowth in hippocampal or cortical cells.
EXAMPLE 9
In vivo Facial Nerve Compression Model
The ability of test compound to provide neuro-protective or neuro-
regenerative effects was investigated in a facial nerve compression model.
Facial nerve motor axons arise exclusively from neurons within the pons in a
well-defined nucleus. Facial nerve compression results in retrograde reactions
proximal to the lesion site and Wallerian degeneration at its distal part, which
causes diminished whisker movement on the lesioned side.
Male Long-Evans rats (150-180 g) were anesthetized with 3-5%
isoflorane for induction and 2% for maintenance during the surgical
procedures. The right facial nerve was exposed and compressed with forceps
for 30 sec at its exit from the stylomastoid foramen. The left facial nerve was
sham-operated and served as an internal control. Nerve compression causes
paralysis of whisker muscles, hence the reduced whisker movement on the
Iesioned side, which is observed immediately after recovery from anesthesia.
The following morphological abnormalities associated with the functional deficit
were observed:
(1) an increase in the number of perineuronal glial cells in the facial
nucleus of the Iesioned side, with the increased observed to peak around D3-6;
(2) a thinner myelin sheath and less myelin basic protein staining in the
compressed facial nerve approximately one week after the lesion;
(3) morphological alterations around the N-M junction and whisker
follicles regions and gradually degeneration of motor neurons in the facial
nuclei.

After recovery from anesthesia, the rats were randomly divided into
groups for dosing with vehicle, aMSH or test compound, with 6 animals per
group. aMSH (s.c., 70µg/each 48hr) was used as a positive control. Test
compounds were dosed p.o. at 20mg/kg bid for 14 days. Restoration of
whisker movement was monitored daily after the operation using two criteria:
(1) frequency of whisker movement on the lesioned side relative to the
the opposite side (sham-operated) which served as the baseline control
(2) semi-quantitative measurements (0 to 4+) on strength of the whisker
muscles, characterized based on observation of the percentage of moved
whiskers, muscle tone of whisker muscles, and the position of the nose.
For all observations, the experimental design was blind to the behavioral
observer.
The results of behavioral assessment showed that both test
compounds, compounds #31 and #84, accelerated the recovery time to restore
whisker muscle movement in the lesioned rats as compared to the vehicle
controls (p as percentage of its own internal control (sham-operated), as listed in Table 8.
EXAMPLE 10
in Vitro Assay: Measurement of regulation of sebaceous lipid synthesis.
STEP A: Preparation of a feeder layer
Semiconfluent cultures of 3T3 mouse fibroblasts (Swiss Albino mouse,
ATCC CCL-92 ) were treated with mitomycin C (4µg/ml) for 3 hours,
trypsinized and seeded at a density of 2.5x105/9.5 cm2 tissue culture plate in
Dulbeccos Minimal Essential Medium (DMEM) containing 10% Colorado Calf
Serum, PNC (100U/ml), STM (100ug/ml), L glutamine (0.3mg/ml), sodium
pyruvate (1mM) and nonessential amino acids (100µM). The cells were
incubated at 37°C for 24 hours prior to their use as a feeder layer for
sebocytes.
STEP B: Isolation of human sebocytes
Human sebocytes were isolated from Dermatome shavings of
postoperative pieces of human skin at 0.4-0.8mm depths (this part of the skin
was previously shown to be enriched in sebaceous glands). Shavings so
obtained were treated with 1% Dispase in Iscoves medium containing 10%
serum for 20 min at 37°C. The tissue was then placed in 0.3% trypsin/1%
EDTA in Phosphate Buffered Saline (PBS) for 10 minutes at 37°C. Following
this incubation the cells were gently scraped from the tissue in Growth medium
(GM) containing OMEM/F12 media mixture (3:1), supplemented with 8% heat
inactivated PBS, 2% heat inactivated human serum (HS), 1mM sodium
pyruvate, epidermal growth factor (10ng/ml), insulin (10µg /ml), hydrocortisone
(0.4µg/ml) and +/-cholera toxin (100µg/ml), L-glutamine and antibiotics.
Cells so obtained were filtered through nylon mesh (100µ pore size),
centrifuged at 750 RPM, re-suspended in GM and counted.
Resultant cells from the above isolation procedure were plated on the
3T3 feeder layers at 2x105/9.5cm2 in growth medium and maintained at 37°C
and 5% CO2 for 3days (Phase I). Following the initial growth period they were
transferred to a transition medium (TM) that consisted of DMEM/F12 media
supplemented with 1mM sodium pyruvate, insulin (10µg/ml), transferrin
(6.7µg/ml) and selenium (5.5µg/ml) (ITS), 2% heat inactivated FBS and 2%
heat inactivated human serum as well as +/- cholera toxin (ch.t.) (100µg/ml), L-
glutamine and antibiotics (Phase II). Three days later the cells were changed
to differentiation medium (DM), DMEM/F12 supplemented with ITS, 3,3',5-
triido-L-thyronine sodium(3nM), 1 %(v/v) trace element mix and the choice of
differentiation agent i.e. bovine pituitary extract (10µg/ml). This medium was
changed every 3 days (Phase III).
STEP D: Testing Stimulators or Inhibitors of Sebocyte Differentiation and Lipid
Production
Hormones, mixture of hormones i.e. bovine pituitary extract or
compounds to be tested were added to the culture at the beginning of phase
III. Two criteria were used to evaluate the effect of these materials on
sebaceous cultures: 1) visual observations and 2) evaluation of sebaceous lipid
accumulation and synthesis. The evaluation of lipid accumulation completed
using the Nile red method. This method relies on visualization of neutral lipids
by Nile red and quantitation by reading of fluorescence at 535 nm excitation,
580 nm emission using a plate reader. The lipid synthesis was evaluated by
radioactive labeling using 14C acetate and quantified by Bio Rad
Phosphoimager (Molecular Imager, FX) using 4.1 Software.
STEP E: Visual observations & Nile Red evaluation of lipid accumulation
Morphological evaluation of lipid accumulation was easily recognized
since the cells enlarged and displayed lipid granules that in bright field light
microscopy appeared as yellowish circles in the cells. Quantitation of
accumulation/inhibition of neutral lipids in sebocytes was accomplished by Nile
red binding assay. Briefly, following exposure of sebocytes to test compounds,
the cells were allowed to interact with 1µM Nile red in Hanks buffered saline

