US2003175883A1PendingUtilityA1
DNA encoding a mammalian LPA receptor and uses thereof
Priority: Dec 10, 1997Filed: Jul 26, 2002Published: Sep 18, 2003
Est. expiryDec 10, 2017(expired)· nominal 20-yr term from priority
Inventors:Jonathan Bard
C07K 14/705A61K 38/00A01K 2217/05
56
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Claims
Abstract
This invention provides an isolated nucleic acid encoding a mammalian LPA receptor, a purified mammalian LPA receptor, vectors comprising isolated nucleic acid encoding an mammalian LPA receptor, cells comprising such vectors, antibodies directed to a mammalian LPA receptor, nucleic acid probes useful for detecting nucleic acid encoding a mammalian LPA receptor, antisense oligonucleotides complementary to unique sequences of nucleic acid encoding mammalian LPA receptor, transgenic, nonhuman animals which express DNA encoding a normal or a mutant mammalian LPA receptor, methods of isolating an mammalian LPA receptor, methods of treating an abnormality that is linked to the activity of the mammalian LPA receptor, as well as methods of determining binding of compounds to mammalian LPA receptors.
Claims
exact text as granted — not AI-modifiedWhat is claimed is:
1 . An isolated nucleic acid encoding a mammalian LPA receptor.
2 . The nucleic acid of claim 1 , wherein the nucleic acid is DNA.
3 . The DNA of claim 2 , wherein the DNA is cDNA.
4 . The DNA of claim 2 , wherein the DNA is genomic DNA.
5 . The nucleic acid of claim 1 , wherein the nucleic acid is RNA.
6 . The nucleic acid of claim 1 , wherein the mammalian LPA receptor is a human LPA receptor.
7 . The nucleic acid of claim 1 , wherein the nucleic acid encodes a mammalian LPA receptor which has substantially the same amino acid sequence as does the mammalian LPA receptor encoded by the plasmid HL-18a (ATCC Accession No. 209448).
8 . The nucleic acid of claim 6 , wherein the nucleic acid encodes a human LPA receptor which has an amino acid sequence identical to that encoded by the plasmid HL-18a (ATCC Accession No. 209448).
9 . The nucleic acid of claim 6 , wherein the human LPA receptor has an amino acid sequence, substantially the same as the amino acid sequence shown in FIG. 3 (Seq. I.D. No. 3).
10 . The nucleic acid of claim 9 , wherein the human LPA receptor has an amino acid sequence identical to the amino acid sequence shown in FIG. 3 (Seq. I.D. No. 3).
11 . A purified mammalian LPA receptor protein.
12 . A vector comprising the nucleic acid of claim 1 .
13 . A vector comprising the nucleic acid of claim 6 .
14 . A vector of claim 12 adapted for expression in a bacterial cell which comprises the regulatory elements necessary for expression of the nucleic acid in the bacterial cell operatively linked to the nucleic acid encoding the mammalian LPA receptor so as to permit expression thereof.
15 . A vector of claim 12 adapted for expression in an amphibian cell which comprises the regulatory elements necessary for expression of the nucleic acid in the amphibian cell operatively linked to the nucleic acid encoding the mammalian LPA receptor so as to permit expression thereof.
16 . A vector of claim 12 adapted for expression in a yeast cell which comprises the regulatory elements necessary for expression of the nucleic acid in the yeast cell operatively linked to the nucleic acid encoding the mammalian LPA receptor so as to permit expression thereof.
17 . A vector of claim 12 adapted for expression in an insect cell which comprises the regulatory elements necessary for expression of the nucleic acid in the insect cell operatively linked to the nucleic acid encoding the mammalian LPA receptor so as to permit expression thereof.
18 . The vector of claim 17 which is a baculovirus.
19 . A vector of claim 12 adapted for expression in a mammalian cell which comprises the regulatory elements necessary for expression of the nucleic acid in the mammalian cell operatively linked to the nucleic acid encoding the mammalian LPA receptor so as to permit expression thereof.
20 . The vector of claim 12 , wherein the vector is a plasmid.
21 . The plasmid of claim 20 designated HL-18a (ATCC Accession No. 209448).
