Claims for Patent: 6,436,639

Summary for Patent: 6,436,639

Title:	Bak promoter expression system
Abstract:	Polynucleotides which regulate the expression of a gene involved in apoptosis is described. Also provided are methods for identifying agents that modulate expression of a gene involved in apoptosis.
Inventor(s):	Kiefer; Michael C. (Clayton, CA), Ossina; Natalya K. (Albany, CA)
Assignee:	Tanox, Inc. (Houston, TX)
Application Number:	09/367,750
Patent Claims:	1. An isolated polynucleotide of about 4066 or fewer nucleotides selected from the group consisting of: (a) an isolated polynucleotide comprising the bak promoter (positions 1-4021 of SEQ ID NO:1); (b) an isolated polynucleotide comprising a fragment of said bak promoter of (a) that has at least basal bak promoter transcriptional activity; (c) an isolated mutated polynucleotide of (a) or (b) comprising a mutation selected from the group consisting of a point mutation and a deletion, said mutated polynucleotide having at least basal bak promoter transcriptional activity; and (d) an isolated polynucleotide comprising a nucleic acid sequence that is fully complementary to said polynucleotide of (a), (b) or (c). 2. The polynucleotide of claim 1, wherein said fragment comprises an ISRE site of said bak promoter (positions 2945-2967 of SEQ ID NO:1). 3. The polynucleotide of claim 1, wherein said fragment comprises an SP1 site and a GAS site of said bak promoter (positions 1646-1662 of SEQ ID NO:1). 4. The polynucleotide of claim 1, wherein said fragment comprises an NF.kappa.B2 site of said bak promoter (positions 1038-1047 of SEQ ID NO:1). 5. The polynucleotide of claim 1, wherein said fragment comprises an NF.kappa.B1 site of said bak promoter (positions 949-958 of SEQ ID NO:1). 6. The polynucleotide of claim 1, wherein said fragment comprises a p53 cluster and an SP1 site of said bak promoter (positions 1431-1473 of SEQ ID NO:1). 7. A method for identifying an agent that regulates bak promoter activity, comprising the steps of: a) introducing into a cell a recombinant polynucleotide comprising a reporter gene operably linked to a bak promoter polynucleotide, wherein said bak promoter polynucleotide is selected from the group consisting of: (i) a polynucleotide comprising a bak promoter (positions 1-4021 of SEQ ID NO:1); (ii) a polynucleotide comprising a fragment of said bak promoter of (i) that has at least basal bak promoter transcriptional activity; and (iii) a mutated polynucleotide of (i) or (ii) comprising a mutation selected from the group consisting of a point mutation and a deletion, said mutated polynucleotide having at least basal bak promoter transcriptional activity; b) determining the level of expression of said reporter gene in said cell of step (a) in the absence of a test agent; c) contacting said cell of step (a) with said test agent; d) determining the level of expression of said reporter gene in said cell after step (c) of contacting; and, e) identifying an agent that regulates the expression of said reporter gene determined in step (d), as compared to the level of expression determined in step (b), wherein a difference in the expression of said reporter gene in step (d) as compared to step (b) indicates that the agent regulates bak promoter activity. 8. A method for identifying an agent that increases the expression of a gene that is operably linked to a bak promoter, comprising the steps of: a) introducing into a cell a recombinant polynucleotide comprising a reporter gene operably linked to a bak promoter polynucleotide wherein said bak promoter polynucleotide is selected from the group consisting of: (i) a polynucleotide comprising a bak promoter (positions 1-4021 of SEQ ID NO:1); (ii) a polynucleotide comprising a fragment of said bak promoter of (i) that has at least basal bak promoter transcriptional activity; and (iii) a mutated polynucleotide of (i) or (ii) comprising a mutation selected from the group consisting of a point mutation and a deletion, said mutated polynucleotide having at least basal bak promoter transcriptional activity; b) determining the level of expression of said reporter gene in said cell of step (a) in the absence of a test agent; c) contacting said cell of step (a) with said test agent; d) determining the level of expression of said reporter gene in said cell after step (c) of contacting; and, e) identifying an agent that increases the expression of said reporter gene determined in step (d), as compared to the level of expression determined in step (b), wherein an increase in the expression of said reporter gene in step (d) as compared to step (b) indicates that the agent increases expression of a gene that is operably linked to a bak promoter. 