WO2010031114A1

WO2010031114A1 - Novel receptor hetero-dimers/-oligomers

Info

Publication number: WO2010031114A1
Application number: PCT/AU2009/001219
Authority: WO
Inventors: Kevin Pfleger; Ruth Marie Seeber; Heng Boon See; Matthew Blake Dalrymple
Original assignee: Dimerix Bioscience Pty Ltd
Priority date: 2008-09-18
Filing date: 2009-09-16
Publication date: 2010-03-25

Abstract

A hetero-dimeric or hetero-oligomeric receptor, comprising at least one metabotropic glutamate receptor subunit associated with at least one orexin receptor subunit.

Description

NOVEL RECEPTOR HETERO-DIMERS/-OLIGOMERS Field of the Invention

The present invention relates to a hetero-dimeric or hetero-oligomeric receptor, comprising at least one metabotropic glutamate receptor subunit associated with at least one orexin receptor subunit.

Background Art

Proteins do not act in isolation in a cell, but in stable or transitory complexes, with protein- protein interactions being key determinants of protein function (Auerbach et al., (2002), Proteomics 2:611-623). Furthermore, proteins and protein complexes interact with other cellular components like DNA, RNA and small molecules. Understanding both the individual proteins involved in these interactions and their interactions are important for a better understanding of biological processes.

The primary physiological function of L-glutamate (GIu) reported by Nicoletti (Nicoletti, F et al (2007) Metabotropic glutamate receptors: beyond the regulation of synaptic transmission. Psychoneuroendocrinology. VoI 32; S40-S45) is as a fundamental regulator of excitatory synaptic activity in the central nervous system. For example, the metabotropic glutamate receptor type-5 (mGluR5) is highly expressed in specific regions of the hippocampus, basal ganglia and amygdala, regions of the brain that have been implicated in processes of emotion and motivation (Palucha A., and Pile, A., (1994) Metabotropic glutamate receptor ligands as possible anxiolytic and antidepressant drugs. Pharmacology and Therapeutics, Vo1 115; 116-147).

In addition to its role in synaptic transmission, a number of reports have suggested a role for metabotropic glutamate pathways in the perception of pain (Neugebauer, V. (2002) Metabotropic glutamate receptors - important modulators of nociception and pain behaviour. Pain, Vol.98; 1-8), anxiety and depression (Swanson, C, et al., (2005) Metabotropic glutamate receptors as novel drug targets for anxiety and stress disorders. Nature Reviews - Drug Discovery, Vol.4; 131-144), and the development of addictive behaviour (Lea IV, P. and Faden, A. (2006) Metabotropic glutamate receptor subtype 5 antagonists MPEP and MTEP. CNS Drug Reviews, Vol.12; 149-166). Narcolepsy with cataplexy is associated with low or undetectable levels of cerebrospinal fluid (CSF) orexin A levels in about 90% of patients (Baumann and Bassetti (2005) Sleep Medicine Reviews 9, 253-268). Mutations of the orexin receptor 2 gene lead to familial canine narcolepsy and a loss of orexin neurons and low CSF orexin A were observed with sporadic canine narcolepsy. Neurological disorders arising from acute traumatic brain injury, Guillain-Barre syndrome and advanced Parkinson's syndrome may also be linked with low or undetectable levels of CSF orexin A levels in some instances. Sakurai has postulated a role for the orexin system in feeding and energy homeostasis as the activity of orexin neurons is inhibited by glucose and leptin, and stimulated by ghrelin, a stomach derived peptide which promotes feeding. This may have implications for the treatment of obesity (Sakurai (2005) Sleep Medicine Reviews 9, 231-241).

The preceding discussion is intended only to facilitate an understanding of the invention. It should not be construed as in any way limiting the scope or application of the following description of the invention, nor should it be construed as an admission that any of the information discussed was within the common general knowledge of the person skilled in the appropriate art at the priority date.

Disclosure of the Invention

The inventors have discovered that the orexin receptor and the metabotropic glutamate receptor associate. This has important implications regarding therapies for ailments associated with either receptor.

Recent studies have shown that GPCRs may not only act as monomers but also as homo- and hetero-dimers which causes altered ligand binding, signalling and endocytosis (Rios et al. (2000) Pharmacol. Ther. 92:71-87). The effect of drugs acting as agonists or antagonists of a specific receptor may therefore depend on the binding partners of this receptor. It may be desirable to limit the effect of a drug to a cellular response mediated by a specific receptor dimer. As Milligan (Milligan G. (2006), Drug Discovery Today 11 :541-549) observes, while homo-dimerisation and -oligomerisation have limited implications for the drug discovery industry, "differential pharmacology, function and regulation of GPCR hetero-dimers and -oligomers suggest means to selectively target GPCRs in different tissues and hint that the mechanism of function of several pharmacological agents might be different in vivo than anticipated from simple ligand screening programmes that rely on heterologous expression of a single GPCR".

The phrase "metabotropic glutamate receptor" or "mGluR" is to be understood to at least include the G protein-coupled receptor metabotropic glutamate receptor 5 (mGluRδ) analogous to that activated by L-glutamate that is, among other roles, a fundamental regulator of excitory synaptic activity in the central nervous system (Nicoletti, F. et al, (2007) Metabotropic glutamate receptors: beyond the regulation of synaptic transmission. Psychoneuroendocrinology 32:S40-S45). The phrase "metabotropic glutamate receptor" or "mGluR" is also to be understood to mean the metabotropic glutamate receptor 1 (mGluRI ), the second member of Group 1 of metabotropic glutamate receptors (Nicoletti, F. et al, (2007) Metabotropic glutamate receptors: beyond the regulation of synaptic transmission. Psychoneuroendocrinology 32:S40-S45). The phrase "metabotropic glutamate receptor" or "mGluR" is also to be understood to mean the metabotropic glutamate receptors 2 (mGluR2) and 3 (mGluR3) which both belong to the second, inhibitory group of metabotropic glutamate receptors (Nicoletti, F. et al, (2007) Metabotropic glutamate receptors: beyond the regulation of synaptic transmission. Psychoneuroendocrinology 32:S40-S45). The phrase "metabotropic glutamate receptor" or "mGluR" is to be further understood to include metabotropic glutamate receptors 4 (mGluR4), 6 (mGluRΘ), 7 (mGluR7) and 8 (mGluRδ) which comprise the third group of metabotropic glutamate receptors (Nicoletti, F. et al, (2007) Metabotropic glutamate receptors: beyond the regulation of synaptic transmission. Psychoneuroendocrinology 32:S40-S45). Finally, the phrase, "metabotropic glutamate receptor" or "mGluR" is to be further understood to include newly discovered mGluR family members. Throughout the examples, metabotropic glutamate receptor and acronym mGluR refers to mGluR5. The phrase "orexin receptor" or "OxR" is to be understood to mean either orexin receptor 1 (OxR1 ; OXR1 ; OX₁R; hypocretin-1 -receptor; hctri) or orexin receptor 2 (OxR2; OXR2; OX₂R; hypocretin-2-receptor; hctr2), being G protein-coupled receptors analogous to those described by Sakuri et al. to be activated by orexin A (OxA; hypocretin-1 ; Hcrt-1) and orexin B (OxB; hypocretin-2; Hcrt-2) (Sakuri et al (1998) Cell 92, 573-585). "Orexin receptor" or "OxR" is to be further understood to include newly discovered orexin receptor family members.

In a first aspect of the invention, there is provided a hetero-dimeric or hetero-oligomeric receptor, comprising at least one metabotropic glutamate receptor subunit associated with at least one orexin receptor subunit. In a second aspect of the invention, there is provided a method for the treatment of a patient suffering from an orexin-related ailment by administering a therapeutically effective amount of a metabotropic glutamate receptor-related compound.

In one embodiment, the metabotropic glutamate receptor-related compound is selective for the metabotropic glutamate receptor relative to the orexin receptor. In one embodiment, the metabotropic glutamate receptor-related compound is coadministered with an orexin receptor-related compound.

In a third aspect of the invention, there is provided a method for the treatment of a patient suffering from a glutamate-related ailment by administering a therapeutically effective amount of an orexin receptor-related compound. In one embodiment, the orexin receptor-related compound is selective for the orexin receptor relative to the metabotropic glutamate receptor.

In one embodiment, the orexin receptor-related compound is co-administered with a metabotropic glutamate receptor-related compound. In a fourth aspect of the invention, there is provided a method for the manufacture of a medicament for the treatment of a patient suffering from an orexin-related ailment comprising use of a therapeutically effective amount of a metabotropic glutamate receptor-related compound.

In one embodiment, the medicament contains an orexin receptor-related compound. In a fifth aspect of the invention, there is provided a method for the manufacture of a medicament for the treatment of a patient suffering from a glutamate-related ailment comprising use of a therapeutically effective amount of an orexin receptor-related compound.

In one embodiment, the medicament contains a metabotropic glutamate receptor-related compound.

In a sixth aspect of the invention, there is provided a method for the treatment of a patient suffering from an orexin-related ailment by administering a therapeutically effective amount of a glutamate-selective binding agent, or fragment thereof.

In a seventh aspect of the invention, there is provided a method for the treatment of a patient suffering from a glutamate-related ailment by administering a therapeutically effective amount of an orexin-selective binding agent, or fragment thereof.

In an eighth aspect of the invention, there is provided a method for the treatment of a patient suffering from a glutamate-related ailment or an orexin-related ailment comprising administering a therapeutically effective amount of a metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/-oligomer, such as a selective allosteric modulator.

In a ninth aspect of the invention, there is provided the use of a therapeutically effective amount of a metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/-oligomer, such as a selective allosteric modulator, for the manufacture of a medicament for the treatment of a patient suffering from a glutamate-related ailment or an orexin-related ailment. In a tenth aspect of the invention, there is provided a method for screening a test compound for potential therapeutic activity against an orexin-related ailment using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) exposing the test compound to a metabotropic glutamate receptor; b) assessing whether and/or the extent to which the activity of the metabotropic glutamate receptor is modulated compared to activity of the metabotropic glutamate receptor in the absence of the test compound; said modulation being indicative of potential therapeutic activity against the orexin- related ailment.

In an eleventh aspect of the invention, there is provided a method for screening a test compound for potential therapeutic activity against a glutamate-related ailment using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) exposing the test compound to an orexin receptor; b) assessing whether and/or the extent to which the activity of the orexin receptor is modulated compared to activity of the orexin receptor in the absence of the test compound; said modulation being indicative of potential therapeutic activity against the glutamate- related ailment.

In a twelfth aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective activity using a detector capable of detecting changes in receptor activity, the method comprising the step of: determining whether, and/or the extent to which, the test compound interacts with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor compared to whether, and/or the extent to which the test compound interacts with the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater affinity and/or potency and/or efficacy when interacting with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

In a thirteenth aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective activity using a detector capable of detecting changes in receptor activity, the method comprising the step of: determining whether, and/or the extent to which, the test compound interacts with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor compared to whether, and/or the extent to which the test compound interacts with the metabotropic glutamate receptor in the absence of the orexin receptor; such that a test compound that exhibits greater affinity and/or potency and/or efficacy when interacting with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer.