22 . A cell comprising the vector of claim 12 .
23 . A cell of claim 22 , wherein the cell is a non-mammalian cell.
24 . A cell of claim 23 , wherein the non-mammalian cell is a Xenopus oocyte cell or a Xenopus melanophore cell.
25 . A cell of claim 22 , wherein the cell is a mammalian cell.
26 . A mammalian cell of claim 25 , wherein the cell is a COS-7 cell, a 293 human embryonic kidney cell, a NIH-3T3 cell, a LM(tk−) cell, a mouse Y1 cell, or a CHO cell.
27 . An insect cell comprising the vector of claim 17 .
28 . An insect cell of claim 27 , wherein the insect cell is an Sf9 cell, an Sf21 cell or a HighFive cell.
29 . A membrane preparation isolated from the cell of any one of claims 22 , 23 , 25 or 27 .
30 . A nucleic acid probe comprising at least 15 nucleotides, which probe specifically hybridizes with a nucleic acid encoding a mammalian LPA receptor, wherein the probe has a unique sequence corresponding to a sequence present within one of the two strands of nucleic acid which encode the mammalian LPA receptor and are contained in plasmid HL-18a (ATCC Accession No. 209448).
31 . A nucleic acid probe comprising at least 15 nucleotides, which probe specifically hybridizes with a nucleic acid encoding a mammalian LPA receptor, wherein the probe has a unique sequence corresponding to a sequence present within (a) the nucleic acid sequence shown in FIG. 2 (Seq. I.D. No. 2) or (b) the reverse complement thereto.
32 . The nucleic acid probe of claim 30 or 31 , wherein the nucleic acid is DNA.
33 . The nucleic acid probe of claim 30 or 31 , wherein the nucleic acid is RNA.
34 . A nucleic acid probe comprising a nucleic acid molecule of at least 15 nucleotides which is complementary to a unique fragment of the sequence of a nucleic acid molecule encoding a mammalian LPA receptor.
35 . A nucleic acid probe comprising a nucleic acid molecule of at least 15 nucleotides which is complementary to the antisense sequence of a unique fragment of the sequence of a nucleic acid molecule encoding a mammalian LPA receptor.
36 . An antisense oligonucleotide having a sequence capable of specifically hybridizing to the RNA of claim 5 , so as to prevent translation of the RNA.
37 . An antisense oligonucleotide having a sequence capable of specifically hybridizing to the genomic DNA of claim 4 .
38 . An antisense oligonucleotide of claim 36 or 37 , wherein the oligonucleotide comprises chemically modified nucleotides or nucleotide analogues.
39 . An antibody capable of binding to a mammalian LPA receptor encoded by the nucleic acid of claim 1 .
40 . An antibody of claim 39 , wherein the mammalian LPA receptor is a human LPA receptor.
41 . An agent capable of competitively inhibiting the binding of the antibody of claim 39 to a mammalian LPA receptor.
42 . The antibody of claim 39 , wherein the antibody is a monoclonal antibody or antisera.
43 . A pharmaceutical composition comprising (a) an amount of the oligonucleotide of claim 36 capable of passing through a cell membrane and effective to reduce expression of a mammalian LPA receptor and (b) a pharmaceutically acceptable carrier capable of passing through the cell membrane.
44 . A pharmaceutical composition of claim 43 , wherein the oligonucleotide is coupled to a substance which inactivates mRNA.
45 . A pharmaceutical composition of claim 44 , wherein the substance which inactivates mRNA is a ribozyme.
46 . A pharmaceutical composition of claim 43 , wherein the pharmaceutically acceptable carrier comprises a structure which binds to a mammalian LPA receptor on a cell capable of being taken up by the cells after binding to the structure.
47 . A pharmaceutical composition of claim 46 , wherein the pharmaceutically acceptable carrier is capable of binding to a mammalian LPA receptor which is specific for a selected cell type.
48 . A pharmaceutical composition which comprises an amount of the antibody of claim 39 effective to block binding of a ligand to a human LPA receptor and a pharmaceutically acceptable carrier.
49 . A transgenic, nonhuman mammal expressing the DNA encoding a mammalian LPA receptor of claim 1 .