solution containing DMSO and Pluronic F127. After 4 hours of incubation,
washing and incubation overnight, the fluorescence was read at 535 excitation
and 580 emission using a fluorescence plate reader. To determine whether
the compounds had an inhibitory effect on cell growth, cell counts were
performed.
Following the procedure described above, select compounds of the
present invention were tested for visual and Nile red evaluation of lipid
accumulation, with results as listed in Table 9.
STEP F: Evaluation of sebaceous lipid synthesis by sebaceous cells
At day 11 of the culture, sebocytes were labeled with 14C acetate at a
final concentration of 2µCi/ml for 24 hours in serum free culture medium. The
cells were than scraped from plates and frozen at -80°C in glass vials. Lipid
extraction was completed using the Bligh- Dyer method (Bligh, E.G. and Dyer,
W.J., Can. J. Biochem. Physiol., 1959, 37, pp 911-916) with slight modification
as detailed herein. Briefly, cells were homogenized in a 2:1 chloroform-
methanol mixture, in the presence of KCI. The organic phase was removed
from the mixture, the separated lipids were dried under argon and spotted to
high performance thin layer chromatography (HPTLC) plates. The plates were
developed by three separate mobile phases. The first was hexane (to the top
of the plate), followed by toluene (to the top) and finally a 70:30:1 mixture of
hexane : ether: acetic acid (half way up the plate-10cm). The plates were than
exposed to radiographic film for visualization of radioactive lipid species. For
visualization of unlabeled lipids the plates were sprayed with 8% cupric acid
and charred on a hot plate. Quantitation of the results was done by Image Pro
Plus 3.0 (Media Cybernetics, Silver Springs, MD).
Following the procedure described above, select compounds of the
present invention were tested for their inhibitory effect on differentiation of
human sebocytes. Visually, the stained cell cultures treated with compound
#74 showed complete disappearance of lipid granules in sebocyte cultures
following a seven day treatment. The same cells examined for lipid synthesis
revealed inhibition of squalene, cholesterol esters and wax esters, as
measured by radioactively labeled lipids separated by HPTLC. The cell panels
were quantified by measurement of the intensity of bands using Image Pro
Plus (version 5.0). % inhibition was calculated based on the difference
between treated samples and vehicle treated controls, with results as listed in
Table 10.
In Vivo Evaluation of the Test Compound Effect on Sebum Production:
Human Skin - SCID Mouse Chimera Model
Severe combined immunodeficient mice (SCID) provide an invaluable
model for skin xenografting. These animals are devoid of both T and B cell
immunity. Human skin grafts in SCID mice retain human cellular tissue
components including skin immune cells, i.e. Langerhans cells, macrophages
and lymphocytes and also part of the engrafted endothelium ( Kaufman R.,et
al., Exp. Dermatol. 1993: 2: 209-216,1993). These properties allow for the
study of physiological and/or pathological responses of human skin cells to a
test compound.
STEP A: Method of transplantation:
C.B-17 scid/scid mice (Taconic, Germantown, N.Y.) were used for
grafting at 5-6 weeks of age. Full thickness human facial skin was shaved to
-0.4 mm using a Forman Dermatome. The skin shavings were washed 3
times in antibiotics and antimycotics (penicillin, streptomycin, fungizone) (Life
technologies) in Dulbecco's Modified Eagle Medium (DMEM, Life
Technologies). Eliptical skin - 2.0-2.5x1.0-1.5 was grafted onto the prepared
graft bed and sutured using 4.0 silk. During surgical procedures the mice were
anaesthetized using a mixture of Ketaset (0.16 mg/g body weight) and Rompun
(8.0 µg/g body weight).
It is well accepted that the wound healing process of the transplanted
skin in the SCID mouse takes one month, at which time the human skin can be
used for experimental purposes. We also found that there is a gradual
regeneration of sebaceous glands in the transplanted human skin and that
these glands are fully regenerated and secrete sebum at 7 weeks as shown by
Sebutape and histo-morphometry. Maximum size of the glands was observed 3
month post transplantation. The glands retained their capability to produce
human specific sebum and the glandular tissue expressed human specific
markers including MC5-R. Since glands reached their maximum size at 2-3
month post transplantation the effects of inhibitors of sebum secretion was
tested at this point.