In a fourteenth aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective antagonism or selective partial agonism or selective negative allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of:

a) determining whether, and/or the extent to which, the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the metabotropic glutamate receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether, or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater antagonistic, partial agonistic or negative allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero- dimer/-oligomer.

In a fifteenth aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective antagonism, selective partial agonism or selective negative allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of:

a) determining whether, and/or the extent to which, the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the orexin receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b). detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether, or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater antagonistic, partial agonistic or negative allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer.

In a sixteenth aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective inverse agonism using a detector capable of detecting changes in receptor activity, the method comprising the steps of:

a) determining whether, and/or the extent to which, the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising a constitutively active metabotropic glutamate receptor; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether, or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater inverse agonistic properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate / orexin receptor hetero-dimer/-oligomer.

In a seventeenth aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer inverse agonism using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising a constitutively active orexin receptor; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether, or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater inverse agonistic properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

In an eighteenth aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective positive allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the metabotropic glutamate receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting an increase in the signal as a determination of whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater positive allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero- dimer/-oligomer.

In a nineteenth aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective positive allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the orexin receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b) detecting an increase in the signal as a determination of whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater positive allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero- dimer/-oligomer. In the aforementioned methods of the invention, the step of determining whether, and/or the extent to which, the test compound interacts with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor; and/or the step of determining whether, and/or the extent to which, the test compound interacts with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor may be performed by way of one or more of the methods described in the applicant's co-pending international patent application "Detection System and Uses Therefor" PCT/AU2007/001722 (published as WO 2008/055313).

In a twentieth aspect of the invention, there are provided selective agonists and/or selective antagonists and/or selective inverse agonists and/or selective allosteric modulators of the metabotropic glutamate receptor/orexin receptor hetero-dimer/- oligomer.

In a twenty-first aspect of the invention, there is provided a cell, or fraction of a cell, in which both a metabotropic glutamate receptor and an orexin receptor are over- expressed. In a twenty-second aspect of the invention, there is provided a cell, or fraction of a cell, in which a metabotropic glutamate receptor is over-expressed with an endogenously expressed orexin receptor.

In a twenty-third aspect of the invention, there is provided a cell, or fraction of a cell, in which an orexin receptor is over-expressed with an endogenously expressed metabotropic glutamate receptor.

Brief Description of the Drawings

Figures 1 to 3 are illustrative of the technique by which the association of the metabotropic glutamate receptor and the orexin receptor was detected.

Figure 1 shows the composition of the agents forming the basis of the system for detecting molecular associations: A first agent comprises a first interacting group coupled to a first reporter component; a second agent comprises a second interacting group coupled to a second reporter component; and a third agent comprises a third interacting group.

Figure 2 shows how the administration of the modulator modulates the association of the second interacting group with the third interacting group, preferably by interacting with the third interacting group, either alone, or simultaneously with the first interacting group.

Figure 3 shows that if the first and third interacting groups are associated, modulation of the association of the second and third interacting groups consequently modulates the proximity of the first and second reporter components thereby modulating the signal that is able to be detected by the detector. Therefore monitoring the signal generated by proximity of the first and second reporter components by the detector constitutes monitoring the association of the first and third agents. If the first and third interacting groups are not associated, the first and second reporter components will remain spatially separated and generation of a detectable signal is unlikely. Figure 4 shows the metabotropic glutamate receptor (mGluR5) as IG1 , Rlucδ as RC1 , beta-arrestin 2 (barr2) as IG2, Venus as RC2 and hemagglutin epitope-tagged OxR2 (HA-OxR2) as IG3. eBRET measurements at 37C were carried out on HEK293FT cells transiently expressing mGluR5/Rluc8 and barr2Λ/enus with either pcDNA3 or HA-OxR2 following treatment with 10^"6M orexin A (OxA). Figure 5 shows the metabotropic glutamate receptor (mGluR5) as IG1, Rlucδ as RC1 , beta-arrestin 2 (barr2) as IG2, Venus as RC2 and hemagglutin epitope-tagged OxR2 (HA-OxR2) as IG3. BRET measurements with coelenterazine h at 37C were carried out on HEK293FT cells transiently co-expressing mGluR5/Rluc8 and barr2Λ/enus with HA- OxR2 aliquoted into all wells of a 96-well plate. Phosphate-buffered saline (PBS) was added to the first two rows and the last two rows of the 96-well plate (48 wells in total) as a vehicle control for the data shown in Figures 6 and 7. Data presented as fluorescence/luminescence.

Figure 6 shows the metabotropic glutamate receptor (mGluRδ) as IG1 , Rlucδ as RC1 , beta-arrestin 2 (barr2) as IG2, Venus as RC2 and hemagglutin epitope-tagged OxR2 (HA-OxR2) as IG3. BRET measurements with coelenterazine h at 37C were carried out on HEK293FT cells transiently co-expressing mGluR5/Rluc8 and barr2/Venus with HA- OxR2 aliquoted into all wells of a 96-well plate. OxA was added to the middle four rows of the 96-well plate (48 wells in total). Data presented as fluorescence/luminescence. Figure 7 shows z'-factor data for the metabotropic glutamate receptor (mGluRδ) as IG1 , Rlucδ as RC1 , beta-arrestin 2 (barr2) as IG2, Venus as RC2 and hemagglutin epitope- tagged OxR2 (HA-OxR2) as IG3. As shown in figures 5 and 6, BRET measurements with coelenterazine h at 37C were carried out on HEK293FT cells transiently co-expressing mGluR5/Rluc8 and barr2Λ/enus with HA-OxR2 aliquoted into all wells of a 96-well plate. Phosphate-buffered saline (PBS) was added to the first two rows and last two rows of the 96-well plate (48 wells in total) as a vehicle control. OxA was added to the middle four rows of the 96-well plate (48 wells in total). Data presented as fluorescence/luminescence.

ABBREVIATIONS barr beta-arrestin.

BRET Bioluminescence resonance energy transfer.

BROM Bromocriptine.

CB Cannabinoid receptor.

CHPG ([RS]-2-chloro-5-hydroxyphenylglycine); mGluR agonist. CSF Cerebrospinal fluid.

DOP Delta opioid. eBRET extended BRET: BRET monitored over extended time periods.

ECFP Enhanced Cyan Fluorescent Protein, which is a variant of the

Aequorea victoria green fluorescent protein gene (GFP). EGFP Enhanced Green Fluorescent Protein is a red-shifted variant of wild-type GFP.

EYFP Enhanced Yellow Fluorescent Protein.

FRET Fluorescence resonance energy transfer.

GIu L-Glutamate. GPCRs G-protein coupled receptors. HA Hemagglutin epitope-tag.

His(6) Histidine tag consisting of 6 consecutive histidine residues.

IG Interacting group.

KOP Kappa opioid. mGluRI Metabotropic glutamate type 1 receptor. mGluR2 Metabotropic glutamate type 2 receptor. mGluR3 Metabotropic glutamate type 3 receptor. mGluR4 Metabotropic glutamate type 4 receptor. mGluR5 Metabotropic glutamate type 5 receptor. mGluRΘ Metabotropic glutamate type 6 receptor. mGluR7 Metabotropic glutamate type 7 receptor. mGluRβ Metabotropic glutamate type 8 receptor.

OR Opioid receptor.

OxA Orexin A.

OxB Orexin B.

OxR Orexin receptor.

PBS Phosphate-buffered saline. pcDNA3 Eukaryotic expression vector.

RC Reporter component.

RET Resonance energy transfer.

Rluc Renilla luciferase.

Rlucδ An improved Renilla luciferase.

TYFP Topaz Yellow Fluorescent Protein.

Venus An improved Yellow Fluorescent Protein

Wt Wild type.

Best Mode(s) for Carrying Out the Invention

General All publications, including patents and patent applications, cited herein, whether supra or infra, are hereby incorporated by reference in their entirety. However, publications mentioned herein are cited for the purpose of describing and disclosing the protocols, reagents and vectors that are reported in the publications and which may be used in connection with the invention. Nothing herein is to be construed as an admission that the invention is not entitled to antedate such disclosure by virtue of prior invention. Furthermore, the practice of the present invention employs, unless otherwise indicated, conventional molecular biology, chemistry and fluorescence techniques, within the skill of the art. Such techniques are well known to the skilled worker, and are explained fully in the literature. See, eg., Coligan, Dunn, Ploegh, Speicher and Wingfield "Current protocols in Protein Science" (1999) Volume I and Il (John Wiley & Sons Inc.); and Bailey, J. E. and Ollis, D. F., Biochemical Engineering Fundamentals, McGraw-Hill Book Company, NY, 1986; Lakowicz, J. R. Principles of Fluorescence Spectroscopy, New York : Plenum Press (1983) for fluorescence techniques.

As used herein and in the appended claims, the singular forms "a," "an," and "the" include the plural unless the context clearly dictates otherwise. Thus, for example, a reference to "a protein" includes a plurality of such proteins, and a reference to "an analyte" is a reference to one or more analytes, and so forth.

Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any materials and methods similar or equivalent to those described herein can be used to practice or test the present invention, the preferred materials and methods are now described.

The invention described herein may include one or more ranges of values (e.g. size, concentration etc). A range of values will be understood to include all values within the range, including the values defining the range, and values adjacent to the range that lead to the same or substantially the same outcome as the values immediately adjacent to that value which defines the boundary to the range.

Throughout this specification, unless the context requires otherwise, the word "comprise" or variations, such as "comprises" or "comprising" will be understood to imply the inclusion of a stated integer, or group of integers, but not the exclusion of any other integers or group of integers.

Specific

As is apparent from the preceding summary of the invention, the invention relates, inter alia, to hetero-dimeric or hetero-oligomeric receptor, comprising at least one metabotropic glutamate receptor subunit associated with at least one orexin receptor subunit.

The terms "hetero-dimer" and "hetero-oligomer", and variations such as "hetero-dimeric" and "hetero-oligomeric", as used herein, refer to an entity within which at least one metabotropic glutamate receptor is associated with at least one orexin receptor.

The phrase "associated with", as used herein, refers to combination via any known direct or indirect stabilising atomic or molecular level interaction or any combination thereof, where the interactions include, without limitation, bonding interactions such as covalent bonding, ionic bonding, hydrogen bonding, co-ordinate bonding, or any other molecular bonding interaction, electrostatic interactions, polar or hydrophobic interactions, or any other classical or quantum mechanical stabilising atomic or molecular interaction.

