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Title:
INHIBITORS OF THE MAP KINASE PATHWAY
Document Type and Number:
WIPO Patent Application WO/2005/007090
Kind Code:
A3
Abstract:
MAP kinases (e.g., ERKI/2) phosphorylate a variety of target proteins including, for example, several immediate early gene products (e.g., Fos, Myc, and Jun family proteins). Certain phosphorylation reactions require binding of the MAP kinase to the DEF domain of the target protein. Inhibitors that block this interaction may be useful therapeutics for human disease, including as antineoplastic agents. Also disclosed are screening assays useful for identifying compounds that inhibit the MAP kinase-DEF domain interaction.

Inventors:
BLENIS JOHN (US)
MURPHY LEON O (US)
Application Number:
US2004/021514
Publication Date:
April 09, 2009
Filing Date:
July 02, 2004
Export Citation:
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Assignee:
HARVARD COLLEGE (US)
BLENIS JOHN (US)
MURPHY LEON O (US)
International Classes:
C12N15/00; C12N5/00; G01N33/50; G01N33/574; G01N33/68; A61K
Foreign References:
US6475778B12002-11-05
US5958721A1999-09-28
Other References:
MURPHY ET AL., NATURE CELL BIOL, vol. 4, August 2002 (2002-08-01), pages 556 - 563
MURPHY ET AL., NATURE CELL BIOL., vol. 4, August 2002 (2002-08-01), pages 556 - 563
Attorney, Agent or Firm:
BIEKER-BRADY, Kristina, Ph., D., P.C. (101 Federal StreetBoston, MA, US)
Download PDF:
Claims:
Claims 1. A method for identifying therapeutic compounds, said method comprising : (i) providing test cells that express a target protein containing a DEF domain and a MAP kinase, (ii) culturing said cells in the presence of a growth factor, cytokine, tumor promoter, or oncogene, (iii) contacting said cells with a candidate compound, and (iv) assessing the binding of said MAP kinase to said DEF domain relative to the binding in the absence of said candidate compound, wherein a candidate compound that inhibits said binding is identified as a therapeutic compound.
2. The method of claim 1, wherein said DEF domain comprises the amino acid sequence F-X-F-P (SEQ ID NO: 1).
3. The method of claim 1, wherein said test cells are selected from the group consisting of fibroblasts, a primary cell line, an immortalized cell line, and a tumor-derived cell line.
4. The method of claim 1, wherein said growth factor, cytokine, tumor promoter, or oncogene is selected from the group consisting of epidermal growth factor (EGF), transforming growth factor a, heparin-binding-like EGF, heregulin, amphiregulin, epiregulin, cripto, PDGF-AA, PDGF-BB or PDGF-CC, insulin, insulin-like growth factors, fibroblast growth factors, colony stimulating factor, heaptocyte growth factor, a chemokine, an interleukin, lysophosphatidic acid, a phorbol ester, okadaic acid, microcystin, vanadate, hydrogen peroxide, calyculin A, Erb2/neu, sis, kit, Ras, Raf, P13-kinase, and PTEN.
5. The method of claim 1, wherein said MAP kinase is extracellular signal- regulated kinase 1/2 (ERK1/2).
6. The method of claim 1, wherein said binding is assessed by detecting a DEF domain-dependent phosphorylation.
7. The method of claim 1, wherein said target protein is a Fos, Myc, or Jun family protein.
8. The method of claim 7, wherein said target protein is c-Fos.
9. The method of claim 8, wherein said step (vi) comprises assessing the phosphorylation of T325 or T331.
10. The method of claim 8, wherein said step (vi) comprises an antibody that specifically binds to phospho-T-325 c-Fos.
11. The method of claim 1, wherein said therapeutic is useful for the treatment of cancer and said target protein comprises the sequence of a protein identified in Tables 1 or 2.
12. The method of claim 1, wherein said therapeutic is useful for the treatment of a cardiovascular disorder and said target protein comprises the sequence of a protein identified in Table 3.
13. The method of claim 1, wherein said therapeutic is useful for the treatment of an inflammatory disorder, and said target protein comprises the sequence of a protein identified in Table 4.
14. The method of claim 1, wherein said therapeutic is useful for the treatment of a metabolic disorder, and said target protein comprises the sequence of a protein identified in Table 5.
15. The method of claim 1, wherein said therapeutic is useful for the treatment of a neuropathy or a behavioral disorder, and said target protein comprises the sequence of a protein identified in Table 6.
16. The method of claim 1, wherein said therapeutic is useful for the treatment of a sleep disorder, and said target protein comprises the sequence of a protein identified in Table 7.
17. A method for identifying a therapeutic compound, said method comprising : (i) providing a sample comprising a target protein comprising a DEF domain, a MAP kinase, and a candidate compound, (ii) contacting said target protein, said MAP kinase, and said candidate compound, (iii) assessing the binding of said MAP kinase to said DEF domain in said sample in the presence of said candidate compound relative to binding in the absence of said candidate compound, wherein a compound that inhibits binding of said MAP kinase to said target protein is identified as a therapeutic compound.
18. The method of claim 17, wherein said DEF domain comprises the amino acid sequence F-X-F-P (SEQ ID NO: 1).
19. The method of claim 17, wherein said MAP kinase is extracellular signal-regulated kinase 1/2 (ERK1/2).
20. The method of claim 17, wherein said binding is assessed by detecting a DEF domain-dependent phosphorylation.
21. The method of claim 17, wherein said target protein is a Fos, Myc, or Jun family protein.
22. The method of claim 21, wherein said target protein is c-Fos.
23. The method of claim 22, wherein said target protein is c-Fos and step (vi) comprises assessing the phosphorylation of T325 or T331.
24. The method of claim 22, wherein said step (vi) comprises an antibody that specifically binds to phospho-T-325 c-Fos.
25. The method of claim 17, wherein said therapeutic is useful for the treatment of cancer and said target protein comprises the sequence of a protein identified in Tables 1 or 2.
26. The method of claim 17, wherein said therapeutic is useful for the treatment of a cardiovascular disorder and said target protein comprises the sequence of a protein identified in Table 3.
27. The method of claim 17, wherein said therapeutic is useful for the treatment of an inflammatory disorder, and said target protein comprises the sequence of a protein identified in Table 4.
28. The method of claim 17, wherein said therapeutic is useful for the treatment of a metabolic disorder, and said target protein comprises the sequence of a protein identified in Table 5.
29. The method of claim 17, wherein said therapeutic is useful for the treatment of a neuropathy or a behavioral disorder, and said target protein comprises the sequence of a protein identified in Table 6.
30. The method of claim 17, wherein said therapeutic is useful for the treatment of a sleep disorder, and said target protein comprises the sequence of a protein identified in Table 7.
31. The method of claim 17, wherein said target protein further comprises a fluorescent moiety.
32. A method for treating a cancer in a mammal, said method comprising administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein.
33. The method of claim 32, wherein said MAP kinase is ERK1/2.
34. The method of claim 32, wherein said target protein is an immediate early gene.
35. The method of claim 32, wherein said target protein is selected from the group consisting of c-Fos, Fra-1, Fra-2, c-Myc, N-Myc, JunD, JunB, and c-Jun.
36. The method of claim 35, wherein said target protein is c-Fos.
37. The method of claim 32, wherein said target protein is a protein identified in Tables 1 or 2.
38. The method of claim 32, wherein said cancer is selected from the group consisting of leukemia, Hodgkin's disease lymphoma, non-Hodgkin's disease lymphoma, fibrosarcoma, liposarcoma, osteogenic sarcoma, colon carcinoma, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, Wilm's tumor, cervical cancer, uterine cancer, testicular cancer, small cell lung carcinoma, non-small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, schwannoma, meningioma, melanoma, neuroblastoma, and retinoblastoma.
39. A method for treating cardiovascular disease in a mammal, said method comprising administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein.
40. The method of claim 39, wherein said MAP kinase is ERK1/2.
41. The method of claim 39, wherein said target protein is a protein identified in Table 3.
42. The method of claim 39, wherein said cardiovascular disease is selected from the group consisting of ischemic heart disease, ventricular heart failure, cardiac hypertrophy, hypertension, and atherosclerosis.
43. A method for treating an inflammatory disorder in a mammal, said method comprising administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein.
44. The method of claim 43, wherein said MAP kinase is ERK1/2.
45. The method of claim 43, wherein said target protein is a protein identified in Table 4.
46. The method of claim 43, wherein said inflammatory disorder is selected from the group consisting of anaphylaxis, septic shock, allergic rhinitis, asthma, atopic dermatitis, and food allergies. Examples of autoimmune disorders include, but are not limited to, type 1 insulin-dependent diabetes mellitus, inflammatory bowel disease, Crohn's disease, ulcerative colitis, dermatitis, meningitis, thrombotic thrombocy topenic purpura, Sjogren's syndrome, encephalitis, uveitis, leukocyte adhesion deficiency, rheumatoid and other forms of immune arthritis, rheumatic fever, Reiter's syndrome, psoriatic arthritis, progressive systemic sclerosis, primary biliary cirrhosis, pemphigus, pemphigoid, necrotizing vasculitis, myasthenia gravis, multiple sclerosis, lupus erythematosus, polymyositis, sarcoidosis, granulomatosis, vasculitis, pernicious anemia, CNS inflammatory disorder, antigen-antibody complex mediated diseases, autoimmune hemolytic anemia, Hashimoto's thyroiditis, Graves disease, habitual spontaneous abortions, Reynard's syndrome, glomerulonephritis, dermatomyositis, chronic active hepatitis, celiac disease, autoimmune complications of AIDS, atrophic gastritis, ankylosing spondylitis and Addison's disease.
47. A method for treating a metabolic disorder in a mammal, said method comprising administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein.
48. The method of claim 47, wherein said MAP kinase is ERK1/2.
49. The method of claim 47, wherein said target protein is a protein identified in Table 5.
50. The method of claim 47, wherein said metabolic disorder is selected from the group consisting of diabetes, obesity, jaundice, polycystic kidney and hepatic disease, pancreatitis, Graves'disease, and Werner's syndrome.
51. A method for treating a neuropathy or behavioral disorder in a mammal, said method comprising administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein.
52. The method of claim 51, wherein said MAP kinase is ERK1/2.
53. The method of claim 51, wherein said target protein is a protein identified in Table 6.
54. The method of claim 51, wherein said neuropathy or behavioral disorder is selected from the group consisting of diabetic neuropathy, muscular dystrophy, Williams Beuren's Syndrome, psychosis, schizophrenia, autism, Down's Syndrome, Parkinson's Disease, Alzheimer's Disease, epilepsy, Cockayne syndrome, bipolar disorders, depression, and opiate addiction.
55. A method for treating a sleep disorder in a mammal, said method comprising administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein.
56. The method of claim 55, wherein said MAP kinase is ERK1/2.
57. The method of claim 55, wherein said target protein is a protein identified in Table 7.
58. The method of claim 55, wherein said sleep disorder is selected from the group consisting of advanced sleep phase disorder, delayed sleep phase disorder, insomnia, and narcolepsy.
59. An antibody that specifically binds to phospho-T-325 c-Fos.
60. The antibody of claim 59, wherein said antibody is polyclonal.
61. The antibody of claim 59, wherein said antibody is monoclonal.
Description:

INHIBITORS OF THE MAP KINASE PATHWAY Field of the Invention This invention relates to the development and use of human therapeutics that inhibit intracellular signaling via the MAP kinase pathways.

Background of the Invention The evolutionarily conserved Ras-MAPK signaling network regulates diverse biological processes such as cell proliferation, differentiation, migration, and survival. Many of the regulators and effectors within this network have been implicated in diverse pathological processes. MAP kinases and their targets have been identified as, for example, potent oncogenes or tumor suppressor genes and proinflammatory mediators.

Normally, the MAPK network is activated when growth factors or hormones bind to cell surface receptors. The extracellular signal is amplified and converted into an appropriate biological response. However, dysfunction of any component of the signaling pathway may result in a pathological condition.

Cancer, for example, is a disease marked by the uncontrolled growth of abnormal cells. Cancerous cells have overcome the barriers imposed in normal cells, which have a finite lifespan, and grow indefinitely. As the growth of cancer cells continues, genetic alterations can accrue and persist so that the cancerous cell displays increasingly aggressive growth phenotypes. If left untreated, metastasis, the spread of cancer cells to distant areas of the body by way of the lymph system or bloodstream, may ensue, destroying healthy tissue and, ultimately, leading to death.

According to a recent American Cancer Society study, at least 1, 268, 000 new cancer cases are expected to be diagnosed in the United States in any given year.

However in cancer cells, mutations in upstream activators of MAPK, such as Ras or Raf, lead to constitutive signaling even in the absence of growth factors.

Constitutively activating mutations in Ras are detected in at least 30% of all human malignancies but are present in especially high levels in colon (50%) and pancreatic cancers (90%). The activation kinetics of the ERK1/2-MAPK signaling pathway have also been associated with distinct biological outcomes. In fibroblasts, sustained ERK1/2 activation over several hours induces entry into S phase of the cell cycle while transient (20-30 min) activation does not.

In various cell types, the ERK1/2 pathway also has a critical role in regulation of nucleotide biosynthesis, transcription, migration, cell survival, differentiation and adaptive responses. Specifically, ERK signaling can control cardiomyocyte cell growth and the response to ventricular heart failure, cell survival in atherosclerosis, various metabolic processes such as glucose uptake, protein synthesis and leptin signaling, regulation of the immune response such as in T cell activation and inflammatory cytokine signaling, and mediating the effect of neurotransmitters that control memory and behavior. ERK signaling also can control the induction of genes that are required for establishing circadium rhythms.

Accordingly, small molecule drugs that can selectively inhibit regulatory proteins within the ERK1/2-MAPK pathway have enormous therapeutic potential.

General MAPK inhibitors, however, are likely to be toxic due to the many metabolic and proliferative functions regulated by this pathway in healthy cells.

ERK1/2 specifically recognizes some physiological substrates through the presence of ERK1/2 docking sites in substrates (Jacobs et al., 1999; Tanoue et al., 2000). At least two classes of docking site have been identified and are known as the D-box and DEF domain.

Substrate docking directs ERK1/2 to phosphorylate specific amino acids known to regulate the biological function of the substrate. Interaction of ERK1/2 with the D-box docking site is required for ERK's initial activation by MEK, as well as its inactivation by phosphatases (Tanoue et al., 2000). By contrast, the DEF domain appears to be mainly found in downstream targets of ERK1/2 (Jacobs et al. , 1999).

Summary of the Invention We have discovered that MAP kinases (e. g. , extracellular signal-regulated kinase 1/2 (ERK1/2)), bind to certain target proteins (e. g. , immediate early gene (IEG) products) through a DEF domain. This specific binding interaction results in the phosphorylation of target residues and a resulting biological effect (e. g., progression through the cell cycle). Blocking the binding events we have identified allows treatment of a variety of human diseases where the interaction of MAP kinases with the DEF domain of the target proteins has a causative biological effect.

) Accordingly, in one aspect, the present invention provides for a method of identifying therapeutic compounds that affect the MAP kinase-DEF domain interaction. The method consists of the steps of : (i) providing test cells that express a target protein having a DEF domain and a MAP kinase, and are capable of progressing through the cell cycle; (ii) culturing the test cells in the presence of a growth factor, cytokine, tumor promoter, or oncogene under conditions that activate the MAP kinase; (iii) contacting the test cells with the candidate compound; (iv) assessing the binding of said MAP kinase to the DEF domain of the target protein relative to the binding in the absence of said candidate compound, wherein a candidate compound that inhibits the binding is identified as a therapeutic compound. Desirably, the test cells are mammalian ; most desirably

human. Suitable test cells include, for example, a primary cell line, an immortalized cell line, or a tumor cell line. Fibroblasts (e. g. , NIH 3T3 cells) are particularly useful test cells, but any mammalian cell type can be used because IEGs are ubiquitously expressed. Useful growth factors, cytokines, tumor promoters, and oncogenes include, for example, epidermal growth factor (EGF) and EGF-related factors including, for example, transforming growth factor a (TGFa), heparin-binding-like EGF, heregulin, amphiregulin, epiregulin, cripto, platelet derived growth factor (PDGF), including PGDF-AA, PGDF-BB, and PGDF-CC, insulin, insulin-like growth factors (IGFs), fibroblast growth factors (FGFs), colony stimulating factor (CSF), and heaptocyte growth factor (HGF).

Useful cytokines include, for example, the chemokines, interleukins, and lysophosphatidic acid (LPA). Useful tumor promoters include, for example, phorbol esters, phosphatase inhibitors such as okadaic acid, microcystin, vanadate, hydrogen peroxide. and calyculin A. Useful oncogenes include, for example, Erb2/neu, sis, kit, Ras, Raf, PI3-kinase, and PTEN. Epidermal growth factor (EGF) and platelet-derived growth factor (PDGF) are particularly useful growth factors. In other embodiments in which the target protein is c-Fos, the binding of the MAP kinase to c-Fos is assessed by measuring the phosphorylation of T325 or T331. Preferably, this is performed using a phospho-T325-specific antibody.

In another aspect, the invention provides a method for identifying a therapeutic compound by (i) providing a sample that contains a polypeptide having a DEF domain, a MAP kinase, and a candidate compound, (ii) contacting the target protein, the MAP kinase, and the candidate compound, and (iii) assessing the binding of the MAP kinase to the DEF domain of the target protein in the sample in the presence of the candidate compound relative to binding in the absence of the candidate compound, wherein a compound that inhibits binding of the MAP kinase to the target protein is identified as a therapeutic compound. In desirable

embodiments, the target protein further contains a fluorescent moiety (e. g., fluorescein).

In preferred embodiments of the previous two aspects, the MAP kinase is ERK1/2. In other desirable embodiments, the target proteins are members of the Fos, Jun, and Myc family proteins. Specifically, desirable target proteins include c-Fos, Fra-1, Fra-2, cMyc, N-Myc, JunD, JunB, c-Jun, in addition to Egr-1 and mPerl. In one embodiment, the target protein contains the sequence of a protein identified in Table 1 or 2 and the identified therapeutic is useful for treating cancer. In another embodiment, the target protein contains the sequence of a protein identified in Table 3 and the identified therapeutic is useful for treating a cardiovascular disease. In another embodiment, the target protein contains the sequence of a protein identified in Table 4 and the identified therapeutic is useful for treating an inflammatory disorder. In another embodiment, the target protein contains the sequence of a protein identified in Table 5 and the identified therapeutic is useful for treating a metabolic disorder. In another embodiment, the target protein contains the sequence of a protein identified in Table 6 and the identified therapeutic is useful for treating a neuropathy or a behavioral disorder.

In another embodiment, the target protein contains the sequence of a protein identified in Table 7 and the identified therapeutic is useful for treating a sleep disorder. In other embodiments, the target protein contains a DEF domain having the amino acid sequence F/Y-X-F/Y-X (SEQ ID NO: 28), preferably the sequence is F/Y-X-F/Y-P (SEQ ID NO: 29), more preferably the sequence is F-X-F-P (SEQ ID NO: 1). Assessment of target residue phosphorylation is desirably performed using a phospho-specific antibody.

In another aspect, the invention provides a method for treating cancer in a mammal (e. g. , human) by administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target

protein. Preferably, the MAP kinase is ERK 1/2, and the target protein is a member of the Fos, Jun, and Myc family proteins including, for example, c-Fos, Fra-1, Fra-2, cMyc, N-Myc, JunD, JunB, and c-Jun. Alternatively, the target protein is one identified in Tables 1 or 2. Alternatively, the compound inhibits the binding of a MAP kinase to a DEF domain having the amino acid sequence F/Y- X-F/Y-X (SEQ ID NO: 28), preferably the sequence is F/Y-X-F/Y-P (SEQ ID NO: 29), more preferably the sequence is F-X-F-P (SEQ ID NO: 1).

In another aspect, the invention provides a method for treating a cardiovascular disease in a mammal (e. g. , human) by administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein. Preferably, the MAP kinase is ERK 1/2, and the target protein is one identified in Table 3. Alternatively, the compound inhibits the binding of a MAP kinase to a DEF domain having the amino acid sequence F/Y-X-F/Y-X (SEQ ID NO: 28), preferably the sequence is F/Y-X-F/Y-P (SEQ ID NO: 29), more preferably the sequence is F-X-F-P (SEQ ID NO: 1).

In another aspect, the invention provides a method for treating an inflammatory disorder in a mammal (e. g. , human) by administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein. Preferably, the MAP kinase is ERK 1/2, and the target protein is one identified in Table 4. Alternatively, the compound inhibits the binding of a MAP kinase to a DEF domain having the amino acid sequence F/Y-X-F/Y-X (SEQ ID NO: 28), preferably the sequence is F/Y-X-F/Y-P (SEQ ID NO: 29), more preferably the sequence is F-X-F-P (SEQ ID NO: 1).

In another aspect, the invention provides a method for treating a metabolic disorder in a mammal (e. g. , human) by administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein. Preferably, the MAP kinase is ERK 1/2, and the target protein is one identified in Table 5. Alternatively, the compound inhibits the binding of a MAP kinase to a DEF domain having the amino acid sequence F/Y- X-F/Y-X (SEQ ID NO: 28), preferably the sequence is F/Y-X-F/Y-P (SEQ ID NO: 29), more preferably the sequence is F-X-F-P (SEQ ID NO: 1).

In another aspect, the invention provides a method for treating a neuropathy or a behavioral disorder in a mammal (e. g. , human) by administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein. Preferably, the MAP kinase is ERK 1/2, and the target protein is one identified in Table 6. Alternatively, the compound inhibits the binding of a MAP kinase to a DEF domain having the amino acid sequence F/Y-X-F/Y-X (SEQ ID NO: 28), preferably the sequence is F/Y-X-F/Y-P (SEQ ID NO: 29), more preferably the sequence is F-X-F-P (SEQ ID NO: 1).

In another aspect, the invention provides a method for treating a sleep disorder in a mammal (e. g. , human) by administering a therapeutically effective amount of a compound that inhibits the binding of a MAP kinase to the DEF domain of a target protein. Preferably, the MAP kinase is ERK 1/2, and the target protein is one identified in Table 7. Alternatively, the compound inhibits the binding of a MAP kinase to a DEF domain having the amino acid sequence F/Y- X-F/Y-X (SEQ ID NO: 28), preferably the sequence is F/Y-X-F/Y-P (SEQ ID NO: 29), more preferably the sequence is F-X-F-P (SEQ ID NO: 1).

Particularly useful DEF domain inhibitors for any of the therapeutic methods are polypeptides having the sequence F/Y-X-F/Y-X (SEQ ID NO: 28;"naked DEF domains") and chimeric proteins that contain a DEF domain inserted into a non-target protein. In preferred embodiments, the DEF domain has the sequence F/Y-X-F/Y-P (SEQ ID NO: 29), more preferably the sequence is F- X-F-P (SEQ ID NO: 1). The most desirable chimeric proteins are based on non- target proteins that affect the pharmacokinetic or pharmacodynamic properties compared to administering the naked DEF domain alone. For example, DEF domains may be incorporated into serum albumin or cereloplasmin.

The compound is administered in an amount, frequency, and duration that is therapeutically effective for treating the diagnosed condition. Desirably, the compound is administered in an amount between 0. 01 and 3000 mg/day, more preferably, in an amount between 0.1 and 2000 mg/day, either orally or by injection (i. e., intravenous, intramuscular, or subcutaneous). Alternatively, the compound can be administered as a 0.5% to 25% topical formulation.

Therapy may be provided in any appropriate location: at home, the doctor's office, a clinic, a hospital's outpatient department, or a hospital. Treatment generally begins at a hospital so that the doctor can observe the therapy's effects closely and make any adjustments that are needed. The duration of the therapy depends on the condition being treated, the age and condition of the patient, the stage and type of the patient's disease, and how the patient's body responds to the treatment. Drug administration may be performed at different intervals (e. g. , daily, weekly, or monthly).

In another aspect, the invention provides an antibody that specifically binds to phospho-T-325 c-Fos. The antibody may be monoclonal or polyclonal.

In another aspect, the invention provides a pharmaceutical formulation that contains a therapeutic compound identified by either of the first two aspects of the invention, and a pharmaceutically acceptable carrier. The pharmaceutical formulation may be suitable for oral administration, injection, or topical application.

By"specifically binds to phospho-T-325 c-Fos, "when referring to the antibodies of this invention, is meant an antibody that binds with high affinity (<10-8M) to native c-Fos in which the threonine at amino acid position 325 is phosphorylated, but does not significantly bind to c-Fos in which the T325 is unphosphorylated. Desirably, the difference in specificity of antibody binding between phospho-T-325 c-Fos and the unphosphorylated form is at least 5-fold, 10-fold, 25-fold, 50-fold, 100-fold, or 1000-fold.