50 . A transgenic, nonhuman mammal comprising a homologous recombination knockout of the native mammalian LPA receptor.
51 . A transgenic, nonhuman mammal whose genome comprises antisense DNA complementary to the DNA encoding a mammalian LPA receptor of claim 1 so placed within the genome as to be transcribed into antisense mRNA which is complementary to mRNA encoding the mammalian LPA receptor and which hybridizes to mRNA encoding the mammalian LPA receptor, thereby reducing its translation.
52 . The transgenic, nonhuman mammal of either claim 49 or 50 , wherein the DNA encoding the mammalian LPA receptor additionally comprises an inducible promoter.
53 . The transgenic, nonhuman mammal of claim 49 or 50 , wherein the DNA encoding the mammalian LPA receptor additionally comprises tissue specific regulatory elements.
54 . A transgenic, nonhuman mammal of claim 49 , 50 , or 51 , wherein the transgenic, nonhuman mammal is a mouse.
55 . A process for identifying a chemical compound which specifically binds to a mammalian LPA receptor which comprises contacting cells containing DNA encoding and expressing on their cell surface the mammalian LPA receptor, wherein such cells do not normally express the mammalian LPA receptor, with the compound under conditions suitable for binding, and detecting specific binding of the chemical compound to the mammalian LPA receptor.
56 . A process for identifying a chemical compound which specifically binds to a mammalian LPA receptor which comprises contacting a membrane fragment from a cell extract of cells containing DNA encoding and expressing on their cell surface the mammalian LPA receptor, wherein such cells do not normally express the mammalian LPA receptor, with the compound under conditions suitable for binding, and detecting specific binding of the chemical compound to the mammalian LPA receptor.
57 . The process of claim 55 or 56 , wherein the mammalian LPA receptor is a human LPA receptor.
58 . The process of claim 55 or 56 , wherein the mammalian LPA receptor has substantially the same amino acid sequence as the mammalian LPA receptor encoded by plasmid HL-18a (ATCC Accession No. 209448).
59 . The process of claim 55 or 56 , wherein the mammalian LPA receptor has substantially the same amino acid sequence as that shown in FIG. 3 (Seq. I.D. No. 3).
60 . The process of claim 55 or 56 , wherein the mammalian LPA receptor has the amino acid sequence shown in FIG. 3 (Seq. I.D. No. 3).
61 . The process of claim 59 , wherein the compound is a compound not previously known to bind to a mammalian LPA receptor.
62 . A compound determined by the process of claim 61 .
63 . A process of claim 59 , wherein the cell is an insect cell.
64 . The process of claim 59 , wherein the cell is a mammalian cell.
65 . The process of claim 64 , wherein the cell is non-neuronal in origin.
66 . The process of claim 65 , wherein the non-neuronal cell is a COS-7 cell, 293 human embryonic kidney cell, a CHO cell, a NIH-3T3 cell, a mouse Y1 cell, or a LM(tk−) cell.
67 . A process of claim 64 , wherein the compound is a compound not previously known to bind to a mammalian LPA receptor.
68 . A compound determined by the process of claim 67 .
69 . A process involving competitive binding for identifying a chemical compound which specifically binds to a mammalian LPA receptor which comprises separately contacting cells expressing on their cell surface the mammalian LPA receptor, wherein such cells do not normally express the mammalian LPA receptor, with both the chemical compound and a second chemical compound known to bind to the receptor, and with only the second chemical compound, under conditions suitable for binding of both compounds, and detecting specific binding of the chemical compound to the mammalian LPA receptor, a decrease in the binding of the second chemical compound to the mammalian LPA receptor in is the presence of the chemical compound indicating that the chemical compound binds to the mammalian LPA receptor.
70 . A process involving competitive binding for identifying a chemical compound which specifically binds to a mammalian LPA receptor which comprises separately contacting a membrane fraction from a cell extract of cells expressing on their cell surface the mammalian LPA receptor, wherein such cells do not normally express the mammalian LPA receptor, with both the chemical compound and a second chemical compound known to bind to the receptor, and with only the second chemical compound, under conditions suitable for binding of both compounds, and detecting specific binding of the chemical compound to the mammalian LPA receptor, a decrease in the binding of the second chemical compound to the mammalian LPA receptor in the presence of the chemical compound indicating that the chemical compound binds to the mammalian LPA receptor.
