madrid2i

Resumen

A new phosphorylation site of mitogen-activated kinase proteins (MAPK) has been found. Phosphorylated MAPKs in said phosphorylation site can be used as a diagnostic marker of pathologies mediated by MAPKs. ® KIPO & WIPO 2008

Reivindicaciones

1. An MAPK protein selected from: a) an MAPK protein comprising a phosphorylated residue in a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of said MAPK protein, or a fragment of said protein comprising said phosphorylated residue, wherein - said different phosphorylation site is the threonine residue in position 123 (Thr123) of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid susceptible of phosphorylation in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and - the phosphorylation at said different phosphorylation site prevents the activation of said MAPK protein and also its activity towards its substrates; and b) an MAPK protein comprising a negative charge or a bulky residue in a phosphorylation site, or at the area surrounding said phosphorylation site, that is different from the phosphorylation site or sites present in the activation segment of said MAPK protein, or a fragment of said protein comprising said phosphorylated residue, wherein - said different phosphorylation site is the threonine residue in position 123 (Thr123) of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid susceptible of phosphorylation in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and - the introduction of a negative charge or a bulky residue at said phosphorylation site, or at the area surrounding said phosphorylation site, prevents the activation of said MAPK protein and also its activity towards its substrates. 2. Protein according to claim 1, wherein said MAPK protein is selected from the ERK, JNK and p38 protein kinases, and their respective isoforms, of any species. 3. Protein according to claim 1 or 2, wherein said MAPK protein is mammal p38. 4. Protein according to claim 3, wherein said MAPK protein is mouse p38, .alpha. isoform and has the amino acid sequence shown in SEQ ID NO: 1. 5. Protein according to claim 1, wherein said MAPK protein comprises an activation segment selected from: - an activation segment comprising the amino acid triad of formula (I) Thr-Xaa-Tyr (I) where Thr is threonine, Tyr is tyrosine, and Xaa is the residue of an amino acid, preferably, of an amino acid selected from aspartic acid, glutamic acid, glutamine, glycine and proline; and - an activation segment comprising the amino acid triad of formula (II) Ser-Glu-Gly (II) where Ser is serine, Glu is glutamic acid, and Gly is glycine. 6. A protein according to claim 1, comprising the amino acid sequence shown in SEQ ID NO: 2. 7. The use of a protein according to claim 1, in the diagnosis of a pathology mediated by an active MAPK, or for determining the risk or predisposition of a subject of developing said pathology, or for evaluating or monitoring the effect of a therapy administered to a subject having said pathology, or for analyzing the stage or severity and/or the evolution of said pathology, as well as in the identification of potentially useful compounds for the treatment of said pathology. 8. Use according to claim 7, wherein said pathology mediated by an active MAPK comprises cancer and cardiac, infectious, neuronal, pulmonary and inflammatory diseases. 9. An in vitro method for detecting in a subject a pathology mediated by an active MAPK, or for analyzing the risk or predisposition of a subject of developing a pathology mediated by an active MAPK, comprising: a) detecting and/or quantifying the level of an MAPK protein according to any of claims 1 to 6 in a biological sample from said subject; and b) comparing said level with the level of a control sample, wherein a reduction in said level with respect to the level of the control sample is indicative of the risk of the subject of developing said pathology mediated by an active MAPK. 10. An in vitro method for evaluating or monitoring the effect of a therapy administered to a subject having said pathology mediated by an active MAPK, or for analyzing the stage or severity and/or the evolution of said pathology mediated by an active MAPK, comprising: a) detecting and/or quantifying the level of an MAPK protein according to any of claims 1 to 6 in a biological sample from said subject; and b) comparing said level with the level of a control sample from the same subject. 