By"DEF domain"is meant a polypeptide having the amino acid sequence: F/Y-Xl-F/Y-X2 (SEQ ID NO: 28), wherein F is phenylalanine, Y is tyrosine, P is proline, and Xi and X2 are any naturally-occurring or non-naturally-occurring amino acids. Desirably, X2 is proline.

By"target protein"is meant any protein that contains a DEF domain capable of binding a target kinase (e. g. , a MAP kinase). Desirable target proteins are phosphorylated by the MAP kinase ERK1/2 following ERK1/2 binding to the DEF domain. Target proteins include, for example, gene products of the immediate early genes from the Fos, Myc, and Jun families, proteins identified in Tables 1-7, or chimeric or synthetic proteins into which a DEF domain has bee inserted by artifice. Specific target proteins include, for example, c-Fos, Fra-l, Fra-2, cMyc, N-Myc, JunD, JunB, c-Jun, Egr-1, and mPerl.

By"target residue (s)" is meant one or more residues of a target protein that are N-terminal to the DEF domain and that are phosphorylated as a result of the binding of a MAP kinase. Target residues include, for example, T325 and T331 of c-Fos. This phosphorylation event is also termed a"DEF domain-dependent phosphorylation." By"primed, "when referring to a target protein, is meant a phosphorylation event that makes a DEF domain available for binding of a MAP kinase. Thus, the amino acid residues that are the subject of a"priming"modification are not the same as the target residues. For example, c-Fos is primed when S362 and/or S374 are phosphorylated or substituted for aspartate or glutamate.

By"target kinase"is meant a protein kinase that is capable of binding a DEF domain and phosphorylating a target residue. Target kinases include the MAP kinases such as ERK1/2, for example. Thus, an"activated target kinase"is one that itself has undergone a post-translational modification causing an increase in kinase activity and/or inducing a change in subcellular localization. For example, in order to be fully activated and translocated from the cytoplasm to the nucleus, ERK1/2 is phosphorylated.

By"DEF domain inhibitor"is meant any chemical compound (i. e., polypeptide or non-peptide) that inhibits the interaction of a target kinase (i. e., ERK1/2 or RSK) with the DEF domain of a target protein.

The term"assessing the binding of a MAP kinase to a DEF domain, "is meant to include any appropriate binding or biochemical assessment which may be either qualitative or quantitative. This term specifically includes, for example, directly assessing the interaction of the MAP kinase and the DEF domain.

Alternatively, assays that measure biochemical outcomes of a MAP kinase-DEF domain binding event are useful. These assays include, for example, measuring the amount of DEF domain-dependent phosphorylation.

As exemplified in detail below, the phosphorylation of T325 and/or T331 of c-fos is dependent upon this binding event.

By"cancer"is meant neoplastic cells multiplying in an abnormal manner.

In a cancer, growth is uncontrolled and progressive, and occurs under conditions that would not elicit, or would halt the multiplication of non-cancerous cells.

Cancer includes, for example, leukemias and lymphomas (Hodgkin's disease, non- Hodgkin's disease), as well as solid tumors such as sarcomas and carcinomas (e. g., fibrosarcoma, liposarcoma, osteogenic sarcoma, colon carcinoma, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, Wilm's tumor, cervical cancer, uterine cancer, testicular cancer, small and/or non-small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, schwannoma, meningioma, melanoma, neuroblastoma, and retinoblastoma.

By"treating cancer"is meant a therapy that measurably slows, stops, or reverses the growth rate of the cancer (i. e. , neoplastic cells) in vivo. Desirably, a slowing of the growth rate is by at least 20%, 30%, 50%, or even 70%, as determined using a suitable assay for determination of cell growth rates (e. g. , a cell growth assay described herein). Typically, a reversal of growth rate is accomplished by initiating or accelerating necrotic or apoptotic mechanisms of cell death in the neoplastic cells, resulting in a shrinkage of the neoplasm. Efficacy of a treatment may be measured by any means known to those skilled in the art including tumor imaging or measurement of neoplastic markers.

By"cardiovascular disease"is meant ischemic heart disease, ventricular heart failure, cardiac hypertrophy, hypertension, and atherosclerosis.

By"inflammatory disorder"is meant any condition that is characterized by inflammation as a primary or secondary symptom. Inflammatory disorders include, for example, allergic or autoimmune disorders, anaphylaxis, and septic shock. Examples of allergic disorders include allergic rhinitis, asthma, atopic dermatitis, and food allergies. Examples of autoimmune disorders include, but are not limited to, type 1 insulin-dependent diabetes mellitus, inflammatory bowel disease, Crohn's disease, ulcerative colitis, dermatitis, meningitis, thrombotic thrombocytopenic purpura, Sjögren's syndrome, encephalitis, uveitis, leukocyte adhesion deficiency, rheumatoid and other forms of immune arthritis, rheumatic fever, Reiter's syndrome, psoriatic arthritis, progressive systemic sclerosis, primary biliary cirrhosis, pemphigus, pemphigoid, necrotizing vasculitis, myasthenia gravis, multiple sclerosis, lupus erythematosus, polymyositis, sarcoidosis, granulomatosis, vasculitis, pernicious anemia, CNS inflammatory disorder, antigen-antibody complex mediated diseases, autoimmune hemolytic anemia, Hashimoto's thyroiditis, Graves disease, habitual spontaneous abortions, Renard's syndrome, glomerulonephritis, dermatomyositis, chronic active hepatitis, celiac disease, autoimmune complications of AIDS, atrophic gastritis, ankylosing spondylitis and Addison's disease.

By"metabolic disorder"is meant a disease that interferes with the normal metabolic function of cells, tissues or organs. Metabolic disorders include, for example, diabetes, obesity, jaundice, polycystic kidney and hepatic disease, pancreatitis, Graves'disease, and Werner's syndrome. Metabolic diseases may also arise as secondary complications of another disease such as one involving a tumor. For example, cachexia or muscle wasting, and metabolic and digestive complications often arise from the presence of pancreatic, colonic, stomach, hepatic and hepatocellular tumors.

By"neuropathy"is meant any condition of the central or peripheral nervous system characterized by axonal loss that may or may not be accompanied by neuronal loss. Neuropathies specifically include conditions affecting sensory and motor neurons and include, for example, diabetic neuropathy, muscular dystrophy, Williams Beuren's Syndrome.

By"behavioral disorder"is meant any condition affecting motivation, emotion, learning, or memory. Behavioral disorders are also meant to broadly encompass neurodegenerative diseases. Thus, behavioral disorders include, for example, psychosis, schizophrenia, autism, Down's Syndrome, Parkinson's Disease, Alzheimer's Disease, epilepsy, Cockayne syndrome, bipolar disorders, and depression. Also included are addictions including, for example, addictions to opiates and barbiturates.

By"sleep disorder"is meant any condition that primarily affects sleep and consciousness. Sleep disorders include, for example, advanced sleep phase syndrome, delayed sleep phase syndrome, insomnia and narcolepsy.

By"a therapeutically effective amount"is meant the amount of a compound required to treat cancer (i. e. , inhibit the growth of the neoplastic cells). The effective amount of active compound (s) used to practice the present invention for therapeutic treatment of neoplasms (i. e. , cancer) varies depending upon the manner of administration, the age, body weight, and general health of the subject.

Ultimately, the attending physician or veterinarian will decide the appropriate amount and dosage regimen.

Brief Description of Drawings FIGURE 1 is a series of photomicrographs showing the differential responsiveness of Swiss 3T3 fibroblasts to growth factors. FIGURE 1A shows quiescent Swiss 3T3 cells (-) that were treated with EGF (25 ng/ml) or PDGF (20

ng/ml) for 20 h and then processed for BrdU incorporation, as described below.

FIGURE 1B shows quiescent Swiss 3T3 cells that were treated with PDGF or EGF for the indicated times and ERK1/2 and RSK kinase activities were determined using immunecomplex kinase assays. The fold activation at each time is indicated above each lane. FIGURE 1 C is the indirect immunofluorescence detection of hyperphosphorylated activated ERK1/2 in Swiss 3T3 cells treated with EGF or PDGF. FIGURE 1D is the indirect immunofluorescence detection of c-Fos in Swiss 3T3 cells treated with EGF or PDGF.

FIGURE 2A is an illustration showing the residues in c-Fos that are phosphorylated by RSK and ERK1/2 in vivo. FIGURE 2B shows the electrophoretic separation of cell extracts from parallel cultures of 208F fibroblasts stably expressing Fos-WT (WT), Fos-AA (AA) or Fos-DD (DD) that were metabolically labelled with 35S-methionine or 32P-orthophosphate and cultured with or without 10% FBS for 15 min. Fos proteins were immunoprecipitated from cell lysates and analysed by SDS-PAGE. The serum-stimulated phosphorylation of Fos-DD was consistently two to threefold greater than Fos-AA and arrows indicate the major mobilities observed after stimulation. FIGURE 2C is Western blot from NIH 3T3 cells transfected with Fos-WT (WT), Fos-AA (AA) or Fos-DD (DD) were serum-starved and then pre-treated with 5 u. M U0126 (- ) or 0. 1% DMSO (-) for 30 min before treatment with EGF (+) for 5 min. FIGURE 2D is an autoradiogram of an in vitro phosphorylation of the indicated (His) 6-Fos proteins by endogenous ERK1/2 from quiescent or EGF-stimulated NIH 3T3 cells. Results shown are representative of three independent experiments. Fos-EE (S362E/S374E) was used as primed c-Fos for in vitro phosphorylation studies.

FIGURE 3A is an illustration that details the DEF domain at the C-terminus of c-Fos. The phosphorylation of Fos-EE in the absence of peptide competitor is expressed as 100%. In vitro phosphorylation of (His) 6-Fos-EE was performed as

described below. The data shown are the means : SEM from three experiments.

FIGURE 3B, is a graph showing the inhibition of Fos-EE phosphorylation by peptides containing a DEF domain (FQFP; SEQ ID NO: 3) or a mutated DEF domain (AQAP; SEQ ID NO: 4). FIGURE 3C is a graph showing the inhibition of Fos-EE phosphorylation by peptides containing the c-Fos DEF domain (FTYP; SEQ ID NO: 2) or a mutated DEF (ATYP; SEQ ID NO: 5). FIGURE 3D shows the results of a Western blot of NIH 3T3 cells transfected with the indicated FLAG-Fos-DD (DD) alleles were left quiescent (-) or were stimulated (+) with EGF for 5 min before lysis. Arrows indicate the major Fos-DD mobilities.

FIGURE 3E is a Western blot of cells transfected with the indicated FLAG-Fos alleles. Arrows show the three major Fos-WT mobilities associated with growth factor stimulation.

FIGURE 4A is an illustration identifying the ERK1/2 phosphorylation sites N-terminal to the DEF domain in c-Fos. Iii vitro phosphorylation of (His) 6-Fos-EE proteins by activated (His) 6-ERK/2 was performed. The phosphorylation of the Fos-EE point mutants is expressed as a percentage of Fos-EE (100%). The data shown are the means ITEM from three experiments. FIGURE 4B is a Western blot of NIH 3T3 cells that were transfected with the indicated FLAG-Fos-DD (DD) alleles. Cells were treated with EGF for 5 min or left untreated. FIGURE 4C is a Western blot of cells transfected with the indicated FLAG-Fos alleles and treated as in Figure 3B.

FIGURE 5A is an illustration identifying the phospho-Thr 325 peptide used to generate the phospho-Thr-325-specific anti-c-Fos antiserum. FIGURE 5B is a Western blot of NIH 3T3 cells that were transfected with the indicated c-Fos alleles or with vector alone. Extracts were prepared from quiescent (-) or EGF- stimulated (+) cells and analyzed using either the anti-c-Fos antibody or the phospho-Thr 325 antiserum. Results shown are representative of three

independent experiments. FIGURE 5C is a Western blot of AB-Raf-ER NIH 3T3 cells transfected with Fos-WT or Fos-AA that were either starved and left untreated (0) or treated with 1 p^M tamoxifen (TAMX) for the indicated times before lysis. The in vivo phosphorylation of Thr 325 in Fos-WT and Fos-AA was analyzed by western blotting using the phospho-Thr 325-specifi antiserum.

FIGURES 5D and 5E are Western blots demonstrating the iii vivo mitogen- regulated phosphorylation of Thr 325 in the context of the Fos DEF domain mutants. NIH 3T3 cells expressing the indicated Fos proteins were treated as in the same manner as for FIGURE 3B, and extracts analyzed for phosphorylation of Thr 325.

FIGURE 6A is a Western blot of quiescent Swiss 3T3 cells were treated with EGF (25 ng/ml) for the indicated times. Lysates were probed for endogenous c-Fos, Thr 325 phosphorylation in c-Fos. FIGURE 6B is a Western blot of Swiss 3T3 cells that were treated with PDGF-BB (20 ng/ml) and processed as in described for Figure 6A. Results shown are representative of three experiments.

FIGURE 6C is a Western blot of quiescent Swiss 3T3 cells that were treated with PDGF for 60 min and then treated with U0126 (5 pM), as indicated, or with DMSO (lanes 3-8) for the remainder of the experiment. The expression and phosphorylation of endogenous c-Fos was visualized as in above. FIGURE 6D is an autoradiogram showing the kinase activities of endogenous ERK1/2 and RSK in cell lysates from Figure 6C. The fold activity is provided above each lane.

FIGURE 7A is a bar graph showing the AP-1 transcriptional activity of the indicated c-Fos alleles in Hela cells. AP-1 luciferase activity in cells expressing Fos-WT is expressed as 100%, and the data shown are from five individual experiments. FIGURE 7B is a photomicrograph showing the expression of endogenous c-Fos in quiescent (-) or serum-stimulated (+) pMV7-infected 208F fibroblasts (vector), assayed by immunofluorescence microscopy. To examine the

expression of Fos-WT, Fos or Fos G418-resistant 208F fibroblasts, cells were serum-starved for 24 h before fixation and processing for immunofluorescence microscopy using the anti-Fos antibody. FIGURE 7C is a Western blot of c-Fos protein expression in quiescent 208F cells. FIGURE 7D is a bar graph showing the anchorage-independent growth of G418-resistant pools of 208F cells stably expressing pMV7 (vector) or the indicated FLAG-Fos alleles.

The data are expressed as a percentage of the number of colonies formed by cells expressing Fos-WT (100%) and represent the mean SEM from six experiments performed in duplicate.

FIGURE 8 is a schematic diagram of the molecular interpretation of ERK1/2 signal duration. Growth factor stimulation (stimulus) causes activation of signaling pathways (signals) that result in rapid transcriptional induction of immediate early genes (response). The duration of ERK1/2 signaling is then interpreted by immediate early gene products that contain DEF domains (signal sensors). ERKl/2-docking to the DEF domain results in sensor phosphorylation.

Docking and phosphorylation alters its biological activity, and this dictates the biological outcome. TF, transcription factor.

FIGURE 9A is a photomicrograph showing the nuclear accumulation of active ERK1/2 and c-Fos in growth factor-treated Swiss 3T3 cells. These photomicrographs are enlargements of the images of Figure 1C in order to visualize the cellular distribution of activated ERK1/2 and nucleolar structures.

FIGURE 9B is a photomicrograph of quiescent Swiss 3T3 cells treated with PDGF for 75 minutes followed by the addition of cyclohexamide (+) or vehicle (-). Cells were processed for c-Fos immunofluorescence 90, 180 or 300 minutes after PDGF stimulation. In control experiments (bottom two panels), cells were incubated with cyclohexamide or vehicle for 20 minutes prior to treatment with PDGF for 90 minutes.

FIGURE 10A is a Western blot of NIH3T3 cells transiently transfected with FosWT or FosDD were left quiescent or treated with EGF (50 ng/ml, 5 min) prior to lysis. An aliquot from each cell extract was incubated in the presence or absence of k protein phosphatase (P'ase) for 30 minutes on ice. Data shown is representative of three separate experiments. FIGURE 1 OB is a Western blot of NIH3T3 cells stably expressing AB-Raf : ER that were transfected with FosWT, FosAA or FosDD. Cells were deprived of serum growth factors, pre-treated with 5 uM U0126 (+) or 0.1% DMSO (-) for 30 minutes prior to treating with tamoxifen (TAMX, 1 RM) for 15 minutes prior to cell lysis. FIGURE 10C is a Western blot showing the phosphorylation of (His) 6-FosWT, AA or EE or MBP by Flag-ERK5.

Active (+) and inactive ERK5 (-) was obtained by coexpressing Flag-ERK5 and HA-MEK5 (D) or control vector, respectively, in 293 cells followed by immunoprecipitation of Flag-ERK5 from cell lysates using the M2 anti-Flag monoclonal antibody. Autophosphorylation (auto-P) of ERK5 in kinase reactions is indicated. Together, these data demonstrate that the phosphorylation of primed c-Fos is regulated by the Raf/Mek/ERK pathway.

FIGURE 11 is a Western blot of quiescent Rat-1 cells that were treated with the indicated concentrations of LPA for various times. The activation kinetics of ERK1/2 demonstrates that c-Fos is a sensor for sustained ERK1/2 signaling in Rat- 1 fibroblasts. The data shown is representative of at least three individual experiments.

FIGURE 12 is a series of cell culture plates, fixed and then stained with Giemsa to visualize foci. The indicated Fos proteins were stably expressed in 208F cells and cultured for four weeks in regular culture medium. Identical data was obtained from five separate experiments. Thus, substituting aspartic acid for T235 and T331 in c-Fos promotes Fos-mediated transformation.

FIGURES 13A-C are a series of Western blots showing the regulation of ectopically expressed Fos family proteins (c-Fos, Fra-1, and Fra-2) by the ERK1/2 pathway in NIH 3T3 cells. Cells were treated with or without EGF (50 ng/mL) for 5 minutes prior to cell lysis. Where indicated, U0126 (5 mM) was added to cells 30 minutes before adding EGF. EGF treatment in the absence of U0126 activated ERK1/2, as demonstrated by the mobility shift to a higher molecular weight.

ERK1/2 activation resulted in phosphorylation of the target amino acid, T325, of c-Fos (Figure 13A). ERK1/2 activation of Fra-1 (Figure 13B) and Fra-2 (Figure 13C) is also demonstrated by the observed mobility shift.

FIGURE 14A is a sequence alignment of c-Fos, Fra-1, and Fra-2 (SEQ ID NO: 25-27, respectively) demonstrating a high degree of sequence identity in the C-termini. Fra-1 and Fra-2 have ERK1/2 and RSK priming phosphorylation sites in addition to DEF domains. FIGURE 14B is a Western blot of NIH 3T3 cells transfected with the indicated constructs and deprived of serum growth factors for 24 hours. These results demonstrate that mutations in the DEF domains of Fra-1 and Fra-2 inhibit the ERKl/2-mediated mobility shift (compare to Figures 13B and 13C).

FIGURE 15A is a Western blot of c-Myc immunoprecipitation from NIH 3T3 cells transfected with pcDNA3 (vector) or c-Myc and deprived of serum growth factors. EGF and U0126 were used to treat cells as described in Figure 13.

FIGURE 15B is a Western blot from cells transfected with the indicated c-Myc proteins. These results characterize the DEF domain in c-Myc and show that S62 phosphorylation depends on an intact DEF domain.

FIGURES 16A-F are Western blots demonstrating the kinetics of immediate early gene expression in Swiss 3T3 cells. Cells were deprived of serum growth factors and treated with EGF (25 ng/mL) or PDGF-BB (20 ng/mL) for the indicated times.

FIGURES 17A-B are Western blots demonstrating the kinetics of Egr-1, JunB, and c-Myc expression in Swiss 3T3 cells. Cells were treated as described in Figure 13. Total levels of c-Myc (Figure 17B) were detected by immunoprecipitating c-Myc prior to Western analysis.

FIGURES 18A-E are Western blots demonstrating that sustained expression of immediate early genes requires ERK1/2 activity. Serum deprived Swiss 3T3 cells were treated with PDGF-BB for 90 minutes before adding DMSO vehicle (0. 1%) orU0126 (5j M).

FIGURE 19A is a Western blot of cells treated with PDGF-BB for either 90 minutes (lanes 2-9) or 120 minutes (lanes 11-13) before adding DMSO vehicle (lanes 3-5) or U0126 (lanes 7-9,12, 13). FIGURE 19B is a Western blot of cells treated with PDGF-BB for 5 hours to induce Fra-1 before adding U0126 for a further 20 or 30 minutes. These figures demonstrate that ERK1/2 signaling is required during G1 for the stabilization of c-Myc.

FIGURE 20 is a series of Western blots from Swiss 3T3 cells treated with various concentrations of PDGF-BB before lysis. These results demonstrate that IEG products act as sensors for subtle differences in ERK1/2 signal duration.

FIGURE 21A is a bar graph of the result from an in vitro kinase assay demonstrating ERK 1/2 activation is sensitive to small differences in growth factor (PDGF) stimulation. FIGURE 21B is a Western blot demonstrating that the c-Fos stabilization observed in Figure 20 following stimulation with 10 ng/ml PDGF is a result of ERK 1/2-dependent phosphorylation of T325. Neither long-term c-Fos stabilization (see Figure 20) nor T325 phosphorylation is observed following 4 ng/ml PDGF stimulation.

FIGURE 22 is a series of Western blots showing Fra-1 hyperphosphorylation throughout G1 requires ERK1/2 signaling.

FIGURE 23 is representative gel and the densitometric quantification of an electrophoretic mobility shift assay (EMSA) for AP-1. Swiss 3T3 cells were treated as indicated and extracted in a hypotonic lysis buffer. The nuclear fraction was isolated and aliquots mixed with a 32P-labelled AP-1 oligonucleotide in a standard EMSA. These results demonstrate that PDGF, but not EGF, treatment significantly increases AP-1 expression and AP-1 DNA binding.

FIGURE 24 is an immunoprecipitation of extracts from 208F cells stably expressing c-Myc or c-Myc F196A and treated with cycloheximide (14 mg/mL) for the indicated times. Following immunoprecipitation of the c-Myc proteins, total levels of c-Myc were detected using Western analysis. These results demonstrate that c-Myc stability requires the DEF domain.

FIGURE 25 is a series of indirect immunofluorescence photomicrographs demonstrating typical results of the screening assays described herein.

Representative fields of view using a 10X objective lens are shown.

FIGURE 26 is a series of Western blots and accompanying densitometric analysis showing the effect of mutating the DEF domain binding site in ERK1/2 on a RSK (non-DEF domain-dependent) phosphorylation and c-Fos (DEF domain- dependent) phosphorylation. The bar graphs represent the raw densitometric analysis, unadjusted for ERK1/2 content.

Detailed Description We have discovered that DEF domains are present in numerous proteins that are important in a variety of human diseases and, by blocking the interaction of a MAP kinase with the DEF domain of a target protein, effective therapy may be provided. Also provided are screening methods for identifying novel therapeutics that inhibit the MAP kinase-DEF domain interaction. This invention provides several advantages over known therapies that directly target the MAP

kinase signaling cascade. Typically, most compounds that inhibit the MAP kinase pathway are non-specific and inhibit more than one enzyme. Also, the targeted kinases, if effectively inhibited, are not available to perform normal physiological functions necessary for cell survival, resulting in toxicity to healthy as well as diseased cells. By contrast, the therapeutic methods of the present invention inhibit the activation of particular target proteins, leaving the MAP kinases enzymatically active and available to phosphorylate other, non-DEF domain- containing proteins. Diseased cells (e. g. , cancerous cells) are often more susceptible to therapy because of the higher concentration of target protein, improving the likelihood of success for this approach.

The principles of the invention are exemplified using the immediate early gene, c-Fos, but is not intended to be limiting. c-Fos functions as a molecular sensor for the duration of extracellular-signal-regulated kinase 1/2 (ERK1/2) signaling. c-Fos is known to be phosphorylated by ERK1/2 and RSK, resulting in increased stability of the protein. Therefore, the biological function of c-Fos differs under conditions where ERK1/2 signaling is sustained, rather than transient. Signaling is transduced by ERK1/2 binding to the DEF domain of c-Fos.

Mutating the DEF domain inhibits c-Fos-mediated signaling and, ultimately, the downstream effects of ERK1/2 activation.

Further, Fos, Myc and Jun family proteins are transcription factors encoded by immediate early protooncogenes. Family members c-Fos, Fra-1, Fra-2, c-Myc, N-Myc, JunD, and JunB are frequently found to be amplified or upregulated in human cancers. Sustained ERK1/2 signaling is required for cell proliferation and ERK1/2 docking to these proteins occurs only when signaling is sustained.

Docking controls the growth-promoting function of these transcription factors.

