Remedies for life style-related diseases or cibophobia and method of screening the same
The present invention provides a novel therapeutic agent for a lifestyle-related disease or cibophobia, which is superior in controlling food intake, and a screening method therefor. More specifically, a therapeutic agent for a lifestyle-related disease containing, as an active ingredient, a substance that suppresses expression or function of GPCR that expresses in hypothalamus, clone 901, a therapeutic agent for cibophobia containing, as an active ingredient, a substance that enhances expression or function of the clone, a screening system consisting of a series of coexpression systems of clone 901 and various G proteins, and a screening method for a substance having a therapeutic activity against a lifestyle-related disease or cibophobia, which includes use of the screening system, are provided.
1 . A therapeutic agent for a lifestyle-related disease comprising, as an active ingredient, a substance that suppresses expression or function of a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2.
2 . A therapeutic agent for a lifestyle-related disease comprising, as an active ingredient, a nucleic acid of the following:
(a) a nucleic acid consisting of a base sequence complementary to a base sequence shown in SEQ ID NO:1, or
(b) a nucleic acid consisting of a base sequence capable of hybridizing with a nucleic acid consisting of a base sequence shown in SEQ ID NO:1 or a primary transcript which generates said base sequence after post-transcriptional processing under physiological conditions of hypothalamus of a subject animal for treatment, and which is capable of inhibiting translation into a polypeptide encoded by the base sequence shown in SEQ ID NO:1 under a hybridized state.
3 . A therapeutic agent for a lifestyle-related disease comprising, as an active ingredient, a substance which shows a specific affinity for a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2 and which inhibits a functional expression of said polypeptide.
4 . The therapeutic agent of claim 3 , wherein the substance is a nucleic acid.
5 . The therapeutic agent of claim 3 , wherein the substance is an antibody.
6 . A therapeutic agent for a lifestyle-related disease comprising, as an active ingredient, an expression vector encoding the nucleic acid of claim 2 .
7 . A therapeutic agent for a lifestyle-related disease comprising, as an active ingredient, a host cell transfected with the expression vector of claim 6 .
8 . The therapeutic agent of any of claims 1 to 7 , which is a feeding suppressant, an anti-obesity agent, an anti-diabetic agent or an anti-hyperlipidemic agent.
9 . A therapeutic agent for cibophobia comprising, as an active ingredient, a substance that enhances expression or function of a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2.
10 . A therapeutic agent for cibophobia comprising, as an active ingredient, a polypeptide of the following:
(a) a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, or
(b) a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, wherein one or more amino acids of the amino acid sequence have been substituted, deleted, inserted, added or modified, which shows a ligand—receptor interaction of the same level as the polypeptide of (a), and which is coupled with a G protein α subunit and shows an activity to promote a GDP/GTP exchange reaction of the subunit.
11 . A therapeutic agent for cibophobia, comprising, as an active ingredient, an expression vector comprising a nucleic acid encoding the polypeptide of claim 10 .
12 . A therapeutic agent for cibophobia comprising, as an active ingredient, a host cell transfected with the expression vector of claim 11 .
13 . A screening system for a substance having a therapeutic activity against a lifestyle-related disease or cibophobia, which comprises, as one constitution unit, a system comprising, as constituent elements, a lipid bilayer membrane comprising a polypeptide of the following:
(a) a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, or
(b) a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, wherein one or more amino acids of the amino acid sequence have been substituted, deleted, inserted, added or modified, which shows a ligand—receptor interaction of the same level as the polypeptide of (a), and which is coupled with a G protein α subunit and shows an activity to promote a GDP/GTP exchange reaction of the subunit, and a polypeptide comprising at least a receptor-binding region of a G protein α subunit belonging to a certain family and a guanine nucleotide-binding region of any G protein α subunit, wherein said constitution unit is present in a receptor-binding region of each family of the G protein α subunit.