STEP B: Method of treatment:
Mice at 2-3 month post transplantation with human facial skin were used
for the studies. The graft area was treated with the test compound at the
desired concentration(s) dissolved in polyethylene glycol-ethanol (20 µl/2 cm2).
Controls were treated with vehicle alone. The test compounds were applied
daily, excluding weekends. Sebum secretion was determined using Sebutape
at 15 days and 30 days following treatment.
STEP C: Termination of the experiment:
The termination of the experiment was determined by preliminary clinical
evaluation of sebum production using SEBUTAPE. At this time human skin
grafts were excised and representative samples were collected for histological
evaluation. More particularly, 2mm punch biopsies were prepared and used for
evaluation of lipid synthesis and total lipid accumulation in the treated tissues.
STEP D: Evaluation of lipid synthesis and total lipid accumulation in the
examined tissues.
The collected 2mm punch biopsies were placed individually into 96 well
plates in Krebs buffer and labeled with 10µCi of 14C acetate for 3 hours.
Following this labeling period the samples were washed in medium and 5
biopsies pooled, weighed, and used for lipid extraction. The lipid extraction and
analysis by HPTLC was the same as described for tissue culture derived cells.
Following the procedure described above, compound #74 of the present
invention was tested for the inhibition of sebaceous gland activity following 11
day treatment of human skin transplanted to the SCID mouse.
Visual evaluation of the sebaceous gland following topical treatment with
0.1% solution of compound #74 resulted in visible shrinkage of the sebaceous
gland and down regulation of sebaceous lipids. Topical treatment for 15 days
with 0.05% and 0.005% solutions was not sufficient to down regulate the lipids.
Numerical evaluation of the inhibition of human sebaceous lipids for these
cells was analyzed using HPTLC, with results as listed in Table 11,12 and 13.
%lnhibition numbers are listed relative to control.
As shown in Tables 12 and 13 above, both total sebaceous lipid
accumulation and de novo synthesis of sebaceous lipids was significantly
decreased following 30 days of topical treatment with an 0.05% and 0.01%
solution of compound #74.