Accordingly, ERK1/2 docking inhibitors may be clinically useful drugs because they will likely to inhibit a specific branch of ERK1/2 signaling and would, therefore, be less toxic than general ERK1/2 inhibitors.

Sustained ERK1/2 Activation Correlates With S Phase Entry Treatment of quiescent Swiss 3T3 fibroblasts with platelet-derived growth factor (PDGF) stimulated S phase entry; whereas, treatment with epidermal growth factor (EGF) did not (Figure 1A). The activation kinetics and amplitude of ERK1/2 and RSK, however, were almost identical following a 5-10 minute exposure to either PDGF or EGF (Figure 1B). In both cases, hyperphosphorylated (active) ERK1/2 was localized to the nucleus (Figure 1C). In contrast to ERK1/2 and RSK activation kinetics, rates of inactivation were faster in cells treated with EGF compared to PDGF (Figure 1B). ERK1/2 signaling remained elevated for at least 240 minutes following PDGF exposure, but returned to basal levels within 30-45 minutes following EGF withdrawal (Figure 1B). Notably, the sustained ERK1/2 activity elicited by PDGF treatment remained localized to the nucleus (Figures 1C and 9A), demonstrating a tight correlation with S phase entry.

Further, c-Fos protein expression was prolonged in cells treated with PDGF compared to those treated with EGF (Figure 1D). This indicates that c-Fos becomes stabilized when ERK1/2 signaling is prolonged, but is unstable when ERK1/2 signaling is transient. c-Fos expression was not affected by either the addition of cycloheximide to cells 75 minutes after PDGF treatment (Figure 9B) or the addition of actinomycin D 20 minutes after PDGF treatment. Thus, the differences in c-Fos expression between PDGF-and EGF-treated cells arises from post-translational control. This conclusion is further supported by studies showing that the transcriptional induction of c-fos and other IEGs by various growth factors is completed within 30-45 minutes.

ERK1/2 and RSK coordinately phosphorylate the extreme C-terminus of c- Fos at Ser 374 and Ser 362, respectively (Figure 2A). Mutating these residues to aspartate (Fos-DD), which mimics phosphorylation, results in enhanced transformation of fibroblasts by comparison to c-Fos having Ser 362 and Ser 374 mutated to alanine (Fos-AA) (Okazaki et al., EMBO J., 14: 5048-5059, 1995 ; Chen et al., Proc. Natl. Acad. Sci. USA, 90: 10952-10956,1993). Thus, increasing the stability of c-Fos is not the only manner in which this transcription factor can regulate cellular transformation. Fos-AA appears to be differentially regulated compared to Fos-DD.

We have discovered that the addition of serum to fibroblasts results in a large, k phosphatase-sensitive electrophoretic mobility shift of Fos-DD, compared to Fos-AA (Figures 2B left, and 10A). This effect correlates with increased incorporation of 32P-orthophosphate that was consistently two to threefold greater for Fos-DD than Fos-AA (Figure 2B, right). This demonstrates that phosphorylation of Ser 362 and Ser 374 prime c-Fos for additional growth factor- regulated phosphorylation. As Fos-DD has greater transforming potential than Fos-AA, the regulation of primed c-Fos is critical for promoting fibroblast proliferation.

Phosphorylation of Primed c-Fos is MEK-dependent NIH 3T3 cells transfected with different Fos proteins were treated with the MEK inhibitor U0126 (Favata et al., J. Biol. Chem., 273: 18623-18632,1998) to determine if the mitogen-regulated phosphorylation of Fos-DD is mediated by the Raf/MEK/MAPK pathway. UO 126 inhibited the growth factor-regulated mobility shift of Fos-WT and Fos-DD (Figure 2C) and ERK1/2 activation (Figure 2C, bottom), indicated that ERK1/2 or downstream signaling molecules regulated primed c-Fos. Identical observations were made using NIH 3T3 cells expressing a

conditionally active form of B-Raf (AB-Raf-ER) and treating these cells with tamoxifen instead of EGF (Figure 10B). To determine whether ERK1/2 could phosphorylate primed c-Fos in vitro, we used different hexahistidine-Fos fusion proteins as substrates. ERK1/2 efficiently phosphorylated Fos-WT (Figure 2D).

The phosphorylation of Fos-AA and primed c-Fos, Fos-EE (S362E/S374E), by ERK1/2 was also easily detected in vitro, but the phosphorylation of Fos-EE compared with Fos-AA was consistently greater (Figure 2d). These results demonstrate that ERK1/2 can phosphorylate sites in c-Fos other than Ser 374 and that this phosphorylation is enhanced after priming of the C terminus. In contrast to ERK1/2, phosphorylation of c-Fos by ERK5 in vitro, another U0126-sensitive proline-directed kinase, was not observed (Figure 10C).

An ERK1/2 Targeting Motif Promotes the Phosphorylation of Primed Fos The preference of ERK1/2 for primed c-Fos (Fos-EE) over Fos-AA demonstrates that C-terminal phosphorylation exposes additional phosphorylation sites and/or a motif that would increase the efficiency of phosphorylation at these sites. Examination of the c-Fos sequence identified a site in the C terminus that has similarity with the ERK1/2 targeting motif, FXFP (SEQ ID NO: 1), known as a DEF domain. In c-Fos, this motif is FTYP (Figure 3A; SEQ ID NO: 2).

Mutating either Phe 343 or Tyr 345 to alanine dramatically inhibited the phosphorylation of primed c-Fos (Fos-EE) by ERK1/2 in vitro (Figure 3A).

ERKl/2-regulated phosphorylation of substrates that contain DEF domains can be competitively inhibited in vitro with a synthetic peptide encompassing the DEF domain found in ELK-1. This peptide (FQFP; SEQ ID NO: 3) inhibited the phosphorylation of primed c-Fos in a concentration-dependent manner (Figure 3B). By contrast, a peptide with a mutant DEF domain (AQAP; SEQ ID NO: 4) was less efficient in inhibiting ERKl/2-mediåted phosphorylation of primed c-Fos.

The ELK-1 peptide was then engineered to contain the c-Fos FTYP DEF domain (SEQ ID NO: 2). This peptide also inhibited primed c-Fos phosphorylation, but a mutant version (ATYP; SEQ ID NO: 5) did not (Figure 3C). In both cases, the ICSO for the FQFP (SEQ ID NO: 3) and FTYP (SEQ ID NO: 2) peptides was approximately 80 pM. The EGF-stimulated mobility shift of Fos-DD in vivo was also inhibited when Phe 343 or Tyr 345 were mutated to alanine (Figure 3D).

These results demonstrate that the initial phosphorylation of c-Fos by ERK1/2 and RSK as the extreme C terminus expose an ERK1/2 docking site that allows ERK1/2 to phosphorylate additional sites.

Based on this model, mutation of Phe 343 or Tyr 345 to alanine should prevent hyperphosphorylation of Fos-WT and not interfere with the priming phosphorylations, which are C-terminal to the DEF domain. Indeed, these mutations prevented the appearance of the slowest mobility, but still allowed a shift to the intermediate mobility (Figure 3E). This indicates that the DEF domain is not involved in directing ERK1/2 to prime c-Fos through Ser 374 phosphorylation. Instead, ERK1/2 docking through the DEF domain results in the hyperphosphorylation of primed c-Fos.

ERK1/2 phosphorylates Thr 325 and Thr 331 in primed c-Fos There are two proline-directed threonine residues (Thr 325 and Thr 331) amino-terminal to the DEF domain in c-Fos (Figure 4A). Mutation of Thr 325 to alanine almost completely inhibited the phosphorylation of Fos-EE by ERK1/2 in vitro, and the additional mutation of Thr 331 to alanine was required to reduce the phosphorylation to background levels (Figure 4A). Thr 325 and Thr 331 were also i phosphorylated in primed c-Fos (Fos-DD) in vivo, as evidenced by the complete loss of the mobility shift in the T325A/T331A mutant (Figure 4B). Individual mutation of Thr 325 or Thr 331 to alanine in the context of Fos-DD only partially

inhibited growth factor-regulated phosphorylation. Substituting alanines for Thr 325 and Thr 331 in the context of Fos-WT prevented the EGF-stimulated to the slowest mobility (Figure 4C). However, EGF treatment was associated with the appearance of the intermediate mobility form, resulting from priming phosphorylation of ERK1/2 and RSK. Collectively, these observation demonstrate an ordered phosphorylation process whereby the initial phosphorylation of c-Fos at Ser 374 and Ser 362 (priming) exposes a DEF domain that mediates the hyperphosphorylation of c-Fos at Thr 325 and Thr 331. Further, Ser 374 phosphorylation is not regulated by docking. This is consistent with the phosphorylation of amino acids N-terminal, but not C-terminal, to DEF domains.

A phosphorylation-specific antiserum for Thr 325 in c-Fos (Figure 5A) was generated to investigate the mitogen-regulated phosphorylation of this residue in primed c-Fos. The antiserum showed little or no reactivity with Fos-WT or Fos- DD expressed in quiescent cells (Figure 5B,-EGF). However, after treatment with EGF, strong reactivity was associated with Fos-WT and Fos-DD, but not with FosT325A or Fos-DDT325A (Figure 5B, +EGF). Priming of the extreme C terminus by ERK1/2 and RSK promotes additional phosphorylation of c-Fos in vivo (Figure 2B). To determine if this is caused by increased phosphorylation of Thr 325, Fos- WT and Fos-AA were expressed to similar levels in the AB-Raf-ER NIH 3T3 cells that were then treated with tamoxifen for varying times (Figure 5C). The phosphorylation of Thr 325 was greater in cells transfected with Fos-WT than those transfected with Fos-AA.

Mutating Phe 343 or Tyr 345 to alanine prevented the hyperphosphorylation of primed c-Fos (Figure 3). Specifically, the regulation of Thr 325 phosphorylation in vivo was inhibited when Phe 343 or Tyr 345 were mutated to alanine, either in the context of Fos-WT (Figure 5D) or Fos-DD (Figure 5E).

In this later experiment, there is a strong correlation between the mobility shift of Fos-DD and increased Thr 325 phosphorylation confirming that the DEF domain in c-Fos increases the efficiency of Thr 325 phosphorylation in vivo.

The phosphorylation of Thr 325 is differentially regulated by ERKl/2-signal duration As shown above, the induction kinetics of c-Fos expression 30-45 min after addition of PDGF or EGF to Swiss 3T3 cells were identical (Figure 1D). This is consistent with a model in which an initial activation of ERK1/2 or RSK is sufficient for induction of c-fos IEG expression. To determine if ERK1/2 signal duration differentially regulates the phosphorylation of Thr 325 in endogenous c- Fos, we prepared extracts from Swiss 3T3 cells treated with EGF or PDGF for different times. Importantly, although c-Fos is present in cells after 45 or 60 min of EGF treatment, Thr 325 phosphorylation was not observed (Figure 6A). This is consistent with inactivation of ERK1/2 occurring before c-Fos is present (Figure 6A). By contrast, phosphorylation of Thr 325 increased 45-60 min after addition of PDGF (Figure 6B). Maximal Thr 325 phosphorylation persisted for at least 120 min (Figure 6B), but was still detected after 240 min. In Rat-1 fibroblasts, treatment with 100 p. M lysophosphatidic acid (LPA) results in sustained ERK1/2 activity and S phase entry, whereas treatment with 0.1-1 FM LPA transiently activates ERK1/2 and no cell cycle progression occurrs. Although treatment of quiescent Rat-1 fibroblasts with mitogenic (100 uM) and no-mitogenic (0.5 pM) concentrations of LPA resulted in a similar induction of c-fos, phosphorylation of Thr 325 only occurred with 100 uM LPA (Figure 11). These findings correlate with the generation of transient and sustained ERK1/2 responses by 0. 5 uM and 100 uM LPA, respectively (Figure 11).

Thus, differential phosphorylation of c-Fos occurs in different cell types and in response to agonists that directly activate tyrosine kinase receptors or heterotrimeric G protein-coupled receptors.

To show that the sustained phase of ERK1/2 signaling was required to mediate the stabilization and hyperphosphorylation of endogenous c-Fos in Swiss 3T3 cells, ERK1/2 and RSK activity was inhibited by adding U0126 to cells that had been treated with PDGF for 60 min (Figure 6D). Under this condition, the phosphorylation of Thr 325 was completely inhibited and the electrophoretic mobility of c-Fos increased (Figure 6C). Importantly, these U0126-induced changes in the biochemical properties of c-Fos also preceded the rapid disappearance of c-Fos protein. This result is consistent with hypophosphorylated c-Fos being unstable. These observation show that the phosphorylation of c-Fos at Thr 325 is tightly correlated with the activation/inactivation kinetics of ERK1/2 in different cell types of provide clear evidence that c-Fos can function as sensor for ERK1/2 signal duration.

The DEF domain and Thr 325/Thr 331 phosphorylation modulates c-Fos function In Hela cells expressing Fos-WT, AP-1 transcription factor activity was consistently three to fourfold above background levels (Figure 7A). Mutation of Thr 325 and Thr 331 to alanine reduced AP-1 activity by about 20%; whereas, mutating the DEF domain (F343A) reduced Fos-WT activity by about 65% (Figure 7A). These observations indicate that docking of ERK1/2 to c-Fos is important in regulating c-Fos transcriptional activity under conditions of growth factor stimulation. To determine if ERK1/2 docking to c-Fos can contribute to c-Fos function independently of the phosphorylation-mediated stabilization, Fos-WT, FoST325A/T331A or FOSY345A (a DEF domain mutant) were stably expressed in 208F

fibroblasts. The expression of the different Fos proteins (Figure 7B, bottom six panels) was equivalent to the level of endogenous c-Fos in serum-stimulated vector-infected cells (Figure 7B, top two panels) and was also localized to the nucleus. Western analysis of c-Fos expression in the quiescent cell lines also showed that they were expressed to similar levels (Figure 7C). The stable expression of Fos-WT promoted anchorage-independent growth in soft agar (Figure 7D), as expected. Mutating Thr 325 and Thr 331 to alanine significantly reduced the growth of 208F cells in soft agar suggesting that phosphorylation of these residues promotes cellular transformation (Figure 7D). However, replacing Thr 325 and Thr 331 with Asp enhanced c-Fos-mediated focus formation (Figure 12). Further, mutating the c-Fos DEF domain (FosY345A) completely inhibited the ability of c-Fos to transform 208F cells; more so than the c-FosT325A/T33lA mutant (Figure 7D). These results demonstrate that ERK1/2 docking to c-Fos contributes to transformation through mechanisms in addition to Thr 325/Thr 331 phosphorylation and that stabilization of c-Fos is not the only factor that regulates c-Fos function, as all proteins were expressed equally. In addition, it also demonstrates that ERK1/2 docking to c-Fos regulates biological activity.

Mutating the DEF domain binding site inhibits target residue phosphorylation In order to confirm the criticality of ERK1/2 docking with the DEF domain to phosphorylation of a target residue, the DEF domain binding site in ERK1/2 was mutated. Six single mutations in ERK1/2 (L198A, Y231A, L232A, L235A, Y261A, and D319N) were expressed in NIH 3T3. Several of these mutations have been previously described as forming the DEF domain binding site in ERK1/2 (Lee et al., Molec. Cell, 14: 43-55,2004). Each ERK1/2 mutant was tagged with HA for later detection. EGF-stimulated kinase activity of wildtype and mutant

ERK1/2 was measured using RSK, a non-DEF domain-containing target protein, and c-Fos, DEF domain-containing target protein, as substrates in a standard 32p_ phosphorylation assay. Fos phosphorylation by ERK1/2 mutants with disrupted DEF domain binding pockets (L198A, Y231A, L232A, L235A, and Y261A) was almost completely absent (Figure 26). By contrast, RSK phosphorylation-a event that does not require ERK1/2 interaction with a DEF domain-was only moderately reduced (Figure 26).

These results demonstrate that DEF domain binding is required for a target residue phosphorylation but that a disruption of the DEF domain inhibition does not abolish all kinase activity. Thus, the DEF domain binding event is separate and distinct from the phosphorylation event. Disruption of DEF domain binding may be used to selectively inhibit phosphorylation of a target protein, without significantly inhibiting the phosphorylation of non-target proteins (i. e. , proteins that do not contain a DEF domain) by the same kinase. Thus, small molecule inhibitors and polypeptide inhibitors (e. g. , naked DEF domains) which specifically inhibit DEF domain binding are useful for selectively inhibiting the phosphorylation of target proteins without causing the adverse effects associated with complete inhibition of a target kinase (e. g. , a MAP kinase).

Mechanism of IEG Activation Through DEF Domain Binding The mechanism described here employs an IEG product, typified by c-Fos, which functions as a molecular sensor that differentiates between differences in ERK1/2 and RSK signal duration, as well as their cytoplasmic/nuclear distribution (Figure 8). As observed for a large number of IEGs, the c-fos gene is transcriptionally induced within minutes of growth factor stimulation and therefore occurs with kinetics that are independent of differences in signal duration. Newly synthesized c-Fos protein has a half-life of about 30-45 min but, when

phosphorylated by ERK1/2 and RSK, the half-life is extended to at least 2h. Thus, when ERK1/2 is rapidly activated (transient signal), c-Fos is present in the nucleus, but is not phosphorylated, and is therefore unstable and degraded (Figure 8). By contrast, delayed inactivation of ERK1/2 (sustained signal) results in the efficient phosphorylation of c-Fos at its extreme C terminus, resulting in its stabilization for several hours. The initial priming phosphorylation in the C- terminus exposes a DEF domain that promotes additional ERKl/2-mediated phosphorylation events, increasing the efficiency of ERK1/2-regulated phosphorylation when ERK1/2 is only sub-maximally active (0.5-4 h after stimulation). Further, when priming and docking are inhibited by point mutation (Fos-AA and FosF343A or FoSY345A, respectively) ERK1/2 and/or RSK signals are unable to alter c-Fos function. These non-phosphorylable c-Fos mutants likely resemble the hypophosphorylated form of c-Fos that is present when ERK1/2 is rapidly inactivated during transient signaling and cells do not enter S phase.

Simply prolonging the half-life of c-Fos will not affect its role in promoting transformation. Instead, the combination of protein stabilization and DEF- mediated regulation allows c-Fos to function as sensor for ERK1/2. If c-Fos is not stabilized during the sustained phase of signaling, ERK1/2 will not target the c-Fos DEF domain. Therefore, stabilizing the IEG product is a critical first step if it is to function as a sensor for sustained ERK1/2 signals. The physiological importance of the c-Fos DEF domain is underscored by the fact that mutations in the DEF domain significantly reduced AP-1 activity and inhibit the transforming activity associated with wild-type c-Fos. However, the effect of mutating the DEF domain is stronger than the effect of mutating the phosphorylation sites that are controlled by this docking site, indicating that the DEF domain can have more than one action. An additional function of the DEF domain ERKl/2-mediated trans- phosphorylation of AP-1 complex proteins.

We described a general mechanism for cellular sensing of ERK1/2 signal strength and timing involving the FTYP (SEQ ID NO: 2) DEF domain present in many IEGs. Putative DEF domains are found in additional AP-1 proteins, such as Fra-l, Fra-2, Jun-B and JunD (Table 1). The proto-oncogene products c-Myc and N-Myc also contain putative DEF domains. The IEG product Egr-1 has a DEF domain and several putative proline-directed phosphorylation sites N-terminal to this domain that could enable Egr-1 to sense sustained signaling in PC12 cells and promote neuronal differentiation. In common with the c-Fos DEF domain, the other DEF domains highlighted in Table 1 show subtle deviation form the FXFP consensus (SEQ ID NO: 1), with respect to the presence of phenylalanine at positions 1 and 3 indicating that tyrosine can be tolerated at either site.

Table 1. DEF Domains in Immediate Early Gene Products __ IEG Amino Acid Sequence ID NO. c-Fos Fra-1 Fra-2'69-GGFYGE-EPLHTP-IVVTSTPAITPGTSNLVFTYPSVLEQ Fos-B JunD JunB 11 c-Jun 56-LPTPPLSPSRRSGLCSPSYV c-Myc '-LTA-AASECIDPSWFPYPLND 14 77-AQSPGAGAASPAGRGHGGAAGA N-Myc 1 16 _ SEQ184_QSPPLSCAVPSNDSSPIYSAAPTFPTPNTD 17 Egr-1 247-PMIPDYLFPQQ 18 MPer1 697-PRGGPQPLPPAPTSVPPAAFPAPLVTPMVALPNYLFPTPSSY 19 DEF domains are in bold and number indicate amino acid position. Sequences are from rat (c-Fos, Fra-1, and Fra-2), mouse (FosB, JunD, c-Jun, c-Myc, and Egr-1), or human (JunB, N-Myc, and mPerl).

Screening Methods to Identify Inhibitors of DEF Domain Binding DEF Domain Binding Assessment using a Phospho-specific Antibody We have developed a cell-based assay which is used to screen small molecule compound libraries (Figure 25). In this assay, rat 208F fibroblasts that stably express c-Fos are cultured in a 384-well plate and deprived of serum growth factors for 24 hours. Cells are then treated with EGF for 15 minutes and fixed with 3.7% formaldehyde. Permeabilized fixed cells are incubated with DAPI to stain the nuclei and an anti-phospho-ERKl/2 mouse monoclonal antibody and an anti-phospho-T325 Fos rabbit polyclonal antibody for 2 hours. Anti-mouse Alexa594-conjugated IgG and an anti-rabbit FITC-conjugated IgG are added to each well and unbound antibody is removed by several washes. The fluorescence intensity of both fluorophores in each well can be detected using an automated epifluorescence microscope or Autoscope (Universal Imaging Systems, Inc. ). A clear increase in T325 phosphorylation was observed in cells treated with EGF (indicating that ERK1/2 docking to Fos has taken place). In the same population of cells, the phosphorylation of ERK1/2 also increased, thus indicating its activation by EGF. Under these conditions, only background levels of fluorescence were detected when both phospho-specific antibodies were omitted, and the secondary antibodies show no cross-species reactivity.

Inhibition of ERK 1/2 docking to the c-Fos DEF domain in vivo could result from compounds that (a) are generally toxic, (b) prevent the activation of ERK1/2, (c) prevent the growth factor regulated translocation of ERK1/2 into the nucleus where c-Fos is localized or (d) directly antagonize ERK1/2 docking to the DEF domain. The assay we have developed naturally excludes the first three possibilities. First, toxicity will be reflected by nuclear integrity as visualized with DAPI staining. Second, inhibition of ERK1/2 activation/activity will be apparent from the phospho-ERKl/2 fluorescence signal. Third, nuclear translocation of ERK1/2 can be verified by manually examining images from wells that show decreased c-Fos phosphorylation. Therefore, candidate compounds that specifically inhibit ERK1/2 binding to the DEF domain are defined as compounds that decrease the phosphorylation of T325 in c-Fos but which have no effect on ERK1/2 activation or its localization. Although this assay is exemplified using rat fibroblasts, it may be performed using any appropriate cell type including, for example, myoblasts, epithelial cells, and hepatocytes.

Interaction-Trap Assays A standard yeast two-hybrid assay may be used to assess the effect of a test compound on the MAP kinase-DEF domain interaction (Mendelsohn and Brent, Curr. Opin. Biotechnol. 5: 482-486,1994). Typically, a vector encoding a synthetic or naturally occurring peptide containing a DEF domain, covalently bound a DNA binding domain (e. g. , GAL4), is transfected into yeast cells containing a reporter gene operably linked to a binding site for the DNA binding domain. Further, a vector encoding either the native MAP kinase of interest, or a synthetic fragment containing the sequence that interacts with the target DEF domain, covalently bound to a transcriptional activator (e. g., GalAD) is also transfected.

The effectiveness of a test compound is then assessed by growing the yeast in the presence of the compound and measuring the level of reporter gene expression.

GSTPulldown Assays The interaction of a MAP kinase with a DEF domain may be examined using a GST-fusion protein binding study. A vector encoding a naturally- occurring or synthetic polypeptide containing the DEF domain of interest is fused to GST and expressed in a host cell (e. g., E. coli or Saccharomyces spp.). The GST fusion protein is then contacted with a MAP kinase in the presence and absence of a test compound. The MAP kinase may be naturally expressed by the host cell or may be expressed from a second vector inserted into the host cell.

Following incubation with the test compound, the host cells are lysed and the GST fusion proteins are recovered using glutathione-Sepharose (GSH-Seph) beads.

Typically, the GST fusion proteins are released from the GSH-Seph by boiling and the proteins visualized by electrophoretic separation on an SDS-PAGE gel. A skilled artisan will readily understand that the GST-Pulldown assay described here can be readily adapted to a cell-free assay by incubating the purified GST fusion protein with a purified recombinant MAP kinase.