Instances of different tissues having different repertoires of hetero-dimers have been reported. For example, 6'guanidinoaltrindole, an analogue of a well-known KOP receptor ligand, has been identified as a DOP-KOP hetero-dimer selective agonist, with efficacy as a spinally selective analgesic, leading to the conclusion that DOP-KOP heterodimers are expressed in the spinal cord, but not in the brain (Waldhoer, M. et al. (2005) A hetero- dimer selective agonist shows in vivo relevance of G-protein coupled receptor dimers. Proc. Natl. Acad. Sci. USA 102:9050-9055). Accordingly, the hetero-dimeric or hetero- oligomeric receptor, comprising at least one metabotropic glutamate receptor subunit associated with at least one orexin receptor subunit represents a novel drug target. As is the case with 6'guanidinoaltrindole, known ligands may exhibit differing abilities to trigger a hetero-dimeric receptor, which may uncover new applications for pre-existing molecules:

- Hilairet S. et al. 2003 (J. Biol. Chem. 278:23731-23737) have recently shown that CB1 antagonists suppress appetite by acting through a CB1/OxR1 hetero- dimer pair.

- It has been shown that somatostatin SSTR5 receptor will hetero-dimerise with a dopamine D2 receptor (Rocheville M. et al. (2000) Science 288:154-157).

As will be apparent from the following examples, the inventors herein have identified and characterised the molecular association of the metabotropic glutamate receptor with the orexin receptor.

It will be apparent to a person skilled in the art that association of the metabotropic glutamate receptor with the orexin receptor enables the use of compounds related to one receptor, including and without limitation, ligands of one receptor (be they agonists, inverse agonists or antagonists) in the treatment of ailments related to the other receptor. Thus, the present invention encompasses a method for the treatment of a patient suffering from an orexin-related ailment by administering a therapeutically effective amount of a metabotropic glutamate receptor-related compound.

The phrase "metabotropic glutamate receptor-related compound" is to be understood to mean a compound that interacts with the metabotropic glutamate receptor; a compound that binds to a compound that interacts with the metabotropic glutamate receptor, including but not limited to glutamate; or a compound that modulates the production of a compound that interacts with the metabotropic glutamate receptor, including but not limited to glutamate. In one form of the invention, the metabotropic glutamate receptor-related compound is a metabotropic glutamate receptor agonist, inverse agonist or antagonist.

In one embodiment, the metabotropic glutamate receptor-related compound is an allosteric modulator of the metabotropic glutamate receptor.

In one embodiment, the metabotropic glutamate receptor-related compound modulates the production of glutamate.

In one embodiment, the metabotropic glutamate receptor-related compound is a glutamate binding agent, or a glutamate binding fragment thereof.

In one embodiment, the glutamate binding agent is an antibody, including a humanised antibody, a polyclonal antibody, a monoclonal antibody, a chimeric antibody, a CDR- grafted antibody and/or an anti-idiotypic antibody.

In one embodiment, the metabotropic glutamate receptor-related compound is selective for the metabotropic glutamate receptor relative to the orexin receptor. In one form of the invention, the metabotropic glutamate receptor-related compound is selective for the metabotropic glutamate receptor relative to the orexin receptor by a factor of at least 10. In one form of the invention, the metabotropic glutamate receptor-related compound is selective for the metabotropic glutamate receptor relative to the orexin receptor by a factor of at least 100. In one form of the invention, the metabotropic glutamate receptor- related compound is selective for the metabotropic glutamate receptor relative to the orexin receptor by a factor of at least 1000. In the context of metabotropic glutamate receptor-related compounds that modulate the production of a compound that interacts with the metabotropic glutamate receptor, the phrase "selective for the metabotropic glutamate receptor relative to the orexin receptor" is to be understood to mean that the compound modulates the production of a compound that interacts with the metabotropic glutamate receptor to a greater extent than it modulates the production of orexin.

In the context of metabotropic glutamate receptor-related compounds that are glutamate binding agents, or glutamate binding fragments thereof, the phrase "selective for the metabotropic glutamate receptor relative to the orexin receptor" is to be understood to mean that the glutamate binding agent, or the glutamate binding fragment thereof, binds glutamate selectively relative to orexin.

In one embodiment, the metabotropic glutamate receptor-related compound is coadministered with an orexin receptor-related compound.

The phrase "orexin receptor-related compound" is to be understood to mean a compound that interacts with the orexin receptor; a compound that binds to a compound that interacts with the orexin receptor, including but not limited to orexin; or a compound that modulates the production of a compound that interacts with the orexin receptor, including but not limited to orexin.

In one embodiment, the orexin receptor-related compound is an agonist, inverse agonist or antagonist of the orexin receptor.

In one embodiment, the orexin receptor-related compound is an allosteric modulator of the orexin receptor.

In one embodiment, the orexin receptor-related compound modulates the production of orexin. In one embodiment, the orexin receptor-related compound is an orexin binding agent, or an orexin binding fragment thereof.

In one embodiment, the orexin binding agent is an antibody, including a humanised antibody, a polyclonal antibody, a monoclonal antibody, a chimeric antibody, a CDR- grafted antibody and/or an anti-idiotypic antibody. The present invention further encompasses a method for the treatment of a patient suffering from a glutamate-related ailment by administering a therapeutically effective amount of an orexin receptor-related compound.

In one embodiment, the orexin receptor-related compound is an agonist, inverse agonist or antagonist of the orexin receptor. In one embodiment, the orexin receptor-related compound is an allosteric modulator of the orexin receptor.

In one form of the invention, the orexin receptor-related compound is a compound that modulates the production of orexin.

In one form of the invention, the orexin receptor-related compound is an orexin binding agent, or an orexin binding fragment thereof.

In one embodiment, the orexin binding agent is an antibody, including a humanised antibody, a polyclonal antibody, a monoclonal antibody, a chimeric antibody, a CDR- grafted antibody and/or an anti-idiotypic antibody.

In one embodiment, the orexin receptor-related compound is selective for the orexin receptor relative to the metabotropic glutamate receptor.

In one form of the invention, the orexin receptor-related compound is selective for the orexin receptor relative to the metabotropic glutamate receptor by a factor of at least 10.

In one form of the invention, the orexin receptor-related compound is selective for the orexin receptor relative to the metabotropic glutamate receptor by a factor of at least 100.

In one form of the invention, the orexin receptor-related compound is selective for the orexin receptor relative to the metabotropic glutamate receptor by a factor of at least 1000.

In the context of orexin receptor-related compounds that modulate the production of a compound that interacts with the orexin receptor, the phrase selective for the orexin receptor relative to the metabotropic glutamate receptor is to be understood to mean that the compound modulates the production of a compound that interacts with the orexin receptor to a greater extent than it modulates the production of glutamate.

In the context of orexin receptor-related compounds that are orexin binding agents, or orexin binding fragments thereof, the phrase selective for the orexin receptor relative to the metabotropic glutamate receptor is to be understood to mean that the orexin binding agent, or the orexin binding fragment thereof, binds orexin selectively relative to glutamate.

In one embodiment, the orexin receptor-related compound is co-administered with a metabotropic glutamate receptor-related compound.

In one form of the invention, the metabotropic glutamate receptor-related compound is an agonist, inverse agonist or antagonist of the metabotropic glutamate receptor. In one embodiment, the metabotropic glutamate receptor-related compound is an allosteric modulator of the metabotropic glutamate receptor.

In one embodiment, the glutamate binding agent is an antibody, including a humanised antibody, a polyclonal antibody, a monoclonal antibody, a chimeric antibody, a CDR- grafted antibody and/or an anti-idiotypic antibody. The present invention further encompasses a method for the manufacture of a medicament for the treatment of a patient suffering from an orexin-related ailment by administering a therapeutically effective amount of a metabotropic glutamate receptor- related compound. In one form of the invention, the metabotropic glutamate receptor-related compound is an agonist, inverse agonist or antagonist of the metabotropic glutamate receptor.

In one embodiment, the medicament contains an orexin receptor-related compound.

In one form of the invention, the orexin receptor-related compound is an agonist, inverse agonist or antagonist of the orexin receptor. The present invention further encompasses a method for the manufacture of a medicament for the treatment of a patient suffering from a glutamate-related ailment by administering a therapeutically effective amount of an orexin receptor-related compound.

In one form of the invention, the orexin receptor-related compound is an agonist, inverse agonist or antagonist of the orexin receptor. In one embodiment, the medicament contains a metabotropic glutamate receptor-related compound.

In one form of the invention, the metabotropic glutamate receptor-related compound is an agonist, inverse agonist or antagonist of the metabotropic glutamate receptor.

Thus, the present invention encompasses a method for the treatment of a patient suffering from an orexin-related ailment by administering a therapeutically effective amount of a glutamate-selective binding agent, or fragment thereof.

The glutamate selective binding agent may be an antibody, including a humanised antibody, a polyclonal antibody, a monoclonal antibody, a chimeric antibody, a CDR- grafted antibody and/or an anti-idiotypic antibody. The present invention further encompasses a method for the treatment of a patient suffering from a glutamate-related ailment by administering a therapeutically effective amount of an orexin-selective binding agent, or fragment thereof.

The orexin-selective binding agent may be an antibody, including a humanised antibody, a polyclonal antibody, a monoclonal antibody, a chimeric antibody, a CDR-grafted antibody and/or an anti-idiotypic antibody.

The present invention further encompasses a method for the treatment of a patient suffering from a glutamate-related ailment or an orexin-related ailment by administering a therapeutically effective amount of a metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero- dimerAoligomer, such as a selective allosteric modulator.

The present invention further encompasses the use of a therapeutically effective amount of a metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/-oligomer, such as a selective allosteric modulator, for the manufacture of a medicament for the treatment of a patient suffering from a glutamate-related ailment or an orexin-related ailment. In one form of the invention, the metabotropic glutamate receptor / orexin receptor hetero- dimer/-oligomer selective agonist, inverse agonist, partial agonist, antagonist or other molecule that interacts with the hetero-dimer/-oligomer, such as an allosteric modulator, is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/- oligomer by a factor of at least 10. In one form of the invention, the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective agonist, inverse agonist, partial agonist, antagonist or other molecule that interacts with the hetero-dimer/- oligomer, such as an allosteric modulator, is selective for the metabotropic glutamate / orexin receptor hetero-dimer/-oligomer by a factor of at least 100. In one form of the invention, the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective agonist, inverse agonist, partial agonist, antagonist or other molecule that interacts with the hetero-dimer/-oligomer, such as an allosteric modulator, is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer by a factor of at least 1000.

The present invention further encompasses a method for the treatment of a patient suffering from a glutamate-related ailment by administering a therapeutically effective amount of a selective orexin receptor / metabotropic glutamate receptor hetero-dimer / - oligomer agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/-oligomer, such as a selective allosteric modulator. In one embodiment, the selective orexin receptor / metabotropic glutamate receptor hetero-dimer / -oligomer agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/- oligomer, such as a selective allosteric modulator is co-administered with a metabotropic glutamate receptor-related compound. In one embodiment, the metabotropic glutamate receptor-related compound is a metabotropic glutamate receptor agonist, inverse agonist or antagonist.