14 . The screening system of claim 13 , wherein the constitution unit comprises an eucaryotic host cell transfected with an expression vector comprising a DNA encoding the polypeptide of (a) or (b), and an expression vector comprising a DNA encoding a polypeptide comprising at least a receptor-binding region of a G protein α subunit belonging to a certain family and a guanine nucleotide-binding region of any G protein α subunit, a homogenate of said cell or a membrane fraction derived from said cell.
15 . The screening system of claim 13 , wherein the constitution unit comprises an eucaryotic host cell transfected with an expression vector comprising a DNA encoding a polypeptide obtained by fusion of a polypeptide comprising, on a C terminal of the polypeptide of (a) or (b), at least a receptor-binding region of a G protein α subunit belonging to a certain family and a guanine nucleotide-binding region of any G protein a subunit, a homogenate of said cell or a membrane fraction derived from said cell.
16 . The screening system of claim 13 , wherein the polypeptide in each constitution unit, which comprises a receptor-binding region of a G protein α subunit and a guanine nucleotide-binding region of any G protein α subunit, further comprises the same effector interaction region and the lipid bilayer membrane further comprises an effector that interacts with said region.
17 . The screening system of any of claims 13 to 16 , wherein the therapeutic activity against lifestyle-related diseases is a feeding suppressive activity, an anti-obesity activity, an anti-diabetic activity or an anti-hyperlipidemic activity.
18 . A screening method for a substance having a therapeutic activity against a lifestyle-related disease or cibophobia, which comprises adding, in each constitution unit of the screening system of claim 13 , a labeled GTP analog in the presence of a test substance and in the absence of the test substance and comparing an amount of the label bound with a guanine nucleotide-binding region under the both conditions.
19 . A screening method for a substance having a therapeutic activity against a lifestyle-related disease or cibophobia, which comprises comparing, in each constitution unit of the screening system of claim 16 , an activity of the effector in the presence of a test substance and in the absence of the test substance.
20 . A method for identifying a G protein α subunit capable of coupling with a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, which comprises adding, in each constitution unit of the screening system of claim 13 , a labeled GTP analog in the presence of a ligand for said polypeptide and in the absence of said ligand, and comparing an amount of the label bound with a guanine nucleotide-binding region among constitution units.
21 . A method for identifying a G protein α subunit capable of coupling with a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, which comprises comparing an activity of the effector in each constitution unit of the screening system of claim 16 in the presence of a ligand for said polypeptide and in the absence of said ligand.
22 . A screening method for a substance having a therapeutic activity against a lifestyle-related disease or cibophobia, which comprises applying the method of claim 18 or 19 only to a system comprising, as a constituent element, a polypeptide comprising a receptor-binding region of the G protein α subunit as identified by the method of claim 20 or 21 .
23 . The method of claim 22 , wherein the G protein α subunit belongs to a Gs family.
24 . A screening system for a ligand for a polypeptide of the following:
(a) a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, or
(b) a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, wherein one or more amino acids of the amino acid sequence have been substituted, deleted, inserted, added or modified, which shows a ligand—receptor interaction of the same level as the polypeptide of (a), and which is coupled with a G protein α subunit belonging to a Gs family and shows an activity to promote a GDP/GTP exchange reaction of the subunit, which comprises, as constituent elements, a lipid bilayer membrane comprising said polypeptide and a polypeptide comprising at least a receptor-binding region of a G protein α subunit belonging to a Gs family and a guanine nucleotide-binding region of any G protein α subunit.
25 . The screening system of claim 24 , which comprises an eucaryotic host cell transfected with an expression vector comprising a DNA encoding the polypeptide of (a) or (b), and an expression vector comprising a DNA encoding a polypeptide comprising at least a receptor-binding region of a G protein α subunit belonging to a Gs family and a guanine nucleotide-binding region of any G protein α subunit, a homogenate of said cell or a membrane fraction derived from said cell.
26 . The screening system of claim 24 , which comprises an eucaryotic host cell transfected with an expression vector comprising a DNA encoding a polypeptide fused with a polypeptide comprising, on a C terminal side of said polypeptide, (a) or (b), at least a receptor-binding region of a G protein α subunit belonging to a Gs family and a guanine nucleotide-binding region of any G protein α subunit, a homogenate of said cell or a membrane fraction derived from said cell.