71 . A process of claim 69 or 70 , wherein the mammalian LPA receptor is a human LPA receptor.
72 . The process of claim 71 , wherein the human LPA receptor has substantially the same amino acid sequence as the human LPA receptor encoded by plasmid HL-18a (ATCC Accession No. 209448).
73 . The process of claim 69 or 70 , wherein the mammalian LPA receptor has substantially the same amino acid sequence as that shown in FIG. 3 (Seq. I.D. No. 3).
74 . The process of claim 69 or 70 , wherein the mammalian LPA receptor has the amino acid sequence shown in FIG. 3 (Seq. I.D. No. 3).
75 . The process of claim 73 , wherein the cell is an insect cell.
76 . The process of claim 73 , wherein the cell is a mammalian cell.
77 . The process of claim 76 , wherein the cell is non-neuronal in origin.
78 . The process of claim 77 , wherein the non-neuronal cell is a COS-7 cell, 293 human embryonic kidney cell, a CHO cell, a NIH-3T3 cell, a mouse Y1 cell, or a LM(tk−) cell.
79 . The process of claim 78 , wherein the compound is a compound not previously known to bind to a mammalian LPA receptor.
80 . A compound determined by the process of claim 79 .
81 . A method of screening a plurality of chemical compounds not known to bind to a mammalian LPA receptor to identify a compound which specifically binds to the mammalian LPA receptor, which comprises
(a) contacting cells transfected with and expressing DNA encoding the mammalian LPA receptor with a compound known to bind specifically to the mammalian LPA receptor; (b) contacting the preparation of step (a) with the plurality of compounds not known to bind specifically to the mammalian LPA receptor, under conditions permitting binding of compounds known to bind the mammalian LPA receptor; (c) determining whether the binding of the compound known to bind to the mammalian LPA receptor is reduced in the presence of the compounds within the plurality of compounds, relative to the binding of the compound in the absence of the plurality of compounds; and if so (d) separately determining the binding to the mammalian LPA receptor of compounds included in the plurality of compounds, so as to thereby identify the compound which specifically binds to the mammalian LPA receptor.
82 . A method of screening a plurality of chemical compounds not known to bind to a mammalian LPA receptor to identify a compound which specifically binds to the mammalian LPA receptor, which comprises
(a) preparing a cell extract from cells transfected with and expressing DNA encoding the mammalian LPA receptor, isolating a membrane fraction from the cell extract, contacting the membrane fraction with a compound known to bind specifically to the mammalian LPA receptor; (b) contacting the preparation of step (a) with the plurality of compounds not known to bind specifically to the mammalian LPA receptor, under conditions permitting binding of compounds known to bind the mammalian LPA receptor; (c) determining whether the binding of the compound known to bind to the mammalian LPA receptor is reduced in the presence of the compounds within the plurality of compounds, relative to the binding of the compound in the absence of the plurality of compounds; and if so (d) separately determining the binding to the mammalian LPA receptor of compounds included in the plurality of compounds, so as to thereby identify the compound which specifically binds to the mammalian LPA receptor.
83 . A method of either of claims 81 or 82 , wherein the mammalian LPA receptor is a human LPA receptor.
84 . A method of either of claims 81 or 82 , wherein the cell is a mammalian cell.
85 . A method of claim 84 , wherein the mammalian cell is non-neuronal in origin.
86 . The method of claim 85 wherein the non-neuronal cell is a COS-7 cell, a 293 human embryonic kidney cell, a LM(tk−) cell, a CHO cell, a mouse Y1 cell, or an NIH-3T3 cell.
87 . A method of detecting expression of a mammalian LPA receptor by detecting the presence of mRNA coding for the mammalian LPA receptor which comprises obtaining total mRNA from the cell and contacting the mRNA so obtained from the nucleic acid probe of claim 30 under hybridizing conditions, detecting the presence of mRNA hybridizing to the probe, and thereby detecting the expression of the mammalian LPA receptor by the cell.