In one embodiment, the selective orexin receptor / metabotropic glutamate receptor hetero-dimer / -oligomer agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/- oligomer, such as a selective allosteric modulator is co-administered with an orexin receptor-related compound.

In one embodiment, the orexin receptor-related compound is an orexin receptor agonist, inverse agonist or antagonist.

The present invention further encompasses a method for the treatment of a patient suffering from an orexin-related ailment by administering a therapeutically effective amount of a selective orexin receptor / metabotropic glutamate receptor hetero-dimer / - oligomer agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/-oligomer, such as a selective allosteric modulator. In one embodiment, the selective orexin receptor / metabotropic glutamate receptor hetero-dimer / -oligomer agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/- oligomer, such as a selective allosteric modulator is co-administered with a metabotropic glutamate receptor-related compound.

In one embodiment, the metabotropic glutamate receptor-related compound is a metabotropic glutamate receptor agonist, inverse agonist or antagonist. In one embodiment, the selective orexin receptor / metabotropic glutamate receptor hetero-dimer / -oligomer agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/- oligomer, such as a selective allosteric modulator is co-administered with an orexin receptor-related compound.

In one embodiment, the orexin receptor-related compound is an orexin agonist, inverse agonist or antagonist.

The present invention further encompasses a method for the manufacture of a medicament for the treatment of a patient suffering from a glutamate-related ailment comprising use of a therapeutically effective amount of a selective orexin receptor / metabotropic glutamate receptor hetero-dimer / -oligomer agonist, inverse agonist or antagonist.

The present invention further encompasses a method for the manufacture of a medicament for the treatment of a patient suffering from a glutamate-related ailment comprising use of a therapeutically effective amount of a selective orexin receptor / metabotropic glutamate receptor hetero-dimer / -oligomer agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/-oligomer, such as a selective allosteric modulator.

In one embodiment, the medicament contains an orexin receptor-related compound. In one embodiment, the orexin receptor-related compound is an orexin agonist, inverse agonist or antagonist.

In one embodiment, the medicament contains a metabotropic glutamate receptor-related compound. In one embodiment, the metabotropic glutamate receptor-related compound is a metabotropic glutamate receptor agonist, inverse agonist or antagonist.

The present invention further encompasses a method for the manufacture of a medicament for the treatment of a patient suffering from an orexin-related ailment comprising use of a therapeutically effective amount of a selective orexin receptor / metabotropic glutamate receptor hetero-dimer / -oligomer agonist, inverse agonist or antagonist.

The present invention further encompasses a method for the manufacture of a medicament for the treatment of a patient suffering from an orexin-related ailment comprising use of a therapeutically effective amount of a selective orexin receptor / metabotropic glutamate receptor hetero-dimer / -oligomer agonist, selective inverse agonist, selective partial agonist, selective antagonist or other molecule that selectively interacts with the hetero-dimer/-oligomer, such as a selective allosteric modulator.

In one embodiment, the medicament contains an orexin receptor-related compound. In one embodiment, the orexin-receptor-related compound is an orexin receptor agonist, inverse agonist or antagonist.

Glutamate-related ailments include ailments that are related to increased or decreased production of glutamate, and/or increased or decreased responsiveness of cells to glutamate. The following list (Nicoletti, F. et al, (2007) Metabotropic glutamate receptors: beyond the regulation of synaptic transmission. Psychoneuroendocrinology. Vol32; S40- S45) provides some examples of clinical conditions that could utilise metabotropic glutamate-related interventions and be a glutamate-related ailment: Anxiety and panic attacks (including general anxiety disorder; GAD); Chronic pain; Drug addiction; Schizophrenia; - Parkinson's disease;

Fragile X syndrome

However, it should be understood that the phrase metabotropic glutamate-related interventions and the phrase a glutamate-related ailment is not limited thereto.

Known metabotropic glutamate receptor-related compounds include; L-glutamate, L- quisqualic acid, ACPD agonist ((1S,3S)-1-aminocyclopentane-1 ,3-dicarboxylic acid); mGluRδ-specific agonist CHPG ([RS]-2-chloro-5-hydroxyphenylglycine); Group 1 mGluR- selective agonist DHPG ([S]-3,5-dihydroxyphenylglycine); Group 2 mGluR-selective agonist Eglumegad (LY-354,740); Group 2 mGluR-selective agonist DCG-IV (2-(2,3- dicarboxycyclopropyl)glycine); Group 3 mGluR-selective agonist L-AP4 (2-amino-4- phosphonobutyrate); Group 2 m-GluR-selective antagonist LY-341 ,495; LY-393,675 antagonist (2-(S)-amino-2-(3-cis-carboxycyclobutyl)-3-(9H-thioxanthen-9-yl) propionic acid); MPEP antagonist (2-methyl-6-(phenylethylnyl)-pyridine); MTEP antagonist (3-[(2- methyl-1 ,3-thiazol-4-yl)ethynyl]pyridine); SIB-1757 antagonist; SIB-1893 antagonist; Fenobam antagonist/inverse agonist ([N-(3-chlorophenyl)-N'-(4,5-dihydro-1-methyl-4-oxo- 1H-imidazole-2-yl)ureal]); the antagonists 3-(5-pyridin-2-yl-2H-tetrazol-2-yl) benzonitrile and 5-[(2-methyl-1 ,3-thiazol-4-yl)ethynyl]-2_I3'-bipyridine; ADX-10059 negative allosteric modulator; ADX-48621 negative allosteric modulator; ADX-63365 positive allosteric modulator.

Orexin-related ailments include aliments that are related to increased or decreased production of orexin, and/or increased or decreased responsiveness of cells to orexin. A major example of an orexin-related ailment is narcolepsy with cataplexy. This is associated with low or undetectable levels of cerebrospinal fluid (CSF) orexin A levels in about 90% of patients (Baumann and Bassetti (2005) Sleep Medicine Reviews 9, 253-

268). Mutations of the orexin receptor 2 gene lead to familial canine narcolepsy, and a loss of orexin neurons and low CSF orexin A were observed with sporadic canine narcolepsy. Neurological disorders arising from acute traumatic brain injury, Guillain-

Barre syndrome and advanced Parkinson's syndrome may also be linked with low or undetectable levels of CSF orexin A levels in some instances. Sakurai has postulated a role for the orexin system in feeding and energy homeostasis as the activity of orexin neurons is inhibited by glucose and leptin, and stimulated by ghrelin, a stomach-derived peptide which promotes feeding. This may have implications for the treatment of obesity (Sakurai (2005) Sleep Medicine Reviews 9, 231-241).

However, it should be understood that the phrase orexin-related ailment is not limited thereto. Known orexin receptor-related compounds include orexin A (OxA; hypocretin-1 ; Hcrt-1 ), orexin B (OxB; hypocretin-2; Hcrt2) and fragments thereof (Lang et al (2004) J Med Chem 47, 1153-1160); OxR1 and OxR2 antagonists including 6,7-dimethoxy-i , 2,3,4- tetrahydroisoquinoline analogues (Hirose M et al (2003) Bioorg. Med. Chem. Lett. 13, 4497-4499), Almorexant ((2R)-2-{(1 S)-6,7-dimethoxy-1 -[2-(4-trifluoromethylphenyl)-ethyl]- 3,4-dihydro-1 H-isoquinolin-2-yl}-N-methyl-2-phenyl-acetamide; ACT-078573; Actelion Pharmaceuticals Ltd., Allschwil, Switzerland; Brisbare-Roch et al (2007) Nature Medicine 13, 150-155); OxR1 antagonists including SB-334867-A (1-(2-methylbenzoxazol-6-yl)-3- [1 ,5]naphthyridin-4-yl urea hydrochloride, SB-674042 (1-(5-(2-fluoro-phenyl)-2-methyl- thiazol-4-yl)-1-((S)-2-(5-phenyl-(1,3,4)oxadiazol-2ylmethyl)-pyrrolidin-1-yl)-methanone), SB-408124 (1-(6,8-difluoro-2-methyl-quinolin-4-yl)-3-(4-dimethylamino-phenyl)-urea) and SB-410220 (1-(5,8-difluoro-quinolin-4-yl)-3-(4-dimethylamino-phenyl)-urea (Haynes et al (2000) Regulatory Peptides 96, 45-51 ; Langmead et al (2004) British Journal of Pharmacology 141 , 340-346); OxR2 antagonists including N-Arylmethyl tert-leucyl 6,7- dimethoxy-1 ,2,3,4-tetrahydroiso-quinoline analogues and N-acyl 6,7-dimethoxy-1 ,2,3,4- tetrahydroisoquinoline analogues (Hirose M et al (2003) Bioorg. Med. Chem. Lett. 13, 4497-4499), and substituted 4-phenyl-[1 ,3]dioxanes, particularly 1-(2,4-dibromo-phenyl)- 3-((4S,5S)-2,2-dimethyl-4-phenyl-[1 ,3]dioxan-5-yl)-urea (McAtee LC et al (2004) Bioorg. Med. Chem. Lett. 14, 4225-4229).

Although glutamate plays an extensive role in mediating the synaptic potential of neurons, there appears to be few studies that have examined potential interactions between orexin and metabotropic glutamate receptor pathways. One exception is a study of lateral hypothalamic brain slices from mice, which measured the effect of metabotropic glutamate receptors (mGluRs) upon the activity of orexin neurons (Acuna-Goycolea et al. (2004)). A mGluR agonist inhibited synaptic input to orexin neurons in a dose-dependent fashion, while an antagonist of Group 3 mGluRs suppressed this effect. Interestingly, when this antagonist was administered alone, an increase in the synaptic activity of orexin neurons was observed, indicating that these neurons may be tonically regulated by mGluRs expressed at their surface.

A similar study was reported the following year, where the effect of OxB treatment upon neurons of the dorsal raphe nucleus (DRN) was investigated (Haj-Dahmane and Shen (2005)). OxB administration caused a significant decrease in synaptic current mediated by glutamate in neurons of the DRN expressing serotonin, which was caused by a presynaptic inhibition of glutamate release. Post-synaptic inhibition of G-protein signalling completely abolished this effect, indicating that orexin receptors mediate this response. Interestingly, OxB depression of glutamate release could be prevented by treatment with a cannabinoid receptor antagonist, leading the authors to propose the activation of postsynaptic orexin receptors results in activation of an endocannabinoid signalling pathway, which acts in a retrograde fashion to prevent presynaptic glutamate release. Apart from these aforementioned reports, a likely physiological scenario in which the metabotropic glutamate and orexin receptors interact involves the regulation and development of addictive, reward-seeking behaviour. The role that mGluRs play in this neural plasticity has been reviewed previously (Dalrymple M. et al (2008) G protein- coupled receptor dimers: Functional consequences, disease states and drug targets. Pharmacol Ther 118:359-371 ), with numerous studies demonstrating that heterodimeric interactions between mGluRs (particularly mGluR5) and the adenosine-2a receptor subtype (A_2aR) are strongly implicated in the development of such behaviours. In addition, there have also been several recent publications that have highlighted the involvement of the orexin system in reinforcing behaviours that produce a rewarding outcome (Bingham, M. et al (2006) Eating, sleeping and rewarding: orexin receptors and their antagonists. Curr Opin Drug Discov Devel 9:551-559; Harris, G. et al., (2007) Lateral hypothalamic orexin neurons are critically involved in learning to associate an environment with morphine reward. Behav Brain Res 183:43-51 ).