EXAMPLE 12
Oral Formulation
As a specific embodiment of an oral composition, 100 mg of the
compound #74 of Example 2 is formulated with sufficient finely divided lactose
to provide a total amount of 580 to 590 mg to fill a size O hard gel capsule.
EXAMPLE 13
Topical Formulations
A: Microemulsion
As a specific embodiment of a microemulsion composition the following
components are blended, with heating as need:
To the blended mixture is then slowly added water (42.9 parts by
weight), with mixing as necessary, to yleld the emulsion.
B: Hvdroalcoholic gel
As a specific embodiment of a hydroalcoholic gel composition the
polypropylene glycol (10 parts by weight), butylene glycol (10 parts by weight),
benzyl alcohol (2 parts by weight), EDTA (0.05 parts by weight) and BHT (0.05
parts by weight) are mixed with water (74.85 parts by weight total). The
mixture is blended until all the components are dissolved. Carbomer (e.g.
Carbopol 934P from Goodrich) (3 parts by weight) is then slowly added with
constant turning to yleld a gel. Compound #74 (0.05 parts by weight) is then
dispersed into the gel with mixing. The gel pH is adjusted to about pH 3-4.
C: Anhydrous Gel
As a specific embodiment of an anhydrous gel isopropanol (20 parts by
weight) is added to butylene glycol (20 parts by weight). BHT (0.05 parts by
weight) and benzyl alcohol (1.0 parts by weight) are then added to the
isopropanol/butylene glycol mixture. To the resulting mixture is then added
Cyclotetrasiloxane (D4) and Organopolysiloxane-11 (e.g. Gransil GSM Gel from
Grant Industries) (58.85 parts by weight) with continuous mixing. Compound
#74 (0.1 parts by weight) is micronized by and dispersed into the gel with
continuous mixing until uniformly distributed.
D: Cream
As a specific embodiment of an o/w (oil/water) cream, the following
components are mixed in the amounts (parts by weight) as noted. The final
mixture is adjusted to about pH 2 with hydrochloric acid.
While the foregoing specification teaches the principles of the present
invention, with examples provided for the purpose of illustration, it will be
understood that the practice of the invention encompasses all of the usual
variations, adaptations and/or modifications as come within the scope of the
following claims and their equivalents.
We Claim:
1. A compound of the formula (II), for use in the treatment of a
disorder mediated by the melanocortin receptor, in particular the
melanocortin-3 receptor, the melanocortin-4 receptor or the
melanocortin-5 receptor, such as a metabolic disorder, a CNS
disorder or a dermatological disorder, for instance obesity,
impaired oral glucose tolerance, elevated blood glucose level, type
II diabetes, Syndrome X, diabetic retinopathy, an acute
neurodegenerative disorder, a chronic neurodegenerative disorder,
a plexopathy, male erectile dysfunction, dry eyes, acne, dry skin,
aged skin, seborrheic dermatitis, rosacea, excessive ear wax,
meibomian gland disorder, pseudofolliculitis, a yeast infection,
dandruff, hiradenitis suppurativa, ocular rosacea or an eccrine
gland disorder, in particular obesity, impaired oral glucose
tolerance, elevated blood glucose level, type II diabetes Syndrome
X, acne, dry skin or seborrheic dermatitis,
wherein
R1 is heteroaryl, heteroaryl-alkyl, heterocycloalkyl,
heterocycloalkyl-alkyl, cycloalkyl, cycloalkyl-alkyl, substituted
aryl or substituted aralkyl,
Wherein the heteroaryl, heteroaryl-alkyl, heterocycloalkyl,
heterocycloalkyl-alkyl, cycloalkyl or cycloalkyl-alkyl group is
optionally independently substituted with one or more halogen,
hydroxyl, alkyl, alkoxy, halogenated alkyl, halogenated alkoxy,
amino, alkylamino or di(alkyl)amino substituents,
wherein the aryl or aralkyl group is independently substituted with
one or more halogen, hydroxy, alkyl, alkoxy, halogenated alkyl,
halogenated alkoxy, amino, alkylamino or di(alkyl)amino
substituents; and either
R is heteroaryl, heteroaryl-alkyl, heterocycloalkyl,
heterocycloalkyl-alkyl, cycloalkyl, cycloalkyl-alkyl, substituted
