DNA encoding a human serotonin (5-HT2) receptor and uses thereof

• US 5,885,785 A
• Filed: 03/08/1996
• Issued: 03/23/1999
• Est. Priority Date: 10/31/1989
• Status: Expired due to Fees

• Current Assignee: Lundbeck Research USA, Inc. (Lundbeck Foundation)
• Original Assignee: Synaptic Pharmaceutical Corporation (Lundbeck Foundation)
• Inventors: Branchek, Theresa, Hartig, Paul R., Kao, Hung-Teh
• Primary Examiner(s): Walsh, Stephen
• Assistant Examiner(s): Kaufman, Claire M.
• Application Number:
  US08/613,044
• Time in Patent Office: 1,110 Days
• Field of Search: 435/7.2, 435/69.1, 435/240.2, 435/325, 435/7.1, 435/7.21, 530/350, 536/23.5
• US Class Current: 435/7.21
• CPC Class Codes: C07K 14/70571 for neuromediators, e.g. serotonin receptor, dopamine receptor

Abstract

The present invention provides an isolated nucleic acid molecule encoding a 5-HT₂ receptor, and an isolated protein which is a human 5-HT₂ receptor. The invention also provides vectors comprising DNA molecules encoding a human 5-HT₂ receptor, and vectors adapted for expression of the 5-HT₂ receptor in bacterial, yeast, or mammalian cells. In addition, the invention provides a DNA probe useful for detecting nucleic acid encoding the 5-HT₂ receptor, a method for determining whether a ligand which is not known to be capable of binding to the 5-HT₂ receptor can bind to the 5-HT₂ receptor, a method for detecting the presence of 5-HT₂ receptor on the surface of a cell, and a method of screening drugs to identify drugs which specifically interact with, and bind to, the 5-HT₂ receptor. The invention herein also concerns an antibody directed to the human 5-HT₂ receptor, such as a monoclonal antibody directed to an epitope of the 5-HT₂ receptor present on the surface of a cell and having an amino acid sequence included within the amino acid sequence shown in FIGS. 2A-2G.

Claims

1. A process for identifying a chemical compound which specifically binds to a human 5-HT₂ receptor, which comprises contacting mammalian cells transfected with and expressing nucleic acid encoding the human 5-HT₂ receptor having the amino acid sequence shown in FIG. 2, wherein the human 5-HT₂ receptor is expressed on the surface of the cells, with the chemical compound under conditions suitable for binding, and detecting specific binding of the chemical compound to the human 5-HT₂ receptor.

11. The process of any one of claims 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, wherein the mammalian cell is a COS-7 cell, Ltk-cell, NIH-3T3 cell, CHO cell, or HeLa cell.

2. A process for identifying a chemical compound which specifically binds to a human 5-HT₂ receptor, which comprises contacting a membrane fraction from mammalian cells transfected with and expressing nucleic acid encoding the human 5-HT₂ receptor having the amino acid sequence shown in FIG. 2, wherein the human 5-HT₂ receptor is expressed on the surface of the cells, with the chemical compound under conditions suitable for binding, and detecting specific binding of the chemical compound to the human 5-HT₂ receptor.

3. A process involving competitive binding for identifying a chemical compound which specifically binds to a human 5-HT₂ receptor, which comprises contacting mammalian cells transfected with and expressing nucleic acid encoding the human 5-HT₂ receptor having the amino acid sequence shown in FIG. 2, wherein the human 5-HT₂ receptor is expressed on the surface of the cells, with both the chemical compound and a second chemical compound known to bind to the human 5-HT₂ receptor, under conditions suitable for binding of both compounds, and detecting specific binding of the chemical compound to the human 5-HT₂ receptor, a decrease in binding of the second chemical compound to the human 5-HT₂ receptor in the presence of the chemical compound indicating that the chemical compound binds to the human 5-HT₂ receptor.

4. A process involving competitive binding for identifying a chemical compound which specifically binds to a human 5-HT₂ receptor, which comprises contacting a membrane fraction from mammalian cells transfected with and expressing nucleic acid encoding the human 5-HT₂ receptor having the amino acid sequence shown in FIG. 2, wherein the human 5-HT₂ receptor is expressed on the surface of the cells, with both the chemical compound and a second chemical compound known to bind to the receptor, under conditions suitable for binding of both compounds, and detecting specific binding of the chemical compound to the human 5-HT₂ receptor, a decrease in binding of the second chemical compound to the human 5-HT₂ receptor in the presence of the chemical compound indicating that the chemical compound binds to the human 5-HT₂ receptor.

5. A method for determining whether a chemical compound specifically binds to and activates a human 5-HT₂ receptor, which comprises contacting mammalian cells producing a second messenger response and transfected with and expressing nucleic acid encoding the human 5-HT₂ receptor having the amino acid sequence shown in FIG. 2, wherein the human 5-HT₂ receptor is expressed on the surface of the cells, with the chemical compound under conditions suitable for activation of the human 5-HT₂ receptor, and measuring the second messenger response in the presence and in the absence of the chemical compound, a change in second messenger response in the presence of the chemical compound indicating that the chemical compound activates the human 5-HT₂ receptor.

9. The process of either of claims 5 or 6, wherein the second messenger response comprises phosphoinositide hydrolysis and the change in second messenger response is an increase in phosphoinositide hydrolysis.

6. A method for determining whether a chemical compound specifically binds to and activates a human 5-HT₂ receptor, which comprises contacting a membrane fraction from mammalian cells producing a second messenger response and transfected with and expressing nucleic acid encoding the human 5-HT₂ receptor having the amino acid sequence shown in FIG. 2, wherein the human 5-HT₂ receptor is expressed on the surface of the cell, with the chemical compound under conditions suitable for activation of the human 5-HT₂ receptor, and measuring the second messenger response in the presence and in the absence of the chemical compound, a change in second messenger response in the presence of the chemical compound indicating that the chemical compound activates the human 5-HT₂ receptor.

7. A method for determining whether a chemical compound specifically binds to and inhibits activation of a human 5-HT₂ receptor, which comprises separately contacting mammalian cells producing a second messenger response and transfected with and expressing nucleic acid encoding the human 5-HT₂ receptor having the amino acid sequence shown in FIG. 2, wherein the human 5-HT₂ receptor is expressed on the surface of the cells, with both the chemical compound and a second chemical compound known to activate the human 5-HT₂ receptor, and with only the second chemical compound, under conditions suitable for activation of the human 5-HT₂ 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 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 human 5-HT₂ receptor.

10. The process of either of claims 7 or 8, wherein the second messenger response comprises phosphoinositide hydrolysis and the change in second messenger response is a smaller increase in phosphoinositide hydrolysis in the presence of both the chemical compound and the second chemical compound than in the presence of only the second chemical compound.

8. A method for determining whether a chemical compound specifically binds to and inhibits a human 5-HT₂ receptor, which comprises separately contacting a membrane fraction from mammalian cells producing a second messenger response and transfected with and expressing nucleic acid encoding the human 5-HT₂ receptor having the amino acid sequence shown in FIG. 2, wherein the human 5-HT₂ receptor is expressed on the surface of the cells, with both the chemical compound and a second chemical compound known to activate the human 5-HT₂ receptor, and with only the second chemical compound, under conditions suitable for activation of the human 5-HT₂ 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 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 human 5-HT₂ receptor.