Claims
- 1. A recombinant nucleic acid molecule which encodes a kappa3 opioid receptor.
- 2. The recombinant nucleic acid molecule of claim 1, wherein the nucleic acid molecule is a DNA molecule.
- 3. The recombinant nucleic acid molecule of claim 2, wherein the DNA molecule is a cDNA molecule.
- 4. The recombinant nucleic acid molecule of claim 1, wherein the nucleic acid molecule is an RNA molecule.
- 5. The recombinant nucleic acid molecule of claim 1, wherein the kappa3 opioid receptor is native kappa3 opioid receptor.
- 6. The recombinant nucleic acid molecule of claim 1, wherein the kappa3 opioid receptor is a variant kappa3 opioid receptor.
- 7. The recombinant nucleic acid molecule of claim 6, wherein the variant kappa3 opioid receptor is an aqueous-soluble variant kappa3 opioid receptor.
- 8. The recombinant nucleic acid molecule of claim 1, wherein the kappa3 opioid receptor is a mammalian kappa3 opioid receptor.
- 9. The recombinant nucleic acid molecule of claim 8, wherein the mammalian kappa3 opioid receptor is a murine kappa3 opioid receptor.
- 10. The recombinant nucleic acid molecule of claim 8, wherein the mammalian kappa3 opioid receptor is a human kappa3 opioid receptor.
- 11. An anti-sense oligonucleotide molecule capable of specifically hybridizing to an mRNA molecule encoding kappa3 opioid receptor, at the portion thereof encoding kappa3 opioid receptor, so as to prevent translation of the mRNA molecule.
- 12. A nucleic acid molecule encoding the anti-sense oligonucleotide molecule of claim 11, wherein the nucleic acid molecule is capable of being expressed in a suitable host cell.
- 13. A vector comprising the recombinant nucleic acid molecule of claim 3.
- 14. The vector of claim 13, wherein the vector is a plasmid.
- 15. The plasmid of claim 14, wherein the plasmid is designated 13-2A4 (ATCC Accession No. ______).
- 16. A host vector system for the production of a kappa3 opioid receptor which comprises the vector of claim 13 in a suitable host cell.
- 17. The host vector system of claim 16, wherein the suitable host cell is a bacterial cell.
- 18. The host vector system of claim 17, wherein the bacterial cell is an E. coli cell.
- 19. The host vector system of claim 16, wherein the suitable host cell is an eucaryotic cell.
- 20. The host vector system of claim 19, wherein the eucaryotic cell is a mammalian cell.
- 21. The host vector system of claim 20, wherein the mammalian cell is a COS cell.
- 22. The host vector system of claim 21, wherein the COS cell is a COS-7 cell.
- 23. A method for producing a kappa3 opioid receptor which comprises growing the host vector system of claim 16 under conditions permitting the production of the kappa3 opioid receptor, and recovering the kappa3 opioid receptor produced thereby.
- 24. The nucleic acid molecule of claim 1, wherein the nucleic acid molecule is labeled with a detectable marker.
- 25. A method for detecting the presence of kappa3 opioid receptor-encoding nucleic acid molecules present in a sample which comprises contacting the sample with the labeled nucleic acid molecule of claim 24 under conditions permitting the labeled nucleic acid molecule to form a complex with kappa3 opioid receptor-encoding nucleic acid molecules present in the sample, and detecting the presence of such complex formed so as to thereby detect the presence of kappa3 opioid receptor-encoding nucleic acid molecules present in the sample.
- 26. A method for quantitatively determining the amount of kappa3 opioid receptor-encoding nucleic acid molecules present in a sample which comprises contacting the sample with a suitable amount of the labeled nucleic acid molecule of claim 24 under conditions permitting the labeled nucleic acid molecule to form a complex with kappa3 opioid receptor-encoding nucleic acid molecules present in the sample, quantitatively determining the amount of complex so formed so as to thereby quantitatively determine the amount of kappa3 opioid receptor-encoding nucleic acid molecules present in the sample.
- 27. A purified native kappa3 opioid receptor.
- 28. The variant kappa3 opioid receptor encoded by the recombinant nucleic acid molecule of claim 6.
