Claims
- 1. A method of identifying an agent that modulates a signal transduction pathway, said method comprising the steps of:
(i) providing a polypeptide comprising an RKIP motif; and (ii) contacting said polypeptide with a candidate agent, wherein binding of the candidate agent to said RKIP motif is indicative that said candidate agent is a signal transduction modulating agent.
- 2. The method of claim 1 wherein said binding is detected using a method selected from the group consisting of surface plasmon resonance, yeast two-hybrid assay, pull-down assay, FRET, fluorescence polarization assay, scintillation proximity assay, transcription assay, kinase assay and transformation assay.
- 3. The method of claim 1 wherein said modulation is an increase in the activity of said signal transduction pathway.
- 4. The method of claim 1 wherein said modulation is a decrease in the activity of said signal transduction pathway.
- 5. A method of identifying an agent that modulates cell growth, said method comprising the steps of:
(i) providing a polypeptide comprising an RKIP motif; and (ii) contacting said polypeptide with a candidate agent, wherein binding of the candidate agent to said RKIP motif is indicative that said candidate agent is a cell growth modulating agent.
- 6. The method of claim 5 wherein said binding is detected using a method selected from the group consisting of surface plasmon resonance, yeast two-hybrid assay, pull-down assay, FRET, fluorescence polarization assay, scintillation proximity assay, transcription assay, kinase assay and transformation assay.
- 7. The method of claim 5 wherein said modulation is an increase in cell growth.
- 8. The method of claim 5 wherein said modulation is a decrease in cell growth.
- 9. The method of claim 8 wherein said modulation occurs in a cell proliferative disease.
- 10. The method of claim 9 wherein said disease is cancer.
- 11. A method of identifying an agent that modulates apoptosis, said method comprising the steps of:
(i) providing a polypeptide comprising an RKIP motif; and (ii) contacting said polypeptide with a candidate agent, wherein binding of the candidate agent to said RKIP motif is indicative that said candidate agent is an apoptosis modulating agent.
- 12. The method of claim 11 wherein said binding is detected using a method selected from the group consisting of surface plasmon resonance, yeast two-hybrid assay, pull-down assay, FRET, fluorescence polarization assay, scintillation proximity assay, transcription assay, kinase assay and transformation assay.
- 13. The method of claim 11 wherein said modulation is an increase in apoptosis.
- 14. The method of claim 13 wherein said modulation occurs in a tumor.
- 15. The method of claim 11 wherein said modulation is a decrease in apoptosis.
- 16. A method of identifying an agent that modulates an RKIP-sensitive pathway, said method comprising the steps of:
(i) providing a polypeptide comprising an RKIP motif; and (ii) contacting said polypeptide with a candidate agent, wherein binding of the candidate agent to said RKIP motif is indicative that said candidate agent is a modulator of an RKIP-sensitive pathway.
- 17. The method of claim 16 wherein said binding is detected using a method selected from the group consisting of surface plasmon resonance, yeast two-hybrid assay, pull-down assay, FRET, fluorescence polarization assay, scintillation proximity assay, transcription assay, kinase assay and transformation assay.
- 18. The method of claim 16 wherein said modulation is an increase the activity of an RKIP-sensitive pathway.
- 19. The method of claim 18 wherein said modulation occurs in a tumor.
- 20. The method of claim 16 wherein said modulation is a decrease in the activity of an RKIP-sensitive pathway.
- 21. A method of identifying an agent that modulates the activity of an RKIP motif-containing polypeptide, said method comprising the steps of:
i) providing an RKIP motif-containing polypeptide and a polypeptide binding partner thereof; and ii) contacting said RKIP motif-containing polypeptide and said binding partner thereof under conditions permitting the binding of said RKIP motif-containing polypeptide to said binding partner thereof; iii) and monitoring the association of said RKIP motif-containing polypeptide and said binding partner thereof in the presence and absence of a candidate agent, wherein an increase or decrease in the binding of said RKIP motif-containing polypeptide to said binding partner thereof is indicative that said candidate agent modulates the activity of an RKIP motif containing polypeptide.
- 22. The method of claim 21 wherein said monitoring is performed using a method selected from the group consisting of: surface plasmon resonance, yeast two-hybrid assay, pull-down assay, FRET, fluorescence polarization assay and scintillation proximity assay.
- 23. A method of identifying an agent that modulates the activity of an RKIP motif-containing polypeptide, said method comprising the steps of:
i) providing a cell comprising a reporter gene construct wherein the expression of said reporter gene is functionally coupled to a control region regulated by an RKIP-sensitive kinase; and ii) measuring the amount of reporter gene expression from said construct in the presence and absence of a candidate agent, wherein an increase or decrease in the expression of said reporter is indicative that said candidate agent modulates the activity of an RKIP motif-containing polypeptide.
- 24. The method of claim 23 wherein the expression of said reporter gene is controlled by an AP-1 sensitive control region which is functionally coupled to said reporter gene.
- 25. The method of claim 23 wherein the expression of said reporter gene is controlled by an NF-κB sensitive control region which is functionally coupled to said reporter gene.
- 26. A method of detecting a condition associated with the activity of an RKIP-sensitive signal transduction pathway comprising:
a) measuring the amount of an RKIP motif-encoding RNA present in a tissue sample; and b) comparing said amount an RKIP motif-encoding RNA present in said sample to the amount of said RKIP motif-encoding RNA present in a control tissue sample present in a control tissue sample, wherein an increase or decrease in the amount of said RKIP motif-encoding RNA relative to the amount of said RKIP motif-encoding RNA in said control tissue sample is indicative of a condition associated with the activity of an RKIP-sensitive signal transduction pathway.