Fluorescence Polarization Assay A variety of well known cell-free techniques may be used to assess the effects of a test compound on the interaction between a MAP kinase and a DEF domain-containing target protein. Fluorescence polarization assays are particularly useful for this purpose. In this assay, a peptide (about 6-12 amino acids) containing a DEF domain at its C-terminus and a fluorophore (e. g. , fluorescein) conjugated to its N-terminus is incubated in the presence and absence of increasing amounts of recombinant MAP kinase (e. g., GST-ERK1 ; 0.01-1 uM) for 10

minutes at room temperature. Aliquots from each reaction are placed in a plate black-walled microtiter (e. g., 384-well) plate and polarization measured using an Analyst plate reader. Increasing concentrations of the MAP kinase causes an increase in polarization. Titrating in the"free"DEF domain-containing peptide (i. e. , un-conjugated) inhibits the change in polarization, whereas the mutated DEF domain peptide does not. The appearance of low polarization, even in the presence of high concentrations of kinase, indicates flexible binding of the DEF domain to the kinase and suggests the presence of the propeller effect. Designing shorter dye-conjugated DEF domain-containing peptides usually alleviates this problem. The effect of standard assay variables, including incubation time, temperature, pH (7.2-8. 5), and buffers, on polarization is readily controlled during routine assay optimization.

This assay is readily adaptable for identifying test compounds that inhibit binding of a MAP kinase to a DEF domain. The use of automated liquid handling systems and plate readers makes this assay readily adaptable to a high-throughput format for screening large numbers of test compounds. For compound screening, the test compound is added to a mixture of the fluorescently labeled DEF domain- containing peptide and the target MAP kinase. Compounds that inhibit the polarization increase (or cause a decrease in polarization) resulting from increasing amounts of the MAP kinase are therapeutic candidates.

Identification of Test Compounds as Potential Therapeutics We have identified a variety of DEF domain-containing target proteins that have been implicated in a variety of diseases. The particular DEF domain or target protein may be substituted for c-Fos in any of the exemplary assays described here.

Further, the lists of target proteins provided are not intended to be limiting. Other target proteins are easily identified based on the availability of a DEF domain.

Test compounds having antineoplastic activity are those that inhibit binding of a MAP kinase (e. g. , ERK 1/2) to the DEF domain of any of the proteins of Table 1 (except mPerl) or Table 2. Test compounds that are useful for treating cardiovascular disorders inhibit MAP kinase binding to the DEF domain of the proteins identified in Table 3. Test compounds that are useful for treating acute and chronic inflammation or inflammatory disorders inhibit MAP kinase binding to the DEF domain of the proteins identified in Table 4. Test compounds that are useful for treating a variety of metabolic disorders inhibit MAP kinase binding to the DEF domain of the proteins identified in Table 5. Test compounds that are useful for treating a variety of nervous system disorders (e. g. , central and peripheral neuropathies) and behavioral disorders (e. g. , psychosis, schizophrenia, autism, Down's Syndrome, Parkinson's Disease, Alzheimer's Disease, epilepsy, Cockayne syndrome, depression, and opiate addiction) inhibit MAP kinase binding to the DEF domain of the proteins identified in Table 6. Test compounds useful for treating sleep disorders inhibit MAP kinase binding to the DEF domain of the Per proteins (Table 1: mPer ; Table 7). Other potentially useful therapeutics inhibit MAP kinase binding to the DEF domain of the PKA-anchoring proteins (AKAPs) (Table 7). In each of Tables 2-7, the alpha-numeric Accession Codes refer to the SWISS-PROT accession numbers. The numeric Accession Codes refer to the GENPEPT accession numbers. In each case, the DEF domain is underscored.

Administration of a DEF Domain Inhibitors or Candidate Compounds for the Treatment of Disease As described above, ERK1/2 activate several IEG products through an interaction with the DEF domain and a subsequent phosphorylation event. It is also well known that activation of certain IEGs, and the proteins identified in

Table 2, cause cellular proliferation and may cause tumor promotion and progression. Accordingly, this invention also provides methods and compositions for antineoplastic (i. e. , cancer) therapy by administering DEF domain inhibitors.

Likewise, therapy for cardiovascular disorders, inflammatory disorders, metabolic disorders, neuropathies and behavioral disorders, and sleep disorders may be provided by inhibiting MAP kinase binding to the DEF domain of one or more of the proteins identified in Table 3,4, 5,6, and 7, respectively. Useful DEF domain inhibitors include compounds that bind to the DEF domain of target proteins and prevent the binding of the target kinases. Also, DEF domain inhibitors include "bait"proteins that bind activated target kinases but do not cause cellular proliferation or tumor promotion and/or progression.

In addition to candidate compounds identified using the screening methods of this invention, DEF domain inhibitors can be created by inserting, by artifice, a DEF domain into a non-target protein. The cellular activation/proliferation pathway described herein is limited by the presence of activated target kinase, not by the availability of target proteins. Thus, a DEF domain that is present in a non- target protein effectively"baits"the target kinase, reducing its availability to phosphorylate the target proteins. DEF domains suitable for therapy have the general structure: F/Y-Xl-F/Y-X2 (SEQ ID NO : 28). Desirably, Xa is proline. Most desirably, the DEF domain is identical to the DEF domain of the target protein to which therapy is directed. For example, Figures 3B and C demonstrate that the"naked"DEF domains FQFP (SEQ ID NO: 3) and FTYP (SEQ ID NO: 2) are effective inhibitors of target protein phosphorylation.

Substitution of phenylalanine for alanine in these polypeptides results in approximately a two-fold reduction in potency.

Accordingly, therapy can be provided by administering pharmaceutical formulations containing a naked DEF domain. Typically, these polypeptides are administered by parenteral injection such as intravenous, intramuscular, or subcutaneous injection. These small polypeptides may be administered in any appropriate formulation including, for example, in a liposomal formulation. The polypeptides may also be injected directly into a solid tumor.

Alternatively, therapy can be achieved by administering a chimeric protein consisting of a DEF domain that is engineered into a non-target protein. Typically, the chimeric protein will"display"the four amino acid DEF domain on a hydrophilic face, making it available to bind to the activated target kinase. The non-target protein can be chosen based upon the desired pharmacokinetic or pharmacodynamic effect and is readily determined by a person of ordinary skill.

For example, a DEF domain inhibitor sequence may be engineered into a serum protein such as albumin or ceruloplasmin in order to prolong the plasma half life.

Alternatively, the DEF domain may be engineered into a protein that promotes uptake into a particular cell type.

Phataaceutical Formulations The peptide agents and candidate compounds of the invention can be administered to a subject, e. g. , a human, directly or in combination with any pharmaceutically acceptable carrier or salt known in the art. Pharmaceutically acceptable salts may include non-toxic acid addition salts or metal complexes that are commonly used in the pharmaceutical industry. Examples of acid addition salts include organic acids such as acetic, lactic, pamoic, maleic, citric, malic, ascorbic, succinic, benzoic, palmitic, suberic, salicylic, tartaric, methanesulfonic, toluenesulfonic, or trifluoroacetic acids or the like; polymeric acids such as tannic acid, carboxymethyl cellulose, or the like; and inorganic acids such as hydrochloric

acid, hydrobromic acid, sulfuric acid phosphoric acid, or the like. Metal complexes include zinc, iron, and the like. One exemplary pharmaceutically acceptable carrier is physiological saline. Other physiologically acceptable carriers and their formulations are known to one skilled in the art and described, for example, in Remington's Pharmaceutical Sciences, (19th edition), ed. A. Gennaro, 1995, Mack Publishing Company, Easton, PA.

Pharmaceutical formulations of a therapeutically effective amount of a peptide agent or candidate compound of the invention, or pharmaceutically acceptable salt-thereof, can be administered orally, parenterally (e. g. intramuscular, intraperitoneal, intravenous, or subcutaneous injection), or by intrathecal or intracerebroventricular injection in an admixture with a pharmaceutically acceptable carrier adapted for the route of administration.

Methods well known in the art for making formulations are found, for example, in Remington's Pharmaceutical Sciences (19th edition), ed. A. Gennaro, 1995, Mack Publishing Company, Easton, PA. Compositions intended for oral use may be prepared in solid or liquid forms according to any method known to the art for the manufacture of pharmaceutical compositions. The compositions may optionally contain sweetening, flavoring, coloring, perfuming, and/or preserving agents in order to provide a more palatable preparation. Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid forms, the active compound is admixed with at least one inert pharmaceutically acceptable carrier or excipient. These may include, for example, inert diluents, such as calcium carbonate, sodium carbonate, lactose, sucrose, starch, calcium phosphate, sodium phosphate, or kaolin. Binding agents, buffering agents, and/or lubricating agents (e. g. , magnesium stearate) may also be used.

Tablets and pills can additionally be prepared with enteric coatings.

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and soft gelatin capsules.

These forms contain inert diluents commonly used in the art, such as water or an oil medium. Besides such inert diluents, compositions can also include adjuvants, such as wetting agents, emulsifying agents, and suspending agents.

Formulations for parenteral administration (i. e., intravenous, intramuscular, and subcutaneous injection) include sterile aqueous or non-aqueous solutions, suspensions, or emulsions. Examples of suitable vehicles include propylene glycol, polyethylene glycol, vegetable oils, gelatin, hydrogenated naphalenes, and injectable organic esters, such as ethyl oleate. Such formulations may also contain adjuvants, such as preserving, wetting, emulsifying, and dispersing agents.

Biocompatible, biodegradable lactide polymer, lactide/glycolide copolymer, or polyoxyethylene-polyoxypropylene copolymers may be used to control the release of the compounds. Other potentially useful parenteral delivery systems for the proteins of the invention include ethylene-vinyl acetate copolymer particles, osmotic pumps, implantable infusion systems, and liposomes.

Liquid formulations can be sterilized by, for example, filtration through a bacteria-retaining filter, by incorporating sterilizing agents into the compositions, or by irradiating or heating the compositions. Alternatively, they can also be manufactured in the form of sterile, solid compositions which can be dissolved in sterile water or some other sterile injectable medium immediately before use.

The amount of active ingredient in the compositions of the invention can be varied. One skilled in the art will appreciate that the exact individual dosages may be adjusted somewhat depending upon a variety of factors, including the protein being administered, the time of administration, the route of administration, the nature of the formulation, the rate of excretion, the nature of the subject's conditions, and the age, weight, health, and gender of the patient. Generally,

dosage levels of between 0.1 Fg/kg to 100 mg/kg of body weight are administered daily as a single dose or divided into multiple doses. Desirably, the general dosage range is between 250 Fg/kg to 5.0 mg/kg of body weight per day. Wide variations in the needed dosage are to be expected in view of the differing efficiencies of the various routes of administration. For instance, oral administration generally would be expected to require higher dosage levels than administration by intravenous injection. Variations in these dosage levels can be adjusted using standard empirical routines for optimization, which are well known in the art. In general, the precise therapeutically effective dosage will be determined by the attending physician in consideration of the above identified factors.

The protein or candidate compound of the invention can be administered in a sustained release composition, such as those described in, for example, U. S.

Patent No. 5,672, 659 and U. S. Patent No. 5,595, 760. The use of immediate or sustained release compositions depends on the type of condition being treated and the desired pharmacokinetic profile. For preventive or long-term treatments, a sustained released composition may be preferred.

The protein or candidate compound of the present invention can be prepared in any suitable manner. The protein or candidate compound can be isolated from naturally occurring sources, recombinantly produced, or produced synthetically, or produced by a combination of these methods. The synthesis of short peptides is well known in the art. See e. g. Stewart et al. , Solid Phase Peptide Synthesis (Pierce Chemical Co. , 2d ed. , 1984).

Methods Cell Culture NIH 3T3 fibroblasts were transfected with Lipofectamine (Invitrogen, Carlsbad, CA) and then cultured for 18 h in DMEM/10% calf serum. Swiss 3T3 fibroblasts expressing a conditionally active form of B-Raf, were cultured in DMEM containing 10% fetal bovine serum (FBS). Before stimulation, cells were cultured in DMEM containing 20 mM HEPES (starving medium) for 24 h (NIH 3T3 or 208F) or 48 h (Swiss 3T3). Rat-1 fibroblasts were cultured for 48 h, washed with starving medium and culture for an additional 24 h in starving medium. For AP-1 assays, Hela cells were transfected with Lipofectamine for 6 h and then cultured for an additional 16 h before cell lysis and assay of luciferase activity (Promega). EGF and PDGF (Invitrogen) were reconstituted in sterile water containing 0.1% BSA. LPA (Aventi Polar Lipids, Alabaster, AL) was reconstituted in 50% ethanol before sonication for 30 min.

Retroviruses used to infect rat 208F cells were produced as described previously (Chen etal., Oncogene, 12: 1493-1502,1996). Neomycin-resistant pools of c-Fos-expressing cells were assayed for anchorage-independent growth or focus formation. Metabolic labeling with 35S-methionine or 32P-orthophosphate (performed in parallel) was performed as described by Chen et al.

Cell lysis and western analysis Cell extracts were prepared as described previously (Richards, et al., Curr.

Biol., 9: 810-820,1999). To analyze shifts in c-Fos mobility, samples were resolved on a 7.5% SDS-polyacrylamide gel electrophoresis (PAGE) gel, transferred to nitrocellulose and probed with anti-c-Fos antibody (Update Biotechnology Inc., Lake Placid, NY). This antibody is specific for c-Fos and does not cross-react with FosB, Fra-1 or Fra-2. For ERK1/2-MAPK western

analysis, a polyclonal anti-ERKl/2 antibody to an anti-phospho-p42/p44 MAPK monoclonal antibody was used (Sigma, St. Louis, MO). Phosphatase treatment of cell extract was performed for 30 min on ice using X protein phosphatase (New England Biolabs, Beverly, MA). To generate antiserum specific for phosphorylated Thr 325 in c-Fos, residues 317-329 (VTELEPLCTPWT) (SEQ ID NO: 20) were synthesized (underlined residue is phospho-Thr at position 325), conjugated to keyhole limpet haemacyanin and injected into rabbits (Research Genetics, Inc. , Huntsville, AL). To determine the specificity of the antiserum, extracts from cells expressing vector or c-Fos proteins were immobilized on nitrocellulose and probed with a solution of this anti-serum (1: 3000) for 12 h at 4 OC.

Recombinant protein purification M15pREP4 cells transformed with pDS56- (His) 6Fos or pETHis6/ERK2 and MEK 1 R4F were cultured at 25 C until an OD600 of 0.7 was attained. Cells were then incubated in the presence of 1mM isopropyl-p-D-thiogalactoside (IPTG) for an additional 12 h at 25 ° C. and then harvested by centrifugation. Pellets were resuspended in column buffer (20 mM Tris-HCl at pH 8.0, 200 mM sodium chloride, 10% glycerol, and 10 mM imidazole) and cells were lysed by passage through a French Press. The (His) 6 proteins were purified using Nickel-NTA- agarose resin (Qiagen, Alencia, CA), dialyzed in column buffer containing 50% glycerol and then stored at-20°C.

In vitro kinase reactions Phosphorylation of (His) 6-Fos by ERK1 immunoprecipitated from NIH3T3 cells, activate (His) 6-ERK2 (ref. 42) or FLAG-ERK5/BMK1, was performed in kinase buffer containing 10 pCi y32P-ATP at 30 C (Chen et al., Mol. Cell. Biol.

10: 3204-3215,1990). Endogenous ERK1 and RSK kinase activities were performed as described previously (Chung et al., Mol. Cell. Biol., 11: 1868-1871, 1991). The HPLC-purified synthetic peptides used in the competition kinase assays were mixed with (His) 6-Fos-EE before addition of activated ERK2 and y- 32P-ATP. The peptides derived from ELK-1 were as follows: RRPRSPAKLSFQFPSFQFP (SEQ ID NO: 21); RRPRSPAKLSAQAPSAQAP (SEQ ID NO: 22); RRPRSPAKLSFTYPSFTYP (SEQ ID NO: 23); RRPRSPAKLSATYPSATYP (SEQ ID NO: 24).

Immunofluorescence Swiss 3T3 cells (1.35 x 105 per 35-mm dish) were cultured on poly-L- lysine-coated glass coverslips for 24 h and serum-starved for 48 h. Cyclohexamide (14 ug/ml) was delivered in dimethyl sulphoxide. After stimulation with growth factors, cells were washed with ice-sold PBS containing 0.1% BSA, fixed with 3.7% formaldehyde for 10 min at room temperature and permeabilized with 0.2% Triton-X100 for 5 min. Analysis of c-Fos expression was performed using a rabbit anti-human c-Fos antibody (1: 500, Upstate Biotechnology Inc.) under conditions described by the manufacturer. Conditions for phospho-p24/p44 MAPK immunofluorescence were identical to those used for c-Fos, except that a monoclonal phospho-MAPK antibody was used (Sigma). Coverslips were mounted in Citifluor (Ted Pella Inc. , Redding, CA) and examined under epifluorescent illumination.

Bromodeoxyuridine (BrdU) incorporation Swiss 3T3 cells were cultured as described for immunofluorescence studies, treated with growth factors and 20 pM BrdU Labeling Reagent (Amersham Life Sceinces Inc. , Piscataway, NJ) for 20 h at 37 °C. For immunofluorescence analysis, a mouse anti-BrdU monoclonal (Amersham Life Sciences Inc.) supplemented with DNAase I (Invitrogen) was used.

Other Embodiments All publications and patent applications cited in this specification are herein incorporated by reference as if each individual publication or patent application were specifically and individually indicated to be incorporated by reference.

Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, it will be readily apparent to those of ordinary skill in the art in light of the teachings of this invention that certain changes and modifications may be made thereto without departing from the spirit or scope of the appended claims.