aryl or substituted aralkyl,
wherein the heteroaryl, heteroaryl-alkyl, heterocycloalkyl,
heterocycloalkyl-alkyl, cycloalkyl or cycloalkyl-alkyl group is
optionally independently substituted with one or more halogen,
hydroxyl, alkyl, alkoxy, halogenated alkyl, halogenated alkoxy,
amino, alkylamino or di(alkyl)amino substituents,
wherein the aryl or aralkyl group is independently substituted with
one or more halogen, hydroxyl, alkyl, alkoxy, halogenated alkyl,
halogenated alkoxy, amino, alkylamino or di(alkyl)amino; and
R4 is aryl, aralkyl, heteroaryl, heterocycloalkyl or cycloalkyl-alkyl,
wherein the ary], aralkyl, heteroaryl, heterocycloalkyl or
cycloalkyl-alkyl group is optionally independently substituted with
one or more halogen, hydroxyl, alkyl, alkoxy, halogenated alkyl,
halogenated alkoxy, amino, alkylamino or di(alkyl)amino
substituents,
or
R2 is aryl, aralkyl, heteroaryl, heteroaryl-alkyl, heterocycloalkyl,
heterocycloalkyl-alkyl, cycloalkyl or cycloalkylalkyl,
wherein the aryl, aralkyl, heteroaryl, heteroaryl-alkyl,
heterocycloalkyl, heterocycloalkyl-alkyl, cycloalkyl or cycloalkyl-
alkyl group is optionally independently substituted with one or
more halogen, hydroxyl, alkyl, alkoxy, halogenated alkyl,
halogenated alkoxy, amino, alkylamino or di(alkyl)amino
substituents; and
R4 is heteroaryl, heterocycloalkyl, cycloalkyl-alkyl, substituted
aryl or substituted aralkyl,
wherein the heteroaryl, heterocycloalkyl or cycloalkyl-alkyl group
is optionally independently substituted with one or more halogen,
hydroxyl, alkyl, alkoxy, halogenated alkyl, halogenated alkoxy,
amino, alkylamino or di(alkyl)amino substituents,
wherein the aryl or aralkyl group is independently substituted with
one or more halogen, hydroxyl, alkyl, alkoxy, halogenated alkyl,
halogenated alkoxy, amino, alkylamino or di(alkyl)amino
substituents, and pharmaceutically acceptable salts or solvates
thereof,
wherein
alkyl is C1-8alkyl;
cycloalkyl is C3-8alkyl;
aralkyl is arylC1-4alkyl;
heteroaryl is:(i) a five or six membered monocyclic aromatic ring
structure containing at least one heteroatom selected from the
group consisting of O, N and S, optionally containing one to three
additional heteroatoms independently selected from the group
consisting of O, N and S, and attached at any carbon atom of the
ring such that the result is a stable structure; or (ii) a nine or ten
membered bicyclic aromatic ring structure containing at least one
heteroatom selected from the group consisting of O, N and S,
optionally containing one to four additionally heteroatoms
independently selected from the group consisting of O, N or S, and
attached at any carbon atom of the ring such that the result is a
stable structure; and
heterocycloalkyl is: (i) a five to seven membered monocyclic,
saturated, partially unsaturated or partially aromatic ring structure
containing at least one heteroatom selected from the group
consisting of O, N and S, optionally containing one to three
additional heteroatoms independently selected from the group
consisting of O, N and S and attached at any carbon atom of the
ring such that the result is a stable structure; or (ii) a nine or ten
membered saturated, partially unsaturated or partially aromatic
bicyclic ring system containing at least one hetroatom selected
from the group consisting of O, N and S, optionally containing one
to four additional heteroatoms independently selected from the
group consisting of O, N and S, and attached at any carbon atom of
the ring such that the result is a stable structure.
2. A compound for use as claimed in claim 1, and the "either" option
is selected, wherein
R1 is substituted aryl or substituted aralkyl,
wherein the aryl or aralkyl is independently substituted with one or
more halogen, hydroxyl, alkyl, alkoxy, trihalomethyl,
trihalomethoxy, amino, alkylamino or di(alkyl)amino substituents;
R is substituted aryl or substituted aralkyl,
wherein the aryl or aralkyl is independently substituted with one or