In one embodiment, the present invention provides a method for the treatment of a patient suffering from an orexin-related ailment by administering a therapeutically effective amount of a metabotropic glutamate receptor-related compound selected from the group: L-glutamate, L-quisqualic acid, ACPD agonist ((1S,3S)-1-aminocyclopentane-

1 ,3-dicarboxylic acid); mGluRδ-specific agonist CHPG ([RS]-2-chloro-5- hydroxyphenylglycine); Group 1 mGluR-selective agonist DHPG ([S]-3,5- dihydroxyphenylglycine); Group 2 mGluR-selective agonist Eglumegad (LY-354,740);

Group 2 mGluR-selective agonist DCG-IV (2-(2,3-dicarboxycyclopropyl)glycine); Group 3 mGluR-selective agonist L-AP4 (2-amino-4-phosphonobutyrate); Group 2 m-GluR- selective antagonist LY-341,495; LY-393,675 antagonist (2-(S)-amino-2-(3-cis- carboxycyclobutyl)-3-(9H-thioxanthen-9-yl) propionic acid); MPEP antagonist (2-methyl-6- (phenylethylnyl)-pyridine); MTEP antagonist (3-[(2-methyl-1 ,3-thiazol-4- yl)ethynyl]pyridine); SIB-1757 antagonist; SIB-1893 antagonist; Fenobam antagonist/inverse agonist ([N-(3-chlorophenyl)-N'-(4,5-dihydro-1-methyl-4-oxo-1 H- imidazole-2-yl)ureal]); the antagonists 3-(5-pyridin-2-yl-2H-tetrazol-2-yl) benzonitrile and 5-[(2-methyl-1 ,3-thiazol-4-yl)ethynyl]-2,3'-bipyridine; ADX-10059 negative allosteric modulator; ADX-48621 negative allosteric modulator; ADX-63365 positive allosteric modulator.

In one form of the invention, the metabotropic glutamate-receptor related compound is a metabotropic glutamate receptor agonist, inverse agonist or antagonist selected from the foregoing group.

In one embodiment, the present invention provides a method for the treatment of a patient suffering from a glutamate-related ailment by administering a therapeutically effective amount of an orexin receptor-related compound selected from the group: orexin A (OxA; hypocretin-1 ; Hcrt-1), orexin B (OxB; hypocretin-2; Hcrt2) and fragments thereof (Lang et al (2004) J Med Chem 47, 1153-1160); OxR1 and OxR2 antagonists including 6,7-dimethoxy-1 ,2,3,4-tetrahydroisoquinoline analogues (Hirose M et al (2003) Bioorg. Med. Chem. Lett. 13, 4497-4499), Almorexant ((2R)-2-{(1S)-6,7-dimethoxy-1-[2-(4- trifluoromethylphenyl)-ethyl]-3,4-dihydro-1H-isoquinolin-2-yl}-N-methyl-2-phenyl- acetamide; ACT-078573; Actelion Pharmaceuticals Ltd., Allschwil, Switzerland; Brisbare- Roch et al (2007) Nature Medicine 13, 150-155); OxR1 antagonists including SB-334867- A (1-(2-methylbenzoxazol-6-yl)-3-[1 ,5]naphthyridin-4-yl urea hydrochloride, SB-674042 (1-(5-(2-fluoro-phenyl)-2-methyl-thiazol-4-yl)-1-((S)-2-(5-phenyl-(1 ,3,4)oxadiazol-

2ylmethyl)-pyrrolidin-1-yl)-methanone), SB-408124 (1-(6,8-difluoro-2-methyl-quinolin-4- yl)-3-(4-dimethylamino-phenyl)-urea) and SB-410220 (1-(5,8-difluoro-quinolin-4-yl)-3-(4- dimethylamino-phenyl)-urea (Haynes et al (2000) Regulatory Peptides 96, 45-51 ; Langmead et al (2004) British Journal of Pharmacology 141 , 340-346); OxR2 antagonists including N-Arylmethyl tert-leucyl 6,7-dimethoxy-1 ,2,3,4-tetrahydroiso-quinoline analogues and N-acyl 6,7-dimethoxy-1 ,2,3,4-tetrahydroisoquinoline analogues (Hirose M et al (2003) Bioorg. Med. Chem. Lett. 13, 4497-4499), and substituted 4-phenyl- [1 ,3]dioxanes, particularly 1-(2,4-dibromo-phenyl)-3-((4S,5S)-2,2-dimethyl-4-phenyl- [1 ,3]dioxan-5-yl)-urea (McAtee LC et al (2004) Bioorg. Med. Chem. Lett. 14, 4225-4229). In one form of the invention, the orexin receptor related compound is an orexin receptor agonist, inverse agonist or antagonist selected from the foregoing group.

The present invention also includes a method for screening a test compound for potential therapeutic activity against an orexin-related ailment using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) exposing the test compound to a metabotropic glutamate receptor; b) assessing whether and/or the extent to which the activity of the metabotropic glutamate receptor is modulated compared to activity of the metabotropic glutamate receptor in the absence of the test compound; said modulation being indicative of potential therapeutic activity against the orexin- related ailment.

The present invention also includes a method for screening a test compound for potential therapeutic activity against an orexin-related ailment, the method comprising the steps of: a) exposing the test compound to a metabotropic glutamate receptor; b) assessing the extent to which the activity of the metabotropic glutamate receptor is modulated; said modulation being indicative of potential therapeutic activity against the orexin- related ailment.

In one embodiment, the method for screening a test compound for potential therapeutic activity against an orexin-related ailment using a detector capable of detecting changes in receptor activity comprises the step of: administering said test compound to an animal.

In one embodiment, the method for screening a test compound for potential therapeutic activity against an orexin-related ailment using a detector capable of detecting changes in receptor activity comprises the steps of: contacting a cell expressing metabotropic glutamate receptor with said test compound; detecting a change in metabotropic glutamate receptor activity.

Methods for assessing the extent to which the activity of a metabotropic glutamate receptor is modulated and detectors capable of detecting changes in receptor activity are well-known to those skilled in the art.

The present invention also includes a method for screening a test compound for potential therapeutic activity against a glutamate-related ailment using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) exposing the test compound to an orexin receptor; b) assessing whether and/or the extent to which the activity of the orexin receptor is modulated compared to activity of the orexin receptor in the absence of the test compound; said modulation being indicative of potential therapeutic activity against the glutamate- related ailment.

The present invention also includes a method for screening a test compound for potential therapeutic activity against a glutamate-related ailment, the method comprising the steps of: a) exposing the test compound to an orexin receptor; b) assessing the extent to which the activity of the orexin receptor is modulated; said modulation being indicative of potential therapeutic activity against the glutamate- related ailment.

In one embodiment, the method for screening a test compound for potential therapeutic activity against a glutamate-related ailment using a detector capable of detecting changes in receptor activity comprises the step of: administering said test compound to an animal.

In one embodiment, the method for screening a test compound for potential therapeutic activity against a glutamate-related ailment using a detector capable of detecting changes in receptor activity comprises the steps of: contacting a cell expressing orexin receptor with said test compound; detecting a change in orexin receptor activity.

Methods for assessing the extent to which the activity of an orexin receptor is modulated and detectors capable of detecting changes in receptor activity are well-known to those skilled in the art.

The present invention comprises a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer selective activity using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound interacts with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor; and b) determining whether, and/or the extent to which the test compound interacts with the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater affinity and/or potency and/or efficacy when interacting with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer. The present invention comprises a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound interacts with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor; and b) if the test compound interacts with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor, determining whether, or the extent to which the test compound interacts with the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater affinity and/or potency and/or efficacy when interacting with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer. In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer selective activity using a detector capable of detecting changes in receptor activity comprises the step of: administering said test compound to an animal.

In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective activity using a detector capable of detecting changes in receptor activity comprises the steps of: contacting a cell expressing orexin receptor and metabotropic glutamate receptor with said test compound; detecting a change in orexin receptor or metabotropic glutamate receptor activity.

The present invention comprises a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound interacts with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor; and b) if the test compound interacts with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor, determining whether, or the extent to which the test compound interacts with the metabotropic glutamate receptor in the absence of the orexin receptor; such that a test compound that exhibits greater affinity and/or potency and/or efficacy when interacting with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

The present invention further provides a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective activity using a detector capable of detecting changes in receptor activity, the method comprising the step of: determining whether, and/or the extent to which, the test compound interacts with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor compared to whether, and/or the extent to which the test compound interacts with the metabotropic glutamate receptor in the absence of the orexin receptor; such that a test compound that exhibits greater affinity and/or potency and/or efficacy when interacting with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective activity using a detector capable of detecting changes in receptor activity comprises the step of: administering said test compound to an animal.

In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective activity using a detector capable of detecting changes in receptor activity comprises the steps of: contacting a cell expressing orexin receptor and metabotropic glutamate receptor with said test compound; detecting a change in orexin receptor or metabotropic glutamate receptor activity. In one aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer selective antagonism or partial agonism, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an antagonist or partial agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the metabotropic glutamate receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an antagonist or partial agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; c) if the test compound is an antagonist or partial agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, determining whether, or the extent to which the test compound is an antagonist or partial agonist of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater antagonistic or partial agonistic properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

In one aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective antagonism or selective partial agonism or selective negative allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the metabotropic glutamate receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether, or the extent to which the test compound is an antagonist, partial agonist, or negative allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater antagonistic, partial agonistic or negative allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero- dimer/-oligomer. In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective antagonism, selective partial agonism or selective negative allosteric modulation using a detector capable of detecting changes in receptor activity comprises the step of: administering said test compound to an animal. In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective antagonism, selective partial agonism or selective negative allosteric modulation using a detector capable of detecting changes in receptor activity comprises the steps of: contacting a cell expressing orexin receptor and metabotropic glutamate receptor with said test compound; detecting a change in orexin receptor or metabotropic glutamate receptor activity.