What is claimed is: TABLE 2: ONCOLOGY<BR> Amino<BR> Accession Code Target Description Target Sequence<BR> Acid<BR> 1-phosphatidylinositol-4,5-bisphosphate phosphodiesterase beta 3 (EC 3.1.4.11) (Phosphoinositide<BR> 1 PIP3_HUMAN phospholipase C) (PLC-beta-3) (Phospholipase C-beta-3). 777 DEEPFDFPKVVLPTL<BR> 26S proteasome non-ATPase regulatory subunit 1 (26S proteasome regulatory subunit S1) (26S<BR> 2 PSD1_HUMAN proteasome subunit p112). 767 TQFWFWFPLSHFLSL<BR> 26S proteasome non-ATPase regulatory subunit 1 (26S proteasome regulatory subunit S1) (26S<BR> 3 PSD1_HUMAN proteasome subunit p112). 807 KPSTFAYPAPLEVPK<BR> 4 RT31_HUMAN 28S ribosomal protein S31, mitochondrial precursor (S31mt) (MRP-S31) (Imogen 38). 307 EGKLWEFPINNEAGF<BR> 5 PDPK_HUMAN 3-phosphoinositide dependent protein kinase-1 (EC 2.7.1.37) (hPDK1). 295 IKLEYDFPEKFFPKA<BR> 6 RS3_HUMAN 40S ribosomal protein S3. 73 VQKRFGFPEGSVELY<BR> 52 kDa repressor of the inhibitor of the protein kinase (p58IKP-interacting protein) (58 kDa<BR> 7 P52K_HUMAN interferon-induced protein kinase-interacting protein) (P52rIKP) (Death associated protein 4). 18 DLAFFRFPRDPARCQ<BR> 8 B53A_HUMAN 53 kDa BRG1-associated factor A (Actin-related protein Baf53a) (ArpNbeta). 283 PTVHYEFPNGYNCDF<BR> 9 AAK1_HUMAN 5'-AMP-activated protein kinase, catalytic alpha-1 chain (EC 2.7.1.-) (AMPK alpha-1 chain). 273 DLPKYLFPEDPSYSS<BR> 10 AAK2_HUMAN 5'-AMP-activated protein kinase, catalytic alpha-2 chain (EC 2.7.1.-) (AmPK alpha-2 chain). 271 DLPSYLFPEDPSYDA<BR> 11 RLA0_HUMAN 60S acidic ribosomal protein P0 (L10E) 253 VETDYTFPLAEKVKA<BR> 12 RL10_HUMAN 60S ribosomal protein L10 (QM protein) (Tumor suppressor QM) (Laminin receptor homolog). 152 RRAKFKFPGRQKIHI<BR> 13 MC3A_MOUSE 80 kda MCM3-associated protein (GANP protein). 184 GLTPFSFPQVTNSSV<BR> 14 ASH3_MOUSE Achaete-scute homolog 3 (bHLH transcriptional regulatory Sgn-1) (Mash-3). 69 DPYPFPFPMPYTNYR<BR> 15 ASH3_HUMAN Achaete-scute homolog 3 (bHLH transcriptional regulatory Sgn-1). 69 EPCPFSFPMPYPNYR<BR> ADAM 10 precursor (EC 3.4.24.-) (A disintegrin and metalloproteinase domain 10) (Mammalian<BR> 16 AD10_HUMAN disintegrin-metalloprotease) (Kuzbanian protein homolog). 286 PTNPFRFPNIGVEKF<BR> 17 AD12_HUMAN ADAM 12 precursor (EC 3.4.24.-) (A disintegrin and metallproteinase domain 12) (Meltrin alpha). 382 ASTGYPFPMVFSSCS<BR> 18 BS69_HUMAN Adenovirus 5 E1A-binding protein (BS69 protein). 229 PDNWFCYPCIPNHEL<BR> 19 CYA4_HUMAN Adenylate cyclase, type IV (EC 4.6.1.1) (ATP pyrophosphate-lyase) (Adenylyl cyclase). 679 ITSLFFFPTSSDCPF<BR> 20 AF4_HUMAN AF-4 protein (Proto-oncogene AF4) (FEL protein). 381 EPSKFPFPTKDSQHV<BR> A-kinase anchor protein 11 (Protein kinase A anchoring protein 11) (PRKA11) (A kinase anchor<BR> 21 AK11_HUMAN protein 220 kDa) (AKAP 220) (hAKAP220). 661 EVCQFSYPQTPASPQ<BR> A-kinase anchor protein 3 (Protein kinase A anchoring protein 3) (PRKA3) (A-kinase anchor protein<BR> 110 kDa) (AKAP 110) (Sperm oocyte binding protein) (Fibrousheathin I) (Fibrous sheath protein of<BR> 22 AKA3_HUMAN 95 kDa) (FSP95). 490 SDISFEYPEDIGNLS<BR> ALK tyrosine kinase receptor precursor (EC 2.7.1.112) (Anaplastic lymphoma kinase) (cD246)<BR> 23 ALK_HUMAN antigen). 264 LECSFDFPCELEYSP<BR> 24 ANR5_HUMAN Ankyrin repeat domain protein 5. 437 VIPEYAFPRRQDGGP<BR> 25 ATR_HUMAN Anthrax toxin receptor precursor (Tumor endothelial marker 8). 421 PEQEYEFPEPRNLNN 26 KRAA_HUMAN A-Raf proto-oncogene serine/threonine-protein kinase (EC 2.7.1.-) (A-raf-1) (Proto-oncogene Pks). 193 DPEHFPFPAPANAPL<BR> ATP-binding cassette, sub-family A, member 3 (ATP-binding cassette transporter 3) (ATP-binding<BR> 27 ABC3_HUMAN cassette 3) (ABC-C transporter). 65 LPLFFTFPPPGDTWE<BR> ATP-dependent DNA helicase II, 70 kDa subunit (Lupus Ku autoantigen protein p70) (Ku70) (70 kDa<BR> subunit of Ku antigen) (Thyroid-lupus autoantigen) (TLAA) (CTC box binding factor 75 kDa subunit)<BR> 28 KU70_HUMAN (CTCBF) (CTC75). 362 PRSLFVYPEESLVIG<BR> AXIN up-regulated gene 1 protein (TGF-beta induced apoptosis protein 3) (TAIP-3) (URAX1<BR> 29 AXU1_HUMAN protein). 88 GITVFYFPRCQGFTS<BR> 30 12751139 B aggressive lymphoma long isoform [Homo sapiens] 240 SSGIFQFPLNLCTKT<BR> 31 11544936 bA271B5.1 (similar to ribosomal protein S7) [Homo sapiens] 183 KDVNFEFPEFQLQTK<BR> 32 ACTY_HUMAN Beta-centractin (Actin-related protein 1B) (ARP1B). 29 QIPKYCFPNYVGRPK<BR> 33 BRC1_HUMAN Breast cancer type 1 susceptibility protein. 825 DTEGFKYPLGHEVNH<BR> Bromodomain adjacent to zinc finger domain protein 1A (ATP-utilizing chromatin assembly and<BR> remodeling factor 1) (hACF1) (ATP-dependent chromatin remodelling protein) (Williams syndrome<BR> 34 BA1A_HUMAN transcription factor-related chromatin remodeling factor 180) (WCRF18 254 QDFSYFFPDDPPTFI<BR> 35 3928855 calcium and DAG-regulated guanine nucleotide exchange factor II [Homo sapiens] 627 EEGPFTFPNGEAVEH<BR> Calcium/calmodulin-dependent protein kinase type II alpha chain (EC 2.7.1.123) (CaM-kinase II<BR> 36 KCCA_HUMAN alpha chain) (CaM kinase II alpha subunit) (CaMK-II alpha subunit). 226 KAGAYDFPSPEWDTV<BR> CASP8 and FADD-like apoptosis regulator precursor (Cellular FLICE-like inhibitory protein) (cFLIP)<BR> (Caspase-eight-related protein) (Casper) (Caspase-like apoptosis regulatory protein (CLARP)<BR> 37 CFLA_MOUSE (MACH-related inducer of toxicity) (MRIT) (Caspase homolog) (C 282 HIQLFLFPKSHDITQ<BR> 38 CARA_HUMAN Caspase recruitment domain protein 10 (CARD-containing MAGUK protein 3) (Carma 3). 98 EALEFYYPEHFTLLT<BR> 39 CARB_HUMAN Caspase recruitment domain protein 11 (CARD-containing MAGUK protein 3) (Carma 1). 86 ESLEFYYPELYKLVT<BR> 40 CARF_HUMAN Caspase recruitment domain protein 15 (Nod2 protein) (Inflammatory bowel disease protein 1). 323 QEFLFVFPFSCRQLQ<BR> 41 CAR6_HUMAN Caspase recruitment domain protein 6. 728 LENSWLFPTRIGGNF<BR> 42 CTD1_HUMAN Catenin delta-1 (p120 catenin) (p120(ctn)) (Cadherin-associated Src substrate) (CAS) (p120(cas)). 202 LPRNFHYPPDGYSRH<BR> Catenin delta-2 (Delta-catenin) (Neural plakophilin-related ARM-repeat protein) (NPRAP)<BR> 43 CTD2_HUMAN (Neurojungin) (GT24). 162 PEGSFQYPASYHSNQ<BR> 44 CEBA-HUMAN CCAAT/enhancer binding protein alpha (C/EBP alpha). 27 SSAAFGFPRGAGPAQ<BR> 45 CEBA_HUMAN CCAAT/enhancer binding protein alpha (C/EBP alpha). 102 GGGDFDYPGAPAGPG<BR> 46 CEBE_HUMAN CCAAT/enhancer binding protein epsilon (C/EBP epsilon). 84 DPRPFAYPPHTFGPD<BR> 47 5020264 Cdc42 GTPase-activating protein [Mus musculus] 673 SPAPFPFPEAPGSLP<BR> 48 ZIZ1_HUMAN Cdc42 guanine nucleotide exchange factor zizimin 1. 1457 RSLIYKFPSTFYEGR<BR> 49 CBL2_HUMAN Cdk5 and abl enzyme substrate 2 (Interactor with cdk3 2) (Ik3-2). 241 SYAKFLYPTNALVTH Clathrin coat assembly protein AP50 (Clathrin coat associated protein AP50) (Plasma membrane<BR> adaptor AP-2 50 kDa protein) (HA2 50 kDa subunit) (Clathrin assembly protein complex 2 medium<BR> 50 A2M1_HUMAN chain) (AP-2 mu 2 chain). 114 EILDFGYPQNSETGA<BR> 51 CTA3_HUMAN Contactin associated proten-like 3 precursor (Cell recognition molecule Caspr3). 126 EESIWGFPGNTNADS<BR> 52 CRKL_HUMAN Crk-like protein. 128 VRTLYDFPGNDAEDL<BR> 53 CRN1_HUMAN Crooked neck-like protein 1 (Crooked neck homolog) (hCrn) (CGI-201) (MSTP021). 800 EYFDYIFPEDAANQP<BR> 54 11385644 CTCL tumor antigen se2-1 [Homo sapiens] 42 TEDDFEFPFAKTINLS<BR> 55 SRA4_RAT CTD-binding SR-like protein RA4 (Fragment). 158 PQAPFGYPGDGMQQP<BR> 56 CG1C_HUMAN Cyclin C. 138 TRFSYAFPKEFPYRM<BR> 57 CCT1_HUMAN Cyclin T1 (Cyclin T) (CycT1). 598 SSLNFSFPSLPTMGQ<BR> Cytochrome P450 2A12 (Ec 1.14.14.1) (CYPIIA12) (Steroid hormones 7-alpha-hydroxylase)<BR> 58 CPAC_MOUSE (Testosterone 7-alpha-hydroxylase). 456 QNFRFKFPRKLEDIN<BR> Cytoplasmic tyrosine-protein kinase BMX (EC 2.7.1.112) (Bone marrow kinase BMX) (Epithelial and<BR> 59 BMX_HUMAN endothelial tyrosine kinase) (ETK) (NTK38). 273 SISWEFPESSSSEE<BR> Cytosolic phospholipase A2 (CPLA2) [Includes: Phospholipase A2 (EC 3.1.1.4) (Phosphatidylcholine<BR> 60 PA24_HUMAN 2-acylhydrolase); Lysophospholipase (EC 3.1.1.5)]. 679 STFNFQYPNQAFKRL<BR> 61 5NTC_HUMAN Cytosolic purine 5'-nucleotidase (EC 3.1.3.5) (5'-nucleotidase cytosolic II). 260 MTYLFDFPHGPKPGS<BR> 62 1616601 disintegrin-metalloprotease MADM [Homo sapiens] 229 PTNPFRFPNISVEKF<BR> DNA (cytosine-5)-methyltransferase 3A (EC 2.1.1.37) (Dnmt3a) (DNA methyltransferase HsaIIIA)<BR> 63 DM3A-HUMAN (DNA MTase HsaIIIA) (M.HsaIIIA). 861 MERVFGFPVHYTDVS<BR> DNA (cytosine-5)-methyltransferase 3B (EC 2.1.1.37) (Dnmt3b) (DNA methyltransferase HsaIIIB)<BR> 64 DM3B_HUMAN (DNA MTase HsaIIIB) (M.HsaIIIB). 850 LERIFGFPVHYTDVS<BR> DNA (cytosine-5)-methyltransferase-like protein 2 (Dnmt2) (DNA methyltransferase homolog<BR> 65 DNM2_HUMAN HsaIIP) (DNA MTase homolog HsaIIP) (M.HsaIIP) (PuMet). 354 FPPEFGFPEKITVKQ<BR> 66 MLH3_HUMAN DNA mismatch repair protein Mlh3 (MutL protein homolog 3). 1285 LGLEFVFPDTSDSLV<BR> 67 DPD2_HUMAN DNA polymerase delta subunit 2 (EC 2.7.7.7). 389 KTDPFIFPECPHVYF<BR> 68 DPE2_HUMAN DNA polymerase epsilon subunit B (EC 2.7.7.7) (DNA polymerase II subunit B). 236 HVNAFGFPPTEPSST<BR> DNA polymerase gamma subunit 2, mitochondrial precursor (EC 2.7.7.7) (Mitochondrial DNA<BR> polymerase accessory subunit) (PoIG-beta) (MtPolB) (DNA polymerase gamma accesory 55 kDa<BR> 69 DPG2_HUMAN subunit) (p55). 287 NKLYYNFPWGKELIE<BR> 70 TP2A_MOUSE DNA topoisomerase II, alpha isozyme (EC 5.99.1.3). 633 HRIQFKYPGPEDDAA<BR> 71 TP2B_HUMAN DNA topoisomerase II, beta isozyme (EC 5.99.1.3). 1444 FGNLFSFPSYSQKSE<BR> 72 STAU_HUMAN Double-stranded RNA-binding protein Staufen homolog. 114 PRYFYPFPVPPLLYQ<BR> Dual specificity mitogen-activated protein kinase kinase 4 (EC 2.7.1.-) (MAP kinase kinase 4) (JNK<BR> 73 MPK4_HUMAN activating kinase 1) (c-Jun N-terminal kinase kinase 1) (JNKK) (SAPK/ERK kinase 1) (SEK1). 301 ATGRFPYPKWNSVFD<BR> Dual specificity protein phosphatase 1 (EC 3.1.3.48) (EC 3.1.3.16) (MAP kinase phoshatase-1)<BR> 74 DUS1_HUMAN (MKP-1) (Protein-tyrosine phosphatase CL100) (Dual specifity protein phosphatase hVH1). 335 TTTVFNFPVSIPVHS Dual specificity protein phosphatase 4 (EC 3.1.3.48) (EC 3.1.3.16) (Mitogen-activated protein kinase)<BR> 75 DUS4_HUMAN phosphatase-2) (MAP kinase phosphatase-2) (MKP-2) (Dual specificity protein phosphatase hVH2). 362 SQFVFSFPVSVGVHS<BR> 76 EDA_HUMAN Ectodysplasin A (Ectodermal dysplasia protein) (EDA protein). 134 ALLNFFFPDEKPYSE<BR> Ectonucleotide pyrophosphatase/phosphodiesterase 2 (E-NPP 2) (Phosphodiesterase I/nucleotide<BR> pyrophosphatase 2) (Phosphodiesterase I alpha) (PD-lalpha) (Autotaxin) [Includes: Alkaline<BR> 77 NPP2_HUMAN phosphodiesterase I (EC 3.1.4.1); Nucleotide pyrophosphatase (EC 3.6.1. 676 MSYGFLFPPYLSSSP<BR> 78 EF1G_HUMAN Elongation factor 1-gamma (EF-1-gamma) (eEF-1B gamma) (PRO1608). 322 WYSEYRFPEELTQTF<BR> 79 GAT2_HUMAN Endothelial transcription factor GATA-2. 167 GSHLFGFPPTPPKEV<BR> Enhancer of filmentation 1 (HEF1) (CRK-associated substrate-related protein) (CAS-L) (CasL)<BR> 80 CASL_HUMAN (PP105) (Neural precursor cell expressed developmentally down-regulated 9). 237 REKDYDFPPPMRQAG<BR> 81 ESR1_MOUSE Estrogen receptor (ER) (Estradiol receptor) (ER-alpha). 48 KPTVFNYPEGAAYEF<BR> 82 ETV2_HUMAN Ets translocation variant 2 (Ets-related protein 71). 25 AKLGFCFPDLALQGD<BR> ETS-domain protein ELK-3 (ETS-related protein NET) (ETS-related protein ERP) (SRF accessory<BR> 83 ELK3_HUMAN protein 2) (SAP-2). 370 PSTLFQFPTLLNGHM<BR> 84 ELK4_HUMAN ETS-domain protein ELK-4 (Serum response factor accessory protein 1) (SAP-1). 394 ANTLFQFPSVLNSHG<BR> 85 ERF_HUMAN ETS-domain transcription factor ERF (Ets2 repressor factor). 138 GGSHFRFPPSTPSEV<BR> 86 ERF_HUMAN ETS-domain transcription factor ERF (Ets2 repressor factor). 5 ADTGFAFPDWAYKPE<BR> 87 ETV3_HUMAN ETS-related protein PE-1 (ETS translocation variant 3) (Fragment). 168 ASSRFHFPPLDTHSP<BR> 88 ETV3_HUMAN ETS-related protein PE-1 (ETS translocation variant 3) (Fragment). 35 GGGGYQFPDWAYKTE<BR> 89 IF33_HUMAN Eukaryotic translation initiation factor 3 subunit 3 (eIF-3 gamma) (eIF3 p40 subunit) (eIF3h). 78 ITNCFPFPQHTEDDA<BR> 90 IF37_HUMAN Eukaryotic translation initiation factor 3 subunit 7 (eIF-3 zeta) (eIF3 p66) (eIF3d). 331 GKERYNFPNPNPFVE<BR> Folate receptor alpha precursor (FR-alpha (Folate receptor 1) (Folate receptor, adult) (Adult folate-<BR> 91 FOL1_HUMAN binding protein (FBP) (Ovarian tumor-associated antigen MOv18) (KB cells FBP.) 176 QPFHFYFPTPTVLCN<BR> 92 FXJ2_HUMAN Forkhead box protein J2 (Fork head homologous X). 404 NNTGFAFPSDWCSNI<BR> 93 FXK1_MOUSE Forkhead box protein K1 (Myocyte nuclear factor (MNF). 170 QQCTFRFPSTAIKIQ<BR> 94 FZD4_HUMAN Frizzled 4 precursor (Frizzled-4) (Fz-4) (hF-z4) (FzE4). 244 DSSRFSYPERPIIFL<BR> 95 GCP6-HUMAN Gamma-tubulin complex component 6 (GCP-6). 1451 LPRAFAFPVDPQVQS<BR> Geranylgeranyl pyrophosphate synthetase (GGPP synthetase) (GGPPSASE) (Geranylgeranyl<BR> diphosphate synthase) [Includes: Dimethylallyltransferase (EC 2.5.1.1); Geranyltranstransferase (EC<BR> 96 GGPP_HUMAN 2.5.1.10); Farnesyltranstransferase (EC 2.5.1.29)]. 209 TEGKFSFPTIHAIWS<BR> 97 KG3A_HUMAN Glycogen synthase kinase-3 alpha (EC 2.7.1.37) (GSK-3 alpha). 350 NYTEFKFPQIKAHPW<BR> 98 GDF3_HUMAN Growth/differentiation factor 3 precursor (GDF-3). 88 DQGFFLYPKKISQAS<BR> 99 4206785 guanine nucleotide-binding protein [Mus musculus] 104 NLPNFDFPPEFYEHA<BR> Guanine nucleotide-binding protein G (S), alpha subunit (Adenylate cyclase-stimulating G alpha<BR> 100 GBAS_HUMAN protein). 136 NVPDFDFPPEFYEHA<BR> 101 HM21_HUMAN High-mobility group protein 2-like 1 (HMGBCG protein). 207 DEESFQYPSQQATVK<BR> high-risk human papilloma viruses E6 oncoproteins targeted protein E6TP1 alpha; putative GAP<BR> 102 4151328 protein alpha [Homo sapiens] 1054 VSYEFKFPFRNNNKW 103 HDA3_HUMAN Histone deacetylase 3 (HD3) (RPD3-2). 194 KYGNYFFPGTGDMYE<BR> 104 DBX1_MOUSE Homeobox protein DBX1. 133 PPKTFAFPYFEGSFQ<BR> 105 HXCC_HUMAN Homeobox protein Hox-C12 (Hox-3F). 20 TGD TFYFPNFRASGA<BR> 106 HMPH_HUMAN Homeobox protein PRH (Hematopoietically expressed homeobox) (Homeobox protein HEX). 97 GGPLYPFPRTVNDYT<BR> 107 HIK2_HUMAN Homeodomain-interacting protein kinase 2 (EC 2.7.1.-). 1067 AQAPYSFPHNSPSHG<BR> 108 IKAP_HUMAN IkappaB kinase complex-associated protein (IKK complex-associated protein) (p150). 599 FPVRFPYPCTQTELA<BR> Inositol-trisphosphate 3-kinase B (EC 2.7.1.127) (Inositol 1,4,5- trisphosphate 3-kinase) (IP3K) (IP3<BR> 109 IP3L_HUMAN 3-kinase) (IP3K-B). 52 RGASFLFPPAESL SP<BR> 110 ITA5_HUMAN Integrin alpha-5 precursor (Fibronectin receptor alpha subunit) (Integrin alpha-F) (VLA-5) (CD49e). 422 QGVVFVFPGGPGGLG<BR> Interstitial collagenase precursor (EC 3.4.24.7) (Matrix metalloproteinase-1) (MMP-1) (Fibroblast<BR> 111 MM01_HUMAN collagenase). 364 IYSSFGFPRTVKHID<BR> 112 13383464 kinesin-related protein HASH [Mus musculus] 46 NTRDFMFPGPNQMSG<BR> 113 KLF4_MOUSE Kruppel-like factor 4 (Gut enriched kruppel-like factor) (Epithelial zinc-finger protein EZF). 134 STCSFSYPIRAGGDP<BR> 114 LAF4_HUMAN LAF-4 protein (Lymphoid nuclear protein related to AF4). 325 EPTKFPFPNKDSQLV<BR> 115 LAMA_HUMAN Lamin A/C (70 kDa lamin). 477 PLL TYRFPPKFTLKA<BR> 116 4324434 large tumor suppressor 1 [Homo sapiens] 334 SSSKFNFPSGRPGMQ<BR> 117 930341 LAR-interacting protein 1a [Homo sapiens] 530 SVPDFRFPMADGHTD<BR> 118 EDG2_HUMAN Lysophosphatidic acid receptor Edg-2 (LPA receptor 1) (LPA-1). 76 VNRRFHFPIYYLMAN<BR> 119 EDG7_HUMAN Lysophosphatidic acid receptor Edg-7 (LPA receptor 3) (LPA-3). 57 KNRKFHFPFYYLLAN<BR> Macrophage metalloelastase precursor (EC 3.4.24.65) (HME) (Matrix metalloproteinase-12) (MMP-<BR> 120 MM12_HUMAN 12) (Macrophage elastase) (ME). 367 SIHSFGFPNFVKKID<BR> MAP kinase-activated protein kinase 2 (EC 2.7.1.-) (MAPK-activated protein kinase 2) (MAPKAP<BR> 121 MKK2_HUMAN kinase 2) (MAPKAPK-2). 280 RMGQYEFPNPEWSEV<BR> Melanoma antigen preferentially expressed in tumors (Preferentially expressed antigen of<BR> 122 MAPE_HUMAN melanoma) (OPA-interacting protein 4) (OIP4). 140 RASLYSFPEPEAAQP<BR> 123 MAG3_HUMAN Melanoma-associated antigen 3 (MAGE-3 antigen) (Antigen MZ2-D). 141 GNWQYFFPVIFSKAS<BR> Metalloproteinase inhibitor 3 precursor (TIMP-3) (Tissue inhibitor of metalloproteinases-3) (MIG-5<BR> 124 TIM3_HUMAN protein). 168 MLSNFGYPGYQSKHY<BR> 125 AMP1_HUMAN Methionine aminopeptidase 1 (EC 3.4.11.18) (MetAP 1) (Peptidase M 1) (Fragment). 191 PLNYYNFPKSCCTSV<BR> Microtubuts-associated protein 1A (MAP 1A) (Proliferation-related protein p80) [Contains: MAP1<BR> 126 MAPA_HUMAN light chain LC2]. 37 KPCCYIFPGGRGDSA<BR> Mitochondrial 28S ribosomal protein S29 (S29mt) (MRP-S29) (Death- associated protein 3) (DAP-3)<BR> 127 RT29_HUMAN (lonizing radiation resistance conferring protein). 113 KNTSFAYPAIRYLLY<BR> 128 3133291 mitogen activated protein kinase activated protein kinase [Homo sapiens] 259 MTGSFEFPEEEWSQI<BR> Mitogen-activated protein kinase 6 (EC 2.7.1.-) (Extracellular signal- regualted kinase 3) (ERK-3)<BR> 129 MK06_HUMAN (MAP kinase isoform p97) (p97-MAPK). 315 YMSIYSFPMDEPISS<BR> 130 M3K6_HUMAN Mitogen-activated protein kinase kinase kinase 6 (EC 2.7.1.-). 800 HRPLFAFPDAVKQIL 131 TAB1_HUMAN Mitogen-activated protein kinase kinase kinase 7 interacting protein 1 (TAK1-binding protein 1). 316 LVRNFGYPLGEMSQP<BR> Mitogen-activated protein kinase kinase kinase kinase 2 (EC 2.7.1.37) (MAPK/ERK kinase kinase<BR> kinase 2) (MEK kinase kinase 2) (MEKKK 2) (Germinal center kinase) (GC kinase) (Rab8 interacting<BR> 132 M4K2_HUMAN protein) (B lymphocyte serine/threonine protein kinase). 