88 . A method of detecting the presence of a mammalian LPA receptor on the surface of a cell which comprises contacting the cell with the antibody of claim 39 under conditions permitting binding of the antibody to the receptor, detecting the presence of the antibody bound to the cell, and thereby detecting the presence of a mammalian LPA receptor on the surface of the cell.
89 . A method of determining the physiological effects of varying levels of activity of mammalian LPA receptors which comprises producing a transgenic, nonhuman mammal of claim 52 whose levels of mammalian LPA receptor activity are varied by use of an inducible promoter which regulates mammalian LPA receptor expression.
90 . A method of determining the physiological effects of varying levels of activity of mammalian LPA receptors which comprises producing a panel of transgenic, nonhuman mammals of claim 52 each expressing a different amount of mammalian LPA receptor.
91 . A method for identifying an antagonist capable of alleviating an abnormality wherein the abnormality is alleviated by decreasing the activity of a mammalian LPA receptor comprising administering a compound to the transgenic, nonhuman mammal of any one of claims 49 , 52 , 53 , or 54 , and determining whether the compound alleviates the physical and behavioral abnormalities displayed by the transgenic, nonhuman mammal as a result of overactivity of a mammalian LPA receptor, the alleviation of the abnormality identifying the compound as an antagonist.
92 . An antagonist identified by the method of claim 91 .
93 . A pharmaceutical composition comprising an antagonist identified by the method of claim 92 and a pharmaceutically acceptable carrier.
94 . A method of treating an abnormality in a subject wherein the abnormality is alleviated by decreasing the activity of a mammalian LPA receptor which comprises administering to the subject an effective amount of the pharmaceutical composition of claim 93 , thereby treating the abnormality.
95 . A method for identifying an agonist capable of alleviating an abnormality in a subject wherein the abnormality is alleviated by increasing the activity of a mammalian LPA receptor comprising administering a compound to the transgenic, nonhuman mammal of any one of claims 49 , 52 , 53 , or 54 , and determining whether the compound alleviates the physical and behavioral abnormalities displayed by the transgenic, nonhuman mammal, the alleviation of the abnormality identifying the compound as an agonist.
96 . An agonist identified by the method of claim 95 .
97 . A pharmaceutical composition comprising an agonist identified by the method of claim 95 and a pharmaceutically acceptable carrier.
98 . A method of treating an abnormality in a subject wherein the abnormality is alleviated by increasing the activity of a mammalian LPA receptor which comprises administering to the subject an effective amount of the pharmaceutical composition of claim 97 , thereby treating the abnormality.
99 . A method for diagnosing a predisposition to a disorder associated with the activity of a specific mammalian allele which comprises:
(a) obtaining DNA of subjects suffering from the disorder; (b) performing a restriction digest of the DNA with a panel of restriction enzymes; (c) electrophoretically separating the resulting DNA fragments on a sizing gel; (d) contacting the resulting gel with a nucleic acid probe capable of specifically hybridizing with a unique sequence included within the sequence of a nucleic acid molecule encoding a mammalian LPA receptor and labeled with a detectable marker; (e) detecting labeled bands which have hybridized to the DNA encoding a mammalian LPA receptor of claim 1 labeled with a detectable marker to create a unique band pattern specific to the DNA of subjects suffering from the disorder; (f) preparing DNA obtained for diagnosis by steps (a)-(e); and (g) comparing the unique band pattern specific to the DNA of subjects suffering from the disorder from step (e) and the DNA obtained for diagnosis from step (f) to determine whether the patterns are the same or different and to diagnose thereby predisposition to the disorder if the patterns are the same.
100 . The method of claim 99 wherein a disorder associated with the activity of a specific mammalian allele is diagnosed.
101 . A method of preparing the purified mammalian LPA receptor of claim 11 which comprises:
(a) inducing cells to express the mammalian LPA receptor;
(b) recovering the mammalian LPA receptor from the induced cells; and
(c) purifying the mammalian LPA receptor so recovered.