- 29. A method for obtaining partially purified polyclonal antibodies capable of specifically binding to kappa3 opioid receptor and thereby competing with opioid binding thereto which method comprises (a) immunizing a subject with the kappa3 opioid receptor encoded by the nucleic acid molecule of claim 1, (b) recovering from the immunized subject serum comprising antibodies capable of specifically binding to kappa3 opioid receptor and thereby competing with opioid binding thereto, and (c) partially purifying the antibodies present in the serum, thereby obtaining partially purified polyclonal antibodies capable of specifically binding to kappa3 opioid receptor and thereby competing with opioid binding thereto.
- 30. The partially purified antibodies produced by the method of claim 29.
- 31. A method for obtaining a purified monoclonal antibody capable of specifically binding to kappa3 opioid receptor and thereby competing with opioid binding thereto which method comprises (a) immunizing a subject with the kappa3 opioid receptor encoded by the nucleic acid molecule of claim 1, (b) recovering from the immunized subject a B cell-containing cell sample, (c) contacting the B cell-containing cell sample so recovered with myeloma cells under conditions permitting fusion of the myeloma cells with the B cells therein so as to form hybridoma cells, (d) isolating from the resulting sample a hybridoma cell capable of producing a monoclonal antibody capable of specifically binding to kappa3 opioid receptor and thereby competing with opioid binding thereto, (e) growing the hybridoma cell so isolated under conditions permitting the production of the monoclonal antibody, and (f) recovering the monoclonal antibody so produced, thereby obtaining a purified monoclonal antibody capable of specifically binding to kappa3 opioid receptor and thereby competing with opioid binding thereto.
- 32. The hybridoma cell produced in step (d) of claim 31.
- 33. The purified monoclonal antibody produced by the method of claim 31.
- 34. An antibody capable of specifically binding to kappa3 opioid receptor and thereby competing with opioid binding thereto, said antibody being labeled with a detectable marker.
- 35. A method for detecting the presence of kappa3 opioid receptor in a sample which comprises contacting the sample with the antibody of claim 34 under conditions permitting the antibody to form a complex with kappa3 opioid receptor present in the sample, and detecting the presence of complex so formed, so as to thereby detect the presence of kappa3 opioid receptor in the sample.
- 36. A method for quantitatively determining the amount of kappa3 opioid receptor in a sample which comprises contacting the sample with the antibody of claim 34 under conditions permitting the antibody to form a complex with kappa3 opioid receptor present in the sample, quantitatively determining the amount of complex so formed, and comparing the amount so determined with a known standard, so as to thereby quantitatively determine the amount of kappa3 opioid receptor in the sample.
- 37. A composition which comprises the antibody of claim 34 in an amount effective to permit imaging cell membrane-bound kappa3 opioid receptor present in a subject, and a pharmaceutically acceptable carrier.
- 38. A method for imaging cell membrane-bound kappa3 opioid receptor present in a subject which comprises administering to the subject an amount of the composition of claim 37 effective to permit imaging cell membrane-bound kappa3 opioid receptor present in the subject under conditions permitting the antibody of the composition to specifically bind to cell membrane-bound kappa3 opioid receptor present in the subject, and imaging the antibody specifically bound to cell membrane-bound kappa3 opioid receptor present in the subject after a suitable period of time, so as to thereby image cell membrane-bound kappa3 opioid receptor present in the subject.
- 39. A composition which comprises the antibody of claim 34 in an amount effective to permit quantitatively determining the amount of cell membrane-bound kappa3 opioid receptor present in a subject, and a pharmaceutically acceptable carrier.
- 40. A method for quantitatively determining the amount of cell membrane-bound kappa3 opioid receptor present in a subject which comprises administering to the subject an amount of the composition of claim 39 effective to permit quantitatively determining the amount of cell membrane-bound kappa3 opioid receptor present in the subject under conditions permitting the antibody of the composition to specifically bind to cell membrane-bound kappa3 opioid receptor in the subject, quantitatively determining the amount of antibody specifically bound to cell membrane-associated kappa3 opioid receptors present in the subject, and comparing the amount so determined with a known standard so as to thereby quantitatively determine the amount of cell membrane-bound kappa3 opioid receptor present in the subject.