In one aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective antagonism or partial agonism, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an antagonist or partial agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the orexin receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b). detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an antagonist or partial agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; c) if the test compound is an antagonist or partial agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer, determining whether, or the extent to which the test compound is an antagonist or partial agonist of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater antagonistic or partial agonistic properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

In one aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective antagonism, selective partial agonism or selective negative allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the orexin receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b). detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether, or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater antagonistic, partial agonistic or negative allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer. In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective antagonism, selective partial agonism or selective negative allosteric modulation using a detector capable of detecting changes in receptor activity comprises the step of: administering said test compound to an animal. In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer selective antagonism, selective partial agonism or selective negative allosteric modulation using a detector capable of detecting changes in receptor activity comprises the steps of: contacting a cell expressing orexin receptor and metabotropic glutamate receptor with said test compound; detecting a change in orexin receptor or metabotropic glutamate receptor activity.

In one aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective inverse agonism, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising a constitutively active metabotropic glutamate receptor; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; c) if the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, determining whether, or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater inverse agonistic properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero- dimer/-oligomer is selective for the metabotropic glutamate / orexin receptor hetero-dimer/-oligomer. In one aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective inverse agonism using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; Hi), a third agent, comprising a constitutively active metabotropic glutamate receptor; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally ; c) determining whether, or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater inverse agonistic properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer is selective for the metabotropic glutamate / orexin receptor hetero-dimer/-oligomer.

In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective inverse agonism using a detector capable of detecting changes in receptor activity comprises the step of: administering said test compound to an animal.

In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective inverse agonism using a detector capable of detecting changes in receptor activity comprises the steps of: contacting a cell expressing orexin receptor and metabotropic glutamate receptor with said test compound; detecting a change in orexin receptor or metabotropic glutamate receptor activity.

In one aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer inverse agonism, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising a constitutively active orexin receptor; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; c) if the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, determining whether, or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater inverse agonistic properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero- dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

In one aspect of the invention, there is provided a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer inverse agonism using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising a constitutively active orexin receptor; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether, or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater inverse agonistic properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

The present invention further provides a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-o!igomer selective positive allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the metabotropic glutamate receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting an increase in the signal as a determination of whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer and optionally; c) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater positive allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate / orexin receptor hetero-dimer/- oligomer. In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer selective positive allosteric modulation using a detector capable of detecting changes in receptor activity comprises the step of: administering said test compound to an animal. In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective positive allosteric modulation using a detector capable of detecting changes in receptor activity comprises the steps of: contacting a cell expressing orexin receptor and metabotropic glutamate receptor with said test compound; detecting a change in orexin receptor or metabotropic glutamate receptor activity.

The present invention further provides a method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective positive allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the orexin receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b) detecting an increase in the signal as a determination of whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater positive allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero- dimer/-oligomer.

In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective positive allosteric modulation using a detector capable of detecting changes in receptor activity comprises the step of: administering said test compound to an animal.

In one embodiment, the method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective positive allosteric modulation using a detector capable of detecting changes in receptor activity comprises the steps of: contacting a cell expressing orexin receptor and metabotropic glutamate receptor with said test compound; detecting a change in orexin receptor or metabotropic glutamate receptor activity.

In a preferred embodiment of the invention, the step of determining whether, and/or the extent to which, the test compound interacts with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor; and/or the step of determining whether, and/or the extent to which, the test compound interacts with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor are performed by way of the methods described in the applicant's co- pending international patent application "Detection System and Uses Therefor", PCT/AU2007/001722 (published as WO 2008/055313). However, for the sake of clarity, it should be understood that the methods of the present invention are not restricted to methods where the step of determining whether, and/or the extent to which, the test compound interacts with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor; and/or the step of determining whether, and/or the extent to which, the test compound interacts with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor are performed by way of the methods described in the applicant's co-pending international patent application "Detection System and Uses Therefor", PCT/AU2007/001722 (published as WO 2008/055313). Altemate methods of determining whether, and/or the extent to which, the test compound interacts with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor; and/or the step of determining whether, and/or the extent to which, the test compound interacts with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor include assays observing a change in coupling to signalling pathways such as a change in G- protein utilisation, ligand binding assays, signalling assays such as those monitoring changes in Ca²⁺, inositol phosphate, cyclic adenosine monophosphate (cAMP), extracellular-signal regulated kinase (ERK) and/or mitogen-activated protein kinase (MAPK), receptor trafficking assays, beta-arrestin translocation assays, enzyme-linked immunosorbent assays (ELISAs) and any other assay that can detect a change in receptor function as a result of receptor heterodimerization.

The present invention includes selective agonists and/or antagonists and/or inverse agonists of the metabotropic glutamate receptor/orexin receptor hetero-dimer/-oligomer. The present invention includes selective agonists and/or selective antagonists and/or selective inverse agonists and/or selective allosteric modulators of the metabotropic glutamate receptor/orexin receptor hetero-dimer/-oligomer.

The present invention comprises a cell, or fraction of a cell, in which both a metabotropic glutamate receptor and an orexin receptor are over-expressed. The present invention comprises a cell, or fraction of a cell, in which a metabotropic glutamate receptor is over-expressed with an endogenously expressed orexin receptor.

The present invention comprises a cell, or fraction of a cell, in which an orexin receptor is over-expressed with an endogenously expressed metabotropic glutamate receptor.

Throughout this specification, unless the context requires otherwise, the phrase "fraction of a cell' includes, without limitation, cell membrane preparations. As would be understood by a person skilled in the art, cell membrane preparations are useful in binding assays, or as antigens against which antibodies, including antibody therapeutics, may be raised.

The present invention comprises a cell in which both a metabotropic glutamate receptor and an orexin receptor are over-expressed.

The present invention comprises a cell in which a metabotropic glutamate receptor is over-expressed with an endogenously expressed orexin receptor.

The present invention comprises a cell in which an orexin receptor is over-expressed with an endogenously expressed metabotropic glutamate receptor. The phrase "over-expressed", as used herein in the context of receptors, refers to an abnormal level of expression of the receptor within the cell relative to the natural level of expression. This may include a level of expression considered to be within the physiological range, but expressed in cells not normally expressing the receptor. This may also include a level of expression considered to be within the physiological range, but in cells not normally expressing the receptors modified in any way, such as by fusion to other proteins or by the addition of immunolabels. Cells in which a receptor is over- expressed may be identified by standard assay techniques well known in the art.

As used herein the term "patient" refers to any animal that may be suffering from one or more of orexin- or glutamate-related ailments. Most preferably the animal is a mammal. The term will be understood to include for example human, farm animals (i.e., cattle, horses, goats, sheep and pigs), household pets (i.e., cats and dogs) and the like.

The phrase "therapeutically effective amount" as used herein refers to an amount sufficient to modulate a biological activity associated with the interaction of orexin receptor agonist, inverse agonist, antagonist or allosteric modulator with the orexin receptor or metabotropic glutamate receptor agonist, inverse agonst, antagonist or allosteric modulator with the metabotropic glutamate receptor or of orexin receptor/ metabotropic glutamate receptor hetero-dimer/oligomer-specific agonist, inverse agonist, antagonist or allosteric modulator with an orexin receptor/ metabotropic glutamate receptor hetero-dimer/oligomer. In the context of aspects of the invention where both a metabotropic glutamate receptor- related compound, such as and without limitation a metabotropic glutamate receptor agonist, inverse agonist or antagonist, and an orexin receptor-related compound, such as and without limitation an orexin receptor agonist, inverse agonist or antagonist, are administered in combination, a therapeutically effective amount of a metabotropic glutamate receptor-related compound or a therapeutically effective amount of an orexin receptor-related compound in combination may be lower than therapeutically effective amounts of metabotropic glutamate receptor-related compound or orexin receptor-related compound when administered alone. That is, the administration of a metabotropic glutamate receptor-related compound and an orexin receptor-related compound in combination may generate a therapeutic effect at what would otherwise be subtherapeutic doses of either.

Medicaments of the invention are, in various aspects, administered by injection, or prepared for oral, pulmonary, nasal or for any other form of administration. Preferably the medicaments are administered, for example, intravenously, subcutaneously, intramuscularly, intraorbital^, ophthalmically, intraventricular^, intracranial^, intracapsularly, intraspinally, intracisternally, intraperitoneally, buccal, rectally, vaginally, intranasally or by aerosol administration.

The mode of administration is in one aspect at least suitable for the form in which the medicament has been prepared. The mode of administration for the most effective response is in one aspect determined empirically and the means of administration described below are given as examples, and do not limit the method of delivery of the composition of the present invention in any way. All the above formulations are commonly used in the pharmaceutical industry and are commonly known to suitably qualified practitioners. The medicaments of the invention in certain aspects include pharmaceutically acceptable nontoxic excipients and carriers and administered by any parenteral techniques such as subcutaneous, intravenous and intraperitoneal injections. In addition the formulations optionally contain one or more adjuvants.

The pharmaceutical forms suitable for injectable use optionally include sterile aqueous solutions (where water-soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion. Alternatively, the compounds of the invention are, in certain aspects, encapsulated in liposomes and delivered in injectable solutions to assist their transport across cell membrane.

Alternatively or in addition such preparations contain constituents of self-assembling pore structures to facilitate transport across the cellular membrane. The carrier, in various aspects, isa solvent or dispersion medium containing, for example, water, ethanol, polyol

(for example, glycerol, propylene glycol and liquid polyethylene glycol, and the like), suitable mixtures thereof, and vegetable oils. Proper fluidity is maintained, for example and without limitation, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants. Prolonged absorption of the injectable compositions is in certain aspects, brought about by the use in the compositions of agents delaying absorption.

Sterile injectable solutions are prepared by incorporating the active compounds in the required amount in an appropriate solvent with various of the other ingredients enumerated above, as required, followed by filtered sterilisation. Generally, dispersions are prepared by incorporating the various sterilised active ingredient into a sterile vehicle that contains the basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, preparation in certain aspects include without limitation vacuum drying and freeze-drying techniques that yield a powder of the active ingredient plus any additional desired ingredient from previously sterile-filtered solution thereof.

Contemplated for use herein are oral solid dosage forms, which are described generally in Martin, Remington's Pharmaceutical Sciences, 18th Ed. (1990 Mack Publishing Co. Easton PA 18042) at Chapter 89, which is herein incorporated by reference. Solid dosage forms include tablets, capsules, pills, troches or lozenges, cachets or pellets. Also, liposomal or proteinoid encapsulation may be used to formulate the present compositions (as, for example, proteinoid microspheres reported in U.S. Patent No. 4,925,673). Liposomal encapsulation may be used and the liposomes may be derivatised with various polymers (E.g., U.S. Patent No. 5,013,556). A description of possible solid dosage forms for the therapeutic is given by Marshall, in Modern Pharmaceutics, Chapter 10, Banker and Rhodes ed., (1979), herein incorporated by reference. In general, the formulation will include the compounds described as part of the invention (or a chemically modified form thereof), and inert ingredients which allow for protection against the stomach environment, and release of the biologically active material in the intestine.