more halogen, hydroxyl, alkyl, alkoxy, trihalomethyl,
trihalomethoxy, amino, alkylamino or di(alkyl)amino substituents;
R4 is aryl, aralkyl or heteroaryl,
Wherein the aryl, aralkyl or heteroaryl group is optionally
independently substituted with one or more halogen, hydroxyl,
alkyl, alkoxy, trihalomethyl, trihalomethoxy, amino, alkylamino or
di(alkyl)amino substituents,
and pharmaceutically acceptable salts or solvates thereof.
3. A compound for use as claimed in claim 1 and the "or" option is
selected, wherein
R1 is substituted aryl or substituted aralkyl,
wherein the aryl or aralkyl group is independently substituted with
one or more halogen, hydroxyl, alkyl, alkoxy, halogenated alkyl,
halogenated alkoxy, amino, alkylamino or di(alkyl)amino
substituents;
R2 is aryl or aralkyl,
wherein the aryl or aralkyl group is optionally independently
substituted with one or more halogen, hydroxyl, alkyl, alkoxy,
halogenated alkyl, halogenated alkoxy, amino, alkylamino or
di(alkyl)amino substituents;
R4 is substituted aryl or substituted aralkyl,
wherein the aryl or aralkyl group is independently substituted with
one or more halogen, hydroxyl, alkyl, alkoxy, halogenated alkyl,
halogenated alkoxy, amino, alkylamino or di(alkyl)amino
substituents;
and pharmaceutical acceptable salts or solvates thereof.
4. A compound for use as claimed in claim 2 wherein
R1 is substituted aryl,
wherein the aryl group is optionally substituted with one to two
halogen, alkyl or alkoxy substituents;
R2 is substituted aryl,
wherein the aryl group is substituted with one to two alkyl or
alkoxy substituents;
R4 is aryl or substituted aryl,
wherein the substituents on the aryl are independently one to
halogen, alkyl or alkoxy substituents;
and pharmaceutically acceptable salts or solvates thereof.
5. A compound for use as claimed in claim 4 wherein
R1 is 2-methoxyphenyl;
R2 is 2-methoxyphenyl;
R4 is phenyl, 4-methoxyphenyl, 2,6-difluorophenyl, 3,5-
difluorophenyl or 2-chloro-6-methylphenyl;
and pharmaceutical acceptable salts or solvates thereof.
6. A compound for use as claimed in claim 5 whichis [2-(2-
methoxyphenyl)-3-(2-methoxyphenyl )-2H- [1,2,4]-thiadiazol-5 -
ylidene]-phenylamine or a pharmaceutical acceptable salt or
solvate thereof.
7. A compound as claimed in any one of claims 1 to 6, and
pharmaceutically acceptable salts or solvates thereof, provides that
when the "either" or the "or" option in claim 1 is selected, when R1
is aryl substituted with one halogen and R4 is aryl substituted with
one halogen, then R2 is not morpholinyl
when the "or" option in claim 1 is selected, when R1 is
methylphenyl and R4 is methoxyphenyl, then R2 is aralkyl,
heteroaryl, heteroaryl-alkyl, heterocycloalkyl, heterocycloalkyl-
alkyl, cycloalkyl, cycloalkyl-alkyl or substituted aryl, wherein the
aralkyl, heteroaryl, heterocycloalkyl or cycloalkyl group is
optionally independently substituted with one or more halogen,
hydroxyl, alkyl, alkoxy, halogenated alkyl, halogenated alkoxy,
amino, alkylamino or di(alkyl)amino substituents and wherein the
aryl group is independently substituted with one or more halogen,
hydroxyl, alkyl, alkoxy, halogenated alkyl, halogenated alkoxy,
amino, alkylamino or di(alkyl)amino substituents;
in the definitions given in claim 3, wherein R1 is methylphenyl and
R4 is methoxyphenyl, then R2 is aralkyl or substituted aryl, wherein
the aralkyl group is optionally independently substituted with one
or more halogen, hydroxyl, alkyl, alkoxy, halogenated alkyl,
halogenated alkoxy, amino, alkylamino or di(alkyl)amino
substituents, wherein the aryl group is independently substituted
with one or more halogen, hydroxyl, alkyl, alkoxy, halogenated
alkyl, halogenated alkoxy, amino, alkylamino or di(alkyl)amino
substituents; and
the salt is not 2-p-bromophenyl-3-morpholino-5-p-
methoxyphenylimino-1,2,4-thiadiazoline hydrobromide.
8. A pharmaceutical composition comprising a compound of the
formula II as claimed in any one of claims 1 to 6 and a
pharmaceutically acceptable carrier.