11. An in vitro method for identifying a potentially useful compound for the treatment of pathologies mediated by active MAPK proteins, comprising: a) placing the candidate compound in contact with an MAPK protein, and b) detecting the phosphorylation of said MAPK protein in a phosphorylation site different from the phosphorylation site or sites present in the activation segment of said MAPK protein, and c) analyzing if said phosphorylation site (i) is Thr123 of mouse p38, .alpha. isoform, in the event that the MAPK protein used was said protein, or a residue of a positionally equivalent amino acid susceptible of phosphorylation in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and if (ii) the phosphorylation in said different phosphorylation site prevents the activation of said MAPK protein; or alternatively, i) placing a candidate compound selected from a compound capable of phosphorylating said MAPK protein or a compound that mimics the effect of said phosphorylation, in contact with an MAPK protein; ii) detecting the phosphorylation of said MAPK protein in a phosphorylation site of the activation segment of said MAPK protein to measure the effect of the candidate compound on the activation of the MAPK, or detecting the effect of mimicking said phosphorylation on said MAPK protein to measure the effect of the candidate compound on the activation of the MAPK; iii) analyzing the activity of the said MAPK protein in the presence of the candidate compound towards its substrates to test the possible inhibition of the docking and/or activity of the MAPK protein to its substrates in the presence of a competing compound; and iv) analyzing if said phosphorylation site (i) at Thr123 of mouse p38, .alpha. isoform, (in the event that the MAPK protein used was said protein) is affected by the candidate compound and if (ii) the phosphorylation in said phosphorylation site prevents the activation of said MAPK protein. 12. A compound capable of binding to an MAPK protein according to any of claims 1 to 6, and/or able to detect said MAPK protein according to any of claims 1 to 6. 13. A compound according to claim 12, characterized in that it is - an antibody; or - a compound capable of binding to a MAPK protein according to any of claims 1 to 6 which binds to said MAPK protein at Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the association of said compound at said phosphorylation site Thr123 causes a decreased phosphorylation of the MAPK protein at the activation segment and thereby prevents its activation and/or its activity towards its substrates; or - a compound capable of binding to the docking region of p38 and able to mimic the introduction of a negative charge in said region, said compound introducing a negative charge or a bulky residue at Thr123, or at its surrounding area, of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the association of said compound at said phosphorylation site Thr123, or at the area surrounding Thr123 prevents the activation of said MAPK protein; or - a compound capable of binding to the docking region of p38 and able to mimic the introduction of a negative charge in said region, said compound introducing a negative charge or a bulky residue at Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the association of said compound at said phosphorylation site Thr123 impairs the activity of said MAPK protein towards its substrates. 14. A compound according to claim 13, wherein said antibody is an antibody that is able of binding to the epitope comprising the amino acid sequence shown in SEQ ID NO: 2. 15. The use of said compound according to any of claims 12 to 14, for analyzing the risk or predisposition of a subject of developing a pathology mediated by an active MAPK, or for evaluating or monitoring the effect of a therapy administered to a subject who has said pathology, or for analyzing the stage or severity and/or the evolution of said pathology, as well as in the identification of potentially useful compounds for the treatment of said pathology. 16. A vector comprising: (i) a nucleic acid sequence encoding a compound phosphorylating a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of an MAPK protein, wherein said different phosphorylation site is Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid susceptible of phosphorylation in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the phosphorylation in said different phosphorylation site prevents the activation of said MAPK protein; or (ii) a nucleic acid sequence encoding a compound preventing the phosphorylation of a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of an MAPK protein; or (iii) a compound phosphorylating a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of an MAPK protein, wherein said different phosphorylation site is Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid susceptible of phosphorylation in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the phosphorylation in said different phosphorylation site prevents the activation of said MAPK protein; or (iv) a compound preventing phosphorylation in a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of an MAPK protein. 17. A pharmaceutical composition comprising a therapeutically effective amount of: (i) a compound phosphorylating a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of an MAPK protein, wherein said different phosphorylation site is Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid susceptible of phosphorylation in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the phosphorylation in said different phosphorylation site prevents the activation of said MAPK protein; or (ii) a compound mimicking the phosphorylation at a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of an MAPK protein, wherein said different phosphorylation site is Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid susceptible of phosphorylation in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the