For the metabotropic glutamate receptor-related compounds or orexin receptor-related compounds of the invention the location of release may be the stomach, the small intestine (the duodenum, the jejunum, or the ileum), or the large intestine. One skilled in the art has available formulations that will not dissolve in the stomach, yet will release the material in the duodenum or elsewhere in the intestine. In one aspect, the release will avoid the deleterious effects of the stomach environment, either by protection of the composition or by release of the compounds beyond the stomach environment, such as in the intestine.

To ensure full gastric resistance, a coating impermeable to at least pH 5.0 is used. Examples of the more common inert ingredients that are used as enteric coatings are cellulose acetate trimellitate (CAT), hydroxypropylmethylcellulose phthalate (HPMCP), HPMCP 50, HPMCP 55, polyvinyl acetate phthalate (PVAP), Eudragit L30D, Aquateric, cellulose acetate phthalate (CAP), Eudragit L, Eudragit S, and Shellac. These coatings may be used as mixed films.

A coating or mixture of coatings can also be used on tablets, which are not intended for protection against the stomach. This coating includes, without limitation, sugar coatings, or coatings that make the tablet easier to swallow. Exemplary capsules consist of a hard shell

(such as gelatin) for delivery of dry therapeutic i.e. powder; for liquid forms, a soft gelatin shell may be used. The shell material of cachets in certain aspects is thick starch or other edible paper. For pills, lozenges, moulded tablets or tablet triturates, moist massing techniques are also contemplated, without limitation. In certain aspects, the therapeutic is included in the formulation as fine multiparticulates in the form of granules or pellets of particle size about 1 mm. The formulation of the material for capsule administration is, in certain aspects, a powder, lightly compressed plugs or even as tablets. In one aspect, the therapeutic could be prepared by compression. Colourants and flavouring agents are optionally included. For example, compounds may be formulated (such as, and without limitation, by liposome or microsphere encapsulation) and then further contained within an edible product, such as for example and without limitation a refrigerated beverage containing colorants and flavouring agents.

The volume of the therapeutic is in one aspect, diluted or increased with an inert material. These diluents could include for example and without limitation, carbohydrates, especially mannitol, alpha-lactose, anhydrous lactose, cellulose, sucrose, modified dextrans and starch. Certain inorganic salts are also optionally used as fillers including calcium triphosphate, magnesium carbonate and sodium chloride. Some commercially available diluents are Fast-Flo, Emdex, STA-Rx 1500, Emcompress and Avicell. In other embodiments, disintegrants are included in the formulation of the therapeutic into a solid dosage form. Materials used as disintegrants include but are not limited to starch including the commercial disintegrant based on starch, Explotab. Sodium starch glycolate, Amberlite, sodium carboxymethylcellulose, ultramylopectin, sodium alginate, gelatin, orange peel, acid carboxymethyl cellulose, natural sponge and bentonite are also contemplated. Another form of the disintegrants is the insoluble cationic exchange resins. Powdered gums are also optionally used as disintegrants and as binders and these include, without limitation, powdered gums such as agar, Karaya or tragacanth. Alginic acid and its sodium salt are also useful as disintegrants.

Binders are contemplated to hold the therapeutic compounds together to form a hard tablet and include, without limitation, materials from natural products such as acacia, tragacanth, starch and gelatin. Other binders include, also without limitation, methylcellulose (MC), ethyl cellulose (EC) and carboxymethyl cellulose (CMC). Polyvinyl pyrrolidone (PVP) and hydroxypropylmethyl cellulose (HPMC) are contemplated for use in alcoholic solutions to granulate the therapeutic. Antifrictional agents are optionally included in the formulation of the therapeutic to prevent sticking during the formulation process. Lubricants are optionally used as a layer between the therapeutic and the die wall, and these include but are not limited to: stearic acid including its magnesium and calcium salts, polytetrafluoroethylene (PTFE), liquid paraffin, vegetable oils and waxes. Exemplary soluble lubricants include sodium lauryl sulfate, magnesium lauryl sulfate, polyethylene glycol of various molecular weights, and Carbowax 4000 and 6000.

Glidants that improve the flow properties of the compound during formulation and to aid rearrangement during compression are optionally added. The glidants include without limitation starch, talc, pyrogenic silica and hydrated silicoaluminate.

To aid dissolution of the therapeutic into the aqueous environment, a surfactant is added in certain embodiments as a wetting agent. Surfactants include, for example and without limitation, anionic detergents such as sodium lauryl sulfate, dioctyl sodium sulfosuccinate and dioctyl sodium sulfonate. Cationic detergents are optionally used and include, without limitation, benzalkonium chloride or benzethomium chloride. The list of potential nonionic detergents that are contemplated in the formulation as surfactants are lauromacrogol 400, polyoxyl 40 stearate, polyoxyethylene hydrogenated castor oil 10, 50 and 60, glycerol monostearate, polysorbate 40, 60, 65 and 80, sucrose fatty acid ester, methyl cellulose and carboxymethyl cellulose. When used, these surfactants are present in the formulation of the compounds either alone or as a mixture in different ratios.

Additives that potentially enhance uptake of the compounds are for instance and without limitation the fatty acids oleic acid, linoleic acid and linolenic acid.

Controlled release formulations are also contemplated. In certain aspects, the compounds are incorporated into an inert matrix that permits release by either diffusion or leaching mechanisms i.e., gums. In some aspects, slowly degenerating matrices may also be incorporated into the formulation. Another form of a controlled release of this therapeutic is by a method based on the Oros therapeutic system (Alza Corp.), i.e. the drug is enclosed in a semipermeable membrane which allows water to enter and push drug out through a single small opening due to osmotic effects. Some enteric coatings have a delayed release effect. In other aspects, a mix of materials is used to provide the optimum film coating. Film coating is carried out, for example and without limitation, in a pan coater or in a fluidized bed or by compression coating.

Also contemplated herein is pulmonary delivery of the compounds. In these aspects, he compounds are delivered to the lungs of a mammal while inhaling and traverses across the lung epithelial lining to the blood stream.

Contemplated for use in the practice of this invention are a wide range of mechanical devices designed for pulmonary delivery of therapeutic products, including but not limited to nebulizers, metered-dose inhalers, and powder inhalers, all of which are familiar to those skilled in the art. Some specific examples of commercially available devices suitable for the practice of this invention are, for example and without limitation, the Ultravent nebulizer, manufactured by Mallinckrodt, Inc., St. Louis, Missouri; the Acorn Il nebulizer, manufactured by Marquest Medical Products, Englewood, Colorado; the Ventolin metered dose inhaler, manufactured by Glaxo Inc., Research Triangle Park, North Carolina; and the Spinhaler powder inhaler, manufactured by Fisons Corp., Bedford, Massachusetts.

All such devices require the use of formulations suitable for the dispensing of the compounds. Typically, each formulation is specific to the type of device employed and may involve the use of an appropriate propellant material, in addition to the usual diluents, adjuvants and/or carriers useful in therapy. Also, the use of liposomes, microcapsules or microspheres, inclusion complexes, or other types of earners is contemplated.

Formulations suitable for use with a nebulizer, either jet or ultrasonic, optionally comprise the compounds suspended in water. The formulation also includes, in one aspect, a buffer and a simple sugar (e.g., for protein stabilization and regulation of osmotic pressure). In one embodiment, the nebulizer formulation also contains a surfactant, to reduce or prevent surface induced aggregation of the compounds caused by atomization of the solution in forming the aerosol.

Formulations for use with a metered-dose inhaler device comprise, in one aspect a finely divided powder containing the compounds suspended in a propellant with the aid of a surfactant. The propellant is any conventional material employed for this purpose, such as and without limitation, a chlorofluorocarbon, a hydrochlorofluorocarbon, a hydrofluorocarbon, or a hydrocarbon, including trichlorofluoromethane, dichlorodifluoromethane, dichlorotetrafluoroethanol, and 1,1,1,2-tetrafluoroethane, or combinations thereof. Suitable surfactants include, without limitation sorbitan trioleate and soya lecithin. Oleic acid is also contemplated as a surfactant in certain aspects.

Formulations for dispensing from a powder inhaler device comprise a finely divided dry powder containing the compound and optionally include a bulking agent, such as and without limitation lactose, sorbitol, sucrose, or mannitol in amounts which facilitate dispersal of the powder from the device, e.g., 50 to 90% by weight of the formulation. In certain embodiments, the compound(s) is/are prepared in particulate form with an average particle size of less than 10 microns, most preferably 0.5 to 5 microns, for most effective delivery to the distal lung.

Nasal delivery of the compounds is also contemplated. Nasal delivery allows the passage of the protein to the blood stream directly after administering the therapeutic product to the nose, without the necessity for deposition of the product in the lung. Formulations for nasal delivery include those with, for example and without limitation, dextran or cyclodextran.

It will be appreciated that in certain aspects, the medicaments of the invention are given as a single dose schedule, or in a multiple dose schedule. A multiple dose schedule is one in which a primary course of delivery for example with 1 to 10 separate doses, is optionally followed by other doses given at subsequent time intervals required to maintain or reinforce the treatment. The dosage regimen is, at least in part, determined by the need of the individual and the judgement of the practitioner.

The invention will now be further described by way of reference only to the following non- limiting examples. It should be understood, however, that the examples following are illustrative only, and should not be taken in any way as a restriction on the generality of the invention described above.

EXAMPLES General methods

Briefly, referring to Figures 1 to 3, the IGs are provided in the form of the two receptors (mGluR and OXR). One of the two is attached to an RC (IG1-RC1, IG3). A second IG (IG2-RC2) is derived from a molecule that interacts with the receptors upon ligand binding (e.g. beta-arrestin, or a mutant thereof). The detection system not only detects the formation of the mGluR-OXR heterodimer but can distinguish whether a ligand or drug acts as an agonist, partial agonist, antagonist, inverse agonist, partial inverse agonist or allosteric modulator at the receptor hetero-dimer.

HEK293FT cells were seeded in 6-well plates at a density of approximately 630,000 cells/well and maintained at 37 ⁰C, 5% CO₂ in Complete Media (DMEM containing 0.3 mg/ml glutamine, 100 IU/ml penicillin and 100 μg/ml streptomycin (Gibco)) supplemented with 10% fetal calf serum (FCS; Gibco). Transient transfections were carried out 24 h after seeding using GeneJuice (Novagen) according to manufacturer instructions. 24 h post-transfection, cells were washed with PBS, detached using 0.05% trypsin/0.53 mM EDTA, resuspended in HEPES-buffered phenol red free Complete Media containing 5% FCS and added to a poly-L-lysine-coated white 96-well microplate (Nunc). 48 h post- transfection, eBRET assays were carried out following pre-incubation of cells with EnduRen™ (Promega) at a final concentration of 30 μM, at 37 ⁰C, 5% CO₂ for 2 h. For original BRET studies, the HEPES-buffered phenol red free Complete Media was replaced with PBS and coelenterazine h (Molecular Probes) added to a final concentration of 5 μM immediately prior to commencing the assay. BRET measurements were taken at 37 °C using the Victor Light plate reader with Wallac 1420 software (Perkin-Elmer). Filtered light emissions were sequentially measured for 2-3 s in each of the 'donor wavelength window' (400-475 nm) and 'acceptor wavelength window' (520-540 nm).