102 . A method of preparing the purified mammalian LPA receptor of claim 11 which comprises:
(a) inserting nucleic acid encoding the mammalian LPA receptor in a suitable vector;
(b) introducing the resulting vector in a suitable host cell;
(c) placing the resulting cell in suitable condition permitting the production of the isolated mammalian LPA receptor;
(d) recovering the mammalian LPA receptor produced by the resulting cell; and
(e) purifying the mammalian LPA receptor so recovered.
103 . A process for determining whether a chemical compound is a mammalian LPA receptor agonist which comprises contacting cells transfected with and expressing DNA encoding the mammalian LPA receptor with the compound under conditions permitting the activation of the mammalian LPA receptor, and detecting an increase in mammalian LPA receptor activity, so as to thereby determine whether the compound is a mammalian LPA receptor agonist.
104 . A process for determining whether a chemical compound is a mammalian LPA receptor antagonist which comprises contacting cells transfected with and expressing DNA encoding the mammalian LPA receptor with the compound in the presence of a known mammalian LPA receptor agonist, under conditions permitting the activation of the mammalian LPA receptor, and detecting a decrease in mammalian LPA receptor activity, so as to thereby determine whether the compound is a mammalian LPA receptor antagonist.
105 . A process of any one of claims 103 or 104 , wherein the mammalian LPA receptor is a human LPA receptor.
106 . A pharmaceutical composition which comprises an amount of a mammalian LPA receptor agonist determined by the process of claim 103 effective to increase activity of a mammalian LPA receptor and a pharmaceutically acceptable carrier.
107 . A pharmaceutical composition of claim 106 , wherein the mammalian LPA receptor agonist is not previously known.
108 . A pharmaceutical composition which comprises an amount of a mammalian LPA receptor antagonist determined by the process of claim 104 effective to reduce activity of a mammalian LPA receptor and a pharmaceutically acceptable carrier.
109 . A pharmaceutical composition of claim 108 , wherein the mammalian LPA receptor antagonist is not previously known.
110 . A process for determining whether a chemical compound specifically binds to and activates a mammalian LPA receptor, which comprises contacting cells producing a second messenger response and expressing on their cell surface the mammalian LPA receptor, wherein such cells do not normally express the mammalian LPA receptor, with the chemical compound under conditions suitable for activation of the mammalian LPA receptor, and measuring the second messenger response in the presence and in the absence of the chemical compound, a change in the second messenger response in the presence of the chemical compound indicating that the compound activates the mammalian LPA receptor.
111 . The process of claim 110 , wherein the second messenger response comprises chloride channel activation and the change in second messenger is an increase in the level of inward chloride current.
112 . A process for determining whether a chemical compound specifically binds to and inhibits activation of a mammalian LPA receptor, which comprises separately contacting cells producing a second messenger response and expressing on their cell surface the mammalian LPA receptor, wherein such cells do not normally express the mammalian LPA receptor, with both the chemical compound and a second chemical compound known to activate the mammalian LPA receptor, and with only the second chemical compound, under conditions suitable for activation of the mammalian LPA receptor, and measuring the second messenger response in the presence of only the second chemical compound and in the presence of both the second chemical compound and the chemical compound, a smaller change in the second messenger response in the presence of both the chemical compound and the second chemical compound than in the presence of only the second chemical compound indicating that the chemical compound inhibits activation of the mammalian LPA receptor.
113 . The process of claim 112 , wherein the second messenger response comprises chloride channel activation and the change in second messenger response is a smaller increase in the level of inward chloride current in the presence of both the chemical compound and the second chemical compound than in the presence of only the second chemical compound.
114 . A process of any one of claims 110 , 111 , 112 or 113 , wherein the mammalian LPA receptor is a human LPA receptor.
115 . The process of claim 114 , wherein the human LPA receptor has substantially the same amino acid sequence as encoded by the plasmid HL-18a (ATCC Accession No. 209448).
116 . The process of claim 114 , wherein the human LPA receptor has substantially the same amino acid sequence as that shown in FIG. 3 (Seq. I.D. No. 3).
117 . The process of claim 114 , wherein the human LPA receptor has an amino acid sequence, identical to the amino acid sequence shown in FIG. 3 (Seq. I.D. No. 3).
118 . The process of any one of claims 110 , 111 , 112 , 113 , 114 , 115 , 116 , or 117 wherein the cell is an insect cell.