9. A method for identifying an agent that decreases the expression of a gene that is operably linked to a bak promoter, comprising the steps of: a) introducing into a cell a recombinant polynucleotide comprising a reporter gene operably linked to a bak promoter polynucleotide, wherein said bak promoter polynucleotide is selected from the group consisting of: (i) a polynucleotide comprising a bak promoter (positions 1-4021 of SEQ ID NO:1); (ii) a polynucleotide comprising a fragment of said bak promoter of (i) that has at least basal bak promoter transcriptional activity; and (iii) a mutated polynucleotide of (i) or (ii) comprising a mutation selected from the group consisting of a point mutation and a deletion said mutated polynucleotide having at least basal bak promoter transcriptional activity; b) determining the level of expression of said reporter gene in said cell of step (a) in the absence of a test agent; c) contacting said cell of step (a) with said test agent; d) determining the level of expression of said reporter gene in said cell after step (c) of contacting; and, e) identifying an agent that decreases the expression of said reporter gene determined in step (d), as compared to the level of expression determined in step (b), wherein a decrease in the expression of said reporter gene in step (d) as compared to step (b) indicates that the agent decreases expression of a gene that is operably linked to a bak promoter. 10. The isolated polynucleotide of claim 1, wherein said fragment is an about 0.7 kb fragment comprising from at least about the first nucleotide of an NF.kappa.B1 site (position 949 of SEQ ID NO:1) through at least about the first transcription start site in said bak promoter (position 1515 of SEQ ID NO:1). 11. The isolated polynucleotide of claim 1, wherein said fragment is an about 1.6 kb fragment comprising from at least about the first nucleotide of said bak promoter (position 1 of SEQ ID NO:1) through at least about the first transcription start site that is 3' of GAS (position 1695 of SEQ ID NO:1). 12. The isolated polynucleotide of claim 1, wherein said fragment is an about 2 kb fragment comprising from at least about the first nucleotide that is 3' of an NF.kappa.B2 site (position 1048 of SEQ ID NO:1) through at least about the last nucleotide of an ISRE in intron 1 of said bak promoter (position 2967 of SEQ ID NO:1). 13. The isolated polynucleotide of claim 1, wherein said fragment has interferon-.gamma. (IFN-.gamma.)-activated bak promoter activity. 14. A recombinant polynucleotide, comprising an isolated polynucleotide having at least basal bak promoter transcriptional activity, operably linked to a coding sequence, wherein said isolated polynucleotide is selected from the group consisting of: (a) an isolated polynucleotide comprising the bak promoter (positions 1-4021 of SEQ ID NO:1); (b) an isolated polynucleotide comprising a fragment of said bak promoter that has at least basal bak promoter transcriptional activity; and (c) a mutated polynucleotide of (a) or (b) comprising a mutation selected from the group consisting of a point mutation and a deletion, said mutated polynucleotide having at least basal bak promoter transcriptional activity. 15. The recombinant polynucleotide of claim 14, wherein said coding sequence is a heterologous polynucleotide with respect to said isolated polynucleotide. 16. A recombinant cell transfected with a recombinant polynucleotide of claim 14. 17. The method of claim 7, wherein said fragment is an about 0.7 kb fragment comprising from at least about the first nucleotide of an NF.kappa.B1 site (position 949 of SEQ ID NO:1) through at least about the first transcription start site in said bak promoter (position 1515 of SEQ ID NO:1). 18. The method of claim 7, wherein said fragment is an about 1.6 kb fragment comprising from at least about the first nucleotide of said bak promoter (position 1 of SEQ ID NO:1) through at least about the first transcription start site that is 3' of GAS (position 1695 of SEQ ID NO:1). 19. The method of claim 7, wherein said fragment is an about 2 kb fragment comprising from at least about the first nucleotide that is 3' of an NF.kappa.B2 site (position 1048 of SEQ ID NO:1) through at least about the last nucleotide of an ISRE in intron 1 of said bak promoter (position 2967 of SEQ ID NO:1). 20. An isolated polynucleotide selected from the group consisting of: (a) an isolated polynucleotide consisting essentially of the bak promoter (positions 1-4021 of SEQ ID NO:1); (b) an isolated polynucleotide comprising a fragment of said bak promoter of (a) that has at least basal bak promoter transcriptional activity; (c) an isolated mutated polynucleotide of (a) or (b) comprising a mutation selected from the group consisting of a point mutation and a deletion, said mutated polynucleotide having at least basal bak promoter transcriptional activity; and (d) an isolated polynucleotide consisting essentially of a nucleic acid sequence that is fully complementary to said polynucleotide of (a), (b) or (c).

Details for Patent 6,436,639

Applicant	Tradename	Biologic Ingredient	Dosage Form	BLA	Approval Date	Patent No.	Expiredate
Merck Sharp & Dohme Corp.	INTRON A	interferon alfa-2b	For Injection	103132	06/04/1986		2017-02-18
Merck Sharp & Dohme Corp.	INTRON A	interferon alfa-2b	For Injection	103132			2017-02-18
Merck Sharp & Dohme Corp.	INTRON A	interferon alfa-2b	Injection	103132			2017-02-18