27 . The screening system of claim 24 , wherein the polypeptide comprising a receptor-binding region of a G protein α subunit belonging to a Gs family and a guanine nucleotide-binding region of any G protein α subunit further comprises any effector interacting region and the lipid bilayer membrane further comprises an effector that interacts with said region.
28 . The screening system of claim 27 , wherein the effector is adenyl cyclase.
29 . The screening system of claim 24 , which is a system for searching a substance having a therapeutic activity against a lifestyle-related disease or cibophobia.
30 . The screening system of claim 29 , wherein the therapeutic activity against a lifestyle-related disease is a feeding suppressive activity, an anti-obesity activity, an anti-diabetic activity or an anti-hyperlipidemic activity.
31 . A screening method for a ligand for a polypeptide of the following:
(a) a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, or
(b) a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, wherein one or more amino acids of the amino acid sequence have been substituted, deleted, inserted, added or modified, which shows a ligand—receptor interaction of the same level as the polypeptide of (a), and which is coupled with a G protein α subunit belonging to a Gs family and shows an activity to promote a GDP/GTP exchange reaction of the subunit, which comprises adding, in the screening system of claim 24 and in the presence of a test substance and in the absence of the test substance, a labeled GTP analog, and comparing the amount of the label bound with a guanine nucleotide-binding region under the both conditions.
32 . A screening method for a ligand for a polypeptide of the following:
(a) a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, or
(b) a polypeptide consisting of an amino acid sequence shown in SEQ ID NO:2, wherein one or more amino acids of the amino acid sequence have been substituted, deleted, inserted, added or modified, which shows a ligand—receptor interaction of the same level as the polypeptide of (a), and which is coupled with a G protein α subunit belonging to a Gs family and shows an activity to promote a GDP/GTP exchange reaction of the subunit, which comprises comparing an activity of the effector in the screening system of claim 27 in the presence of a test substance and in the absence of the test substance.
33 . The screening method of claim 32 , which comprises comparing an amount of cAMP in an eucaryotic host cell in the presence of a test substance and in the absence of the test substance.
34 . The screening system of claim 31 , which is a system for searching a substance having a therapeutic activity against a lifestyle-related disease or cibophobia.
35 . The screening system of any of claims 18 , 19 , 34 , 38 , and 39 , wherein the therapeutic activity against a lifestyle-related disease is a feeding suppressive activity, an anti-obesity activity, an anti-diabetic activity or an anti-hyperlipidemic activity.
36 . A therapeutic agent for a lifestyle-related disease comprising, as an active ingredient, a substance having a therapeutic activity against a lifestyle-related disease, which is obtained by the screening method of any of claims 13 - 16 , 18 , 19 , 29 , 34 , 38 , and 39 .
37 . A therapeutic agent for cibophobia comprising, as an active ingredient, a substance having a therapeutic activity against cibophobia, which is obtained by the screening method of any of claims 13 - 16 , 18 , 19 , 29 , 34 , 38 and, 39 .
38 . The screening system of claim 32 , which is a system for searching a substance having a therapeutic activity against a lifestyle-related disease or cibophobia.
39 . The screening system of claim 33 , which is a system for searching a substance having a therapeutic activity against a lifestyle-related disease or cibophobia.
40 . The screening system of claim 22 , wherein the therapeutic activity against a lifestyle-related disease is a feeding suppressive activity, an anti-obesity activity, an anti-diabetic activity or an anti-hyperlipidemic activity.
41 . A therapeutic agent for a lifestyle-related disease comprising, as an active ingredient, a substance having a therapeutic activity against a lifestyle-related disease, which is obtained by the screening method of claim 22 .
42 . A therapeutic agent for cibophobia comprising, as an active ingredient, a substance having a therapeutic activity against cibophobia, which is obtained by the screening method of claim 22.