- 27. The method of claim 26 wherein said measuring is performed by a method selected from the group consisting of RT-PCR, RNase protection, in situ hybridization and Northern hybridization.
- 28. A method of detecting a condition associated with the activity of an RKIP-sensitive signal transduction pathway comprising:
a) measuring the amount of an RKIP motif-containing polypeptide present in a tissue sample; and b) comparing said amount of an RKIP motif-containing polypeptide present in said sample to the amount of an RKIP motif-containing polypeptide present in a control tissue sample, wherein an increase or decrease in the amount of said RKIP motif-containing polypeptide relative to the amount of said RKIP motif-containing polypeptide in said control tissue sample is indicative of a condition associated with the activity of an RKIP-sensitive signal transduction pathway.
- 29. The method of claim 28 wherein said condition is cancer.
- 30. The method of claim 28 wherein said measuring is performed by measuring the binding of an antibody to said RKIP motif-containing polypeptide or an antigen-binding fragment thereof.
- 31. A method of identifying an agent that modulates the RKIP-sensitive phosphorylation of a polypeptide, comprising the steps of:
a) providing a system that permits phosphorylation of said polypeptide by an RKIP-sensitive kinase; and b) detecting phosphorylation of said polypeptide in the presence and absence of a candidate agent wherein an increase or decrease in phosphorylation is indicative that said agent is a modulator of the RKIP-sensitive phosphorylation of said polypeptide.
- 32. The method of claim 31 wherein said RKIP-sensitive kinase is selected from the group consisting of: Raf, MEK, and ERK.
- 33. The method of claim 31 wherein said RKIP-sensitive kinase is Raf-1.
- 34. The method of claim 31 wherein said RKIP-sensitive kinase is IκB Kinase alpha (IKK-alpha).
- 35. The method of claim 31 wherein said RKIP-sensitive kinase is IkB Kinase beta (IKK-beta).
- 36. The method of claim 31 wherein said RKIP-sensitive kinase is NK-κB-inducing kinase (NIK).
- 37. The method of claim 31 wherein said RKIP-sensitive kinase is transforming growth factor beta-activated kinase 1 (TAK-1).
- 38. The method of claim 31 wherein said RKIP-sensitive kinase is MEKK3.
- 39. The method of claim 31 wherein said RKIP-sensitive kinase is A-Raf.
- 40. The method of claim 31 wherein said RKIP-sensitive kinase is B-Raf.
- 41. A method of inhibiting the activity of an RKIP-sensitive kinase, comprising the step of contacting said RKIP-sensitive kinase with an amount of an agent which inhibits the activity of said RKIP-sensitive kinase sufficient to inhibit said activity.
- 42. The method of claim 41 wherein said agent is a polypeptide.
- 43. The method of claim 41 wherein said polypeptide comprises an RKIP motif.
- 44. The method of claim 41 wherein said kinase is a MAPK/ERK kinase.
- 45. The method of claim 41 wherein said agent binds to Raf-1.
- 46. The method of claim 41 wherein said agent binds IkB Kinase alpha (IKK-alpha).
- 47. The method of claim 41 wherein said agent binds IkB Kinase beta (IKK-beta).
- 48. The method of claim 41 wherein said agent binds NK-κB-inducing kinase (NIK).
- 49. The method claim 41 wherein said agent binds transforming growth factor beta-activated kinase 1 (TAK-1).
- 50. The method claim 41 wherein said agent binds to MEKK3.
- 51. The method claim 41 wherein said agent binds to A-Raf.
- 52. The method claim 41 wherein said agent binds to B-Raf.
- 53. A method of treating a disorder that is associated with inappropriate expression or activity of an RKIP family polypeptide comprising administering a pharmaceutical composition comprising an agent that modulates the activity of an RKIP family polypeptide to an individual in need of treatment for a cell proliferative disorder.
- 54. A method of treating a disorder that is associated with inappropriate activity of an RKIP-sensitive signal transduction pathway comprising administering a pharmaceutical composition comprising an agent that modulates the activity of an RKIP family polypeptide to an individual in need of treatment for a disorder that is associated with inappropriate activity of an RKIP-sensitive signal transduction pathway.
- 55. A polypeptide consisting essentially of an RKIP motif.
- 56. An RKIP motif fusion protein.
- 57. An RKIP motif-containing fusion protein.
- 58. An isolated nucleic acid encoding an RKIP motif cassette.
- 59. The isolated nucleic acid of claim 43 wherein said nucleic acid encoding an RKIP motif cassette comprises a vector sequence.
- 60. The isolated nucleic acid of claim 44 wherein said nucleic acid encoding an RKIP motif cassette is linked to sequences encoding a heterologous amino acid sequence such that said vector encodes a fusion protein comprising an RKIP motif.
Parent Case Info
[0001] This application is a continuation-in-part application of U.S. patent application Ser. No. 09/654,281, filed Sep. 1, 2000, which claims priority to U.S. Patent Application No. 60/151,992, filed Sep. 1, 1999.
Provisional Applications (1)
|
Number |
Date |
Country |
|
60151992 |
Sep 1999 |
US |
Continuation in Parts (1)
|
Number |
Date |
Country |
Parent |
09654281 |
Sep 2000 |
US |
Child |
10382970 |
Mar 2003 |
US |