737 PELTFDFPIETVVCL<BR> 133 MDR1_HUMAN Multidrug resistance protein 1 (P-glycoprotein 1) (CD243 antigen). 1038 GEVVFNYPTRPDIPV<BR> 134 MDR1_HUMAN Multidrug resistance protein 1 (P-glycoprotein 1) (CD243 antigen). 395 RNVHFSYPSRKEVKI<BR> 135 MDR3_HUMAN Multidrug resistance protein 3 (P-glycoprotein 3). 88 TAGNFSFPVNFSLSL<BR> 136 MDR3_HUMAN Multidrug resistance protein 3 (P-glycoprotein 3). 397 NDVHFSYPSRANVKI<BR> 137 DRNL_HUMAN Muscle-specific DNase I-like precursor (EC 3.1.21.-) (DNase X) (XIB). 242 TAAAFDFPTSFQLTE<BR> 138 6959304 MYB-binding protein 1A [Homo sapiens] 497 TKHPFSFPLENQARE<BR> 139 2706549 myc-intron-binding protein-1 [Mus musculus] 135 SEDLFPFPMHGHSGG<BR> 140 MY15_HUMAN Myosin XV (Unconventional myosin-15). 751 RGAAFGFPGASPRAS<BR> Neutrophil collagenase precursor (EC 3.4.24.34) (Matrix metalloproteinase-8) (MMP-8) (PMNL<BR> 141 MM08_HUMAN collagenase) (PMNL-CL). 364 DISNYGFPSSVQAID<BR> Nuclear factor NF-kappa-B p105 subunit (DNA-brinding factor KBF1) (EBP- 1) [Contains: Nuclear<BR> 142 KBF1_HUMAN factor NF-kappa-B p50 subunit]. 405 SFPHYGFPTYGGITF<BR> Nuclear factor NF-kappa-B p105 subunit (DNA-binding factor KBF1) (EBP- 1) [Contains: Nuclear<BR> 143 KBF1_HUMAN factor NF-kappa-B p50 subunit]. 400 TGPGYSFPHYGFPTY<BR> 144 RI14_HUMAN Nuclear factor RIP140 (Nuclear receptor interacting protein 1). 996 DNRTFSYPGVVKTPV<BR> 145 RI14_HUMAN Nuclear factor RIP140 (Nuclear receptor interacting protein 1). 905 NDLEFKYPAGHGSAS<BR> 146 N153_HUMAN Nuclear pore complex protein Nup153 (Nucleoporin Nup153) (153 kDa nucleoporin). 421 RESGFSYPNFSLPAA<BR> 147 NCR1_HUMAN Nuclear receptor co-respressor 1 (N-CoR1) (N-CoR). 23 HSVQYTFPNTRHQQE<BR> 148 RORG_HUMAN Nuclear receptor ROR-gamma (Nuclear receptor RZR-gamma). 540 PPSPFSFPMNPGGWS<BR> 149 DD21_HUMAN Nucleolar RNA helicase II (Nucleolar RNA helicase Gu) (RH II/Gu) (DEAD-box protein 21). 136 RGVTFLFPIQAKTFH<BR> Nucleoside diphosphate kinase homolog 5 (NDK-H 5) (NDP kinase homolog 5) (nm23-H5) (Testis-<BR> 150 NDK5_HUMAN specific nm23 homolog) (Inhibitor of p53-induced apoptosis-beta) (IPIA-beta). 137 REIRFMFPEVIVEPI<BR> 151 OXE2_HUMAN Olfactory receptor 51E2 (Prostate specific G-protein coupled receptor) (HPRAJ). 150 RGSLFFFPLPLLIKR<BR> 152 ORC4_HUMAN Origin recognition complex subunit 4. 217 LMNSFGFPQYVKIFK<BR> 153 ORC5_HUMAN Origin recognition complex subunit 5. 24 ERHHFSFPSIFIYGH<BR> 154 2645205 p160 myb-binding protein [Mus musculus] 495 TKQHFSFPLDDRNRG<BR> 155 PAXI_HUMAN Paxillin. 114 EEHVYSFPNKQKSAE<BR> Peptidyl-prolyl cis-trans isomerase like 2 (EC 5.2.1.8) (PPIase) (Rotamase) (Cyclophilin-60)<BR> 156 CYP6_HUMAN (Cyclophilin-like protein Cyp-60). 45 SLQPFVYPVCTPDGI<BR> Peripheral plasma membrane protein CASK (EC 2.7.1.-) (hCASK) (Calcium/calmodulin-dependent<BR> 157 CSKP_HUMAN serine protein kinase) (Lin-2 homolog). 763 HPDRFAYPIPHTTRP<BR> Peroxisomal 3,2-trans-enoyl-CoA isomerase (EC 5.3.3.8) (Dodecenoyl-CoA delta-isomerase)<BR> (D3,D2-enoyl-CoA isomerase) (DBI-related protein 1) (DRS-1) (Hepatocellular carcinoma-<BR> 158 PECI_HUMAN associated antigen 88). 246 GCSSYTFPKIMSPAK 159 PPAR_HUMAN Peroxisome proliferator activated receptor alpha (PPAR-alpha). 417 PDDIFLFPKLLQKMA<BR> Peroxisome proliferator activated receptor delta (PPAR-delta) (PPAR-beta) (Nuclear hormone<BR> 160 PPAS_MOUSE receptor 1) (NUC1). 389 PDSQYLFPKLLOKMA<BR> Phosphatidylinositol 3-kinase regulatroy alpha subunit (PI3-kinase p85-alpha subunit) (Ptdins-3-<BR> 161 P85A_MOUSE kinase p85-alpha) (PI3K). 628 GLDLFVFPYRVVATA<BR> Phosphatidylinositol-3,4,5-trisphosphate 3-phosphatase PTEN (EC 3.1.3.67) (Mutated in multiple<BR> 163 PTEN_HUMAN advanced cancers 1). 237 KFMYFEFPQPLPVCG<BR> Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit, beta isoform (EC 2.7.1.153) (PI3-<BR> 164 P11B_HUMAN kinase p110 subunit beta) (Ptdlns-3-kinase p110) (PI3K) (PI3Kbeta). 608 ELLDFNYPDQYVREY<BR> Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit. delta isofomr (EC 2.7.1. 153) (PI3-<BR> 165 P11D_HUMAN kinase p110 subunit delta) (Ptdlns- 3-kinase p110) (PI3K) (p110delta). 581 ELLDFSFPDCHVGSF<BR> Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit, delta isoform (EC 2.7.1. 153) (PI3-<BR> 166 P11D_HUMAN kinase p110 subunit delta (Ptdlns- 3-kinase p110) (PI3K) (p110delta). 161 AWLQYSFPLQLEPSA<BR> 167 940231 phosphodiesterase A' subunit [Homo sapiens] 366 ADEYFTFPKGPVDET<BR> 168 22758919 phosphoinositide 3 kinase P110delta [Mus musculus] 580 ELLDFSFPDCYVGSF<BR> 169 2696236 phospholipase B [Rattus norvegicus] 42 TLKNFSFPCKPKKLE<BR> 170 PLEK_HUMAN Pleckstrin (Platelet p47 protein).213 PDAFYYFPDSGFFCE<BR> 171 3002588 Plenty of SH3s; POSH [Mus musculus] 376 TGPAFTFPSDVPYQA<BR> 172 PAB5_HUMAN Polyadenylate-binding protein 5 (Poly(A)-binding protein 5) (PABP 5). 176 YVGRFKFPEERAAEV<BR> 173 PAB5_HUMAN Polyadenylate-binding protein 5 (Poly(A)-binding protein 5) (PABP 5). 62 GYVNFRFPADAEWAL<BR> 174 15667468 polyploidy associated protein kinase PAPK-A [Mus musculus] 402 SELEFQFPDDKDPVW<BR> 175 DP 1_HUMAN Polyposis locus protein 1 (TB2 protein). 56 NLIGFGYPAYISIKA<BR> 176 CIQ3_HUMAN Potassium voltage-gated channel subfamily KQT member 3 (Potassium channel KQT-like 3). 587 KGSAFTFPSQQSPRN<BR> 177 MV10_HUMAN Potentail helicase MOV-10. 601 KKGEYVEPAKKKLQE<BR> 178 A11A_HUMAN Potential phospholipid-transporting ATPase IS (EC 3.6.3.1) (Fragment). 552 KNVCFIFPQELYQFF<BR> 179 4097902 potential transcriptional repressor Not4hp [Mus musculus] 485 VYNSFGFPGQAARYP<BR> PR-domain zinc finger protein 2 (Retinoblastoma protein-interacting zinc-finger protein) (MTE-<BR> 180 PRD2_HUMAN binding protein) (MTB-ZF). 354 VFETFMFPCQHCERK<BR> 181 DDX6_HUMAN Probable ATP-dependent RNA helicase p54 (ONcogene RCK) (DEAD-box protein 6). 405 VVINFDFPKLAFTYL<BR> 182 GPR4_HUMAN Probable G protein-coupled receptor GPR4 (GPR19). 186 VFVGFLFPWALMLLS<BR> Probable G protein-coupled receptor GPR68 (Ovarian cancer G protein- coupled receptor 1) (OGR-<BR> 183 GP68_HUMAN 1). 190 FLVGFLFPICLLLAS<BR> 184 K682_HUMAN Probable RNA-binding protein KIAA0682. 446 AFITFMFPEHAVKAY<BR> 185 MN1_HUMAN Probable tumor suppressor protein MN1. 455 QEARFDFPGSAGVDR<BR> Probable ubiquitin carboxyl-terminal hydrolase FAF-X (EC 3.1.2.15) (Ubiquitin thiolesterase FAF-X)<BR> (Ubiquitin-specific processing protease FAF-X) (Deubiquitinating enzyme FAF-X) (Fat facets protein<BR> 186 FAFX_HUMAN related, X-linked) (Ubiquitin-specific protease 9, X chro 1805 FNDYFEFPRELDMEP Probabie ubiquitin carboxyi-terminal nydrolase FAF-@ (EC 3.1.2.15) (Ubiquitin thioiesterase FAF-Y)<BR> (Ubiquitin-specific processing protease FAF-Y) (Deubiquitinating enzyme FAF-Y) (Fat facets protein<BR> 187 FAFY_HUMAN related, Y-linked) (Ubiquitin-specific protease 9, Y chro 1616 RDDVFGYPHQFEDKP<BR> 188 PDC2_HUMAN Programmed cell death protein 2 (Zinc finger protein Rp-8). 183 PDHNFLFPEFEIVIE<BR> 189 3858885 profilferation potential-related protein [Mus musculus] 742 TDTLFVFPSREDATP<BR> 190 KPCM_HUMAN Protein kinase C, mu type (EC 2.7.1.-) (nPKC-mu) (Protein kinase D). 794 QNAAFMYPPNPWKEI<BR> 191 KPCN_HUMAN Protein kinase C, nu type (EC 2.7.1.-) (nPKC-nu) (Protein kinase EPK2). 787 QNAAFMYPPNPWREI<BR> 192 15080775 protein kinase NYD-SP5 [Homo sapiens] 97 DQKSFIFPQESEGTF<BR> Protein phosphatase 1, regulatory subunit 3D (Protein phosphatase 1, regulatory subunit 6) (Protein<BR> 193 PP3D_HUMAN phosphatase 1 binding subunit R6). 234 DVFTFGFPVPPFLLE<BR> 194 10567793 protein tyrosine phosphatase BK [Mus musculus] 262 RDRRFHFPEETPETP<BR> 195 1144002 protein tyrosine phosphatase D30 [Rattus norvegicus] 261 RDRPFHFPEETPETP<BR> 196 23268287 protein tyrosine phosphatase receptor-like protein J [Mus musculus] 172 TNNSFAFPESNETQA<BR> Protein tyrosine phosphatase, non-receptor type 13 (EC 3.1.3.48) (Protein-tyrosine phosphatase 1E)<BR> (PTP-E1) (hPTPE1) (PTP-BAS) (Protein-tyrosine phosphatase PTPL1) (Fas-associated protein-<BR> 197 PTND_HUMAN tyrosine phosphatase 1) (FAP-1). 1704 ICTMFYYPQKIPNKP<BR> Proteinase activated receptor 2 precursor (PAR-2) (Thrombin receptor- like 1) (Coagulation factor II<BR> 198 PAR2_HUMAN receptor-like 1). 247 AIGVFLFPAFLTASA<BR> Protein-tyrosine phosphatase eta precursor (EC 3.1.3.48) (R-PTP-eta( (HPTP beta-like tyrosine<BR> 199 PTPJ_MOUSE phosphatase). 172 TNNTFAFPESNETQA<BR> Protein-tyrosine phosphatase zeta precursor (EC 3.1.3.48) (R-PTP- zeta) (Phosphacan) (3F8<BR> 200 PTPZ_RAT chondroitin sulfate proteoglycan) (3H1 keratan sulfate proteoglycan). 1565 TSTDFSFPDVNEKDA<BR> Protein-tyrosine phosphatase zeta precursor (EC 3.1.3.48) (R-PTP- zeta (Phosphacan) (3F8<BR> 201 PTPZ_RAT chondroitin sulfate proteoglycan) (3H1 keraten sulfate proteoglycan). 720 HYSTFAYPPTEVTSH<BR> 202 FAT2_HUMAN Protocadherin Fat 2 precursor (hFat2) Multiple epidermal growth faclor-like domains 1). 4222 LYGGFPFPLEMENKR<BR> 203 8216989 putative cell cycle control protein [Homo sapiens] 100 NNQLFRFPATSPLKT<BR> 204 GP40 HUMAN Putative G protein-coupled receptor GPR40. 244 NVASFLYPNLGGSWR<BR> 205 RBM9_HUMAN Putative RNA-binding protein 9 (RNA binding motif protein 9). 251 IIPGFPYPTAATTAA<BR> 206 6007826 rab escrot protein-2 [Mus musculus] 370 GNTPFIFPLYGHGEI<BR> Rab proteins geranylgeranyltransferase component A 1 (Rab escort protein 1) (REP-1)<BR> 207 RAE1_HUMAN (Choroideraemia protein) (TCD protein). 369 GNTPFLFPLYGQGEL<BR> Rab proteins geranylgeranyltransferase component A 2 (Rab escort protein 2) (REP-2)<BR> 208 RAE2_HUMAN (Choroideraemia-like protein). 371 GNTPFLFPLYGQGEI<BR> 209 RBPL_HUMAN Recombining binding protein suppressor of hairless-like protein (Transcription factor RBP-L). 311 HKCAFQFPGSPPGGG<BR> 210 RB_HUMAN Retinoblastoma-associated protein (PP110) (P105-RB) (RB). 786 PRSPYKFPSSPLRIP<BR> 211 RBB5_HUMAN Retinoblastoma-binding protein 5 (RBBP-5) (Retinoblastoma-binding protein RBQ-3). 100 CDQRFRFPSPILKVQ<BR> Retinoblastoma-binding protein 8 (RBBP-8) (CtBP interacting protein) (CtIP) (Retinoblastoma-<BR> 212 RBBB_HUMAN interacting protein and myosin-like) (RIM). 467 KENAFPFPMDNQFSM 213 RBL2_HUMAN Retinoblastoma-like protein 2 (130 kDa retinoblastoma-associated protein) (PRB2) (P130) (RBR-2). 170 FQDIFKYPQEEQPRQ<BR> 214 12053793 retinoid-acid induced protein 1 [Homo sapiens] 1830 PISLFSFPPLLPQQF<BR> Rho guanine nucleotide exchange factor 2 (GEF-H1 protein) (Proliferating cell nucleolar antigen<BR> 215 ARH2_HUMAN p40). 505 KDQKYIFPTLDKPSV<BR> Rho-GTPase-activating protein 7 (Rho-type GTPase-activating protein 7) (Deleted in liver cancer 1<BR> protein) (Dlc-1) (HP protein) (StAR-related lipid transfer protein 12) (StARD 12) (START domain-<BR> 216 RHG7_HUMAN containing protein 12). 34 LYEDFLFPIDISLVK<BR> 217 RBMS_HUMAN RNA-binding protein with multiple splicing (RBP-MS). 163 PPPAFTYPASLHAQM<BR> Runt-related transcription factor 2 (Core-binding factor, alpha 1 subunit) (CBF-alpha 1) (Acute<BR> myeloid leukemia 3 protein) (Oncogene AML-3) (Polyomavirus enhancer binding protein 2 alpha A<BR> 218 RUN2_HUMAN subunit) (PEBP2-alpha A) (PEA2-alpha A) (SL3-3 enhancer factor 1 450 SSGSYQFPMVPGGDR<BR> 219 SEN2_HUMAN Sentrin-specific protease 2 (EC 3.4.22.-) (Sentrin/SUMO-specific protease SENP2). 66 AASLFGFPFQLTTKP<BR> 220 SEN2_HUMAN Sentrin-specific protease 2 (EC 3.4.22.-) (Sentrin/SUMO-specific protease SENP2). 437 VFSTFFYPKLKSGGY<BR> 221 SEP1_HUMAN Septin 1 (LARP) (Serologically defined breast cancer antigen NY-BR-24). 195 EIHIYQFPECDSDED<BR> Septin 4 (Peanut-like proten 2) (Brain protein H5) (Cell division control-related protein 2) (hCDCREL-<BR> 222 SEP4_HUMAN 2) (Bradeion beta) (CE5B3 beta) (Cerebral protein-7) (hucep-7). 314 GIKIYQFPDCDSDED<BR> 223 SEP5_HUMAN Septin 5 (Peanut-like protein 1) (Cell division control related protein 1) (CDCREL-1). 214 GIHVYQFPECDSDED<BR> 224 SEP7_HUMAN Septin 7 (CDC10 protein homolog). 200 KIKIYEFPETDDEEE<BR> Serine/threonine kinase 6 (EC 2.7.1.37) (Serine/threonine kinase 15) (Aurora/IPL 1-related kinase 1)<BR> 225 STK6_HUMAN (Aurora-related kinase 1) (hARK1) (Aurora-A) (Breast-tumor-amplified kinase). 342 SRVEFTFPDFVTEGA<BR> 226 STKD_MOUSE Serine/threonine protein kinase 13 (EC 2.7.1.37) (Aurora/IpI1/Eg2 protein 1). 225 RQVDFKFPSSVPAGA<BR> 227 KPT3_HUMAN Serine/threonine protein kinase PCTAIRE-3 (EC 2.7.1.-). 371 EFRTYSFPCYLPQPL<BR> Serine/threonine protein phosphatase 2A, 72/130 kDa regulatory subunit B (PP2A, subunit B, B"-<BR> PR72/PR130) (PP2A, subunit B, B72/B130 isoforms) (PP2A, subunit B, PR72/PR130 isoforms)<BR> 228 2ACA_HUMAN (PP2A, subunit B, R3 isoform). 708 NIPRFYFPEGLPDTC<BR> 229 ST19_HUMAN Serine/threonine-protein kinase 19 (EC 2.7.1.37) (RP1 protein) (G11 protein). 96 SLHSYPFPGTIKSRD<BR> Serine/threonine-protein kinase MAK (EC 2.7.1.-) (Male germ cell- associated kinase) (Protein<BR> 230 MAK_MOUSE kinase RCK). 432 KESPFRFPDSGLPVS<BR> 231 MAK_HUMAN Serine/threonine-protein kinase MAK (EC 2.7.1.-) (Male germ cell- associated kinase). 232 SSMNFRFPQCVPINL<BR> 232 KPT2_HUMAN Serine/threonine-protein kinase PCTAIRE-2 (EC 2.7.1.-). 421 EFKNYNFPKYKPEPL<BR> 233 ULK1_HUMAN Serine/threonine-protein kinase ULK1 (EC 2.7.1.-) (Unc-51-like kinase 1). 627 AVPSFDFPKTPSSQN<BR> 234 ST5A_HUMAN Signal transducer and activator of transcription 5A. 664 SYLIYVFPDRPKDEV<BR> 235 ST5B_HUMAN Signal transducer and activator of transcription 5B. 664 NYLIYVFPDRPKDEV<BR> 236 CBL_HUMAN Signal transduction protein CBL (Proto-oncogene c-CBL). 333 REGFYLFPDGRNQNP<BR> 237 CBLB_HUMAN Signal transduction protein CBL-B (SH3-binding protein CBL-B). 966 ILREFAFPPPVSPRL<BR> 238 6649242 splicing coactivator subunit SRm300 [Homo sapiens] 2225 SSSSFPFPCKAWPSG<BR> Splicing factor 3B subunit 3 (Spliceosome associated protein 130) (SAP 130) (SF3b130) (Pre-<BR> 239 S3B3_HUMAN mRNA splicing factor SF3b 130 kDa subunit). 1162 SFRSYYFPVKNVIDG 240 STAC_HUMAN Stac protein (SRC homology 3 and cysteine-rich domain protein). 253 SNSVFTYPENGTDDF<BR> 241 TX18_HUMAN T-box transcription factor TBX18 (T-box protein 18) (Fragment). 186 GVKAFSFPETVFTTV<BR> 242 TX20_HUMAN T-box transcription factor TBX20 (T-box protein 20) (Fragment). 205 EFRTFIFPETVFTAV<BR> 243 TBX3_HUMAN T-box transcription factor TBX3 (T-box protein 3). 597 FGSLFPYPYTYMAAA<BR> 244 TBX6_HUMAN T-box transcription factor TBX6 (T-box protein 6). 235 GMASFRFPETTFISV<BR> 245 T2AY_HUMAN TFIIA-alpha and beta like factor (ALF). 106 SSANFTFPGYPIHVP<BR> TFIIH basal transcription factor complex p34 subunit (Basic transcription factor 2 34 kDa subunit)<BR> 246 TFH3_HUMAN (BTF2-p34) (General transcription factor IIH polypeptide 3). 65 QESRFLYPGKNGRLG<BR> 247 TOB1_HUMAN Tob1 protein (Transducer of erbB-2 1). 270 NAKEFIFPNMQGQGS<BR> 248 GAT3_HUMAN Trans-acting T-cell specific transcription factor GATA-3. 147 SPHLFTFPPTPPKDV<BR> 249 GAT4_HUMAN Transcription factor GATA-4 (GATA binding factor-4). 104 VSPRFSFPGTTGSLA<BR> 250 GAT5_HUMAN Transcription factor GATA-5 (GATA binding factor-5). 365 EPEDFAFPSTAPSPQ<BR> 251 TF65_MOUSE Transcription factor p65 (Nuclear factor NF-kappa-B p65 subunit). 345 APQPYTFPASLSTIN<BR> 252 TCL5_HUMAN Transcription factor-like 5 protein (Cha transcription factor) (HPV- 16 E2 binding protein 1) (E2BP-1). 203 TALQFTYPLFTTNAC<BR> 253 TF1A_HUMAN Transcription intermediary factor 1-alpha (TIF1-alpha) (Tripartite motif protein 24). 729 PPENYDFPVVIVKQE<BR> 254 ERG_MOUSE Transcriptional regulator ERG (Fragment). 52 GGAAFIFPNTSVYPE<BR> Transitional endoplasmic reticulum ATPase (TER ATPase) (15S Mg(2+)- ATPase p97 subunit)<BR> 255 TERA_HUMAN (Valosin containing protein) (VCP) [Contains: Valosin]. 767 GFGSFRFPSGNQGGA<BR> 256 E2BE_HUMAN Translation initiation factor eIF-2B epsilon subunit (eIF-2B GDP-GTP exchange factor). 220 GLRRFAFPLSLFQGS<BR> Translocon-associated protein, alpha subunit precursor (TRAP-alpha) (Signal sequence receptor<BR> 257 SSRA_HUMAN alpha subunit) (SSR-alpha). 120 LDASFRYPQDYQFYI<BR> 258 TRIO_HUMAN Triple functional domain protein (PTPRF interacting protein). 2437 RPGSFTFPGDSDSLQ<BR> 259 1113923 tyrosine phosphatase-like protein IA-2a [Rattus norvegicus] 947 SKDKFEFPLTPVGEE<BR> Tyrosine-protein kinase BTK (EC 1.7.1.112) (Bruton's tyrosine kinase) (Agammaglobulinaemia<BR> 260 BTK_HUMAN tyrosine kinase) (ATK) (B cell progenitor kinase) (BPK). 94 IIERFPYPFQVVYDE<BR> 261 JAK2_HUMAN Tyrosine-protein kinase JAK2 (EC 1.7.1.112) (Janus kinase 2) (JAK-2). 114 YRIRFYFPRWYCSGS<BR> Tyrosine-protein kinase JAK3 (C 2.7.1.112) (Janus kinase 3) (JAK-3) (Leukocyte janus kinase) (L-<BR> 262 JAK3_HUMAN JAK). 100 YRIRFYFPNWFGLEK<BR> 263 KSYK_HUMAN Tyrosine-protein kinase SYK (EC 2.7.1.112) (Spleen tyrosine kinase).<BR> <P>Tyrosine-protein kinase transmembrane receptor ROR1 precursor (EC 2.7.1.112) (Neurotrophic<BR> 264 ROR1_HUMAN tyrosine kinase, receptor-related 1).<BR> <P>Tyrosine-protein kinase transmembrane receptor ROR2 precursor (EC 2.7.1.112) (Neurotrophic<BR> 265 ROR2_HUMAN tyrosine kinase, receptor-related 2). 230 SFCHFVFPLCDARSR<BR> Ubiquitin carboxyl-terminal hydrolase 26 (EC 3.1.2.15) (Ubiquitin thiolesterase 26) (Ubiquitin-specific<BR> 266 UBPQ_MOUSE processing protease 26) (Deubiquitinating enzyme 26). 223 SKPGFGFPFETNYPE<BR> Ubiquitin carboxyl-terminal hydrolase 8 (EC 3.1.2.15) (Ubiquitin thiolesterase 8) (Ubiquitin-specific<BR> 267 UBP8_HUMAN processing protease 8) (Deubiquitinating enzyme 8). 