phosphorylation in said different phosphorylation site prevents the activation of said MAPK protein; or (iii) a compound preventing phosphorylation in a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of an MAPK protein; or (iv) a vector according to claim 16; or (v) a compound capable of binding to a MAPK protein according to any of claims 1 to 6 which binds to said MAPK protein at Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the association of said compound at said phosphorylation site Thr123 causes a decreased phosphorylation of the MAPK protein at the activation segment and thereby prevents its activation and/or its activity towards its substrates; or (vi) a compound capable of binding to the docking region of p38 and able to mimic the introduction of a negative charge in said region, said compound introducing a negative charge or a bulky residue at Thr123, or at its surrounding area, of mouse p38, a isoform, or a residue of a positionally equivalent amino acid in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the association of said compound at said phosphorylation site Thr123, or at the area surrounding Thr123 prevents the activation of said MAPK protein; or (vii) a compound capable of binding to the docking region of p38 and able to mimic the introduction of a negative charge in said region, said compound introducing a negative charge or a bulky residue at Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the association of said compound at said phosphorylation site Thr123 impairs the activity of said MAPK protein towards its substrates, together with, optionally, a pharmaceutically acceptable carrier. 18. Composition according to claim 17, comprising a kinase. 19. Composition according to claim 18, wherein said kinase is the GRK2 kinase or a functionally active fragment thereof. 20. The use of: (i) a compound phosphorylating a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of an MAPK protein, wherein said different phosphorylation site is Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid susceptible of phosphorylation in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the phosphorylation in said different phosphorylation site prevents the activation of said MAPK protein; or (ii) a compound mimicking the phosphorylation at a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of an MAPK protein, wherein said different phosphorylation site is Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid susceptible of phosphorylation in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the phosphorylation in said different phosphorylation site prevents the activation of said MAPK protein; or (iii) a compound preventing phosphorylation in a phosphorylation site that is different from the phosphorylation site or sites present in the activation segment of an MAPK protein; or (iv) a vector according to claim 16; or (v) a compound capable of binding to a MAPK protein according to any of claims 1 to 6 which binds to said MAPK protein at Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the association of said compound at said phosphorylation site Thr123 causes a decreased phosphorylation of the MAPK protein at the activation segment and thereby prevents its activation and/or its activity towards its substrates; or (vi) a compound capable of binding to the docking region of p38 and able to mimic the introduction of a negative charge in said region, said compound introducing a negative charge or a bulky residue at Thr123, or at its surrounding area, of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the association of said compound at said phosphorylation site Thr123, or at the area surrounding Thr123 prevents the activation of said MAPK protein; or (vii) a compound capable of binding to the docking region of p38 and able to mimic the introduction of a negative charge in said region, said compound introducing a negative charge or a bulky residue at Thr123 of mouse p38, .alpha. isoform, or a residue of a positionally equivalent amino acid in another MAPK protein as it is defined by multiple alignment of amino acid sequences, and the association of said compound at said phosphorylation site Thr123 impairs the activity of said MAPK protein towards its substrates, in the manufacture of a pharmaceutical composition for the treatment of a pathology mediated by active MAPKs. 21. A kit comprising an MAPK protein according to any of claims 1 to 6, or a compound that is able of binding to and/or detecting said MAPK protein according to any of claims 12 to 14. 22. Kit according to claim 21, useful for the diagnosis of a pathology mediated by an active MAPK, or for determining the risk or predisposition of a subject of developing said pathology, or for evaluating or monitoring the effect of a therapy administered to a subject who has said pathology, or for analyzing the stage or severity and/or the evolution of said pathology, as well as in the identification of potentially useful compounds for the treatment of said pathology. 23. Kit according to claim 21 or 22, wherein said MAPK protein is a phosphorylated mammal p38 kinase in Thr123 of mammal p38, a isoform.

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