119 . The process of any one of claims 110 , 111 , 112 , 113 , 114 , 115 , 116 , or 117 wherein the cell is a mammalian cell.
120 . The process of claim 119 , wherein the mammalian cell is non-neuronal in origin.
121 . The process of claim 120 , wherein the non-neuronal cell is a COS-7 cell, CHO cell, 293 human embryonic kidney cell, NIH-3T3 cell or LM(tk−) cell.
122 . The process of claim 119 , wherein the compound is a compound not previously known to bind to a mammalian LPA receptor.
123 . A compound determined by the process of claim 122 .
124 . A pharmaceutical composition which comprises an amount of a mammalian LPA receptor agonist determined by the process of claim 110 or 111 effective to increase activity of a mammalian LPA receptor and a pharmaceutically acceptable carrier.
125 . A pharmaceutical composition of claim 124 , wherein the mammalian LPA receptor agonist is not previously known.
126 . A pharmaceutical composition which comprises an amount of a mammalian LPA receptor antagonist determined by the process of any one of claims 112 or 113 effective to reduce activity of a mammalian LPA receptor and a pharmaceutically acceptable carrier.
127 . A pharmaceutical composition of claim 126 , wherein the mammalian LPA receptor antagonist is not previously known.
128 . A method of screening a plurality of chemical compounds not known to activate a mammalian LPA receptor to identify a compound which activates the mammalian LPA receptor which comprises:
(a) contacting cells transfected with and expressing the mammalian LPA receptor with the plurality of compounds not known to activate the mammalian LPA receptor, under conditions permitting activation of the mammalian LPA receptor; (b) determining whether the activity of the mammalian LPA receptor is increased in the presence of the compounds; and if so (c) separately determining whether the activation of the mammalian LPA receptor is increased by each compound included in the plurality of compounds, so as to thereby identify the compound which activates the mammalian LPA receptor.
129 . A method of claim 128 , wherein the mammalian LPA receptor is a human LPA receptor.
130 . A method of screening a plurality of chemical compounds not known to inhibit the activation of a mammalian LPA receptor to identify a compound which inhibits the activation of the mammalian LPA receptor, which comprises:
(a) contacting cells transfected with and expressing the mammalian LPA receptor with the plurality of compounds in the presence of a known mammalian LPA receptor agonist, under conditions permitting activation of the mammalian LPA receptor; (b) determining whether the activation of the mammalian LPA receptor is reduced in the presence of the plurality of compounds, relative to the activation of the mammalian LPA receptor in the absence of the plurality of compounds; and if so (c) separately determining the inhibition of activation of the mammalian LPA receptor for each compound included in the plurality of compounds, so as to thereby identify the compound which inhibits the activation of the mammalian LPA receptor.
131 . A method of claim 130 , wherein the mammalian LPA receptor is a human LPA receptor.
132 . A method of any one of claims 128 , 129 , 130 , or 131 , wherein the cell is a mammalian cell.
133 . A method of claim 132 , wherein the mammalian cell is non-neuronal in origin.
134 . The method of claim 133 , wherein the non-neuronal cell is a COS-7 cell, a 293 human embryonic kidney cell, a LM(tk−) cell or an NIH-3T3 cell.
135 . A pharmaceutical composition comprising a compound identified by the method of any one of claims 128 or 129 effective to increase mammalian LPA receptor activity and a pharmaceutically acceptable carrier.
136 . A pharmaceutical composition comprising a compound identified by the method of any one of claims 130 or 131 effective to decrease mammalian LPA receptor activity and a pharmaceutically acceptable carrier.
137 . A method of treating an abnormality in a subject wherein the abnormality is alleviated by increasing the activity of a mammalian LPA receptor which comprises administering to the subject an amount of compound which is a mammalian LPA receptor agonist effective to treat the abnormality.
138 . A method of treating an abnormality in a subject wherein the abnormality is alleviated by decreasing the activity of a mammalian LPA receptor which comprises administering to the subject an amount of compound which is a mammalian LPA receptor antagonist effective to treat the abnormality.Join the waitlist — get patent alerts
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