330 ISLDFTYPSLEESIP<BR> 268 UB4B_HUMAN Ubiquitin conjugation factor E4 B (Ubiquitin-fusion degradation protein 2). 537 DSDYFKYPLMALGEL 269 27434480 ubiquitin ligase E3 alpha-II [Homos apiens] 80 EDPAFGFPKLEQANK<BR> Ubiquitin-protein ligase EDD (EC 6.3.2.-) (Hyperplastic discs protein homolog) (hHYD) (Progestin<BR> 270 EDD_HUMAN induced protein). 274 DISYFGYPSFRRSSL<BR> 271 SYV2 HUMAN Valyl-tRNA synthetase 2 (EC 6.1.1.9) (Valine-tRNA ligase 2) (ValRS 2) (G7a). 1001 GFQAYDFPAVTTAQY<BR> Valut poly (ADP-ribose) polymerase (EC 2.4.2.30) (VPAPP) (193-kDa vault protein) (PARP-<BR> 272 PPOV_HUMAN related/lalphal-related H5/proline-rich) (PH5P). 921 YKELFSYPKHITSNT<BR> 273 VINE_HUMAN Vinexin (SH3-containing adaptor molecule-1) (SCAM-1). 565 RRTGFSFPTQEPRPQ<BR> 274 VINE_HUMAN Vinexin (SH3-containing adaptor molecule-1) (SCAM-1). 352 GRRDFVYPSSTRDPS<BR> 275 WRN_HUMAN Werner syndrome helicase. 577 KSLCFQYPPVYVGKI<BR> Wiskott-Aldrich syndrome protein family member 2 (WASP-family protein member 2) (Verprolin<BR> 276 WAS2_HUMAN homology domain-containing protein 2). 270 PPAEFSYPVDNQRGS<BR> Wiskott-Aldrich syndrome protein family member 2 (WASP-_HUMAN-family protein member 2) (Verprolin<BR> 277 WAS2_HUMAN homology domain-containing protein 2). 218 KLGPFGYPPTLVYQN<BR> 278 WBP2_MOUSE WW domain binding protein 2 (WBP-2). 149 PSGAYVFPPPVANGM<BR> Zinc finger protein 44 (Zinc finger protein KOX7) (Gonadotropin inducible transcription repressor-2)<BR> 279 ZN44_HUMAN (GIOT-2). 342 CGKGFDFPGSARIHE<BR> 280 HRX_HUMAN Zinc finger protein HRX (ALL-1) (Trithorax-like protein). 792 LNPTFTFPSHSLTQS<BR> 281 Z287_HUMAN Zinc finger protein ZNF287. 44 NFRNFPYPDLAGPRK<BR> TABLE 3: CARDIOVASCULAR<BR> Amino<BR> Accession Code Target Description Target Sequence<BR> Acid<BR> 1 ACHB_HUMAN Acetylcholine receptor protein, beta chain precursor. 395 PPSDFLFPKPNRFQP<BR> 2 CYA4_HUMAN Adenylate cyclase type IV (EC 4.6.1.1) (ATP pyrophosphate-lyase) (Adenylyl cyclase). 679 ITSLFFFPTSSDCPF<BR> 3 GAT2_HUMAN Endothelial transcription factor GATA-2. 167 GSHLFGFPPTPPKEV<BR> 4 CXA1_HUMAN Gap junction alpha-1 protein (Connexin 43) (Cx43) (Gap junction 43 kDa heart protein). 330 HAQPFDFPDDNQNSK<BR> Guanine nucleotide-binding protein G (S), alpha subunit (Adenylate cyclase-stimulating G alpha<BR> 5 GBAS_HUMAN protein). 136 NVPDFDFPPEFYEHA<BR> 6 IKAP_HUMAN IkappaB kinase complex-associated protein (IKK complex-associated protein) (p150). 599 FPVRFPYPCTQTELA<BR> 7 LGR6_HUMAN Leucine-rich repeat-containing G protein-coupled receptor 6. 751 GLETYGFPSVTLISC<BR> 8 MYHD_HUMAN Myosin heavy chain, skeletal muscle, extraocular (MyHC-eo). 306 STNPFDFPFVSQGEV<BR> NDRG4 protein (Brain development-related molecule 1) (Vascular smooth muscle cell associated<BR> 9 NDR4_HUMAN protein-8 (SMAP-8). 80 FPQGYQFPSMEQLAA<BR> 10 ACHO_RAT Neuronal acetylcholine receptor protein, beta-3 chain precursor. 358 PMDRFSFPDGKESDT<BR> 11 P2X1_HUMAN P2X purinoceptor 1 (ATP receptor) (P2X1) (Purinergic receptor). 86 DVADYVFPAQGDNSF<BR> 12 P2X7_HUMAN P2X purinoceptor 7 (ATP receptor) (P2X7) (Purinergic receptor) (P2Z receptor). 89 DTADYTFPLQGNSFF<BR> 13 GPR4_HUMAN Probable G protein-coupled receptor GPR4 (GPR19). 186 VFVGFLFPWALMLLS Probable G protein-coupled receptor GPR68 (Ovarian cancer G protein- coupled receptor 1) (OGR-<BR> 14 GP68_HUMAN 1). 190 FLVGFLFPICLLLAS<BR> 15 GP17_HUMAN Probable P2Y purinoceptor GPR17 (P2Y-like receptor) (R12). 225 LAVAFTFPFITTVTC<BR> 16 GP40_HUMAN Putative G protein-coupled receptor GPR40. 244 NVASFLYPNLGGSWR<BR> 17 FK79_HUMAN Putative P2Y purinoceptor FKSG79. 244 APYHFSFPLDFLVKS<BR> 18 LGR7_HUMAN Relaxin receptor 1 (Leucine-rich repeat-containing G protein-coupled receptor 7). 734 KPDLFTYPCEMSLIS<BR> 19 AG2R_HUMAN Type-1 angiotensin II receptor (AT1) (AT1AR). 200 NILGFLFPFLIILTS<BR> Vascular non-inflammatory molecule 2 precursor (Vanin 2) (Glycosylphosphatidyl inositol-anchored<BR> 20 VNN2_HUMAN protein GPI-80) (FOAP-4 protein). 435 FGTEYVFPEVLLTEI<BR> 21 16904210 very large G protein-coupled receptor 1 [Mus musculus] 1971 PYGVFIFPNKTRPLS<BR> 22 VWF_HUMAN Von Willebrand factor precursor (vWF). 879 DGLKYLFPGECQYVL<BR> Williams-Beuren syndrome chromosome region 14 protein (WS basic-helix- loop-helix leucine zipper<BR> 23 WS14_HUMAN protein) (WS-bHLH) (Mlx interactor). 431 FSPRFPFPTYVPPAPG<BR> TABLE 4: INFLAMMATION<BR> Amino<BR> Accession Code Target Description Target Sequence<BR> Acid<BR> Adaptor-related protein complex 1, mu 2 subunit (Mu-adaptin 2) (Adaptor protein complex AP-1 mu-<BR> 2 subunit) (Golgi adaptor HA1/AP1 adaptin mu-2 subunit) (Clathrin assembly protein assembly<BR> 1 A1M2_HUMAN protein complex 1 medium chain 2) (AP-mu chain family member mu1B). 116 ELMDFGFPQTTDSKI<BR> 2 AIF1_HUMAN Allograft inflammatory factor-1 (AIF-1) (lonized calcium-binding adapter molecule 1) (G1). 97 SGETFSYPDFLRMML<BR> 3 APL3_HUMAN Apolipoprotein L3 (Apolipoprotein L-III) (ApoL-III) (TNF-inducible protein CG12-1) (CG12_1). 23 QVVTFTFPFGFQGIS<BR> 4 CABI_HUMAN Calcineurin-binding protein Cabin 1 (Calcineurin inhibitor) (CAIN). 947 FYCLYSFPSKKSKAR<BR> 5 CEBA_HUMAN CCAAT/enhancer binding protein alpha (C/EBP alpha). 102 GGGDFDYPGAPAGPG<BR> 6 CEBE_RAT CCAAT/enhancer binding protein epsilon (C/EBP epsilon) (C/EBP-related protein 1). 84 DPRPFAYPSHTFGPD<BR> 7 CO4_HUMAN Complement C4 precursor [Contains: C4A anaphylatoxin]. 915 ARGSFEFPVGDAVSK<BR> 8 CCR3_HUMAN C-X-C chemokine receptor type 3 (CXC-R3) (CXCR-3) (CKR-L2) (CD183 antigen). 201 THCQYNFPQVGRTAL<BR> 9 GAT2_HUMAN Endothelial transcription factor GATA-2. 167 GSHLFGFPPTPPKEV<BR> FYN-binding protein (FYN-T-binding protein) (FYB-120/130) (p120/p130) (SLP-76 associated<BR> 10 FYB_HUMAN phosphoprotein (SLAP-130). 125 SKPTFPWPPGNKPSL<BR> 11 28274770 HMG-box protein SOX21 [Mus musculus] 88 KKDKFAFPVPYGLGS<BR> 12 IKAP_HUMAN IkappaB kinase complex-associated protein (IKK complex-associated protein) (p 150). 599 FPVRFPYPCTQTELA<BR> Interferon regulatory factor 4 (IRF-4) (Lymphocyte specific interferon regulatory factor) (LSIRF) (NF-<BR> 13 IRF4_HUMAN EM5) (Multiple myeloma oncogene 1). 288 DQVLFPYPEDNGQRK<BR> 14 ILF1_HUMAN Interleukin enhancer-binding factor 1 (Cellular transcription factor ILF-1). 134 RVCTFRFPSTNIKIT<BR> 15 IRA2_HUMAN Interleukin-1 receptor-associated kinase-2 (EC 2.7.1.-) (IRAK-2). 280 QFHSFIYPYMANGSL<BR> 16 BAT2_HUMAN Large proline-rich protein BAT2 (HLA-B-associated transcript 2) (G2). 249 MMPPFMYPPYLPFPP Lipopolysaccharide-responsive and beige-like anchor protein (CDC4-like protein) (Beige-like<BR> 17 LRBA_HUMAN protein). 203 PDAFFNFPGKSAAAI<BR> Macrophage metalloelastase precursor (EC 3.4.24.65) (HME) (Matrix metalloproteinase-12) (MMP-<BR> 18 MM12_HUMAN 12) (Macrophage elastase) (ME). 367 SIHSFGFPNFVKKID<BR> NDRG4 protein (Brain development-related molecule 1) (Vascular smooth muscle cell associated<BR> 19 NDR4_HUMAN protein-8) (SMAP-8). 80 FPQGYQFPSMEQLAA<BR> 20 NOCT_HUMAN Nocturnin (CCR4 protein homolog). 404 RLPSFNYPSDHLSLV<BR> Nuclear factor NF-kappa-B p105 subunit (DNA-binding factor KBF1) (EBP- 1) [Contains: Nuclear<BR> 21 KBF1_HUMAN factor NF-kappa-B p50 subunit]. 405 SFPHYGFPTYGGITF<BR> Nuclear factor NF-kappa-B p105 subunit (DNA-binding factor KBF1) (EBP- 1) [Contains: Nuclear<BR> 22 KBF1_HUMAN factor NF-kappa-B p50 subunit]. 400 TGPGYSFPHYGFPTY<BR> Proteinase activated receptor 2 precursor (PAR-2) (Thrombin receptor-like 1) (Coagulation factor II<BR> 23 PAR2_HUMAN receptor-like 1). 247 AIGVFLFPAFLTASA<BR> 24 LGR7_HUMAN Relaxim receptor 1 (Leucine-rich repeat-containing G protein-coupled receptor 7). 734 KPDLFTYPCEMSLIS<BR> 25 RRA_HUMAN Retinoic acid receptor alpha (RAR-alpha). 22 PPYAFFFPPMLGGLS<BR> 26 ST5A_HUMAN Signal transducer and activator of transcription 5A. 664 SYLIYVFPDRPKDEV<BR> 27 ST5B_HUMAN Signal transducer and activator of transcription 5B. 664 NYLIYVFPDRPKDEV<BR> 28 SX12_HUMAN SOX-12 protein (SOX-22 protein). 281 GTSHFEFPDYCTPEV<BR> 29 15277232 TNFalpha-inducible ATP-binding protein [Homo sapiens] 570 YTVRFTFPDPPPLSP<BR> 30 TLR1_HUMAN Toll-like receptor 1 precursor (Toll/interleukin-1 receptor-like) (TIL). 308 VSDVFGFPQSYIYEI<BR> 31 TLR4_HUMAN Toll-like receptor 4 precursor (hToll). 71 SYSFFSFPELQVLDL<BR> 32 GAT3_HUMAN Trans-acting T-cell specific transcription factor GATA-3. 147 SPHLFTFPPTPPKDV<BR> 33 TF65_MOUSE Transcription factor p65 (Nuclear factor NF-kappa-B p65 subunit). 345 APQPYTFPASLSTIN<BR> 34 SX11_HUMAN Transcription factor SOX-11. 407 LGSHFEFPDYCTPEL<BR> 35 SX21_HUMAN Transcription factor SOX-21. 88 KKDKFAFPVPYGLGG<BR> 36 SOX4_HUMAN Transcription factor SOX-4. 440 SGSHFEFPDYCTPEV<BR> 37 SOX6_HUMAN Transcription factor SOX-6. 285 AQQGFLFPPGITYKP<BR> Translocon-associated protein, alpha subunit precursor (TRAP-alpha) (Signal sequence receptor<BR> 38 SSRA_HUMAN alpha subunit) (SSr-alpha). 120 LDASFRYPQDYQFY1<BR> Tumor necrosis factor receptor superfamily member 11B precursor (Osteoprotegerin)<BR> 39 T11B_HUMAN (Osteoclastogenesis inhibitory factor). 351 HSKTYHFPKTVTQSL<BR> Tumor necrosis factor receptor superfamily member 13B (Transmembrane activator and CAML<BR> 40 T13X_HUMAN interactor). 228 ETCSFCFPECRAPTQ<BR> 41 AG2R_HUMAN Type-1 angiotensin II receptor (AT1) (AT1AR). 200 NILGFLFPFLIILTS<BR> 42 JAK2_HUMAN Tyrosine-protein kinase JAK2 (EC 2.7.1.112) (Janus kinase 2) (JAK-2). 114 YRIRFYFPRWYCSGS<BR> Tyrosine-protein kinase JAK3 (EC 2.7.1.112) (Janus kinase 3) (JAK-3) (Leukocyte janus kinase) (L-<BR> 43 JAK3_HUMAN JAK). 100 YRIRFYFPNWFGLEK<BR> 44 VNN3_HUMAN Vascular non-inflammatory molecule 3 precursor (Vanin 3). 185 PEIQFDFPKDSELVT<BR> 45 VNN3_HUMAN Vascular non-inflammatory molecule 3 precursor (Vanin 3). 436 FGTRYVFPQIILSGS 46 VINE_HUMAN Vinexin (SH3-containing adaptor molecule-1) (SCAM-1). 565 RRTGFSFPTQEPRPQ<BR> 47 VINE_HUMAN Vinexin (SH3-containing adaptor molecule-1) (SCAM-1). 352 GRRDFVYPSSTRDPS<BR> Zinc finger protein 40 (Human immunodeficiency virus type I enhancer- binding protein 1) (HIV-EP1)<BR> (Major histocompatibility complex binding protein 1) (MBP-1) (Positive regulatory domain II binding<BR> 48 ZEP1_HUMAN factor 1) (PRDII-BF1). 1255 KSEKFSWPQRSETLS<BR> TABLE 5: METABOLIC DISORDERS<BR> Accession Code Amino<BR> Target Description Target Sequence<BR> Acld<BR> 1-phosphatidylinositol-4,5-bisphosphate phosphodiesterase beta 3 (EC 3.1.4.11) (Phosphoinositide<BR> 1 PIP3_HUMAN phospholipase C) (PLC-beta-3) (Phospholipase C-beta-3). 777 DEEPFDFPKVVLPTL<BR> 2 AAK1_HUMAN 5'-AMP-activated protein kinase, catalytic alpha-1 chain (EC 2.7.1.-) (AMPK alpha-1 chain). 273 DLPKYLFPEDPSYSS<BR> 3 AAK2_HUMAN 5'-AMP-activated protein kinase, catalytic alpha-2 chain (EC 2.7.1.-) (AMPK alpha-2 chain). 271 DLPSYLFPEDPSYDA<BR> 4 ACDV_HUMAN Acyl-CoA dehydrogenase, very-long-chain specific, mitochondrial precursor (EC 1.3.99.-) (VLCAD). 84 TDQVFPYPSVLNEEQ<BR> 5 ANDR_HUMAN Androgen receptor (Dihydrotestosterone receptor). 359 SRDYYNFPLALAGPP<BR> 6 ANDR_HUMAN Androgen receptor (Dihydrotestosterone receptor). 547 LPIDYYFPPQKTCLI<BR> 7 APB_HUMAN Apolipoprotein B-100 precursor (Apo B-100) [Contains: Apolipoprotein B-48 (Apo B-48)]. 4198 NFPRFQFPGKPGIYT<BR> 8 APL3_HUMAN Apolipoprotein L3 (Apolipoprotein L-III) (ApoL-III) (TNF-inducible protein CG12-1) (CG12 1). 23 QVVTFTFPFGFQGIS<BR> Aquaporin-CHIP (Water channel protein for red blood cells and kidney proximal tubule) (Aquaporin<BR> 9 AQP1_HUMAN 1) (AQP-1) (Urine water channel). 31 SALGFKYPVGNNQTA<BR> ATP-binding cassette, sub-family A, member 1 (ATP-binding cassette transporter 1) (ATP-binding<BR> 10 ABC1_HUMAN cassette 1) (ABC-1) (Cholesterol efflux regulatory protein). 78 NNPCFRYPTPGEAPG<BR> 11 ABF2_HUMAN ATP-binding cassette, sub-family F, member 2 (Iron inhibited ABC transporter 2) (HUSSY-18). 379 KTLSFYFPPCGKIPP<BR> Beta-hexosaminidase alpha chain precursor (EC 3.2.1.52) (N-acetyl- beta-glucosaminidase) (Beta-N.<BR> <P>12 HEXA_HUMAN acetylhexosaminidase) (Hexosaminidase A). 212 PYESFTFPELMRKGS<BR> 13 AB11_HUMAN Bile salt export pump (ATP-binding cassette, sub-family B, member 11). 1081 VDCKFTYPSRPDSQV<BR> 14 BIEA_HUMAN Biliverdin reductase A precursor (EC 1.3.1.24) (Biliverdin-IX alpha-reductase). 158 EEERFGFPAFSGISR<BR> 15 CEBA_BOVIN CCAAT/enhancer binding protein alpha (C/EBP alpha). 27 SSAAFGFPRGAGPSQ<BR> 16 CEBA_HUMAN CCAAT/enhancer binding protein alpha (C/EBP alpha). 27 SSAAFGFPRGAGPAQ<BR> 17 CEBE_HUMAN CCAAT/enhancer binding protein epsilon (C/EBP epsilon). 84 DPRPFAYPPHTFGPD<BR> 18 13562153 channel-kinase 1 [Homo sapiens] 1339 QRKEFNFPEAGSSSG<BR> 19 CLC3_HUMAN Chloride channel protein 3 (ClC-3). 236 NIFSYLFPKYSTNEA<BR> 20 CLC3_HUMAN Chloride channel protein 3 (ClC-3). 280 EEVSYYFPLKTLWRS<BR> 21 CLC6_HUMAN Chloride channel protein 6 (ClC-6). 223 RKIQFNFPYFRSDRD<BR> 22 ClCL_HUMAN Chloride channel protein CLC-KB (ClC-K2). 442 ETLSFIFPEGIVAGG<BR> 23 CLI6_HUMAN Chloride intracellular channel 6. 638 KYRDFEFPSEMTGIW<BR> 24 CETP_HUMAN Cholesteryl ester transfer protein precursor (Lipid transfer protein I). 474 LQMDFGFPEHLLVDF Cyclic-nucleotide-gated cation channel 4 (CNG channel 4) (CNG-4) (CNG4) (Cyclic nucleotide-gated<BR> 25 CNG4_HUMAN cation channel modulatory subunit). 296 PWKKYQFPQSIDPIT<BR> 26 C561_HUMAN Cytochrome b561 (Cytochrome b-561). 139 GFSFFLFPGASFSLR<BR> 27 CX42_HUMAN Cytochrome c oxidase subunit IV isoform 2, mitochondrial precursor (EC 1.9.3.1) (COX IV-2). 122 WQRVYVFPPKPITLT<BR> Cytochrome P450 27, mitochondrial precursor (EC 1.14.-.-) (Cytochrome P-450C27/25) (Sterol 26-<BR> hydroxylase) (Sterl 27-hydroxylase) (Vitamin D(3) 25-hydroxylase) (5-beta-cholestane-3-alpha, 7-<BR> 28 CP27_HUMAN alpha, 12-alpha-triol 27-hydroxylase). 412 EVDGFLFPKNTQFVF<BR> 29 12832024 dJ686C3.1.3 (isocitrate dehydrogenase 3 (NAD+) bela (isoform C)) [Homo sapiens] 7 WSSLFPFPVSPSCCF<BR> Ectonucleoside triphosphate diphosphohydrolase 4 (EC 3.6.1.6) (NTPDase4) (Uridine-<BR> 30 ENP4_HUMAN diphosphatase (UDPase) (Lysosomal apyrase-like protein of 70 kDa). 500 FHRGFSFPVNYKSLK<BR> Excitatory amino acid transporter 1 (Sodium-dependent glutamate/aspartate transporter 1) (Glial<BR> 31 EAA1_HUMAN glutamate transporter (GLAST-1). 77 EVKYFSFPGELLMRM<BR> 32 EAA4_HUMAN Excitatory amino acid transporter 4 (Sodium-dependent glutamate/aspartate transporter). 85 QIKYFSFPGELLMRM<BR> 33 EAA5_HUMAN Excitatory amino acid transporter5 (Retinal glutamate transporter). 46 EISYFQFPGELMRM<BR> Fructose-1,6-bisphosphatase (EC 3.1.3.11) (D-fructose-1,6bisphosphate 1-phosphohydrolase)<BR> 34 F16P_HUMAN (FBPase). 258 YGGIFLYPANKKSPN<BR> Fructose-1,6-bisphosphatase isozyme 2 (EC 3.1.3.11) (D-fructose-1,6- bisphosphate 1-<BR> 35 F16Q_HUMAN phosphohydrolase) (FBPase). 259 YGGIFLYPANQKSPK<BR> Galactosylceramide sulfotransferase (EC 2.8.2.11) (GalCer sulfotransferase) (Cerebroside<BR> sulfotransferase) (3'- phosphoadenylylsulfate:galactosylceramide 3'-sulfotransferase) (3'-<BR> 36 CST_HUMAN phosphoadenosine-5'phosphosulfate: GalCer sulfotransferase). 102 HRLKFAFPNGRNDFD<BR> Glandular kallikrein 1 precursor (EC 3.4.21.35) (Tissue kallikrein) (Kidney/pancreas/salivary gland<BR> 37 KLK1_HUMAN kallikrein). 162 EPENFSFPDDLQCVD<BR> Glutamate decarboxylase, 65 kDa isoform (EC 4.1.1.15) (GAD-65) (65 kDa glutamic acid<BR> 38 DCE2_HUMAN decarboxylase). 131 KVIDFHYPNELLQEY<BR> 39 GSHR_HUMAN Glutathione reductase, mitochondrial precursor (EC 1.8.1.7) (GR) (GRase). 125 DHADYGFPSCEGKFN<BR> Glycine dehydrogenase [decarboxylating], mitochondrial precursor (EC 1.4.4.2) (Glycine<BR> 40 GCSP_HUMAN decarboxylase) (Glycine cleavage system P- protein). 260 SGVLFQYPDTEGKVE<BR> Glycogen debranching enzyme (Glycogen debrancher) [Includes: 4-alpha- glucanotransferase (EC<BR> 2.4.1.25) (Oligo-1,4-1,4-glucantransferase); Amylo-alpha-1,6-glucosidase (EC 3.2.1.33) (Amylo-1,6-<BR> 41 GDE_HUMAN glucosidase) (Dextrin 6-alpha-D-glucosidase)]. 426 VTRYFTFPFEEIDFS<BR> 42 KG3A_HUMAN Glycogen synthase kinase-3 alpha (EC 2.7.1.37) (GSK-3 alpha). 350 NYTEFKFPQIKAHPW<BR> 43 SYG_HUMAN Glycyl-tRNA synthetase (EC 6.1.1.14) (Glycine--tRNA ligase) (GlyRS). 601 QRTFFSFPAVVAPFK<BR> Guanine deaminase (EC 3.5.4.30 (Guanase) (Guanine aminase) (Guanine aminohydrolase) (GAH)<BR> 44 GUAD_HUMAN (p51-nedasin). 102 WLTKYTFPAEHRFQN<BR> 45 HO1_HUMAN Heme oxygenase 1 (EC 1.14.99.3) (HO-1). 163 GLAFFTFPNIASATK<BR> Hepatocyte nuclear factor 1-beta (HNF-1B) (Variant hepatic nuclear factor 1) (VHNF-1)<BR> 46 HNFB_HUMAN (Homeoprotein LFB3) (Transcription factor 2) (TCF-2). 200 DQLLFLFPEFSQQSH<BR> 47 HXK4_HUMAN Hexokinase D (EC 2.7.1.1) (Hexokinase type IV) (HK IV) (HK4) (Glucokinase). 146 LGFTFSFPVRHEDID 48 HXK3_HUMAN Hexokinase type III (EC 2.7.1.1) (HK III). 163 LGFSFSFPCHQTGLD<BR> 49 HXK1_HUMAN Hexokinase, type I (EC 2.7.1.1) (HK I) (Brain form hexokinase). 598 LGFTFSFPCQQTSLD<BR> 50 HXK2_HUMAN Hexokinase, type II (EC 2.7.1.1) (HK II) (Muscle form hexokinase). 598 LGFTFSFPCQQNSLD<BR> 51 IKAP_HUMAN IkappaB kinase complex-associated protein (IKK complex-associated protein) (p150). 599 FPVRFPYPCTQTELA<BR> 52 IRA2_HUMAN Interleukin-1 receptor-associated kinase-2 (EC 2.7.1.-) (IRAK-2). 280 QFHSFIYPYMANGSL<BR> 53 OBRG_HUMAN Leptin receptor gene-related protein (OB-R gene related protein) (OB-RGRP). 78 VVSAFGFPVILARVA<BR> 54 LIPL_HUMAN Lipoprotein lipase precursor (EC 3.1.1.34) (LPL). 