The BRET signal observed between interacting proteins is normalized by subtracting the background BRET ratio. This can be done in one of two ways (see Pfleger et al. (2006) Cell Signal 18:1664-1670; Pfleger et al. (2006) Nat Protoc 1 :336-344): 1) the ratio of the 520-540 nm emission over the 400-475 nm emission for a cell sample containing only the donor construct is subtracted from the same ratio for a sample containing the interacting acceptor and donor fusion proteins; 2) the ratio of the 520-540 nm emission over the 400- 475 nm emission for a cell sample treated with vehicle is subtracted from the same ratio for a second aliquot of the same cell sample treated with ligand. In example 1 , the second calculation will be used and the signal is described as the 'ligand-induced BRET ratio'. Alternatively, and particularly when illustrating z'-factor data (Zhang et al (1999) J Biomol Screen 4:67-73), the BRET signal observed between interacting proteins can be shown in conjunction with (as oppose to being subtracted by) the background BRET ratio to evaluate error associated with the BRET signal observed between interacting proteins and the error associated with the background BRET ratio independently. In this case, data are shown as 'fluorescence/luminescence' being the ratio of the 520-540 nm emission over the 400-475 nm emission for a particular cell sample.

EXAMPLE 1 MEASUREMENT OF DETECTABLE SIGNALS INDICATIVE OF THE MOLECULAR ASSOCIATION OF THE METABOTROPIC GLUTAMATE RECEPTOR WITH THE OREXIN RECEPTOR

Referring now to Figure 4, eBRET signals were measured from cells transiently expressing mGluR5/Rluc8 and barr2Λ/enus with either pcDNA3 or hemagglutin epitope- tagged OxR2R (HA-OxR2) following treatment with either 10^"6M orexin (OxA).

Prior to ligand treatment (added at 0 minutes), a baseline eBRET signal was recorded for each of the combinations. OxA treatment of cells co-expressing mGluR5/Rluc8 and barr2A/enus with pcDNA3 did not result in a ligand-induced BRET signal. A BRET signal was observed following OxA treatment of cells co-expressing mGluR5/Rluc8, barr2/Venus and HA-OxR2. This signal reached a maximum of approximately 0.06.

This example demonstrates that a signal resulting from the proximity of RC1 and RC2 is detected specifically for the combination where the metabotropic glutamate receptor 5 (mGluR5) as IG1 , Rlucδ as RC1 , beta-arrestin 2 (barr2) as IG2, Venus as RC2 and hemagglutin epitope-tagged OxR2 (HA-OxR2) as IG3, and when the modulator, in this case OxA, modulates the association of IG2 and IG3 as a result of interacting specifically with IG3.

This example demonstrates that the inventors have identified the molecular association of the metabotropic glutamate receptor with the orexin receptor.

This example also demonstrates that IG3 can be tagged, such as by the addition of a hemagglutin (HA) epitope-tag, however, this tag does not constitute a reporter component and does not interfere with and/or contribute to the signal generated by the proximity of RC1 and RC2. Such tagging enables additional information to be ascertained, such as the relative expression level of IG3.

EXAMPLE 2 MEASUREMENT OF A DETECTABLE SIGNAL INDICATIVE OF THE MOLECULAR ASSOCIATION OF THE METABOTROPIC GLUTAMATE RECEPTOR

WITH THE OREXIN RECEPTOR WITH A Z'-FACTOR OF 0.52

Referring now to Figures 5, 6 and 7. BRET signals using coelenterazine h were measured from cells transiently co-expressing mGluRδ/Rlucδ and barτ2/Venus with HA-

OxR2 aliquoted into all wells of a 96-well plate. Phosphate-buffered saline (PBS) was added to the first two rows and the last two rows of the 96-well plate (48 wells in total) as a vehicle control. OxA was added to the middle four rows of the 96-well plate (48 wells in total). Data presented as fluorescence/luminescence.

OxA treatment of cells co-expressing mGluR5/Rluc8 and barr2/Venus with HA-OxR2 resulted in higher fluorescence/luminescence ratios (Figure 6) than those observed following treatment with phosphate-buffered saline (PBS) vehicle control (Figure 5). Analysis of the fluorescence/luminescence ratios comparing 48-wells treated with OxA (defined as 'signal' with respect to z'-factor calculation) and 48-wells treated with PBS (defined as 'background' with respect to z'-factor calculation) results in a z'-factor of 0.52 (Figure 7) using the calculation described by Zhang et al., 1999 (J Biomol Screen 4:67- 73). Means are shown as solid lines and 3 standard deviations from the mean are shown as dotted lines.

This example demonstrates that a signal resulting from the proximity of RC1 and RC2 is detected specifically for the combination where mGluR5 is IG1 , Rluc8 is RC1 , beta- arrestin 2 (barr2) is IG2, Venus is RC2 and HA-0xR2 is IG3, and when the modulator, in this case OxA, modulates the association of IG2 and IG3 as a result of interacting specifically with IG3.

This example also demonstrates that the molecular association of the metabotropic glutamate receptor with the orexin receptor is detected in a manner that results in a z¹- factor of 0.52 and is therefore amenable to high-throughput screening.

This example further demonstrates an additional method of representing BRET data that can be used in demonstrating a detectable signal indicative of the molecular association of the metabotropic glutamate receptor and the orexin receptor.

Claims

The Claims Defining the Invention are as Follows:

1. A hetero-dimeric or hetero-oligomeric receptor, comprising at least one metabotropic glutamate receptor subunit associated with at least one orexin receptor subunit.

2. A method for the treatment of a patient suffering from an orexin-related ailment by administering a therapeutically effective amount of a metabotropic glutamate receptor-related compound.

3. A method according to claim 2 characterised in that the metabotropic glutamate receptor-related compound is selective for the metabotropic glutamate receptor relative to the orexin receptor.

4. A method for the treatment of a patient suffering from a glutamate-related ailment by administering a therapeutically effective amount of an orexin receptor-related compound.

5. A method according to claim 4 characterised in that the orexin receptor-related compound is selective for the orexin receptor relative to the metabotropic glutamate receptor.

6. A method for screening a test compound for potential therapeutic activity against an orexin-related ailment using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) exposing the test compound to a metabotropic glutamate receptor; b) assessing whether and/or the extent to which the activity of the metabotropic glutamate receptor is modulated compared to activity of the metabotropic glutamate receptor in the absence of the test compound; said modulation being indicative of potential therapeutic activity against the orexin- related ailment.

7. A method for screening a test compound for potential therapeutic activity against a glutamate-related ailment using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) exposing the test compound to an orexin receptor; b) assessing whether and/or the extent to which the activity of the orexin receptor is modulated compared to activity of the orexin receptor in the absence of the test compound; said modulation being indicative of potential therapeutic activity against the glutamate-related ailment.

8. A method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective activity using a detector capable of detecting changes in receptor activity, the method comprising the step of: determining whether, and/or the extent to which, the test compound interacts with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor compared to whether, and/or the extent to which the test compound interacts with the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater affinity and/or potency and/or efficacy when interacting with the orexin receptor while the orexin receptor is associated with the metabotropic glutamate receptor is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

9. A method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective activity using a detector capable of detecting changes in receptor activity, the method comprising the step of: determining whether, and/or the extent to which, the test compound interacts with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor compared to whether, and/or the extent to which the test compound interacts with the metabotropic glutamate receptor in the absence of the orexin receptor; such that a test compound that exhibits greater affinity and/or potency and/or efficacy when interacting with the metabotropic glutamate receptor while the metabotropic glutamate receptor is associated with the orexin receptor is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

10. A method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective antagonism or selective partial agonism or selective negative allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component;

Ni). a third agent, comprising the metabotropic glutamate receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether, or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater antagonistic, partial agonistic or negative allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero- dimer/-oligomer.

11. A method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective antagonism, selective partial agonism or selective negative allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the orexin receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b). detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether, or the extent to which the test compound is an antagonist, partial agonist or negative allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater antagonistic, partial agonistic or negative allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

12. A method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective inverse agonism using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising a constitutively active metabotropic glutamate receptor; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally ; c) determining whether, or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater inverse agonistic properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate / orexin receptor hetero-dimer/-oligomer.

13. A method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer inverse agonism using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether, and/or the extent to which, the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising a constitutively active orexin receptor; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b) detecting a decrease in the signal as a determination of whether and/or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether, or the extent to which the test compound is an inverse agonist of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater inverse agonistic properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer.

14. A method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective positive allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the orexin receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the metabotropic glutamate receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the metabotropic glutamate receptor; b) detecting an increase in the signal as a determination of whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater positive allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate / orexin receptor hetero-dimer/- oligomer.

15. A method for screening a test compound for metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer selective positive allosteric modulation using a detector capable of detecting changes in receptor activity, the method comprising the steps of: a) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer, by contacting said test compound with a system comprising: i). a first agent, comprising the metabotropic glutamate receptor coupled to a first reporter component; ii). a second agent, comprising an interacting group coupled to a second reporter component; iii). a third agent, comprising the orexin receptor; iv). an agonist of the orexin receptor, the metabotropic glutamate receptor and/or the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer; wherein proximity of the first and second reporter components generates a signal; and wherein the modulator modulates the association of the interacting group with the orexin receptor; b) detecting an increase in the signal as a determination of whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer and optionally; c) determining whether and/or the extent to which the test compound is a positive allosteric modulator of the metabotropic glutamate receptor in the absence of the orexin receptor and the orexin receptor in the absence of the metabotropic glutamate receptor; such that a test compound that exhibits greater positive allosteric modulator properties when interacting with the metabotropic glutamate receptor / orexin receptor hetero-dimer/-oligomer is selective for the metabotropic glutamate receptor / orexin receptor hetero- dimer/-oligomer.

16. Selective agonists and/or selective antagonists and/or selective inverse agonists and/or selective allosteric modulators of the metabotropic glutamate receptor/orexin receptor hetero-dimer/-oligomer.

17. A cell, or fraction of a cell, in which both a metabotropic glutamate receptor and an orexin receptor are over-expressed.

18. A cell, or fraction of a cell, in which a metabotropic glutamate receptor is over- expressed with an endogenously expressed orexin receptor.

19. A cell, or fraction of a cell, in which an orexin receptor is over-expressed with an endogenously expressed metabotropic glutamate receptor.