The present invention is directed to novel 1,2,4-
thiadiazole derivatives of the formula (II) wherein R1, R2-
and R4, are ring containing radicals, useful as agonists or
antagonists of the melanocortin receptor. More particularly,
the compounds of the present invention are useful for the
treatment of metabolic CNS and dermatological disorders.

Documents:

606-KOLNP-2004-FORM-27.pdf

606-kolnp-2004-granted-abstract.pdf

606-kolnp-2004-granted-assignment.pdf

606-kolnp-2004-granted-claims.pdf

606-kolnp-2004-granted-correspondence.pdf

606-kolnp-2004-granted-description (complete).pdf

606-kolnp-2004-granted-examination report.pdf

606-kolnp-2004-granted-form 1.pdf

606-kolnp-2004-granted-form 18.pdf

606-kolnp-2004-granted-form 2.pdf

606-kolnp-2004-granted-form 26.pdf

606-kolnp-2004-granted-form 3.pdf

606-kolnp-2004-granted-form 5.pdf

606-kolnp-2004-granted-reply to examination report.pdf

606-kolnp-2004-granted-specification.pdf


Patent Number 233878
Indian Patent Application Number 606/KOLNP/2004
PG Journal Number 16/2009
Publication Date 17-Apr-2009
Grant Date 16-Apr-2009
Date of Filing 11-May-2004
Name of Patentee ORTHO-MCNEIL PHARMACEUTICAL CORPORATION
Applicant Address ROUTE 202, RARITAN, NJ
Inventors:
# Inventor's Name Inventor's Address
1 MAGDALENA EISINGER 30 PINE TERRACE, DEMAREST, NJ 07627
2 LOUIS J. FITZPATRICK 205 HEATHERFIELD DRIVE, SAUDERTON, PA 19964
3 DANIEL H. LEE 361 DUTTON ROAD, SUDBURY, MA 01776
4 KEVIN PAN 409 GREENE LANE, PHOENIXVILLE, PA 19460
5 CARLOS PLATA-SALAMAN 1313 SQUIRE DRIVE, AMBLER, PA 19002
6 ALLEN B. REITZ 109 GREENBRIAR ROAD, LANDSDALE, PA 19466
7 VIRGINIA L. SMITH-SWINTOSKY 3163 LINE LEXINGTON ROAD, HATFIELD, PA 19440
8 BOYU ZHAO 105 WEYMOUTH CIRCLE, LANSDALE, PA 19446
PCT International Classification Number C07D 285/08, 417/12
PCT International Application Number PCT/US2002/35340
PCT International Filing date 2002-11-04
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 60/337,927 2001-11-08 U.S.A.