142 MEEEFNYPLDNVHLL<BR> 55 LRP2_HUMAN Low-density lipoprotein receptor-related protein 2 precursor (Megalin) (Glycoprotein 330) (gp330). 1025 QCGLFSFPCKNGRCV<BR> 56 LOL1_HUMAN Lysyl oxidase homolog 1 precursor (EC 1.4.3.-) (Lysyl oxidase-like protein 1) (LOL). 173 YPQQFPYPQAPFVSQ<BR> 57 MDHC_HUMAN Malate dehydrogenase, cytoplasmic (EC 1.1.1.37). 281 DDLLYSFPVVIKNKT<BR> 58 MUTA_HUMAN Methylmalonyl-CoA mutase, mitochondrial precursor (EC 5.4.99.2) (MCM). 227 VRNTYIFPPEPSMKI<BR> Mitochondrial 28S ribosomal protein S29 (S29mt) (MRP-S29) (Death- associated protein 3) (DAP-3)<BR> 59 RT29_HUMAN (Ionizing radiation resistance conferring protein). 113 KNTSFAYPAIRYLLY<BR> 60 14141575 mitochondrial ribosomal protein L23 [Homo sapiens] 2 HGQTFTFPDLFPEKD<BR> 61 MOT1_HUMAN Monocarboxylate transporter 1 (MCT 1). 30 IGFSYAFPKSITVFF<BR> 62 MOT2_HUMAN Monocarboxylate transporter 2 (MCT 2). 30 IGFSYAFPKAVTVFF<BR> 63 MOT3_HUMAN Monocarboxylate transporter 3 (MCT 3). 29 TGFAYGFPKAVSVFF<BR> 64 MOT4_HUMAN Monocarboxylate transporter 4 (MCT 4) (MCT 3). 32 TGFSYAFPKAVSVFF<BR> 65 MYHD_HUMAN Myosin heavy chain, skeletal muscle, extraocular (MyHC-eo). 306 STNPFDFPFVSQGEV<BR> N-acetylglucosaminyl-phosphatidylinositol biosynthetic protein (GlcNAc-Pl synthesis protein)<BR> 66 PIGA_HUMAN (Phosphatidylinositol-glycan biosynthesis, class A protein) (PIG-A). 37 MVSDFFYPNMGGVES<BR> NADH-ubiquinone oxidoreductase B18 subunit (EC 1.6.5.3) (EC 1.6.99.3) (Complex I-B18) (Cl-B18)<BR> 67 NB8M_HUMAN (Cell adhesion protein SQM1). 24 FPPDYGFPERKEREM<BR> 68 NI2M_HUMAN NADH-ubiquinone oxidoreductase B22 subunit (EC 1.6.5.3) (EC 1.6.99.3) (Complex I-B22) (Cl-B22). 75 HPQPYIFPDSPGGTS<BR> 69 MAOX_HUMAN NADP-dependent malic enzyme (EC 1.1.1.40) (NADP-ME) (Malic enzyme 1). 452 GNNSYVFPGVALGVV<BR> 70 NPH4_HUMAN Nephrocystin 4 (Nephroretinin). 669 TFQFYRFPPATTPRL<BR> 71 NCO2_HUMAN Nuclear receptor coactivator 2 (NCoA-2) (Transcriptional intermediary factor 2). 1295 NAQQFPFPPNYGISQ<BR> 72 NCR1_HUMAN Nuclear receptor co-repressor 1 (N-CoR1) (N-CoR). 23 HSVQYTFPNTRHQQE<BR> Nuclear receptor co-repressor 2 (N-CoR2) (Silencing mediator of retinoic acid and thyroid hormone<BR> receptor) (SMRT) (SMRTe) (Thyroid-, retinoic-acid-receptor-associated co-repressor) (T3 receptor-<BR> 73 NCR2_HUMAN associating factor) (TRAC) (CTG repeat protein 26). 2160 PAPLYSFPGASCPVL<BR> 74 RORG_HUMAN Nuclear receptor ROR-gamma (Nuclear receptor RZR-gamma). 540 PPSPFSFPMNPGGWS<BR> 75 STT3_HUMAN Oligosaccharyl transferase STT3 subunit homolog (B5) (Integral membrane protein 1) (TMC). 365 QLLVFMFPVGLYYCF<BR> 76 PNK4_HUMAN Pantothenate kinase 4 (EC 2.7.1.33) (Pantothenic acid kinase 4) (hPanK4). 513 CLNEFNFPDPYSKVK<BR> 77 PPAR_HUMAN Peroxisome proliferator activated receptor alpha (PPAR-alpha). 417 PDDIFLFPKLLQKMA<BR> Peroxisome proliferator activated receptor delta (PPAR-delta) (PPAR-beta) Nuclear hormone<BR> 78 PPAS_MOUSE receptor 1) (NUC1). 389 PDSQYLFPKLLQKMA Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit, delta isoform (EC 2.7.1.153) (PI3-<BR> 79 P11D_HUMAN kinase p110 subunit delta) (Ptdlns- 3-kinase p110) (PI3K) (p110delta). 581 ELLDFSFPDCHVGSF<BR> 80 PIGO_HUMAN Phosphatidylinositol-glycan biosynthesis, class O protein (PIG-O). 190 FSKAFFFPSFNVRDL<BR> 81 940231 phosphodiesterase A' subunit [Homo sapiens] 366 ADEYFTFPKGPVDET<BR> Phosphoribosyl pyrophosphate synthetase-associated protein 1 (PRPP synthetase-associated<BR> 82 KPRA_HUMAN protein 1) (39 kDa phosphoribosypyrophosphate synthetase-associated protein) (PAP39). 140 IQGFFSFPVDNLRAS<BR> 83 PIT1_HUMAN Pituitary-specific positive transcription factor 1 (Pit-1) (Growth hormone factor 1) (GHF-1). 75 TPCLYKFPDHTLSHG<BR> 84 PKHD_HUMAN Polycystic kidney and hepatic disease 1 precursor (Fibrocystin) (Polyductin) (Tigmin). 3321 DKNKFYFPSLQPRKD<BR> 85 PKHD_HUMAN Polycystic kidney and hepatic disease 1 precursor (Fibrocystin) (Polyductin) (Tigmin). 2512 NLVAFPFPHAAILED<BR> 86 PKHD_HUMAN Polycystic kidney and hepatic disease 1 precursor (Fibrocystin) (Polyductin) (Tigmin). 2370 RYGLFVYPKFQPPWD<BR> 87 P1L1_HUMAN Polycystic kidney disease 1-like 1 protein (Polycystin 1L1). 2687 PGLLFHFPRRSQKDC<BR> Polycystic kidney disease and receptor for egg jelly related protein precursor (PKD and REJ<BR> 88 PKDR_HUMAN homolog). 1880 GYALYFFPEQQRFNS<BR> 89 PKD1_HUMAN Polycystin 1 precursor (Autosomal dominant polycystic kidney disease protein 1). 3060 SHVRFVFPEPTADVN<BR> Polypeptide N-acetylgalactosaminyltransferase (EC 2.4.1.41) (Protein- UDP<BR> acetylgalactosaminyltransferase) (UDP-GalNAc:polypeptide, N- acetylgalactosaminyltransferase)<BR> 90 PAGT_HUMAN (GalNAc-T1). 348 KATPYTFPGGTGQII<BR> 91 DP1_HUMAN Polyposis locus protein 1 (TB2 protein). 56 NLIGFGYPAYISIKA<BR> 92 CIK1_HUMAN Potassium voltage-gated channel subfamily A member 1 (Potassium channel Kv1.1) (HUKI) (HBK1). 153 VWLLFEYPESSGPAR<BR> Potassium voltage-gated channel subfamily A member 5 (Potassium channel Kv1.5) (HK2)<BR> 93 CIK5_HUMAN (HPCN1). 236 VWLIFEYPESSGSAR<BR> Potassium voltage-gated channel subfamily H member 7 (Ether-a-go-go related gene potassium<BR> 94 KCH7_HUMAN channel 3) (HERG-3) (Ether-a-go-go related protein 3) (Eag related protein 3). 156 KFFGFKFPGLRVLTY<BR> 95 S24C_HUMAN Protein transport protein Sec24C (SEC24-related protein C). 942 ETNVFFYPRLLPLTK<BR> 96 6007826 rab escort protein-2 [Mus musculus] 370 GNTPFIFPLYGHGEI<BR> Renal sodium-dependent phosphate transport protein 2 (Sodium/phosphate cotransporter 2)<BR> (Na(+)/Pi cotransporter 2) (Renal sodium-phosphate transport protein 2) (Renal Na(+)-dependent<BR> 97 NPT2_HUMAN phosphate cotransporter 2). 43 GTSAYAFPSLGPVAL<BR> 98 RRA_HUMAN Retinoic acid receptor alpha (RAR-alpha). 22 PPYAFFFPPMLGGLS<BR> 99 RRG1_HUMAN Retinoic acid receptor gamma-1 (RAR-gamma-1) 23 AGFPFAFPGALRGSP<BR> 100 RRG1_HUMAN Retinoic acid receptor gamma-1 (RAR-gamma-1). 370 PSQPYMFPRMLMKIT<BR> 101 12053793 retinoid-acid induced protein 1 [Homo sapiens] 1830 PISLFSFPPLLPQQF<BR> Sialic acid binding Ig-like lectin 5 precursor (Siglec-5) (Obesity- binding protein 2) (OB binding<BR> 102 SIL5_HUMAN protein-2) (OB-BP2) (CD33 antigen-like 2) (CD170 antigen). 39 VPCSFSYPWRSWYSS<BR> 103 S6A9_HUMAN Sodium- and chloride-dependent glycine transporter 1 (GlyT1) (GlyT-1). 120 GGGAFMFPYFIMLIF<BR> Sodium/bile acid cotransporter (Na(+)/bile acid cotransporter) (Na(+)/taurocholate transport protein)<BR> 104 NTCP_HUMAN (Sodium/taurocholate cotransporting polypeptide). 279 IGPLFFFPLLYMIFQ 105 NAH5_HUMAN Sodium/hydrogen exchanger 5 (Na(+)/H(+) exchanger 5) (NHE-5). 835 PRSSFAFPPSLAKAG<BR> 106 S6A7_HUMAN Sodium-dependent proline transporter. 525 EYGSYRFPPWAELLG<BR> Solute carrier family 21 member 11 (Sodium-independent organic anion transporter D) (Organic<BR> anion transporting polypeptide D) (OATP-D) (Organic anion transporter polypeptide related protein<BR> 107 S21B_HUMAN 3) (OATP-RP3) (OATPRP3) (PGE1 transporter). 279 SLLMFGFPQSLPPHS<BR> 108 SYTA_MOUSE Synaptotagmin X (SytX). 296 FDELFQFPVVYDQLS<BR> 109 SYTB_HUMAN Synaptotagmin XI (SytXI). 163 FSVDYNFPKKALVVT<BR> Thyroid hormone receptor-associated protein complex 240 kDa component (Trap240) (Activator-<BR> 110 T240_HUMAN recruited cofactor 250 kDa component) (ARC250). 703 GDEEFLFPDKKDRQN<BR> Thyroid hormone receptor-associated protein complex 240 kDa component (Trap240) (Activator-<BR> 111 T240_HUMAN recruited cofactor 250 kDa component) (ARC250). 614 AWKYYKFPKKKDVEF<BR> 112 TRIB_HUMAN Thyroid receptor interacting protein 12 (TRIP 12). 1536 KTCPFFFPFDTRQML<BR> 113 15277232 TNFalpha-inducible ATP-binding protein [Homo sapiens] 570 YTVRFTFPDPPPLSP<BR> Transitional endoplasmic reticulum ATPase (TER ATPase) (15S Mg(2+)- ATPase p97 subunit)<BR> 114 TERA_HUMAN (Valosin containing protein) (VCP) [Contains: Valosin]. 767 GFGSFRFPSGNQGGA<BR> 115 TUL2_HUMAN Tubby related protein 2 (Tubby-like protein 2). 498 FTMDFCFPFSPLQAF<BR> 116 TUSP_HUMAN Tubby superfamily protein. 1518 YILDFQYPFSAVQAF<BR> Tumor necrosis factor receptor superfamily member 11B precursor (Osteoprotegerin)<BR> 117 T11B_HUMAN (Osteoclastogenesis inhibitory factor) 351 HSKTYHFPKTVTQSL<BR> Tumor necrosis factor receptor superfamily member 13B (Transmembrane activator and CAML<BR> 118 T13X_HUMAN interactor). 228 ETCSFCFPECRAPTQ<BR> 119 JAK2_HUMAN Tyrosine-protein kinase JAK2 (EC2.7.1.112) (Janus kinase 2) (JAK-2). 114 YRIRFYFPRWYCSGS<BR> UDP-glucuronosyltransferase 2B15 precursor, microsomal (EC2.4.1.17) (UDPGT) (UDPGTH-3)<BR> 120 UDBF_HUMAN (HLUG4). 184 NGGGFLFPPSYVPVV<BR> Vasopressin V1b receptor (V1bR) (AVPR V1b) (Vasopressin V3 receptor) (AVPR V3) (Antidiuretic<BR> 121 V1BR_HUMAN hormone receptor 1b). 186 CWADFGFPWGPRAYL<BR> 122 16904210 very large G protein-coupled receptor 1 [Mus musculus] 1971 PYGVFIFPNKTRPLS<BR> 123 WRN_HUMAN Werner syndrome helicase. 577 KSLCFQYPPVYVGKI<BR> 124 WFS1_HUMAN Wolframin. 413 FFVIFSFPIASKDCI<BR> Zinc finger protein 44 (Zinc finger protein KOX7) (Gonadotropin inducible transcription repressor-2)<BR> 125 ZN44_HUMAN (GIOT-2). 342 CGKGFDFPGSARIHE<BR> TABLE 6: NEUROPATHIES<BR> Amino<BR> Accession Code Target Description Target Sequence<BR> Acid<BR> 5-hydroxytryptamine 1B receptor (5-HT-1B) (Serotonin receptor) (5-HT-1D-beta) (Serotonin 1D beta<BR> 1 5H1B_HUMAN receptor) (S12). 213 TVGAFYFPTLLLIAL<BR> 2 5H2C_HUMAN 5-hydroxytryptamine 2C receptor (5-HT-2C) (Serotonin receptor) (5HT-1C). 42 DGGRFKFPDGVQNWP 5-hydroxytryptamine 3 receptor precursor (5-HT-3) (Serotonin-gated ion channel receptor) (5-<BR> 3 5HT3_HUMAN HT3R). 158 SLDIYNFPFDVQNCS<BR> 4 AUT2_HUMAN Autism susceptibility gene 2 protein. 1057 GGERFPYPSFHWDPI<BR> 5 BIR1_HUMAN Baculoviral IAP repeat-containing protein 1 (Neuronal apoptosis inhibitory protein). 649 CAHWFQYPFDPSFDD<BR> 6 BRA1_HUMAN Brain link protein-1 precursor. 318 GVRSFGFPRPQQAAY<BR> 7 BAI1_HUMAN Brain-specific angiogenesis inhibitor 1 precursor. 341 ELQQFGFPAPQTGDP<BR> 8 CBL2_HUMAN Cdk5 and abl enzyme substrate 2 (Interactor with cdk3 2) (Ik3-2). 241 SYAKFLYPTNALVTH<BR> 9 DYHB_HUMAN Ciliary dynein heavy chain 11 (Axonemal beta dynein heavy chain 11). 2649 TVFAFNFPSLDALNT<BR> Dimethylaniline monooxygenase [N-oxide forming] 5 (EC 1.14.13.8) (Hepatic flavin-containing<BR> 10 FMO5_HUMAN monooxygenase 5) (FMO 5) (Dimethylaniline oxidase 5). 329 TGYSFDFPFLEDSVK<BR> 11 DIS1_HUMAN Disrupted in schizophrenia 1 protein. 61 VGTLFRFPGGVSGEE<BR> 12 DOPO_MOUSE Dopamine beta-monooxygenase precursor (EC 1.14.17.1) (Dopamine beta- hydroxylase) (DBH). 309 GAKAFYYPKEAGVPF<BR> 13 DSCA_HUMAN Down syndrome cell adhesion molecule precursor (CHD2). 80 TLQIFPFPPSSFSTL<BR> 14 DSCA_HUMAN Down syndrome cell adhesion molecule precursor (CHD2). 598 FIQPFEFPRFSIGQR<BR> 15 DSR3_HUMAN Down syndrome critical region protein 3 (Down syndrome critical region protein A). 89 TEIPFEFPLHLKGNK<BR> 16 DYSF_HUMAN Dysferlin (Dystrophy associated fer-1-like protein) (Fer-1 like protein 1). 1867 FNWRFIFPFDYLPAE<BR> Ephrin type-A receptor 7 precursor (EC 2.7.1.112) (Tyrosine-protein kinase receptor EHK-3) (Eph<BR> 17 EPA7_HUMAN homology kinase-3) (Receptor protein- tyrosine kinase HEK11). 596 LYFHFKFPGTKTYID<BR> 18 ERC6_HUMAN Excision repair protein ERCC-6 (Cockayne syndrome protein CSB). 687 SLFDFIFPGKLGTLP<BR> Excitatory amino acid transporter 3 (Sodium-dependent glutamate/aspartate transporter 3)<BR> 19 EAA3_HUMAN (Excitatory amino-acid carrier 1) (Neuronal and epithelial glutamate transporter). 48 EKFYFAFPGEILMRM<BR> Forkhead box protein G1A (Forkhead-related protein FKHL2) (Transcription factor BF-2) (Brain<BR> 20 FXGA_HUMAN factor 2) (BF2) (HFK2). 391 GQTSYFFPHVPHPSM<BR> Fragile X mental retardation 2 protein (Protein FMR-2) (FMR2P) (Ox19 protein) (Fragile X E mental<BR> 21 FMR2_HUMAN retardation syndrome protein). 639 STDEFTWPKPNITSS<BR> Fragile X mental retardation 2 protein (Protein FMR-2) (FMR2P) (Ox19 protein) (Fragile X E mental<BR> 22 FMR2_HUMAN retardation syndrome protein). 193 LEDFFVYPAEQPQIG<BR> 23 FCMD_HUMAN Fukutin precursor (Fukuyama-type congenital muscular dystrophy protein). 388 KKFKYLFPKFTLCWT<BR> 24 GAE_HUMAN Gamma-aminobutyric-acid receptor epsilon subunit precursor (GABA(A) receptor). 212 SFSSFSYPENEMIYK<BR> 25 GAAT_HUMAN Gamma-aminobutyric-acid receptor theta subunit precursor (GABA(A) receptor). 609 KWSRFLFPLAFGLFN<BR> 26 SGCG_HUMAN Gamma-sarcoglycan (Gamma-SG) (35 kDa dystrophin-associated glycoprotein) (35DAG). 81 GESEFLFPLYAKEIH<BR> 27 KG3A_HUMAN Glycogen synthase kinase-3 alpha (EC 2.7.1.37) (GSK-3 alpha). 350 NYTEFKFPQIKAHPW<BR> 28 17384611 kainate receptor subunit KA2a [Homo sapiens] 945 DASSFFFPPISSCPP<BR> 29 OPRK_HUMAN Kappa-type opioid receptor (KOR-1). 340 CFRDFCFPLKMRMER<BR> 30 13383464 kinesin-related protein HASH [Mus musculus] 46 NTRDFMFPGPNQMSQ<BR> 31 MGR1_HUMAN Metabotropic glutamate receptor 1 precursor (mGluR1). 3 GLLLFFFPAIFLEVS<BR> 32 9988950 monooxygenase X [Homo sapiens] 9 RYPDFSFPYFPQDYF<BR> 33 MY15_HUMAN Myosin XV (Unconventional myosin-15). 751 RGAAFGFPGASPRAS 34 MTR2_HUMAN Myotubularin-related protein 2. 178 NLMKYAFPVSNNLPL<BR> 35 2736151 mytonic dystrophy kinase-related Cdc42-binding kinase [Rattus norvegicus] 297 HKERFQFPTQVIDVS<BR> Neural cell adhesion molecule 1, 120 kDa isoform precursor (N-CAM 120) (NCAM-120) (CD56<BR> 36 NCA2_HUMAN antigen). 425 TCEVFAYPSATISWF<BR> 37 NTC3_HUMAN Neurogenic locus notch homolog protein 3 precursor (Notch 3). 1618 ERLDFPYPLRDVRGE<BR> Neurogenin 1 (Neurogenic differentiation factor 3) (NeuroD3) (Neurogenic basic-helix-loop-helix<BR> 38 NGN1_HUMAN protein). 213 GDPVFSFPSLPKDLL<BR> Neurotensin receptor type 1 (NT-R-1) (High-affinity levocabastine- insensitive neurotensin receptor)<BR> 39 NTR1_HUMAN (NTRH). 241 TFMSFIFPMVVISVL<BR> Neutral amino acid transporter B (0) (ATB(0)) (Sodium-dependent neutral amino acid transporter<BR> 40 AAAT_HUMAN type 2) (RD114/simian type D retrovirus receptor) (Baboon M7 virus receptor). 85 RLSAFVFPGELLLRL<BR> 41 OAA3_HUMAN Olfactory receptor 10A3 (HTPCRX12). 160 TTWVFSFPFCGPNEI<BR> 42 OAA5_HUMAN Olfactory receptor 10A5 (HP3) (Olfactory receptor-like protein JCG6). 161 TTWLFSFPFCGTNKV<BR> 43 OXB2_HUMAN Olfactory receptor 51B2 (HOR5'beta3). 275 SYIYFLFPPLMNPVI<BR> 44 OXB4_HUMAN Olfactory receptor 51B4 (HOR5'beta1). 274 SYVHFLFPPFVNPII<BR> 45 OXI1_HUMAN Olfactory receptor 51I1 (HOR5'beta11). 155 FTTLFPFPFVVKRLP<BR> 46 O6B1_HUMAN Olfactory receptor 6B1 (Olfactory receptor 7-3) (OR7-3). 203 ALVIFLFPLFITVLS<BR> 47 PAN1_HUMAN Pannexin 1. 187 SESHFKYPIVEQYLK<BR> 48 RELN_HUMAN Reelin precursor (EC 3.4.21.-). 1894 LMDEFYFPQTTNILF<BR> 49 RELN_HUMAN Reelin precursor (EC 3.4.21.-). 1967 EDNWFFYPGGNIGLY<BR> 50 SACS_HUMAN Sacsin. 1944 NCTMFRFPLRNAEMA<BR> SAM domain and HD domain-containing protein 1 (Dendritic cell-derived IFNG-induced protein)<BR> 51 SAD1_HUMAN (DCIP) (Monocyte protein 5) (MOP-5). 151GGGYYVFPGASHNRF<BR> 52 KPT3_HUMAN Serine/threonine protein kinase PCTAIRE-3 (EC 2.7.1-). 371 EFRTYSFPCYLPQPL<BR> 53 SAMP_HUMAN Serum amyloid P-component precursor 9SAP) (9.5S alpha-1-glycoprotein). 24 SGKVFVFPRESVTDH<BR> SH3 and multiple ankyrin repeat domains protein 2 (Shank2) (Proline- rich synapse associated<BR> protein 1) (ProSAP1) (Cortactin-binding protein 1) (CortBP1) (GKAP/SAPAP interacting protein)<BR> 54 SHK2_RAT (SPANK-3). 131 SLSTFEYPGPRRKLY<BR> similar to dJ309K20.4 (KIAA0765, putative brain nuclearly targeted protein (HRIHFB2091, RNA<BR> 55 20073220 recognition motif (RNP, RRM or RBD domain) containing protein)) [Mus musculus] 631 NGPPFNFPGNFGGPN<BR> 56 20071167 Similar to Per1 interacting protein [Mus musculus] 551 PMNPFRFPKEAASLF<BR> 57 S6A1_HUMAN Sodium- and chloride-dependent GABA transporter 1. 523 TMGNYVFPKWGQGV@<BR> 58 SX14_HUMAN Transcription factor SOX-14. 88 KKDRYVFPLPYLGDT<BR> Tyrosine-protein kinase transmembrane receptor ROR1 precursor (EC 2.7.1.112) (Neurotrophic<BR> 59 ROR1_HUMAN tyrosine kinase, receptor-related 1). 785 RYPNYMFPSQGITPQ<BR> Tyrosine-protein kinase transmembrane receptor ROR1 precursor (EC 2.7.1.112) (Neurotrophic<BR> 60 ROR1_HUMAN tyrosine kinase, receptor-related 1). 226 SLCHYAFPYCDETSS<BR> Tyrosine-protein kinase transmembrane receptor ROR2 precursor (EC 2.7.1.112) (Neurotrophic<BR> 61 ROR2_HUMAN tyrosine kinase, receptor-related 2). 230 SFCHFVFPLCDARSR Williams-Beuren syndrome chromosome region 14 protein (WS basic-helix- loop-helix leucine zipper<BR> 62 WS14_HUMAN protein) (WS0-bHLH) (Mix interactor). 431 FSPRFPFPTVPPAPG<BR> 63 WFS1_HUMAN Wolframin. 413 FFVIFSFPIASKDCI<BR> TABLE 7: MISCELLANEOUS DEF DOMAIN-CONTAINING PROTEINS<BR> Amino<BR> Accession Code Target Description Target Sequence<BR> Acid<BR> 1 15425674 Per1 interacting protein of the suprachiamatic nucleus [Rattus norvegicus] 1023 SLNPFRFPKEAASLF<BR> 2 PER1_HUMAN Period circadian protein 1 (Circadian pacemaker protein Rigui) (hPER). 922 VLPNYLFPTPSSYPY<BR> 3 PER2_HUMAN Period circadian protein 2. 907 MLPSYSFPSGTPNLP<BR> 4 PER3_HUMAN Period circadian protein 3 (hPER3). 843 PYPAFPFPYLDTFMT<BR> A-kinase anchor protein 11 (Protein kinase A anchoring protein 11) (PRKA11) (A kinase anchor<BR> 5 AK11_HUMAN protein 220 kDa) (AKAP 220) (hAKAP220). 661 EVCQFSYPQTPASPQ<BR> A-kinase anchor protein 3 (Protein kinase A anchoring protein 3) (PRKA3) (A-kinase anchor protein<BR> 110 kDa) (AKAP 110) (Sperm oocyte binding protein) (Fibrousheathin I) (Fibrous sheath protein of<BR> 6 AKA3_HUMAN 95 kDa) (FSP95). 490 SDISFEYPEDIGNLS