Claims
- 1. A peptide of formula I
- 2. A peptide according to claim 1, wherein N1 and N2 are independently selected from nothing and the polar residues C, N, Q, S, T and Y.
- 3. A peptide according to claim 2, wherein N1 is a natural or unnatural amino acid.
- 4. A peptide according to claim 3, wherein N1 is threonine.
- 5. A peptide according to claim 2, wherein N2 is a natural or unnatural amino acid.
- 6. A peptide according to claim 3, wherein N1 is serine.
- 7. A peptide according to claim 1, wherein up to 6 amino acid residues are deleted from the N-terminal end of the peptide of formula I.
- 8. A peptide according to claim 7, wherein from 3-5 amino acid residues are deleted from the N-terminal end of the peptide of formula I.
- 9. A peptide according to claim 8, wherein 4 amino acid residues are deleted from the N-terminal end of the peptide of formula I.
- 10. A peptide according to claim 7, wherein N2 is a natural or unnatural amino acid.
- 11. A peptide according to claim 10, wherein N2 is serine.
- 12. A peptide according to claim 1, wherein 7 or 8 amino acid residues are deleted from the N-terminal end of the peptide of formula I.
- 13. A peptide of formula
- 14. A variant according to claim 13, wherein the serine residue corresponding to p21(153Ser), is replaced by an alanine residue.
- 15. A peptide according to claim 13, selected from;
- 16. A peptide of formula II;
- 17. A peptide according to claim 16, wherein X5 is selected from isoleucine and glycine.
- 18. A peptide according to claim 16, wherein X1 and X4 are both basic amino acid residues and X3 is a basic or polar residue.
- 19. A peptide according to claim 18, wherein X1 is histidine and X4 is arginine, and X3 is lysine or cysteine.
- 20. A peptide of formula;
- 21. A peptide of formula;
- 22. A peptide of formula;
- 23. A peptide according to claim 16, wherein X2 is alanine.
- 24. A peptide according to claim 16, wherein X5 is isoleuc Seq. I.D. # ine.
- 25. A peptide according to claim 20, selected from the group consisting of:
- 26. A peptide of the formula III or IV;
- 27. A peptide according to claim 26, wherein X2 is alanine.
- 28. A peptide according to claim 26, wherein X5 isoleucine.
- 29. A peptide according to claim 26 of the formula IV
- 30. The peptide of claim 26, wherein the peptide is in a cyclic form by virtue of a linkage between the C-terminal residue and the residue 3 upstream to it.
- 31. A peptide according to claim 30, wherein X2 is Ala and X5 is Ile.
- 32. A peptide according to claim 26, wherein F′ is para-fluoro-Phe and H′ is Ala or nothing.
- 33. The peptide of claim 26, wherein K′ is Abu; R1 is Gln; R2 is Cit or Ser; and X5 is Ala.
- 34. A peptide according to claim 26 selected from the group consisting of:
- 35. An assay for identifying candidate substances capable of binding to a cyclin associated with a G1 control CDK enzyme and/or inhibition of said enzyme, comprising;
(a) bringing into contact i) a p21 derived peptide as defined in claim 1, ii) said cyclin or portion thereof or cyclin groove, iii) said CDK or portion thereof and iv) said candidate substance, under conditions wherein, in the absence of the candidate substance being an inhibitor of the cyclin/CDK interaction, the p21 derived peptide would bind to said cyclin or portion thereof or cyclin groove, and (b) monitoring any change in the expected binding of the p21 derived peptide and the cyclin or portion thereof or cyclin groove.
- 36. An assay for the identification of compounds that interact with a cyclin or a cyclin when complexed with the physiologically relevant CDK, comprising;
(a) incubating a candidate compound and peptide of formula I; X1X2X3RX4LX5F (formula II) (SEQ ID No. 2) wherein X1, X3, X4 and X5 may be any amino acid and X2 is serine or alanine; and variants thereof or a peptide of the formula III or IV: H′X′2K′R1R2L′X′5F′ (formula III) (SEQ ID No. ______) or H′X′2K′R1R2L° F′X′5 (formula IV) (SEQ ID No. ______) or a variant thereof, wherein H′ is His, nothing, D-His, Ala, Thi, Hse, Phe, or Dab; X′2 is Ala, Ser, Abu, Val; K′ is Lys, Arg, or Abu; R1 is Arg, Lys, or Gln; and R2 is Arg, forms a cyclic peptide with the C-terminal residue, Ser, or Cit; L′ is Leu or Ile; X′5 is Ile, Leu, Gly, or Ala; F′ is Phe, para-fluoroPhe, meta-fluoroPhe, L-Psa, 2-Nap,Dhp, or D-Psa. and a cyclin or cyclin/CDK complex; (b) detecting binding of either the candidate compound or the peptide of formula II or III with cyclin.
- 37. An assay for candidate compounds that interact with a cyclin by virtue of forming associations with at least two of the amino acids corresponding to the cyclin A amino acids L253, I206 and R211.
- 38. An assay according to claim 37, wherein the candidate compound additionally forms associations with at least one of the amino acids corresponding to the cyclin A amino acids E223, E224, D284, D283, L253, I206 and R211.