- 41. A method for determining a subject's potential sensitivity to a kappa3 opioid receptor-specific agent which comprises quantitatively determining the amount of cell membrane-bound kappa3 opioid receptor present in the subject by the method of claim 40, and comparing the amount of cell membrane-bound kappa3 opioid receptor so determined with the amount of cell membrane-bound kappa3 opioid receptor present in a subject having a known sensitivity to the agent, so as to thereby determine the subject's potential sensitivity to the kappa3 opioid receptor-specific agent.
- 42. The method of claim 41, wherein the kappa3 opioid receptor-specific agent is a kappa3 opioid receptor-specific analgesic agent.
- 43. A method for determining the affinity of an agent for kappa3 opioid receptor which comprises (a) contacting a predetermined amount of kappa3 opioid receptor with (i) a predetermined amount of the agent together with (ii) a predetermined amount of a detectable known ligand of kappa3 opioid receptor, said known ligand being a known ligand of kappa3 opioid receptor with a known affinity therefore, under conditions which would permit the formation of a complex between kappa3 opioid receptor and the detectable known ligand in the absence of the agent, (b) quantitatively determining the amount of kappa3 opioid receptor-detectable known ligand complex so formed, (c) comparing the amount of kappa3 opioid receptor-detectable known ligand complex determined in step (b) with the amount of kappa3 opioid receptor-detectable known ligand complex formed in the absence of the agent, and (d) determining the affinity of the agent for kappa3 opioid receptor based on the comparison in step (c).
- 44. A method for determining the affinity of an agent for kappa3 opioid receptor which comprises (a) contacting kappa3 opioid receptor with the agent under conditions which would permit the formation of a complex between kappa3 opioid receptor and a known ligand thereof, (b) contacting a predetermined amount of the agent-receptor complex with a predetermined amount of the antibody of claim 34 under conditions which would permit the formation of a complex between kappa3 opioid receptor and the antibody in the absence of the agent, (c) quantitatively determining the amount of kappa3 opioid receptor-antibody complex so formed, and (d) determining the affinity of the agent for kappa3 opioid receptor based on the amount of kappa3 opioid receptor-antibody complex determined in step (c).
- 45. A method for determining whether a known ligand of kappa3 opioid receptor is an agonist thereof which comprises (a) contacting the kappa3 opioid receptor with (i) a predetermined amount of the ligand and (ii) a predetermined amount of GTP or analog thereof under conditions which would permit the binding of the ligand to the kappa3 opioid receptor in the absence of GTP or analog thereof, (b) quantitatively determining the percentage of such predetermined amount of ligand bound to the kappa3 opioid receptor, and (c) comparing the percentage so determined with the percentage of such predetermined amount of ligand bound to the kappa3 opioid receptor in the absence of GTP or analog thereof, a lower percentage in the presence of GTP or analog thereof relative to the percentage in the absence of GTP or analog thereof indicating that the ligand is an agonist of kappa3 opioid receptor.
- 46. A method for determining whether a ligand of kappa3 opioid receptor is an antagonist thereof which comprises (a) contacting the kappa3 opioid receptor with (i) a predetermined amount of the ligand and (ii) a predetermined amount of GTP or analog thereof under conditions which would permit the binding of the ligand to the kappa3 opioid receptor in the absence of GTP or analog thereof, (b) quantitatively determining the percentage of such predetermined amount of ligand bound to the kappa3 opioid receptor, and (c) comparing the percentage so determined with the percentage of such predetermined amount of ligand bound to the kappa3 opioid receptor in the absence of GTP or analog thereof, an equality between the percentage in the presence of GTP or analog thereof and the percentage in the absence of GTP or analog thereof indicating that the ligand is an antagonist of kappa3 opioid receptor.
Government Interests
[0001] This invention was made with support under Grant Nos. DA02615 RO1 and 1K02 DA00138 from the National Institute on Drug Abuse. Accordingly, the U.S. government has certain rights in the invention.
Divisions (1)
|
Number |
Date |
Country |
Parent |
08986209 |
Dec 1997 |
US |
Child |
10331028 |
Dec 2002 |
US |