- 39. An assay according to claim 37, wherein the candidate additionally forms associations with at least one of the amino acids corresponding to the cyclin A amino acids W217, V219, V221, S408, E411, Y225, I213, L214, G257, R250, Q254, T207 and L214.
- 40. An assay according to claim 37, wherein the candidate compound additionally forms associations with at least one of the amino acids corresponding to the cyclin A amino acids G222, Y225, 1281, E223, E220, V279, A212, V215, L218, Q406, S408, M210, L253, L218, 1239, V256 and M200.
- 41. An assay according to claim 35, wherein the cyclin is selected from cyclin A, cyclin E or cyclin D.
- 42. An assay according to claim 41 wherein the cyclin is cyclin A.
- 43. An assay according to claim 35, comprising use of a three dimensional model of a cyclin and a candidate compound.
- 44. An assay according to claim 35, wherein at least one of the assay components is bound to a solid phase.
- 45. An assay according to claim 44, wherein the p21 derived peptide is labeled such as to emit a signal when bound to said cyclin.
- 46. An assay according to claim 44, wherein the cyclin is labeled such as to emit a signal when bound to the p21 derived peptide.
- 47. An assay according to claim 45, wherein one of the assay components is labeled with a fluorescence emitter and the signal is detected using fluorescence polarization techniques.
- 48. A method of using a cyclin in a drug screening assay comprising:
(a) selecting a candidate compound by performing rational drug design with a three-dimensional model of said cyclin, wherein said selecting is performed in conjunction with computer modeling; (b) contacting the candidate compound with the cyclin; and (c) detecting the binding affinity of the candidate compound for the cyclin groove; wherein a potential drug is selected on the basis of its having a greater affinity for the cyclin groove than that of a peptide of formula II; X1X2X3RX4LX5F (formula II) (SEQ ID No. 2) wherein X1, X3, X4 and X5 may be any amino acid and X2 is serine or alanine; and variants thereof or a peptide of formula III or IV: H′X′2K′R1R2L′X′5F′ (formula III) (SEQ ID No. ______) or H′X′2K′R1R2L° F′X′5 (formula IV) (SEQ ID No. ______) or a variant thereof, wherein H′ is His, nothing, D-His, Ala, Thi, Hse, Phe, or Dab; X′2 is Ala, Ser, Abu, Val; K′ is Lys, Arg, or Abu; R1 is Arg, Lys, or Gln; and R2 is Arg, forms a cyclic peptide with the C-terminal residue, Ser, or Cit; L′ is Leu or Ile; X′5 is Ile, Leu, Gly, or Ala; F′ is Phe, para-fluoroPhe, meta-fluoroPhe, L-Psa, 2-Nap,Dhp, or D-Psa.
- 49. A method of using a cyclin in a drug screening assay comprising:
(a) selecting a candidate compound by performing rational drug design with a three-dimensional model of said cyclin, wherein said selecting is performed in conjunction with computer modeling; (b) contacting the candidate compound with the cyclin; and (c) detecting whether said the candidate compound forms associations with at least the amino acids corresponding to the cyclin A amino acids L253, 1206 and R211.
- 50. A method according to claim 49, further comprising detection of whether the candidate compound additionally forms associations with at least one of the amino acids corresponding to the cyclin A amino acids E223, E224, D284, D283, L253, I206 and R211.
- 51. A method according to claim 50, further comprising detection of whether the candidate compound additionally forms associations with at least one of the amino acids corresponding to the cyclin A amino acids W217, V219, V221, S408, E411, Y225, I213, L214, G257, R250, Q254, T207 and L214.
- 52. A method according to claim 50, further comprising detection of whether the candidate compound additionally forms associations with at least one of the amino acids corresponding to the cyclin A amino acids G222, Y225, I281, E223, E220, V279, A212, V215, L218, Q406, S408, M210, L253, L218, I239, V256 and M200.
- 53. An assay for identifying candidate substances capable of inhibiting CDK in a cell, comprising;
(a) contacting a cell comprising a cyclin or portion thereof or cyclin groove, and a CDK or portion thereof, with a candidate substance under conditions where, in the absence of the candidate substance, the cyclin or portion thereof or cyclin groove and CDK or portion thereof would interact, and (b) monitoring any change in the activity of the CDK or portion thereof, wherein inhibition of CDK activity is indicated by one or more of: G0 and/o G1/S cell cycle arrest; cell cycle-related apoptosis; suppression of E2F transcription factor activity; hypophosphorylation of cellular pRb; and in vitro anti-proliferative effects.
- 54. Use of a peptide defined in claim 1 in the preparation of a medicament for use in (a) inhibition of CDK2 or (b) in the treatment of proliferative disorders such as cancers and leukaemias where inhibition of CDK2 would be beneficial.
Priority Claims (1)
Number |
Date |
Country |
Kind |
9928323.6 |
Nov 1999 |
GB |
|
RELATED APPLICATIONS
[0001] This application is a continuation-in-part of U.S. Ser. No. 09/726,470, filed Nov. 29, 2000, and claims priority to Great Britain application Serial No. 9928323.6 filed Nov. 30, 1999, the contents each of which are entirely incorporated by reference.
Continuation in Parts (1)
|
Number |
Date |
Country |
Parent |
09726470 |
Nov 2000 |
US |
Child |
10441952 |
May 2003 |
US |